Hepcidin-Induced Iron Deficiency Is Related to Transient Anemia and Hypoferremia in Kawasaki Disease Patients

PubMed Central

Huang, Ying-Hsien; Kuo, Ho-Chang; Huang, Fu-Chen; Yu, Hong-Ren; Hsieh, Kai-Sheng; Yang, Ya-Ling; Sheen, Jiunn-Ming; Li, Sung-Chou; Kuo, Hsing-Chun

2016-01-01

Kawasaki disease (KD) is a type of systemic vasculitis that primarily affects children under the age of five years old. For sufferers of KD, intravenous immunoglobulin (IVIG) has been found to successfully diminish the occurrence of coronary artery lesions. Anemia is commonly found in KD patients, and we have shown that in appropriately elevated hepcidin levels are related to decreased hemoglobin levels in these patients. In this study, we investigated the time period of anemia and iron metabolism during different stages of KD. A total of 100 patients with KD and 20 control subjects were enrolled in this study for red blood cell and hemoglobin analysis. Furthermore, plasma, urine hepcidin, and plasma IL-6 levels were evaluated using enzyme-linked immunosorbent assay in 20 KD patients and controls. Changes in hemoglobin, plasma iron levels, and total iron binding capacity (TIBC) were also measured in patients with KD. Hemoglobin, iron levels, and TIBC were lower (p < 0.001, p = 0.009, and p < 0.001, respectively) while plasma IL-6 and hepcidin levels (both p < 0.001) were higher in patients with KD than in the controls prior to IVIG administration. Moreover, plasma hepcidin levels were positively and significantly correlated with urine hepcidin levels (p < 0.001) prior to IVIG administration. After IVIG treatment, plasma hepcidin and hemoglobin levels significantly decreased (both p < 0.001). Of particular note was a subsequent gradual increase in hemoglobin levels during the three weeks after IVIG treatment; nevertheless, the hemoglobin levels stayed lower in KD patients than in the controls (p = 0.045). These findings provide a longitudinal study of hemoglobin changes and among the first evidence that hepcidin induces transient anemia and hypoferremia during KD’s acute inflammatory phase. PMID:27187366

Effects of calorie restriction plus fish oil supplementation on abnormal metabolic characteristics and the iron status of middle-aged obese women.

PubMed

Utami, Fasty Arum; Lee, Hsiu-Chuan; Su, Chien-Tien; Guo, Yu-Ru; Tung, Yu-Tang; Huang, Shih-Yi

2018-02-21

The increasing prevalence of obesity and sedentary lifestyles has led to a higher incidence of metabolic syndrome (MetS) worldwide as well as in Taiwan. Middle-aged women are at a greater risk of MetS, type 2 diabetes, and cardiovascular disease than men because they have more subcutaneous fat and larger waist circumferences compared with men with equal visceral fat levels. In this study, we investigated the effects of calorie restriction (CR) and fish oil supplementation (CRF) on middle-aged Taiwanese women with MetS. An open-label, parallel-arm, controlled trial was conducted for 12 weeks. A total of 75 eligible participants were randomly assigned to the CR or CRF group. Both the dietary intervention groups were further divided into two age groups: ≤45 and >45 years. Changes in MetS severity, inflammatory status, iron status, and red blood cell fatty acid profile were evaluated. A total of 71 participants completed the trial. Both dietary interventions significantly ameliorated MetS and improved the participants' inflammatory status. CR significantly increased the total iron-binding capacity (TIBC) whereas CRF increased hepcidin levels in women aged >45 years. Furthermore, CRF significantly increased the n-6/n-3 and arachidonic acid/docosahexaenoic acid ratios. Both interventions improved the anthropometric and MetS characteristics, including body weight, blood glucose and triglyceride levels, and the score of the homeostasis model assessment of insulin resistance and quantitative insulin sensitivity check index. In conclusion, the 12-week dietary interventions improved the abnormal metabolic status of middle-aged obese women. CRF was demonstrated to be more effective in ameliorating postprandial glucose level and TIBC in women aged >45 years than in those aged ≤45 years.

The influence of combined magnesium and vanadate administration on the level of some elements in selected rat organs: V-Mg interactions and the role of iron-essential protein (DMT-1) in the mechanism underlying altered tissues iron level.

PubMed

Scibior, Agnieszka; Adamczyk, Agnieszka; Gołębiowska, Dorota; Niedźwiecka, Irmina; Fornal, Emilia

2014-04-01

The effect of 12 week co-administration of sodium metavanadate (SMV) and magnesium sulfate (MS) on the levels of some elements in selected rats' organs and an attempt to elucidate a role of divalent metal transporter 1 (DMT-1) in the mechanism(s) of the SMV-induced disorders in some tissue Fe homeostasis were studied. SMV taken up separately or in combination with MS may pose a risk of the rise and shortage of the total hepatic and splenic Fe and Cu contents, respectively, cerebral Fe deficiency, splenic Ca deposition, and the hepatic, renal, and cerebral DMT-1 down-regulation. When administered alone, SMV may also cause the decrease in the total renal Fe and Cu contents. A visible protective effect of Mg against the renal and cerebral V accumulation and the decrease in the renal Fe and Cu contents during the SMV-MS co-administration together with our previous findings suggest a beneficial role of Mg at SMV exposure. Further, the SMV-induced fall in total iron binding capacity (TIBC), reported previously, and its correlations with the hepatic, splenic, and cerebral Fe levels allow us to suggest that diminished TIBC could be partly involved in the mechanism(s) responsible for the dramatic redistribution of Fe in those tissues. Finally, DMT-1, which potentially could participate in the hepatic non-transferrin Fe-bound uptake, does not play a significant role in this process indicating the need for studying other Fe transporters to more precisely elucidate molecular mechanism(s) underlying the hepatic Fe loading in our experimental conditions.

Effects of cranberry (Vaccinum macrocarpon) supplementation on iron status and inflammatory markers in rowers.

PubMed

Skarpańska-Stejnborn, Anna; Basta, Piotr; Trzeciak, Jerzy; Michalska, Alicja; Kafkas, M Emin; Woitas-Ślubowska, Donata

2017-01-01

The aim of this study was to analyze the effect of supplementation with cranberry ( Vaccinum macrocarpon ) on the levels of pro-inflammatory cytokines, hepcidin and selected markers of iron metabolism in rowers subjected to exhaustive exercise. This double-blind study included 16 members of the Polish Rowing Team. The subjects were randomly assigned to the supplemented group ( n  = 9), receiving 1200 mg of cranberry extract for 6 weeks, or to the placebo group ( n  = 7). The participants performed a 2000-m test on a rowing ergometer at the beginning and at the end of the preparatory camp. Blood samples were obtained from the antecubital vein prior to each exercise test, one minute after completing the test, and after a 24-h recovery period. The levels of hepcidin, interleukin 6 (IL-6), tumor necrosis factor alpha (TNF-alpha), ferritin, iron, soluble transferrin receptor (sTfR) and myoglobin were determined, along with total iron-binding capacity (TIBC), unbound iron-binding capacity (UIBC) and total antioxidant capacity (TAC). Both prior and after the supplementation, a significant post-exercise increase in the concentration of IL-6 was observed in both groups. At the end of the study period, cranberry-supplemented athletes presented with significantly higher resting, post-exercise and post-recovery levels of TAC than the controls. However, a significant exercise-induced increase in the concentrations of TNF-alpha, myoglobin and hepcidin was observed solely in the control group. Supplementation with cranberry extract contributed to a significant strengthening of antioxidant potential in individuals exposed to strenuous physical exercise. However, supplementation did not exert direct effects on other analyzed parameters: inflammatory markers and indices of iron metabolism (TNF-alpha, hepcidin and myoglobin).

Enhanced plasma protein carbonylation in patients with myelodysplastic syndromes.

PubMed

Hlaváčková, Alžběta; Štikarová, Jana; Pimková, Kristýna; Chrastinová, Leona; Májek, Pavel; Kotlín, Roman; Čermák, Jaroslav; Suttnar, Jiří; Dyr, Jan Evangelista

2017-07-01

Myelodysplastic syndromes (MDS) represent a heterogeneous group of pre-leukemic disorders, characterized by ineffective hematopoiesis and the abnormal blood cell development of one or more lineages. Oxidative stress, as an important factor in the carcinogenesis of onco-hematological diseases, is also one of the known factors involved in the pathogenesis of MDS. An increase of reactive oxygen species (ROS) may lead to the oxidation of DNA, lipids, and proteins, thereby causing cell damage. Protein carbonylation caused by ROS is defined as an irreversible post-translational oxidative modification of amino acid side chains, and could play an important role in signaling processes. The detection of protein carbonyl groups is a specific useful marker of oxidative stress. In this study, we examined 32 patients divided into three different subtypes of MDS according to the World Health Organization (WHO) classification criteria as refractory anemia with ringed sideroblasts (RARS), refractory cytopenia with multilineage dysplasia (RCMD), refractory anemia with excess blasts-1,2 (RAEB-1,2). We found significant differences in protein carbonylation between the group of all MDS patients and healthy controls (P=0.0078). Furthermore, carbonylated protein levels were significantly elevated in RARS patients compared to healthy donors (P=0.0013) and to RCMD patients (P=0.0277). We also found a significant difference in the total iron binding capacity (TIBC) between individual subgroups of MDS patients (P=0.0263). Moreover, TIBC was decreased in RARS patients compared to RCMD patients (P=0.0203). TIBC moderately negatively correlated with carbonyl levels (r=-0.5978, P=0.0054) in the MDS patients as a whole. Additionally we observed changes in the carbonylated proteins of RARS patients in comparison with healthy controls and their negative controls. Using tandem mass spectrometry (LC-MS/MS) we identified 27 uniquely carbonylated proteins of RARS patients, which were generated by ROS and could influence the pathophysiology of low-risk MDS. These data indicate that increased protein carbonylation is related with RARS as low-risk MDS subgroup. We suggest that this type of post-translational modification in MDS disease is not "only" a consequence of oxidative stress, but also plays an active role in the pathophysiology and iron metabolism within the RARS subgroup of MDS. Measurement of plasma carbonyl levels and the isolation of carbonylated plasma proteins, followed by their identification, could serve as a potential diagnostic and prognostic tool in MDS. Copyright © 2017 Elsevier Inc. All rights reserved.

Relationship between high serum ferritin level and glaucoma in a South Korean population: the Kangbuk Samsung health study.

PubMed

Gye, Hyo Jung; Kim, Joon Mo; Yoo, Chungkwon; Shim, Seong Hee; Won, Yu Sam; Sung, Ki Chul; Lee, Mi Yeon; Park, Ki Ho

2016-12-01

To investigate the association between serum ferritin levels and glaucoma in a South Korean population. This retrospective cross-sectional study included 164 029 subjects who underwent screening at Kangbuk Samsung Hospital Health Screening Center between August 2012 and July 2013. All subjects underwent a physical examination, answered sociodemographic and behavioural questions, and provided samples for laboratory analyses. A digital fundus photograph of both eyes was taken, and all photographs were reviewed by ophthalmologists. The ophthalmologists determined if an eye had glaucoma based on criteria set forth by the International Society of Geographical and Epidemiological Ophthalmology and the appearance of the retinal nerve fibre layer and optic disc. The mean serum ferritin level was 56.98 ng/mL in women and 223.82 ng/mL in men. After adjusting for age, serum iron, total iron-binding capacity (TIBC), transferrin saturation, white blood cell (WBC) count, high-sensitivity C-reactive protein (HsCRP) and total vitamin D level, males in the highest quartile for serum ferritin level had a higher OR for glaucoma than males in the lowest quartile (OR=1.176, 95% CI 1.030 to 1.342, p=0.016); we did not observe this relationship among women. Other markers of iron metabolism, such as iron level, transferrin saturation and TIBC, and inflammation measures, including WBC, HsCRP and total vitamin D, were not associated with glaucoma. High serum ferritin level was associated with a high risk of glaucoma in men, but not in women. Because serum ferritin is related to oxidative stress and inflammation, it might play a role in glaucoma development. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

[Iron nutrition in Mapuche infants fed with human milk (2d phase)].

PubMed

Franco, E; Hertrampf, E; Rodríguez, E; Illanes, J C; Palacios, L; Llaguno, S; Lettelier, A

1990-01-01

Blood hemoglobin, serum iron, total iron binding capacity (TIBC) and serum ferritin were measured in 140 healthy rural mapuche (southern Chile's indigenous ethnic group) infants aged 8 to 15 months: 90 had been exclusively breast fed for the first 5 or 6 months of life, then solid foods were introduced but cow's milk was never given to them. The remaining 50, which were all weaned at nearly 4 months of age and then given cow's milk and solid foods at the corresponding age, were designated as controls. Anemia was detected in 4.5% of breast fed infants and in 38% of controls. Evidence of iron deficient erythropoiesis was found in 5% and 81% of cases and controls, respectively. Human milk apparently protects this ethnic group from iron deficiency anemia and this protection seems to be better in mapuche infants than in other groups of chilean infants, because these late have shown 30% incidence of anemia around the first year of life in other studies. More studies on differences in iron nutritional state among mapuche and non mapuche are needed and are under way.

Iron Plays a Certain Role in Patterned Hair Loss

PubMed Central

Park, Song Youn; Na, Se Young; Kim, Jun Hwan; Cho, Soyun

2013-01-01

Role of iron in hair loss is not clear yet. The purpose of this study was to evaluate the relationship between iron and hair loss. Retrospective chart review was conducted on patients with female pattern hair loss (FPHL) and male pattern hair loss (MPHL). All patients underwent screening including serum ferritin, iron, and total iron binding capacity (TIBC), CBC, ESR and thyroid function test. For normal healthy controls, age-sex matched subjects who had visited the hospital for a check-up with no serious disease were selected. A total 210 patients with FPHL (n = 113) and MPHL (n = 97) with 210 healthy controls were analyzed. Serum ferritin concentration (FC) was lower in patients with FPHL (49.27 ± 55.8 µg/L), compared with normal healthy women (77.89 ± 48.32 µg/L) (P < 0.001). Premenopausal FPHL patients turned out to show much lower serum ferritin than age/sex-matched controls (P < 0.001). Among MPHL patients, 22.7% of them showed serum FC lower than 70 µg/L, while no one had serum FC lower 70 µg/L in healthy age matched males. These results suggest that iron may play a certain role especially in premenopausal FPHL. The initial screening of iron status could be of help for hair loss patients. PMID:23772161

Iron plays a certain role in patterned hair loss.

PubMed

Park, Song Youn; Na, Se Young; Kim, Jun Hwan; Cho, Soyun; Lee, Jong Hee

2013-06-01

Role of iron in hair loss is not clear yet. The purpose of this study was to evaluate the relationship between iron and hair loss. Retrospective chart review was conducted on patients with female pattern hair loss (FPHL) and male pattern hair loss (MPHL). All patients underwent screening including serum ferritin, iron, and total iron binding capacity (TIBC), CBC, ESR and thyroid function test. For normal healthy controls, age-sex matched subjects who had visited the hospital for a check-up with no serious disease were selected. A total 210 patients with FPHL (n = 113) and MPHL (n = 97) with 210 healthy controls were analyzed. Serum ferritin concentration (FC) was lower in patients with FPHL (49.27 ± 55.8 µg/L), compared with normal healthy women (77.89 ± 48.32 µg/L) (P < 0.001). Premenopausal FPHL patients turned out to show much lower serum ferritin than age/sex-matched controls (P < 0.001). Among MPHL patients, 22.7% of them showed serum FC lower than 70 µg/L, while no one had serum FC lower 70 µg/L in healthy age matched males. These results suggest that iron may play a certain role especially in premenopausal FPHL. The initial screening of iron status could be of help for hair loss patients.

Lymphocyte ceruloplasmin and Behçet's disease.

PubMed

Oliveira, Rita; Banha, João; Martins, Fátima; Paixão, Eleonora; Pereira, Dina; Barcelos, Filipe; Teixeira, Ana; Patto, José Vaz; Costa, Luciana

2006-01-01

Behçet's disease (BD) is a rare chronic inflammatory disorder of unknown aetiology. However, it has been postulated that a dysregulation of the prooxidant/antioxidant balance may be important to its pathogenesis. Ceruloplasmin (CP) is an acute phase protein expressed at the surface of peripheral blood lymphocytes (PBL) with antioxidant properties and with a relevant role in iron (Fe) metabolism. To study CP expression at the surface of PBL (PBLCP) in patients with BD. We measured serum CP and PBLCP obtained from BD patients (n=10) and respective controls (n=10) using nephelometry and flow cytometry techniques, respectively. Additionally, haematological parameters, biochemical Fe metabolism markers [serum Fe, serum ferritin, serum transferrin, total Fe binding capacity (TIBC), transferrin saturation] and non-specific markers of inflammation [serum C reactive protein (CRP), beta2-microglobulin] were measured in all individuals. Despite the absence of significant differences between the two study groups when comparing serum CP, a significant difference in PBLCP was found in BD patients mainly due to a significant decrease of CP expression at the surface of CD3-CD56+ lymphocytes. Also, a significant decrease of PBLCP was observed in patients treated with azathioprine compared to patients that were not being treated with this drug. According to this study, we suggest that the significant decrease of PBLCP observed in BD patients might be due to azathioprine treatment and not directly related to the pathophysiology of BD.

Threonine modulates immune response, antioxidant status and gene expressions of antioxidant enzymes and antioxidant-immune-cytokine-related signaling molecules in juvenile blunt snout bream (Megalobrama amblycephala).

PubMed

Habte-Tsion, Habte-Michael; Ren, Mingchun; Liu, Bo; Ge, Xianping; Xie, Jun; Chen, Ruli

2016-04-01

A 9-week feeding trial was conducted to investigate the effects of graded dietary threonine (Thr) levels (0.58-2.58%) on the hematological parameters, immune response, antioxidant status and hepatopancreatic gene expression of antioxidant enzymes and antioxidant-immune-cytokine-related signaling molecules in juvenile blunt snout bream. For this purpose, 3 tanks were randomly arranged and assigned to each experimental diet. Fish were fed with their respective diet to apparent satiation 4 times daily. The results indicated that white blood cell, red blood cell and haemoglobin significantly responded to graded dietary Thr levels, while hematocrit didn't. Complement components (C3 and C4), total iron-binding capacity (TIBC), immunoglobulin M (IgM), superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT) increased with increasing dietary Thr levels up to 1.58-2.08% and thereafter tended to decrease. Dietary Thr regulated the gene expressions of Cu/Zn-SOD, Mn-SOD and CAT, GPx1, glutathione S-transferase mu (GST), nuclear factor erythroid 2-related factor 2 (Nrf2), heat shock protein-70 (Hsp70), tumor necrosis factor-alpha (TNF-α), apolipoprotein A-I (ApoA1), glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and fructose-bisphosphate aldolase B (ALDOB); while the gene expression of peroxiredoxin II (PrxII) was not significantly modified by graded Thr levels. These genes are involved in different functions including antioxidant, immune, and defense responses, energy metabolism and protein synthesis. Therefore, this study could provide a new molecular tool for studies in fish immunonutrition and shed light on the regulatory mechanisms that dietary Thr improved the antioxidant and immune capacities of fish. Copyright © 2015 Elsevier Ltd. All rights reserved.

Relative iron "overload" in offspring of patients with type 2 diabetes mellitus: a new component in the conundrum of insulin resistance syndrome?

PubMed

Psyrogiannis, Agathoklis; Kyriazopoulou, Venetsana; Symeonidis, Argiris; Leotsinidis, Michalis; Vagenakis, Apostolos G

2003-01-01

There are a few reports suggesting that subtle disturbances of iron metabolism are frequently found in patients with type 2 diabetes (DM2), but it is not known if these disturbances precede or accompany the diabetic state. We investigated the serum iron indices in 41 offspring of DM2 parents (group I) with normal glucose tolerance, and in 49 offspring whose parents had no history of DM2 and were matched for sex, age, body mass index (BMI), waist to hip ratio (WHR) and blood pressure (group II). Serum iron, ferritin, total iron binding capacity (TIBC), transferrin saturation, serum triglycerides, cholesterol, Apo-B, high density lipoprotein (HDL) and glucose and insulin values during an oral glucose tolerance test were measured. Insulin resistance was assessed using the homeostasis model assessment (HOMA - Insuline resistence index-IRI). In comparison to controls (group II), the offspring of DM2 subjects (group I) had higher fasting serum triglycerides (mean +/- SD 2.25+/-2.08 vs. 1.6+/-0.8 mmol/L, p<0,05), lower HDL cholesterol (0.96 +/- 0.2 vs. 1.1 +/- 0.2 mmol/L, p<0.001), higher total cholesterol (5.5 +/- 1.1 vs. 5.1 +/- 0.8 mmol/L, p < 0.05), higher apo-B-lipoprotein (133.2+/-34.3 vs. 125.5+/-30.5 mg/dl, p<0.05), higher LDL-C (3.7 +/- 0.8 vs. 3.2 +/- 0.6 mmol/L), higher gamma-GT (28+/-10 vs. 17+/-5.6 iu/L, p<0.01) higher insulin in the Area Under the Curve (204.7+/-140.8 v. 153.1 +/- 63.0 microU/ml, p<0.05) and higher HOMA-IRI (2.84+/-1.39 vs. 1.67+/-0.77, p<0.001), higher serum ferritin concentrations (98.3+/-57.7 vs. 62.0+/-41.1 ng/ml, p<0.01), higher serum iron concentration (20.2+/-6.0 micromol/L vs. 14.5+/-4.3, p<0.001) and higher transferrin saturation index (31.3+/-8.4 vs. 22.6+/-7.3, p<0.0001). By single linear analysis in the offspring of DM2 parents, there was a positive correlation of IRI with transferrin saturation (r=0.400, p<0.01), fibrinogen (r=0.377, p=0.025) and ferritin concentration (r=0.344, p=0.041), and a negative correlation with TIBC (r=-0.477, p < 0.0001), while stepwise multiple regression analysis, IRI showed a positive correlation with fibrinogen (b=0.64, t=3.746, p<0.001), triglycerides (b=0.37, t=2.619, p<0.01) and ferritin (b=0.20, t=1.827, p=0.05). No correlation of IRI, with any of the above parameters was seen in the offspring of normal parents. By logistic regression analysis the parameters characterizing the offspring of parents with DM2 were IRI (OR 14.9 CI 2.4-91.0) serum iron (OR 44.2 CI 6.9-281), TIBC (OR 6.1 CI 1.01-37.0 and gamma-GT (OR 29.6 CI 5.0-174). In conclusion, the data indicate that the iron load, is significantly increased in offspring of DM2 subjects with unaffected glucose tolerance. Furthermore, ferritin concentration is related to insulin resistance. Hence, the relative iron "overload" in offspring of type 2 diabetics is present along with insulin resistance and might worsen the hepatic insulin insensitivity already present in these patients.

Cross sectional, comparative study of serum erythropoietin, transferrin receptor, ferritin levels and other hematological indices in normal pregnancies and iron deficiency anemia during pregnancy.

PubMed

Sharma, Jai B; Bumma, Sirisha D; Saxena, Renu; Kumar, Sunesh; Roy, Kallol K; Singh, Neeta; Vanamail, P

2016-08-01

To test the correlation of the serum erythropoietin levels, serum transferrrin receptor levels and serum ferritin levels along with other hematological parameters in normal pregnant and anemic pregnant patients. In a prospective study, 120 pregnant women were recruited between 18 and 36 weeks of gestation; 53 normal pregnant patients, 67 anemic pregnant patients, in which, 17 had mild, 30 had moderate anemia, 20 had severe anemia. A blood sample was taken. The various hematological parameters, hemoglobin (Hb), mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC), total iron binding capacity (TIBC), serum ferritin, percentage saturation of iron, serum erythropoietin (SEPO) levels, serum transferrin receptors (STfRS) were performed. For statistics, Student's 't' test, Pearson's Chi test, Mann Whitney test and Bartlett test were used as per data. MCV was significantly reduced in anemic pregnancies as compared to non-anemic pregnancies (80.2±9.6 vs 94.12±9.8fl, p=0.001), MCHC was also reduced in them (30.2±3.38% vs 34.2±2.33%, p=0.176), TIBC was significantly increased in anemic pregnancies (343.31±28.54% vs 322.88±23.84%, p=0.001), serum ferritin was significantly reduced (24.9±10.48μg/L vs 31.03±9.98μg/L, p=0.001), percentage saturation of iron was also reduced (53.85±13.21% vs 62.04±15.79%, p=0.0024), serum erythropoietin levels were significantly higher in anemic women (26.24±26.61mU/ml vs 18.12±19.08mU/ml, p=0.064). The levels were significantly higher in severe anemia (46.5±46.8mU/ml than in moderate anemia 27.4±28.1mU/ml and mild anemia 22.8±22.8mU/ml. Serum transferrin receptors were significantly higher in anemic pregnancies than in non-anemic pregnancies (1.40±0.0802μg/ml vs 1.08±0.641μg/ml, p=0.019) with rise being higher in severe anemia (2.28±0.986μg/ml) than in moderate (1.4±0.816μg/ml) and mild anemia (1.16±0.702μg/ml). Various hematological parameters especially sTfR, serum erythropoietin, serum ferritin and sTfR/log ferritin levels correlate with the severity of anemia. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

REVIEWS OF TOPICAL PROBLEMS: Order-disorder transformations and phase equilibria in strongly nonstoichiometric compounds

NASA Astrophysics Data System (ADS)

Gusev, Aleksandr I.

2000-01-01

Data on order-disorder phase transformations in strongly nonstoichiometric carbides and nitrides MXy (X=C, N) of Group IV and V transition metals at temperatures below 1300-1400 K are reviewed. The order-parameter functional method as applied to atomic and vacancy ordering in strongly nonstoichiometric MXy compounds and to phase equilibrium calculations for M-X systems is discussed. Phase diagram calculations for the Ti-C, Zr-C, Hf-C, V-C, Nb-C, Ta-C, Ti-N, and Ti-B-C systems (with the inclusion of the ordering of nonstoichiometric carbides and nitrides) and those for pseudobinary carbide M(1)C-M(2)C systems are presented. Heat capacity, electrical resistivity and magnetic susceptibility changes at reversible order-disorder phase transformations in nonstoichiometric carbides are considered.

Interference of medical contrast media on laboratory testing.

PubMed

Lippi, Giuseppe; Daves, Massimo; Mattiuzzi, Camilla

2014-01-01

The use of contrast media such as organic iodine molecules and gadolinium contrast agents is commonplace in diagnostic imaging. Although there is widespread perception that side effects and drug interactions may be the leading problems caused by these compounds, various degrees of interference with some laboratory tests have been clearly demonstrated. Overall, the described interference for iodinate contrast media include inappropriate gel barrier formation in blood tubes, the appearance of abnormal peaks in capillary zone electrophoresis of serum proteins, and a positive bias in assessment of cardiac troponin I with one immunoassay. The interference for gadolinium contrast agents include negative bias in calcium assessment with ortho-cresolphthalein colorimetric assays and occasional positive bias using some Arsenazo reagents, negative bias in measurement of angiotensin converting enzyme (ACE) and zinc (colorimetric assay), as well as positive bias in creatinine (Jaffe reaction), total iron binding capacity (TIBC, ferrozine method), magnesium (calmagite reagent) and selenium (mass spectrometry) measurement. Interference has also been reported in assessment of serum indices, pulse oximetry and methaemoglobin in samples of patients receiving Patent Blue V. Under several circumstances the interference was absent from manufacturer-supplied information and limited to certain type of reagents and/or analytes, so that local verification may be advisable to establish whether or not the test in use may be biased. Since the elimination half-life of these compounds is typically lower than 2 h, blood collection after this period may be a safer alternative in patients who have received contrast media for diagnostic purposes.

Interference of medical contrast media on laboratory testing

PubMed Central

Lippi, Giuseppe; Daves, Massimo; Mattiuzzi, Camilla

2014-01-01

The use of contrast media such as organic iodine molecules and gadolinium contrast agents is commonplace in diagnostic imaging. Although there is widespread perception that side effects and drug interactions may be the leading problems caused by these compounds, various degrees of interference with some laboratory tests have been clearly demonstrated. Overall, the described interference for iodinate contrast media include inappropriate gel barrier formation in blood tubes, the appearance of abnormal peaks in capillary zone electrophoresis of serum proteins, and a positive bias in assessment of cardiac troponin I with one immunoassay. The interference for gadolinium contrast agents include negative bias in calcium assessment with ortho-cresolphthalein colorimetric assays and occasional positive bias using some Arsenazo reagents, negative bias in measurement of angiotensin converting enzyme (ACE) and zinc (colorimetric assay), as well as positive bias in creatinine (Jaffe reaction), total iron binding capacity (TIBC, ferrozine method), magnesium (calmagite reagent) and selenium (mass spectrometry) measurement. Interference has also been reported in assessment of serum indices, pulse oximetry and methaemoglobin in samples of patients receiving Patent Blue V. Under several circumstances the interference was absent from manufacturer-supplied information and limited to certain type of reagents and/or analytes, so that local verification may be advisable to establish whether or not the test in use may be biased. Since the elimination half-life of these compounds is typically lower than 2 h, blood collection after this period may be a safer alternative in patients who have received contrast media for diagnostic purposes. PMID:24627717

TIBC, UIBC and Transferrin

MedlinePlus

... 28 weeks to delivery) Primary Aldosteronism (Conn Syndrome) Prostate Cancer Protein in Urine (Proteinuria) Reactive Arthritis Rheumatoid Arthritis ... Blood Count (CBC) Hemoglobin Hematocrit Reticulocytes Soluble Transferrin Receptor Conditions Anemia Hemochromatosis Elsewhere On The Web American ...

Hereditary hemochromatosis.

PubMed

Walker, E M; Wolfe, M D; Norton, M L; Walker, S M; Jones, M M

1998-01-01

Genetic (hereditary) hemochromatosis is probably the most common autosomal recessive disorder found in white Americans, of whom about 5/1,000 (0.5 percent) are homozygous for the associated gene. The hemochromatosis gene is probably located close to the HLA-A locus on the short arm of chromosome 6. Homozygous individuals may develop severe and potentially lethal hemochromatosis, especially after age 39. Hereditary hemochromatosis involves an increased rate of iron absorption from the gut with subsequent progressive storage of iron in soft organs of the body. Excess iron storage eventually produces pituitary, pancreatic, cardiac, and liver dysfunction and death may result from cardiac arrhythmias, congestive heart failure, and/or hepatic failure or cancer. Early diagnosis can prevent these excess iron-induced problems. Iron overload owing to HLA-linked hereditary hemochromatosis can be distinguished from other causes of hemochromatosis by liver biopsies and interpretations. Patients at risk for genetic hemochromatosis should be screened, identified, and treated as early as age 20 to prevent or minimize the deadly complications of hemochromatosis. Population screening should include measurements of serum iron concentration, total iron binding capacity (TIBC), percent saturation of transferrin, and serum ferritin concentrations. Family members of hereditary hemochromatosis patients are at increased risk and should be tested. Screening, identification and early treatment (phlebotomies, sometimes in combination with the use of Desferal or other iron-chelating agents) may help prevent or reduce iron-related organ damage and premature deaths. Early diagnosis and treatment will reduce the population of aging individuals with severe, complicated hemochromatosis and dramatically reduce medical costs (billions of U.S. dollars per annum) associated with the management of this disease.

Effects of feeding and fasting on wolf blood and urine characteristics

USGS Publications Warehouse

DelGiudice, G.D.; Seal, U.S.; Mech, L.D.

1987-01-01

Feeding and fasting trials were conducted with 2 groups (A and B) of 4 gray wolves (Canis lupus) each during January 1980. The groups were fed for 9 days and fasted for 10 days in a cross-over design. Blood and urine samples and weight data were collected every 2-3 days during each trial. Hemoglobin (Hb) concentrations, red blood cell (RBC) counts, and hematocrits (HCT) were elevated in both groups during fasting. White blood cell (WBC) counts, serum urea nitrogen (SUN), triiodothyronine (T3), and insulin concentrations decreased during fasting in Groups A and B. Mean corpuscular hemoglobin concentration (MCHC), serum cholesterol, triglyceride, and iron (Fe) concentrations were diminished in fasted Group A wolves compared to fed Group B. Creatine phosphokinase (CPK) concentrations were elevated in fed Group A wolves. Serum creatinine (C) concentrations were reduced in both groups during feeding. Urinary urea: creatinine (U:C), potassium:creatine (K:C), and sodium:creatinine (Na:C, pooled Group A and B data) ratios decreased in fasted wolves. Differences were not found between fed and fasted wolves for mean corpuscular volume (MCV), serum cortisol, glucose, calcium (Ca), bilirubin, serum glutamate-oxaloacetate transaminase (SGOT), serum glutamate-pyruvate transaminase (SGOT), serum glutamate-pyruvate transaminase (SGPT), alkaline phosphatase, and luteinizing hormone (LH) concentrations, total iron binding capacity (TIBC), and urinary calcium: creatine (Ca:C) ratios. Analysis of multiple blood or urine samples collected from free-ranging wolves would be useful in enabling researches and managers to identify the nutritional status and general health of wolves over time.

Underestimation of the coexistence of iron deficiencies and thalassemia minors: a single institution experience in Taiwan.

PubMed

Lin, Chung-King; Chen, Ling-Ping; Chang, Hsiu-Lin; Sung, Yung-Chuan

2014-08-01

Some physicians neglect the possible coexistence of an iron deficiency with a thalassemia minor and do not treat the iron deficiency accordingly. This motivated us to conduct this study. We retrospectively reviewed the records of 3892 patients who visited our clinics and had hemoglobin (Hb) electrophoreses performed in our hematologic laboratory from August 1, 2007 to December 31, 2012. The thalassemia minors were identified by characteristic complete blood count (CBC) parameters obtained from an autoanalyzer and Hb electrophoresis, and some cases were confirmed with molecular tests. Then, we checked iron studies [ferritin and/or serum iron with total iron-binding capacity (TIBC)] to determine the coexistence of an iron deficiency with a thalassemia minor and a response to iron, if such treatments were given. We found 792 cases with thalassemia minors, and excluded those without iron studies, with 661 cases as our sample. A total of 202/661 cases (31%) also had iron deficiencies. They had lower red blood cell (RBC) counts, Hb, and ferritin levels as compared to those thalassemia minor cases without coexistence of iron deficiencies. We concluded that the thalassemia minor patients did not have iron overload complications in our population. On the contrary, iron deficiencies commonly coexist in the clinical visits. We propose that if Hb < 11.5 g/dL in a case of thalassemia minor, one should screen for iron deficiency simultaneously. The sensitivity is 79.8% and the specificity is 82.6%. Therefore, physicians should be aware of this coexisting condition, and know how to recognize and treat it accordingly. Copyright © 2014. Published by Elsevier B.V.

Iron, zinc and iodide status in Mexican children under 12 years and women 12-49 years of age. A probabilistic national survey.

PubMed

Villalpando, Salvador; García-Guerra, Armando; Ramírez-Silva, Claudia Ivonne; Mejía-Rodríguez, Fabiola; Matute, Guadalupe; Shamah-Levy, Teresa; Rivera, Juan A

2003-01-01

To describe the epidemiology of iron, zinc and iodide deficiencies in a probabilistic sample of Mexican women and children and explore its association with some dietary and socio-demographic variables. We carried out in 1999 an epidemiological description of iron (percent transferrin saturation, PTS, < 16%), serum zinc (< 65 ug/dl) and iodide (< 50 ug/l urine) deficiencies in a probabilistic sample of 1,363 Mexican children under 12 years and of 731 women of child-bearing age. Serum iron, Total Iron Binding Capacity (TIBC) and zinc were measured by atomic absorption spectrometry, and urinary iodide by a colorimetric method. Logistic regression models explored determinants for such micromineral deficiencies. Iron deficiency was higher (67%) in infants < 2 years of age. Prevalence declined (34-39%) at school age. The prevalence for iron deficiency in women was 40%. Zinc deficiency was higher in infants < 2 years of age (34%) than in school-age children (19-24%). Prevalence in women was 30%, with no rural/urban difference. In women the likelihood of iron deficiency decreased as SEL improved (p = 0.04) and increased with the intake of cereals (p = 0.01). The likelihood of low serum zinc levels was greater in women and children of low socioeconomic level (SEL) (p < 0.02 and p = 0.001) iodide deficiency was negligible in both children and women. The data shows high prevalence of iron deficiency-specially in infants 12 to 24 months of age. It is suggested that in older children and women 12 to 49 years of age that iron bioavailability is low. The prevalence of zinc deficiency was also very high. The English version of this paper is available too at: http://www.insp.mx/salud/index.html.

Cognitive function, iron status, and hemoglobin concentration in obese dieting women.

PubMed

Kretsch, M J; Fong, A K; Green, M W; Johnson, H L

1998-07-01

To determine the relationships between cognitive function and iron status in dieting obese women. Longitudinal weight loss study (repeated measures within-subject design) with 3 weeks of baseline, 15 weeks of 50% caloric restriction, and 3 weeks of weight stabilization. Dietary iron was fed at twice the US Recommended Dietary Allowance with half of the iron from food sources and half from an oral supplement. This was a free-living study with the exception that subjects came to the research center for one meal per day and were provided all other meals and snacks to take home. Healthy, premenopausal, obese women (mean BMI=31.5) were recruited through local newspaper, poster and radio advertising. Twenty-four women volunteers were recruited and 14 completed the study. Cognitive function, iron and hematological status, height, body weights and body composition were measured at baseline; at weeks 5, 10, and 15 of the energy restriction period; and at the end of weight stabilization. Computerized cognitive tests included: Bakan vigilance task, two finger tapping, simple reaction time, immediate word recall, and a focused attention task. Iron status and hematological measures included: serum iron, total iron binding capacity (TIBC), transferrin saturation, serum ferritin, hemoglobin (Hb), hematocrit, red cell count, MCV, MCH, MCHC, and RDW. A significant reduction in Hb, hematocrit, and red blood cell count occurred across the study. Hb at the end of the study was positively correlated (r=0.72, P < 0.01) with mean performance on a measure of sustained attention. Transferrin saturation also correlated positively to sustained attention task performance for those subjects whose Hb declined across the study (r=0.86, P < 0.01). These findings suggest that dieting diminishes iron status in obese women, even when sufficient dietary iron is available, and that the inability to sustain attention may be an early sign of developing iron deficiency in dieting women.

Crystal structure and chemotherapeutic efficacy of the novel compound, gallium tetrachloride betaine, against breast cancer using nanotechnology.

PubMed

Salem, Ahmed; Noaman, Eman; Kandil, Eman; Badawi, Abdelfattah; Mostafa, Nihal

2016-08-01

The objective of this study was to investigate the antitumor efficacy of a novel synthesized compound, betaine gallium-tetrachloride (BTG), alone or combined with ZnO-nanoparticles (BTG + ZnO-NPs) on the incidence of 7, 12-dimethylbenz-anthrathene-induced mammary tumor in female rats. Crystal and molecular structure of the prepared BTG were identified using X-ray crystallography. In vitro study revealed BTG more cytotoxic than BTG + ZnO-NPs on human breast cancer (MCF-7) cell line. In vivo study demonstrated that the blood antioxidant status of tumor-bearing rats (DMBA group) was significantly lower than normal noticeable by a significant decrease in GSH content, GPx, SOD, and CAT activities associated with a significantly high MDA content. Both treatments have significantly elevated SOD and CAT activities with a concomitant decrease of MDA level compared to DMBA group. However, BTG + ZnO-NPs accentuated the decrease of GSH regarding DMBA group. The results showed also that both treatments significantly activate caspase-3 enzyme and apoptosis in mammary glands. Their administration to tumor-bearing rats was found to significantly reduce plasma iron and iron-binding capacity (TIBC) compared to DMBA group. Regarding liver function, both treatments significantly reduced the increase of ALT and AST activities compared to DMBA group. However, BTG + ZnO-NPs decreased albumin below normal level. Histopathological studies showed that normalization of tissue structures was higher in BTG than BTG + ZnO-NPs treatment. According to the results obtained, it is observed that the antitumor effect of BTG alone was as strong as BTG + ZnO-NPs and even more efficient in some aspects accordingly, a combination is not needed. Thus, the novel synthetic gallium derivatives may potentially present a new hope for the development of breast cancer therapeutics, which should attract further scientific and pharmaceutical interest.

Effect of short-term intravenous ascorbic acid on reducing ferritin in hemodialysis patients.

PubMed

Jalalzadeh, M; Shekari, E; Mirzamohammadi, F; Ghadiani, M H

2012-05-01

Resistance to recombinant erythropoietin (rEPO) in hemodialysis patients may be due to inadequate iron recruitment and defect in iron use. In this cross over randomized clinical trial, 30 hemodialysis patients with serum ferritin levels of ≥500 ng/ml, hemoglobin ≤11.0 g/dl, and transferrin saturation (TSAT) of 20% or less were administrated intravenous iron (50-100 mg/wk) and rEPO (120-360 U/kg/wk) for 6 months. Patients were excluded if there was a clear explanation for rEPO hyporesponsiveness. Patients were divided into two groups. Group1 received standard care and 500 mg of intravenous ascorbic acid (IVAA) with each dialysis session in the first week of each month for a total of 3 months. Group 2 received standard care only. After 2 month washout period, groups were crossed over. Each month hemoglobin (Hb) was assessed. Iron, TIBC (transferrin iron binding capacity), TSAT, iPTH (intact parathyroid hormone), liver enzymes, albumin and cholesterol levels were measured every 3 months. After 3 months of intervention, Hb significantly increased from 10.11 to 12.19 g/dl (P <0 0.001; 95% confidence interval [CI] 2.7-1.4) and TSAT increased from 18.9 to 28.1% (P = 0.008; 95% CI 0.09-3), while ferritin and serum iron declined significantly from 1391 to 938 ng/ml (P = 0.001; 95% CI 216-689), 97.2 to 64.6 (P = 0.001; 95% CI 14.8-50.4) in the study group. Change of Hb over time in IVAA group was significant (P < 0.0005). There were significant differences between two groups in change of Hb level over time (P < 0.0005) and treatment effect (P = 0.002). Baseline laboratory tests were similar in the two groups and there was no carry over effect at phase 2. We showed that low amount of IVAA could reduce ferritin level and enhance Hb and TSAT, suggesting improved iron utilization.

Low capacity of erythrocytes to bind with immune complexes via C3b receptor in patients with systemic lupus erythematosus: correlation with pathological proteinuria

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nojima, Y.; Terai, C.; Minota, S.

1985-01-01

Erythrocytes from 51 patients with systemic lupus erythematosus and 75 controls were tested for the capacity to bind aggregated human gamma-globulin labeled with radioiodine in the presence of complement. Both in patients and controls, a trimodal distribution of binding capacity was observed. Low (less than 9% of the added radioactivity), intermediate (9-17%), and high binding (more than 17%) were observed in 13, 58, and 29% in controls and in 49, 43 and 8% in lupus patients. The low binding capacity of erythrocytes persisted even after patients entered remission following steroid therapy. A genetic control of binding capacity was supported bymore » familial surveys. Prevalence of pathological proteinuria was significantly higher in patients with low binding capacity than those with intermediate or high binding capacity (16/25 vs 7/26, P less than 0.01). These results indicate that an impaired physiological disposal of immune complexes via the erythrocyte C3b receptor in lupus patients may contribute to the development of renal involvement.« less

The productive cellulase binding capacity of cellulosic substrates.

PubMed

Karuna, Nardrapee; Jeoh, Tina

2017-03-01

Cellulosic biomass is the most promising feedstock for renewable biofuel production; however, the mechanisms of the heterogeneous cellulose saccharification reaction are still unsolved. As cellulases need to bind isolated molecules of cellulose at the surface of insoluble cellulose fibrils or larger aggregated cellulose structures in order to hydrolyze glycosidic bonds, the "accessibility of cellulose to cellulases" is considered to be a reaction limiting property of cellulose. We have defined the accessibility of cellulose to cellulases as the productive binding capacity of cellulose, that is, the concentration of productive binding sites on cellulose that are accessible for binding and hydrolysis by cellulases. Productive cellulase binding to cellulose results in hydrolysis and can be quantified by measuring hydrolysis rates. In this study, we measured the productive Trichoderma reesei Cel7A (TrCel7A) binding capacity of five cellulosic substrates from different sources and processing histories. Swollen filter paper and bacterial cellulose had higher productive binding capacities of ∼6 µmol/g while filter paper, microcrystalline cellulose, and algal cellulose had lower productive binding capacities of ∼3 µmol/g. Swelling and regenerating filter paper using phosphoric acid increased the initial accessibility of the reducing ends to TrCel7A from 4 to 6 µmol/g. Moreover, this increase in initial productive binding capacity accounted in large part for the difference in the overall digestibility between filter paper and swollen filter paper. We further demonstrated that an understanding of how the productive binding capacity declines over the course of the hydrolysis reaction has the potential to predict overall saccharification time courses. Biotechnol. Bioeng. 2017;114: 533-542. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

In Vitro Binding Capacity of Bile Acids by Defatted Corn Protein Hydrolysate

PubMed Central

Kongo-Dia-Moukala, Jauricque Ursulla; Zhang, Hui; Irakoze, Pierre Claver

2011-01-01

Defatted corn protein was digested using five different proteases, Alcalase, Trypsin, Neutrase, Protamex and Flavourzyme, in order to produce bile acid binding peptides. Bile acid binding capacity was analyzed in vitro using peptides from different proteases of defatted corn hydrolysate. Some crystalline bile acids like sodium glycocholate, sodium cholate and sodium deoxycholate were individually tested using HPLC to see which enzymes can release more peptides with high bile acid binding capacity. Peptides from Flavourzyme defatted corn hydrolysate exhibited significantly (p < 0.05) stronger bile acid binding capacity than all others hydrolysates tested and all crystalline bile acids tested were highly bound by cholestyramine, a positive control well known as a cholesterol-reducing agent. The bile acid binding capacity of Flavourzyme hydrolysate was almost preserved after gastrointestinal proteases digestion. The molecular weight of Flavourzyme hydrolysate was determined and most of the peptides were found between 500–180 Da. The results showed that Flavourzyme hydrolysate may be used as a potential cholesterol-reducing agent. PMID:21541043

Biological variability of transferrin saturation and unsaturated iron binding capacity

PubMed Central

Adams, PC; Reboussin, DM; Press, RD; Barton, JC; Acton, RT; Moses, GC; Leiendecker-Foster, C; McLaren, GD; Dawkins, FW; Gordeuk, VR; Lovato, L; Eckfeldt, JH

2007-01-01

Background Transferrin saturation is widely considered the preferred screening test for hemochromatosis. Unsaturated iron binding capacity has similar performance at lower cost. However, the within-person biological variability of both these tests may limit their ability at commonly used cut points to detect HFE C282Y homozygous patients. Methods The Hemochromatosis and Iron Overload Screening (HEIRS) Study screened 101,168 primary care participants for iron overload using tansferrin saturation, unsaturated iron binding capacity, ferritin and HFE C282Y and H63D genotyping. Transferrin saturation and unsaturated iron binding capacity were performed at initial screening and again when selected participants and controls returned for a clinical examination several months later. A missed case was defined as a C282Y homozygote who had transferrin saturation below cut point (45 % women, 50 % men) or unsaturated iron binding capacity above cut point (150 μmol/L women, 125 μmol/L men) at either the initial screening or clinical examination, or both, regardless of serum ferritin. Results There were 209 C282Y previously undiagnosed homozygotes with transferrin saturation and unsaturated iron binding capacity testing done at initial screening and clinical examination. Sixty-eight C282Y homozygotes (33%) would have been missed at these transferrin saturation cut points (19 men, 49 women, median SF 170 μg/L, first and third quartiles 50 and 474 μg/L), and 58 homozygotes (28 %) would have been missed at the unsaturated iron binding capacity cut points (20 men, 38 women, median SF 168 μg/L, quartiles 38 and 454 μg/L). There was no advantage to using fasting samples. Conclusions The within-person biological variability of transferrin saturation and unsaturated iron binding capacity limit their usefulness as an initial screening test for expressing C282Y homozygotes. PMID:17976429

Haematological values in pregnant women in Port Harcourt, Nigeria II: Serum iron and transferrin, total and unsaturated iron binding capacity and some red cell and platelet indices.

PubMed

Amah-Tariah, F S; Ojeka, S O; Dapper, D V

2011-12-20

Previous studies on the normal values of serum iron, unsaturated iron binding capacity, total iron binding capacity, serum transferrin, percent transferrin saturation, red cell distribution width, and various platelet indices: Platelet count, mean platelet volume, platelet distribution width, plateletcrit and platelet larger cell ratio in pregnant subjects in Nigeria are relatively scanty. Present study aims to determine the values of these parameters in apparently healthy pregnant subjects residing in Port Harcourt south eastern Nigeria; and help establish normal reference ranges of these parameters for the population under reference. Cross sectional prospective study involving 220 female subjects attending for the first time, the ante-natal clinics of a tertiary health care facility in Port Harcourt. Subjects were divided into 73, 75 and 72 subjects in the first, second and third trimester of pregnancy respectively. Serum iron and unsaturated iron binding capacity, red cell distribution width, platelet count and platelet distribution width were determined by automated methods; total iron binding capacity, serum transferrin concentrations, percent transferrin saturation, mean platelet volume and plateletcrit were calculated using appropriate formulas. The values of serum iron, unsaturated iron binding capacity, total iron binding capacity and serum transferrin concentrations were found to show significant variations between the various trimesters of pregnancy. However, while serum iron showed significant decreases during pregnancy; unsaturated iron binding capacity, total iron binding capacity and serum transferrin concentrations were found to show significant increases during pregnancy amongst our subjects (p<0.05). By contrast the values of red cell distribution width, platelet count, mean platelet volume, platelet distribution width, plateletcrit and platelet larger cell ratio did not show any significant differences at the different trimesters of pregnancy in our subjects (p>0.05). The present study reports, for the first time, normative values for these parameters in apparently healthy pregnant subjects in Port Harcourt south eastern Nigeria. Apparently, increases in unsaturated and total iron binding capacity and serum transferrin values seen amongst our subjects with increasing gestation may perhaps be a mechanism to ensure a fetal adequate iron delivery on account of the decreasing serum iron concentration with gestation in our subjects. The study suggests that values of serum transferrin are perhaps a more useful screening tool for iron deficiency anemia during pregnancy amongst our subjects.

Tropical soils in Mato Grosso, Brazil, retain high phosphorus (P) binding capacity after 30 years of intensive fertilization and will remain a P sink for another 50-160 years.

NASA Astrophysics Data System (ADS)

Porder, S.; Roy, E.; Willig, E.; Martinelli, L. A.; Pegorini, L.; Richards, P.; Spera, S. A.; Vazquez, F. F.

2016-12-01

Intensification of tropical agriculture is one way to meet increasing global food demand, but tropical soils often require more phosphorus (P) fertilizer than those in the world's traditional breadbaskets. Recent studies from Europe suggest that P fertilizer additions will eventually saturate soil P binding capacity, and can build a soil P bank upon which future crop production can draw. We tested this hypothesis in Mato Grosso, Brazil, where highly mechanized agriculture produces 9% of the world's soy harvest on soils with high P binding capacity. In this region, P fertilizer inputs typically exceed harvests by 10kg P/ha, and our expectation was that total P and available P would increase, and P binding capacity would decrease, with time in cultivation. To test this hypothesis, we measured P availability, binding, and accumulation on 31 fields ranging from 0-31 years in intensive production. We also estimated the number of years in production that would be required to saturate the soils with P, since after that time P additions could be reduced to equal harvest P removal. As expected, our data show increasing P availability, and decreasing P binding capacity, over time. A multiple regression including only soil [SiO2] (a proxy for both mineralogy and texture) and years in production explained 87, 63 and 91% of the observed variation in total P, Bray-extractable P, and P sorption capacity, respectively. However, the effect of [SiO2], and thus texture and mineralogy, was 1.7, 1.2, and 4.9 times more important in predicting our dependent variables than was years in production. Despite fertilizer inputs in excess of harvest removals, the reduction in P binding capacity is slow, and we estimate it will take between 50-160 years for fertilizer inputs to saturate the P binding capacity of these soils. These results suggest that the P tax imposed by high P binding soils in the tropics will impose substantial material costs to tropical farmers in the coming decades, and may influence their capacity to intensify food production to meet growing food demands.

Evaluation of the mobile phone electromagnetic radiation on serum iron parameters in rats.

PubMed

Çetkin, Murat; Demirel, Can; Kızılkan, Neşe; Aksoy, Nur; Erbağcı, Hülya

2017-03-01

Electromagnetic fields (EMF) created by mobile phones during communication have harmful effects on different organs. It was aimed to investigate the effects of an EMF created by a mobile phone on serum iron level, ferritin, unsaturated iron binding capacity and total iron binding capacity within a rat experiment model. A total of 32 male Wistar albino rats were randomly divided into the control, sham, mobile phone speech (2h/day) and stand by (12 h/day) groups. The speech and stand by groups were subjected to the EMF for a total of 10 weeks. No statistically significant difference was observed between the serum iron and ferritin values of the rats in the speech and stand by groups than the control and sham groups (p>0.05). The unsaturated iron binding capacity and total iron capacity values of the rats in the speech and stand by groups were significantly lower in comparison to the control group (p<0.01). It was found that exposure to EMF created by mobile phones affected unsaturated iron binding capacity and total iron binding capacity negatively.

Intravenous iron-dextran: studies on unsaturated iron-binding capacity

PubMed Central

Cox, J. S. G.; Moss, G. F.; Bremner, I.; Reason, Janet

1968-01-01

A method is described for measuring the plasma unsaturated iron-binding capacity in the presence of very high concentrations of iron as iron-dextran. The procedure utilizes 59Fe to label the apotransferrin with subsequent separation of ionic iron from transferrin-bound iron on an ion exchange or Sephadex G.25 column. The unsaturated iron-binding capacity has been measured in rabbits and dogs after intravenous injection of iron-dextran and in human subjects after total dose infusion of iron-dextran. No evidence of saturation of the unsaturated iron-binding capacity was found even when the plasma iron values were greater than 40,000 μg Fe/100 ml. PMID:5697365

Phase composition and tribomechanical properties of Ti-B-C nanocomposite coatings prepared by magnetron sputtering

NASA Astrophysics Data System (ADS)

Sánchez-López, J. C.; Abad, M. D.; Justo, A.; Gago, R.; Endrino, J. L.; García-Luis, A.; Brizuela, M.

2012-09-01

Protective nanocomposite coatings based on hard ceramic phases (TiC, TiB2) combined with amorphous carbon (a-C) are of interest because of their adequate balance between mechanical and tribological performances. In this work, Ti-B-C nanocomposite coatings were prepared by co-sputtering of graphite and TiB2 targets. Varying the discharge power ratio applied to the graphite and TiB2 targets from 0 to 2, the a-C content in the coatings could be tuned from 0 to 60%, as observed by means of Raman and x-ray photoelectron spectroscopy (XPS). The microstructural characterization demonstrated a progressive decrease in crystallinity from an initial nanocrystalline (nc) TiB2-like structure to a distorted TiBxCy ternary compound with increasing C concentration. X-ray absorption near-edge structure measurements on the B K-edge helped to determine a hexagonal arrangement around the B atoms in the ternary TiBxCy phase. A fitting analysis of the C 1s XPS peak allowed us to evaluate the relative amount of a-C and TiBxCy components. A drastic change in hardness (from 52 to 13 GPa) and friction coefficient values (from 0.8 to 0.2) is noticed when moving from nc-TiB2 to TiBC/a-C nanocomposites. The fraction of a-C necessary to decrease the friction below 0.2 was found to be 45%. Raman observation of the wear tracks determined the presence of disordered sp2-bonded carbon phase associated with the diminution of the friction level.

Antioxidative capacity and binding affinity of the complex of green tea catechin and beta-lactoglobulin glycated by the Maillard reaction.

PubMed

Perusko, Marija; Al-Hanish, Ayah; Mihailovic, Jelena; Minic, Simeon; Trifunovic, Sara; Prodic, Ivana; Cirkovic Velickovic, Tanja

2017-10-01

Major green tea catechin, epigallocatechin-3-gallate (EGCG), binds non-covalently to numerous dietary proteins, including beta-lactoglobulin of cow's milk. The effects of glycation of proteins via Maillard reaction on the binding capacity for polyphenols and the antiradical properties of the formed complexes have not been studied previously. Binding constant of BLG glycated by milk sugar lactose to EGCG was measured by the method of fluorophore quenching. Binding of EGCG was confirmed by CD and FTIR. The antioxidative properties of the complexes were examined by measuring ABTS radical scavenging capacity, superoxide anion scavenging capacity and total reducing power assay. Glycation of BLG does not significantly influence the binding constant of EGCG for the protein. Conformational changes were observed for both native and glycated BLG upon complexation with EGCG. Masking effect of polyphenol complexation on the antioxidative potential of the protein was of the similar degree for both glycated BLG and native BLG. Copyright © 2017 Elsevier Ltd. All rights reserved.

Thermodynamics of Ligand Binding to a Heterogeneous RNA Population in the Malachite Green Aptamer

PubMed Central

Sokoloski, Joshua E.; Dombrowski, Sarah E.; Bevilacqua, Philip C.

2011-01-01

The malachite green aptamer binds two closely related ligands, malachite green (MG) and tetramethylrosamine (TMR), with near equal affinity. The MG ligand consists of three phenyl rings emanating from a central carbon, while TMR has two of the three rings connected by an ether linkage. The binding pockets for MG and TMR in the aptamer, known from high-resolution structure, differ only in the conformation of a few nucleotides. Herein, we applied isothermal titration calorimetry (ITC) to compare the thermodynamics for binding of MG and TMR to the aptamer. Binding heat capacities were obtained from ITC titrations over the temperature range of 15 to 60 °C. Two temperature regimes were found for MG binding: one from 15 to 45 °C where MG bound with a large negative heat capacity and an apparent stoichiometry (n) of ~0.4, and another from 50 to 60 °C where MG bound with positive heat capacity and n~1.1. The binding of TMR, on the other hand, revealed only one temperature regime for binding, with a more modest negative heat capacity and n~1.2. The large difference in heat capacity between the two ligands suggests that significantly more conformational rearrangement occurs upon the binding of MG than TMR, which is consistent with differences in solvent accessible surface area calculated for available ligand-bound structures. Lastly, we note that binding stoichiometry of MG was improved not only by raising the temperature, but also by lowering the concentration of Mg2+ or increasing the time between ITC injections. These studies suggest that binding of a dynamical ligand to a functional RNA requires the RNA itself to have significant dynamics. PMID:22192051

Protein Binding Capacity of Different Forages Tannin

NASA Astrophysics Data System (ADS)

Yusiati, L. M.; Kurniawati, A.; Hanim, C.; Anas, M. A.

2018-02-01

Eight forages of tannin sources(Leucaena leucocephala, Arachis hypogaea, Mimosa pudica, Morus alba L, Swietenia mahagoni, Manihot esculenta, Gliricidia sepium, and Bauhinia purpurea)were evaluated their tannin content and protein binding capacity. The protein binding capacity of tannin were determined using precipitation of bovine serum albumin (BSA). Swietenia mahagonihas higest total tannin level and condensed tannin (CT) compared with other forages (P<0.01). The Leucaena leucocephala has highest hydrolysable tannin (HT) level (P<0.01). The total and condensed tannin content of Swietenia mahagoni were 11.928±0.04 mg/100 mg and 9.241±0.02mg/100mg dry matter (DM) of leaves. The hydrolysable tannin content of Leucaena leucocephala was 5.338±0.03 mg/100 mg DM of leaves. Binding capacity was highest in Swietenia mahagoni and Leucaena leucocephala compared to the other forages (P<0.01). The optimum binding of BSA to tannin in Leucaena leucocephala and Swietenia mahagoniwere1.181±0.44 and 1.217±0.60mg/mg dry matter of leaves. The present study reports that Swietenia mahagoni has highest of tannin content and Leucaena leucocephala and Swietenia mahagoni capacity of protein binding.

Iron-binding antioxidant capacity is impaired in diabetes mellitus.

PubMed

Van Campenhout, Ann; Van Campenhout, Christel; Lagrou, Albert R; Moorkens, Greta; De Block, Christophe; Manuel-y-Keenoy, Begoña

2006-05-15

Increased lipid peroxidation contributes to diabetic complications and redox-active iron is known to play an important role in catalyzing peroxidation reactions. We aimed to investigate if diabetes affects the capacity of plasma to protect against iron-driven lipid peroxidation and to identify underlying factors. Glycemic control, serum iron, proteins involved in iron homeostasis, plasma iron-binding antioxidant capacity in a liposomal model, and non-transferrin-bound iron were measured in 40 type 1 and 67 type 2 diabetic patients compared to 100 nondiabetic healthy control subjects. Iron-binding antioxidant capacity was significantly lower in the plasma of diabetic subjects (83 +/- 6 and 84 +/- 5% in type 1 and type 2 diabetes versus 88 +/- 6% in control subjects, p < 0.0005). The contribution of transferrin, ceruloplasmin, and albumin concentrations to the iron-binding antioxidant capacity was lost in diabetes (explaining only 4.2 and 6.3% of the variance in type 1 and type 2 diabetes versus 13.9% in control subjects). This observation could not be explained by differences in Tf glycation, lipid, or inflammatory status and was not associated with higher non-transferrin-bound iron levels. Iron-binding antioxidant capacity is decreased in diabetes mellitus.

Quantitative determination of cesium binding to ferric hexacyanoferrate: Prussian blue.

PubMed

Faustino, Patrick J; Yang, Yongsheng; Progar, Joseph J; Brownell, Charles R; Sadrieh, Nakissa; May, Joan C; Leutzinger, Eldon; Place, David A; Duffy, Eric P; Houn, Florence; Loewke, Sally A; Mecozzi, Vincent J; Ellison, Christopher D; Khan, Mansoor A; Hussain, Ajaz S; Lyon, Robbe C

2008-05-12

Ferric hexacyanoferrate (Fe4III[FeII(CN)6]3), also known as insoluble Prussian blue (PB) is the active pharmaceutical ingredient (API) of the drug product, Radiogardase. Radiogardase is the first FDA approved medical countermeasure for the treatment of internal contamination with radioactive cesium (Cs) or thallium in the event of a major radiological incident such as a "dirty bomb". A number of pre-clinical and clinical studies have evaluated the use of PB as an investigational decorporation agent to enhance the excretion of metal cations. There are few sources of published in vitro data that detail the binding capacity of cesium to insoluble PB under various chemical and physical conditions. The study objective was to determine the in vitro binding capacity of PB APIs and drug products by evaluating certain chemical and physical factors such as medium pH, particle size, and storage conditions (temperature). In vitro experimental conditions ranged from pH 1 to 9, to cover the range of pH levels that PB may encounter in the gastrointestinal (GI) tract in humans. Measurements of cesium binding were made between 1 and 24h, to cover gastric and intestinal tract residence time using a validated atomic emission spectroscopy (AES) method. The results indicated that pH, exposure time, storage temperature (affecting moisture content) and particle size play significant roles in the cesium binding to both the PB API and the drug product. The lowest cesium binding was observed at gastric pH of 1 and 2, whereas the highest cesium binding was observed at physiological pH of 7.5. It was observed that dry storage conditions resulted in a loss of moisture from PB, which had a significant negative effect on the PB cesium binding capacity at time intervals consistent with gastric residence. Differences were also observed in the binding capacity of PB with different particle sizes. Significant batch to batch differences were also observed in the binding capacity of some PB API and drug products. Our results suggest that certain physiochemical properties affect the initial binding capacity and the overall binding capacity of PB APIs and drug products during conditions that simulated gastric and GI residence time. These physiochemical properties can be utilized as quality attributes to monitor and predict drug product quality under certain manufacturing and storage conditions and may be utilized to enhance the clinical efficacy of PB.

Phase equilibrium in system Ti-Si-C-B and synthesis of MAX phase layers in vacuum under the influence of electron beam

NASA Astrophysics Data System (ADS)

Smirnyagina, N. N.; Khaltanova, V. M.; Dasheev, D. E.; Lapina, A. E.

2017-05-01

Composite layers on the basis of carbides and borides the titan and silicon on titanic alloy VТ-1 are generated at diffused saturation by electron beam treatment in vacuum. Formation in a composite of MAX phase Ti3SiC2 is shown. Thermodynamic research of phase equilibrium in systems Ti-Si-C and Ti-B-C in the conditions of high vacuum is executed. The thermodynamics, formation mechanisms of superfirm layers borides and carbides of the titan and silicon are investigated.

Thermodynamic modelling of phase equilibrium in system Ti-B-Si-C, synthesis and phases composition of borides and carbides layers on titanic alloyVT-1 at electron beam treatment in vacuum

NASA Astrophysics Data System (ADS)

Smirnyagina, N. N.; Khaltanova, V. M.; Lapina, A. E.; Dasheev, D. E.

2017-01-01

Composite layers on the basis of carbides and borides the titan and silicon on titanic alloy VT-1 are generated at diffused saturation in vacuum. Formation in a composite of MAX phase Ti3SiC2 is shown. Thermodynamic research of phase equilibrium in systems Ti-Si-C and Ti-B-C in the conditions of high vacuum is executed. The thermodynamics, formation mechanisms of superfirm layers borides and carbides of the titan and silicon are investigated.

Contributions of molecular size, charge distribution, and specific amino acids to the iron-binding capacity of sea cucumber (Stichopus japonicus) ovum hydrolysates.

PubMed

Sun, Na; Cui, Pengbo; Jin, Ziqi; Wu, Haitao; Wang, Yixing; Lin, Songyi

2017-09-01

This study investigated the contributions of molecular size, charge distribution and specific amino acids to the iron-binding capacity of sea cucumber (Stichopus japonicus) ovum hydrolysates (SCOHs), and further explored their iron-binding sites. It was demonstrated that enzyme type and degree of hydrolysis (DH) significantly influenced the iron-binding capacity of the SCOHs. The SCOHs produced by alcalase at a DH of 25.9% possessed the highest iron-binding capacity at 92.1%. As the hydrolysis time increased, the molecular size of the SCOHs decreased, the negative charges increased, and the hydrophilic amino acids were exposed to the surface, facilitating iron binding. Furthermore, the Fourier transform infrared spectra, combined with amino acid composition analysis, revealed that iron bound to the SCOHs primarily through interactions with carboxyl oxygen of Asp, guanidine nitrogen of Arg or nitrogen atoms in imidazole group of His. The formed SCOHs-iron complexes exhibited a fold and crystal structure with spherical particles. Copyright © 2017 Elsevier Ltd. All rights reserved.

A nationwide multicentre study in Turkey for establishing reference intervals of haematological parameters with novel use of a panel of whole blood

PubMed Central

Ozarda, Yesim; Ichihara, Kiyoshi; Bakan, Ebubekir; Polat, Harun; Ozturk, Nurinnisa; Baygutalp, Nurcan K.; Taneli, Fatma; Guvenc, Yesim; Ormen, Murat; Erbayraktar, Zubeyde; Aksoy, Nurten; Sezen, Hatice; Demir, Meltem; Eskandari, Gulcin; Polat, Gurbuz; Mete, Nuriye; Yuksel, Hatice; Vatansev, Husamettin; Gun, Fatma; Akin, Okhan; Ceylan, Ozlem; Noyan, Tevfik; Gozlukaya, Ozgul; Aliyazicioglu, Yuksel; Kahraman, Sevim; Dirican, Melahat; Tuncer, Gul Ozlem; Kimura, Shogo; Eker, Pinar

2017-01-01

Introduction A nationwide multicentre study was conducted to establish well-defined reference intervals (RIs) of haematological parameters for the Turkish population in consideration of sources of variation in reference values (RVs). Materials and methods K2-EDTA whole blood samples (total of 3363) were collected from 12 laboratories. Sera were also collected for measurements of iron, UIBC, TIBC, and ferritin for use in the latent abnormal values exclusion (LAVE) method. The blood samples were analysed within 2 hours in each laboratory using Cell Dyn and Ruby (Abbott), LH780 (Beckman Coulter), or XT-2000i (Sysmex). A panel of freshly prepared blood from 40 healthy volunteers was measured in common to assess any analyser-dependent bias in the measurements. The SD ratio (SDR) based on ANOVA was used to judge the need for partitioning RVs. RIs were computed by the parametric method with/without applying the LAVE method. Results Analyser-dependent bias was found for basophils (Bas), MCHC, RDW and MPV from the panel test results and thus those RIs were derived for each manufacturer. RIs were determined from all volunteers’ results for WBC, neutrophils, lymphocytes, monocytes, eosinophils, MCV, MCH and platelets. Gender-specific RIs were required for RBC, haemoglobin, haematocrit, iron, UIBC and ferritin. Region-specific RIs were required for RBC, haemoglobin, haematocrit, UIBC, and TIBC. Conclusions With the novel use of a freshly prepared blood panel, manufacturer-specific RIs’ were derived for Bas, Bas%, MCHC, RDW and MPV. Regional differences in RIs were observed among the 7 regions of Turkey, which may be attributed to nutritional or environmental factors, including altitude. PMID:28694726

Binding characteristics and protective capacity of cyanidin-3-glucoside and its aglycon to calf thymus DNA.

PubMed

Zhang, Chao; Guo, Xiaofei; Cai, Wenqian; Ma, Yue; Zhao, Xiaoyan

2015-04-01

The binding characteristics and protective capacity of cyanidin (Cy) and cyanidin-3-glucoside (C3G) to calf thymus DNA were explored for the first time. The Cy and C3G gave a bathochromic shift to the ultraviolet-visible spectra of the DNA, indicating the formation of the DNA-Cy and DNA-C3G complexes. The complexes were formed by an intercalative binding mode based on the results of the fluorescence spectra and competitive binding analysis. Meanwhile, the Cy and C3G protected the DNA from the damage induced by the hydroxyl radical. The binding capacity and protective capacity of the C3G were stronger than that of the Cy. Furthermore, the formation of the DNA-anthocyanin complexes was spontaneous when the hydrogen bond and hydrophobic force played a key role. Hence, the Cy and C3G could protect the DNA automatically from the damage induced by the hydroxyl radical. © 2015 Institute of Food Technologists®

Effect of Iron Chelation Therapy on Glucose Metabolism in Non-Transfusion-Dependent Thalassaemia.

PubMed

Chuansumrit, Ampaiwan; Pengpis, Pimprae; Mahachoklertwattana, Pat; Sirachainan, Nongnuch; Poomthavorn, Preamrudee; Sungkarat, Witaya; Kadegasem, Praguywan; Khlairit, Patcharin; Wongwerawattanakoon, Pakawan

2017-01-01

To compare insulin sensitivity, β-cell function and iron status biomarkers in non-transfusion-dependent thalassaemia (NTDT) with iron excess during pre- and post-iron chelation. Subjects with NTDT, aged older than 10 years, with serum ferritin >300 ng/ml, were included. Iron chelation with deferasirox (10 mg/kg/day) was prescribed daily for 6 months. Ten patients with a median age of 17.4 years were enrolled. The comparison between pre- and post-chelation demonstrated significantly lower iron load: median serum ferritin (551.4 vs. 486.2 ng/ml, p = 0.047), median TIBC (211.5 vs. 233.5 µg/dl, p = 0.009) and median non-transferrin binding iron (5.5 vs. 1.4 µM, p = 0.005). All patients had a normal oral glucose tolerance test (OGTT) both pre- and post-chelation. However, fasting plasma glucose was significantly reduced after iron chelation (85.0 vs.79.5 mg/dl, p = 0.047). MRI revealed no significant changes of iron accumulation in the heart and liver after chelation, but there was a significantly lower iron load in the pancreas, assessed by higher T2* at post-chelation compared with pre-chelation (41.9 vs. 36.7 ms, p = 0.047). No adverse events were detected. A trend towards improving insulin sensitivity and β-cell function as well as a reduced pancreatic iron load was observed following 6 months of iron chelation (TCTR20160523003). © 2016 S. Karger AG, Basel.

Effects of salts on protein-surface interactions: applications for column chromatography.

PubMed

Tsumoto, Kouhei; Ejima, Daisuke; Senczuk, Anna M; Kita, Yoshiko; Arakawa, Tsutomu

2007-07-01

Development of protein pharmaceuticals depends on the availability of high quality proteins. Various column chromatographies are used to purify proteins and characterize the purity and properties of the proteins. Most column chromatographies require salts, whether inorganic or organic, for binding, elution or simply better recovery and resolution. The salts modulate affinity of the proteins for particular columns and nonspecific protein-protein or protein-surface interactions, depending on the type and concentration of the salts, in both specific and nonspecific manners. Salts also affect the binding capacity of the column, which determines the size of the column to be used. Binding capacity, whether equilibrium or dynamic (under an approximation of a slow flow rate), depends on the binding constant, protein concentration and the number of the binding site on the column as well as nonspecific binding. This review attempts to summarize the mechanism of the salt effects on binding affinity and capacity for various column chromatographies and on nonspecific protein-protein or protein-surface interactions. Understanding such salt effects should also be useful in preventing nonspecific protein binding to various containers. Copyright 2007 Wiley-Liss, Inc.

The presence of high-affinity, low-capacity estradiol-17β binding in rainbow trout scale indicates a possible endocrine route for the regulation of scale resorption

USGS Publications Warehouse

Persson, Petra; Shrimpton, J.M.; McCormick, S.D.; Bjornsson, Bjorn Thrandur

2000-01-01

High-affinity, low-capacity estradiol-17β (E2) binding is present in rainbow trout scale. The Kd and Bmax of the scale E2 binding are similar to those of the liver E2 receptor (Kd is 1.6 ± 0.1 and 1.4 ± 0.1 nM, and Bmax is 9.1 ± 1.2 and 23.1 ± 2.2 fmol x mg protein-1, for scale and liver, respectively), but different from those of the high-affinity, low-capacity E2 binding in plasma (Kd is 4.0 ± 0.4 nM and Bmax is 625.4 ± 63.1 fmol x mg protein-1). The E2 binding in scale was displaced by testosterone, but not by diethylstilbestrol. Hence, the ligand binding specificity is different from that of the previously characterized liver E2 receptor, where E2 is displaced by diethylstilbestrol, but not by testosterone. The putative scale E2 receptor thus appears to bind both E2 and testosterone, and it is proposed that the increased scale resorption observed during sexual maturation in both sexes of several salmonid species may be mediated by this receptor. No high-affinity, low-capacity E2 binding could be detected in rainbow trout gill or skin.

Large heat capacity change in a protein-monovalent cation interaction.

PubMed

Guinto, E R; Di Cera, E

1996-07-09

Current views about protein-ligand interactions state that electrostatic forces drive the binding of charged species and that burial of hydrophobic and polar surfaces controls the heat capacity change associated with the reaction. For the interaction of a protein with a monovalent cation the electrostatic components are expected to be significant due to the ionic nature of the ligand, whereas the heat capacity change is expected to be small due to the size of the surface area involved in the recognition event. The physiologically important interaction of Na+ with thrombin was studied over the temperature range from 5 to 45 degrees C and the ionic strength range from 50 to 800 mM. These measurements reveal an unanticipated result that bears quite generally on studies of molecular recognition and protein folding. Binding of Na+ to thrombin is characterized by a modest dependence on ionic strength but a large and negative heat capacity change of -1.1 +/- 0.1 kcal mol-1 K-1. The small electrostatic coupling can be explained in terms of a minimal perturbation of the ionic atmosphere of the protein upon Na+ binding. The large heat capacity change, however, is difficult to reconcile with current views on the origin of this effect from surface area changes or large folding transitions coupled to binding. It is proposed that this change is linked to burial of a large cluster of water molecules in the Na+ binding pocket upon Na+ binding. Due to their reduced mobility and highly ordered structure, water molecules sequestered in the interior of a protein must have a lower heat capacity compared to those on the surface of a protein or in the bulk solvent. Hence, a binding or folding event where water molecules are buried may result in significant heat capacity changes independent of changes in exposed hydrophobic surface or coupled conformational transitions.

Histamine-binding capacities of different natural zeolites: a comparative study.

PubMed

Selvam, Thangaraj; Schwieger, Wilhelm; Dathe, Wilfried

2018-06-07

Two different natural zeolites from Cuba and Mexico, which are already being used as contemporaneous drugs or dietary supplements in Germany and Mexico, respectively, are applied in a comparative study of their histamine-binding capacities as a function of their particle sizes. The zeolites are characterized using X-ray diffraction (XRD), scanning electron microscopy (SEM) and N 2 -sorption measurements (BET surface areas). The Cuban zeolite contains clinoptilolite and mordenite as major phases (78% zeolite), whereas the Mexican one contains only clinoptilolite (65% zeolite). Both zeolites are apparently free from fibrous materials according to SEM. Both zeolites adsorb significant amount of histamine under the experimental conditions. Nevertheless, the results showed that the histamine-binding capacity of the Cuban zeolite is higher than the Mexican one and the smaller the particle size of zeolite, the higher the histamine-binding capacity. This difference could be due to the variation in their mineralogical compositions resulting in varied BET surface areas. Thus, the high histamine-binding capacities of Cuban zeolites seem to be due at least partly to the presence of the large-pore zeolite mordenite, providing high total pore volumes, which will be discussed in detail. For the first time, we have shown that the mineralogical compositions of natural zeolites and their particle sizes play a key role in binding histamine, which is one of the most important regulators in human physiology.

In vitro hypoglycemic and cholesterol lowering effects of dietary fiber prepared from cocoa (Theobroma cacao L.) shells.

PubMed

Nsor-Atindana, John; Zhong, Fang; Mothibe, Kebitsamang Joseph

2012-10-01

Three dietary fiber (DF) powders; soluble dietary fiber (SDF), insoluble dietary fiber (IDF) and total dietary fiber (TDF) were prepared from cocoa bean shells (CBS) by enzymatic treatment. These DFs were evaluated for their effects on glucose adsorption, glucose diffusion, starch hydrolysis, cholesterol binding, sodium cholate binding and oil binding capacities using in vitro model systems by simulating gastric intestinal conditions. The results showed that SDF generally exhibited significantly (p < 0.05) higher glucose adsorption capacity (GAC), α-amylase inhibition activity, cholesterol and sodium cholate binding capacity, but less significant (>0.05) glucose dialysis retardation index (GDRI) and oil binding capacity, when compared with IDF and TDF which both showed similar effects. Moreover, it was discovered that the three CBS dietary fiber powders contained intrinsic antioxidants (phenolic compounds). The study suggested that CBS could be an alternative cheap source of DF with additional benefits. Thus, CBS fibers could be incorporated as low calorie bulk ingredients in high-fiber diet to reduce calorie and cholesterol levels and control blood glucose level.

Increases thermal stability and cellulose-binding capacity of Cryptococcus sp. S-2 lipase by fusion of cellulose binding domain derived from Trichoderma reesei.

PubMed

Thongekkaew, Jantaporn; Ikeda, Hiroko; Iefuji, Haruyuki

2012-03-30

To improve the thermal stability and cellulose-binding capacity of Cryptococcus sp. S-2 lipase (CSLP), the cellulose-binding domain originates from Trichoderma reesei cellobiohydrolase I was engineered into C-terminal region of the CSLP (CSLP-CBD). The CSLP and CSLP-CBD were successfully expressed in the Pichia pastoris using the strong methanol inducible alcohol oxidase 1 (AOX1) promoter and the secretion signal sequence from Saccharomyces cerevisiae (α factor). The recombinant CSLP and CSLP-CBD were secreted into culture medium and estimated by SDS-PAGE to be 22 and 27 kDa, respectively. The fusion enzyme was stable at 80 °C and retained more than 80% of its activity after 120-min incubation at this temperature. Our results also found that the fusion of fungal exoglucanase cellulose-binding domain to CSLP is responsible for cellulose-binding capacity. This attribute should make it an attractive applicant for enzyme immobilization. Copyright © 2012 Elsevier Inc. All rights reserved.

The presence of high-affinity, low-capacity estradiol-17β binding in rainbow trout scale indicates a possible endocrine route for the regulation of scale resorption

USGS Publications Warehouse

Persson, Petra; Shrimpton, J. Mark; McCormick, Stephen D.; Bjornsson, Bjorn Thrandur

2000-01-01

High-affinity, low-capacity estradiol-17β (E2) binding is present in rainbow trout scale. The Kd and Bmax of the scale E2 binding are similar to those of the liver E2 receptor (Kd is 1.6 ± 0.1 and 1.4 ± 0.1 nM, and Bmax is 9.1 ± 1.2 and 23.1 ± 2.2 fmol × mg protein-1, for scale and liver, respectively), but different from those of the high-affinity, low-capacity E2 binding in plasma (Kd is 4.0 ± 0.4 nM and Bmax is 625.4 ± 63.1 fmol × mg protein−1). The E2 binding in scale was displaced by testosterone, but not by diethylstilbestrol. Hence, the ligand binding specificity is different from that of the previously characterized liver E2 receptor, where E2 is displaced by diethylstilbestrol, but not by testosterone. The putative scale E2 receptor thus appears to bind both E2 and testosterone, and it is proposed that the increased scale resorption observed during sexual maturation in both sexes of several salmonid species may be mediated by this receptor. No high-affinity, low-capacity E2 binding could be detected in rainbow trout gill or skin.

Increases thermal stability and cellulose-binding capacity of Cryptococcus sp. S-2 lipase by fusion of cellulose binding domain derived from Trichoderma reesei

DOE Office of Scientific and Technical Information (OSTI.GOV)

Thongekkaew, Jantaporn, E-mail: jantaporn_25@yahoo.com; Ikeda, Hiroko; Iefuji, Haruyuki

Highlights: Black-Right-Pointing-Pointer The CSLP and fusion enzyme were successfully expressed in the Pichia pastoris. Black-Right-Pointing-Pointer The fusion enzyme was stable at 80 Degree-Sign C for 120-min. Black-Right-Pointing-Pointer The fusion enzyme was responsible for cellulose-binding capacity. Black-Right-Pointing-Pointer The fusion enzyme has an attractive applicant for enzyme immobilization. -- Abstract: To improve the thermal stability and cellulose-binding capacity of Cryptococcus sp. S-2 lipase (CSLP), the cellulose-binding domain originates from Trichoderma reesei cellobiohydrolase I was engineered into C-terminal region of the CSLP (CSLP-CBD). The CSLP and CSLP-CBD were successfully expressed in the Pichia pastoris using the strong methanol inducible alcohol oxidase 1 (AOX1)more » promoter and the secretion signal sequence from Saccharomyces cerevisiae ({alpha} factor). The recombinant CSLP and CSLP-CBD were secreted into culture medium and estimated by SDS-PAGE to be 22 and 27 kDa, respectively. The fusion enzyme was stable at 80 Degree-Sign C and retained more than 80% of its activity after 120-min incubation at this temperature. Our results also found that the fusion of fungal exoglucanase cellulose-binding domain to CSLP is responsible for cellulose-binding capacity. This attribute should make it an attractive applicant for enzyme immobilization.« less

Current Understanding of the Binding Sites, Capacity, Affinity, and Biological Significance of Metals in Melanin

PubMed Central

Hong, Lian; Simon, John D.

2008-01-01

Metal chelation is often invoked as one of the main biological functions of melanin. In order to understand the interaction between metals and melanin, extensive studies have been carried out to determine the nature of the metal binding sites, binding capacity and affinity. These data are central to efforts aimed at elucidating the role metal binding plays in determining the physical, structural, biological, and photochemical properties of melanin. This article examines the current state of understanding of this field. PMID:17580858

Comparative study of thiophilic functionalised matrices for polyclonal F(ab')2 purification.

PubMed

Kumpalume, Peter; Slater, Nigel K H

2004-01-02

Thiophilic adsorbents have been developed using divinyl sulfone or epoxy activated Streamline quartz base matrix. Their capacity and selectivity for binding polyclonal F(ab')2 fragments generated by whole serum proteolysis was tested. Except for epoxy activated guanidine, all the adsorbents displayed high selectivity for F(ab')2 with dynamic binding capacities ranging from 3 to 10 mg/ml of adsorbent. Thiol immobilised ligands adsorbed more F(ab')2 and the recovery was equal to or more than that from amino immobilised ligands. All adsorbents showed good selectivity for IgG and the dynamic binding capacities were better than for F(ab')2.

Effects of retinoids on differentiation, lipid metabolism, epidermal growth factor, and low-density lipoprotein binding in squamous carcinoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ponec, M.; Weerheim, A.; Havekes, L.

The relationship among keratinocyte differentiation capacity, lipid synthesis, low-density lipoprotein (LDL) metabolism, plasma membrane composition, and epidermal growth factor (EGF) binding has been studied in SCC-12F2 cells. The differentiation capacity of the cells, i.e., ionophore-induced cornified envelope formation, was inhibited by various retinoids and stimulated by hydrocortisone. Retinoids that caused a significant reduction of cornified envelope formation, i.e., retinoic acid and 13-cis-retinoic acid, caused only minor changes in lipid synthesis and plasma membrane composition. Arotinoid ethylsulfone, having a minor effect on cornified envelope formation, caused a drastic inhibition of cholesterol synthesis resulting in changes in the plasma membrane composition. Hydrocortisonemore » stimulated cornified envelope formation but had only minor effects on lipid synthesis and plasma membrane composition. Of all retinoids tested, only arotinoid ethylsulfone caused a drastic increase in EGF binding, while hydrocortisone had no effect. These results clearly demonstrate that the plasma membrane composition is not related to keratinocyte differentiation capacity, but most likely does determine EGF binding. Furthermore, EGF binding does not determine keratinocyte differentiation capacity.« less

High-throughput process development: determination of dynamic binding capacity using microtiter filter plates filled with chromatography resin.

PubMed

Bergander, Tryggve; Nilsson-Välimaa, Kristina; Oberg, Katarina; Lacki, Karol M

2008-01-01

Steadily increasing demand for more efficient and more affordable biomolecule-based therapies put a significant burden on biopharma companies to reduce the cost of R&D activities associated with introduction of a new drug to the market. Reducing the time required to develop a purification process would be one option to address the high cost issue. The reduction in time can be accomplished if more efficient methods/tools are available for process development work, including high-throughput techniques. This paper addresses the transitions from traditional column-based process development to a modern high-throughput approach utilizing microtiter filter plates filled with a well-defined volume of chromatography resin. The approach is based on implementing the well-known batch uptake principle into microtiter plate geometry. Two variants of the proposed approach, allowing for either qualitative or quantitative estimation of dynamic binding capacity as a function of residence time, are described. Examples of quantitative estimation of dynamic binding capacities of human polyclonal IgG on MabSelect SuRe and of qualitative estimation of dynamic binding capacity of amyloglucosidase on a prototype of Capto DEAE weak ion exchanger are given. The proposed high-throughput method for determination of dynamic binding capacity significantly reduces time and sample consumption as compared to a traditional method utilizing packed chromatography columns without sacrificing the accuracy of data obtained.

Identification and properties of steroid-binding proteins in nesting Chelonia mydas plasma.

PubMed

Ikonomopoulou, M P; Bradley, A J; Whittier, J M; Ibrahim, K

2006-11-01

We report for the first time the presence of a sex steroid-binding protein in the plasma of green sea turtles Chelonia mydas, which provides an insight into reproductive status. A high affinity, low capacity sex hormone steroid-binding protein was identified in nesting C. mydas and its thermal profile was established. In nesting C. mydas testosterone and oestradiol bind at 4 degrees C with high affinity (K (a) = 1.49 +/- 0.09 x 10(9) M(-1); 0.17 +/- 0.02 x 10(7) M(-1)) and low binding capacity (B (max) = 3.24 +/- 0.84 x 10(-5) M; 0.33 +/- 0.06 x 10(-4) M). The binding affinity and capacity of testosterone at 23 and 36 degrees C, respectively were similar to those determined at 4 degrees C. However, oestradiol showed no binding activity at 36 degrees C. With competition studies we showed that oestradiol and oestrone do not compete for binding sites. Furthermore, in nesting C. mydas plasma no high-affinity binding was observed for adrenocortical steroids (cortisol and corticosterone) and progesterone. Our results indicate that in nesting C. mydas plasma temperature has a minimal effect on the high-affinity binding of testosterone to sex steroid-binding protein, however, the high affinity binding of oestradiol to sex steroid-binding protein is abolished at a hypothetically high (36 degrees C) sea/ambient/body temperature. This suggests that at high core body temperatures most of the oestradiol becomes biologically available to the tissues rather than remaining bound to a high-affinity carrier.

21 CFR 862.1415 - Iron-binding capacity test system.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Iron-binding capacity test system. 862.1415 Section 862.1415 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test...

Cooperative binding of anti-tetanus toxin monoclonal antibodies: Implications for designing an efficient biclonal preparation to prevent tetanus toxin intoxication.

PubMed

Lukic, Ivana; Filipovic, Ana; Inic-Kanada, Aleksandra; Marinkovic, Emilija; Miljkovic, Radmila; Stojanovic, Marijana

2018-05-15

Oligoclonal combinations of several monoclonal antibodies (MAbs) are being considered for the treatment of various infectious pathologies. These combinations are less sensitive to antigen structural changes than individual MAbs; at the same time, their characteristics can be more efficiently controlled than those of polyclonal antibodies. The main goal of this study was to evaluate the binding characteristics of six biclonal equimolar preparations (BEP) of tetanus toxin (TeNT)-specific MAbs and to investigate how the MAb combination influences the BEPs' protective capacity. We show that a combination of TeNT-specific MAbs, which not only bind TeNT but also exert positive cooperative effects, results in a BEP with superior binding characteristics and protective capacity, when compared with the individual component MAbs. Furthermore, we show that a MAb with only partial protective capacity but positive effects on the binding of the other BEP component can be used as a valuable constituent of the BEP. Copyright © 2018 Elsevier Ltd. All rights reserved.

Study of the Thermodynamics of Chromium(III) and Chromium(VI) Binding to Fe3O4 and MnFe2O4 nanoparticles

PubMed Central

Luther, Steven; Brogfeld, Nathan; Kim, Jisoo; Parsons, J.G.

2013-01-01

Removal of chromium(III) or (VI) from aqueous solution was achieved using Fe3O4, and MnFe2O4 nanomaterials. The nanomaterials were synthesized using a precipitation method and characterized using XRD. The size of the nanomaterials was determined to be 22.4 ± 0.9 nm (Fe3O4) and 15.5 ± 0.5 nm (MnFe2O4). The optimal binding pH for chromium(III) and chromium(VI) were pH 6 and pH 3. Isotherm studies were performed, under light and dark conditions, to determine the capacity of the nanomaterials. The capacities for the light studies with MnFe2O4 and Fe3O4 were determined to be 7.189 and 10.63 mg/g, respectively, for chromium(III). The capacities for the light studies with MnFe2O4 and Fe3O4 were 3.21 and 3.46 mg/g, respectively, for chromium(VI). Under dark reaction conditions the binding of chromium(III) to the MnFe2O4 and Fe3O4 nanomaterials were 5.74 and 15.9 mg/g, respectively. The binding capacity for the binding of chromium(VI) to MnFe2O4 and Fe3O4 under dark reaction conditions were 3.87 and 8.54 mg/g, respectively. The thermodynamics for the reactions showed negative ΔG values, and positive ΔH values. The ΔS values were positive for the binding of chromium(III) and for chromium(VI) binding under dark reaction conditions. The ΔS values for chromium(VI) binding under the light reaction conditions were determined to be negative. PMID:23558081

Equilibrium binding behavior of magnesium to wall teichoic acid.

PubMed

Thomas, Kieth J; Rice, Charles V

2015-10-01

Peptidoglycan and teichoic acids are the major cell wall components of Gram-positive bacteria that obtain and sequester metal ions required for biochemical processes. The delivery of metals to the cytoplasmic membrane is aided by anionic binding sites within the peptidoglycan and along the phosphodiester polymer of teichoic acid. The interaction with metals is a delicate balance between the need for attraction and ion diffusion to the membrane. Likewise, metal chelation from the extracellular fluid must initially have strong binding energetics that weaken within the cell wall to enable ion release. We employed atomic absorption and equilibrium dialysis to measure the metal binding capacity and metal binding affinity of wall teichoic acid and Mg2+. Data show that Mg2+ binds to WTA with a 1:2Mg2+ to phosphate ratio with a binding capacity of 1.27 μmol/mg. The affinity of Mg2+ to WTA was also found to be 41×10(3) M(-1) at low metal concentrations and 1.3×10(3) M(-1) at higher Mg2+ concentrations due to weakening electrostatic effects. These values are lower than the values describing Mg2+ interactions with peptidoglycan. However, the binding capacity of WTA is 4 times larger than peptidoglycan. External WTA initially binds metals with positive cooperativity, but metal binding switches to negative cooperativity, whereas interior WTA binds metals with only negative cooperativity. The relevance of this work is to describe changes in metal binding behavior depending on environment. When metals are sparse, chelation is strong to ensure survival yet the binding weakens when essential minerals are abundant. Copyright © 2015 Elsevier B.V. All rights reserved.

Norepinephrine transporter function and desipramine: residual drug effects versus short-term regulation.

PubMed

Ordway, Gregory A; Jia, Weihong; Li, Jing; Zhu, Meng-Yang; Mandela, Prashant; Pan, Jun

2005-04-30

Previous research has shown that exposure of norepinephrine transporter (NET)-expressing cells to desipramine (DMI) downregulates the norepinephrine transporter, although changes in the several transporter parameters do not demonstrate the same time course. Exposures to desipramine for <1 day reduces only radioligand binding and uptake capacity while transporter-immunoreactivity is unaffected. Recent demonstration of persistent drug retention in cells following desipramine exposures raises the possibility that previous reported changes in the norepinephrine transporter may be partly accountable by residual drug. In this study, potential effects of residual desipramine on norepinephrine transporter binding and uptake were re-evaluated following exposures of PC12 cells to desipramine using different methods to remove residual drug. Using a method that minimizes residual drug, exposure of intact PC12 cells to desipramine for 4h had no effect on uptake capacity or [(3)H]nisoxetine binding to the norepinephrine transporter, while exposures for > or =16 h reduced uptake capacity. Desipramine-induced reductions in binding to the transporter required >24 h or greater periods of desipramine exposure. This study confirms that uptake capacity of the norepinephrine transporter is reduced earlier than changes in radioligand binding, but with a different time course than originally shown. Special pre-incubation procedures are required to abolish effects of residual transporter inhibitor when studying inhibitor-induced transporter regulation.

Adsorption of plasmid DNA on anion exchange chromatography media.

PubMed

Tarmann, Christina; Jungbauer, Alois

2008-08-01

Anion exchange chromatography (AEC) is a useful and effective tool for DNA purification, but due to average pore sizes between 40 and 100 nm most AEC resins lack truly useful binding capacities for plasmid DNA (pDNA). Equilibrium binding capacities and uptake kinetics of AEC media including conventional media (Source 30 Q, Q Sepharose HP), a polymer grafted medium (Fractogel EMD DEAE (M)), media with large pores (Celbeads DEAE, PL SAX 4000 A 30 microm) and a monolithic medium (CIM-DEAE) were investigated by batch uptake or shallow bed experiments at two salt concentrations. Theoretical and experimental binding capacities suggest that the shape of the pDNA molecule can be described by a rod with a length to diameter ratio of 20:1 and that the molecule binds in upright position. The arrangement of DNA like a brush at the surface can be considered as entropy driven, kind of self-assembly process which is inherent to highly and uniformly charged DNA molecules. The initial phase of adsorption is very fast and levels off, associated with a change in mass transfer mechanism. Feed concentrations higher than 0.1 mg/mL pDNA pronounce this effect. Monolithic media showed the fastest adsorption rate and highest binding capacity with 13 mg pDNA per mL.

Pentoxifylline for Anemia in Chronic Kidney Disease: A Systematic Review and Meta-Analysis

PubMed Central

Bolignano, Davide; D’Arrigo, Graziella; Pisano, Anna; Coppolino, Giuseppe

2015-01-01

Background Pentoxifylline (PTX) is a promising therapeutic approach for reducing inflammation and improving anemia associated to various systemic disorders. However, whether this agent may be helpful for anemia management also in CKD patients is still object of debate. Study Design Systematic review and meta-analysis. Population Adults with CKD (any KDOQI stage, including ESKD patients on regular dialysis) and anemia (Hb<13 g/dL in men or < 12 g/dL in women). Search Strategy and Sources Cochrane CENTRAL, EMBASE, Ovid-MEDLINE and PubMed were searched for studies providing data on the effects of PTX on anemia parameters in CKD patients without design or follow-up restriction. Intervention PTX derivatives at any dose regimen. Outcomes Hemoglobin, hematocrit, ESAs dosage and resistance (ERI), iron indexes (ferritin, serum iron, TIBC, transferrin and serum hepcidin) and adverse events. Results We retrieved 11 studies (377 patients) including seven randomized controlled trials (all comparing PTX to placebo or standard therapy) one retrospective case-control study and three prospective uncontrolled studies. Overall, PTX increased hemoglobin in three uncontrolled studies but such improvement was not confirmed in a meta-analysis of seven studies (299 patients) (MD 0.12 g/dL, 95% CI -0.22 to 0.47). Similarly, there were no conclusive effects of PTX on hematocrit, ESAs dose, ferritin and TSAT in pooled analyses. Data on serum iron, ERI, TIBC and hepcidin were based on single studies. No evidence of increased rate of adverse events was also noticed. Limitations Small sample size and limited number of studies. High heterogeneity among studies with respect to CKD and anemia severity, duration of intervention and responsiveness/current therapy with iron or ESAs. Conclusions There is currently no conclusive evidence supporting the utility of pentoxifylline for improving anemia control in CKD patients. Future trials designed on hard, patient-centered outcomes with larger sample size and longer follow-up are advocated. PMID:26237421

Comparative haematological parameters of HbAA and HbAS genotype children infected with Plasmodium falciparum malaria in Yemen.

PubMed

Albiti, Anisa H; Nsiah, Kwabena

2014-04-01

Sickle haemoglobin (HbS) is known to offer considerable protection against falciparum malaria. However, the mechanism of protection is not yet completely understood. In this study, we investigate how the presence of the sickle cell trait affects the haematological profile of AS persons with malaria, in comparison with similarly infected persons with HbAA. This study is based on the hypothesis that the sickle cell trait plays a protective role against malaria. Children from an endemic malaria transmission area in Yemen were enrolled in this study. Hematological parameters were estimated using manual methods, the percentage of parasite density on stained thin smear was calculated, haemoglobin genotypes were determined on paper electrophoresis, ferritin was measured using enzyme-linked immunosorbent assay, serum iron and TIBC were assayed using spectrophotometer, transferrin saturation index was calculated by dividing serum iron by TIBC and expressing the result as a percentage. Haematological parameters were compared in HbAA- and HbAS-infected children. Falciparum malaria parasitaemia was confirmed in the blood smears of 62 children, 44 (55.7%) of AA and 18 (37.5%) AS, so there was higher prevalence in HbAA children (P = 0.047). Parasite density was lower in HbAS- than HbAA-infected children (P = 0.003). Anaemia was prominent in malaria-infected children, with high proportions of moderate and severe forms in HbAA (P = 0.001). The mean levels of haemoglobin, packed cell volume, reticulocyte count, platelets count, lymphocytes, eosinophils, and serum iron were significantly lower while total leukocytes, immature granulocytes, monocytes, erythrocyte sedimentation rate, transferrin saturation, and serum ferritin were significantly higher in HbAA-infected children than HbAS-infected children. Infection with Plasmodium falciparum malaria caused more significant haematological alterations of HbAA children than HbAS. This study supports the observation that sickle cell trait seems to be a beneficial genetic factor that resists malaria, since inheriting it protects against significant haematological consequences of malaria.

Alteration of the Copper-Binding Capacity of Iron-Rich Humic Colloids during Transport from Peatland to Marine Waters.

PubMed

Muller, François L L; Cuscov, Marco

2017-03-21

Blanket bogs contain vast amounts of Sphagnum-derived organic substances which can act as powerful chelators for dissolved iron and thus enhance its export to the coastal ocean. To investigate the variations in quantity and quality of these exports, adsorptive cathodic stripping voltammetry (CSV) was used to characterize the metal binding properties of molecular weight-fractionated dissolved organic matter (MW-fractionated DOM) in the catchment and coastal plume of a small peat-draining river over a seasonal cycle. Within the plume, both iron- and copper-binding organic ligands showed a linear, conservative distribution with increasing salinity, illustrating the high stability of peatland-derived humic substances (HS). Within the catchment, humic colloids lost up to 50% of their copper-binding capacity, expressed as a molar ratio to organic carbon, after residing for 1 week or more in the main reservoir of the catchment. Immediately downstream of the reservoir, the molar ratio [L 2 ]/[C org ], where L 2 was the second strongest copper-binding ligand, was 0.75 × 10 -4 when the reservoir residence time was 5 h but 0.34 × 10 -4 when it was 25 days. Residence time did not affect the carbon specific iron-binding capacity of the humic substances which was [L]/[C org ] = (0.80 ± 0.20) × 10 -2 . Our results suggest that the loss of copper-binding capacity with increasing residence time is caused by intracolloidal interactions between iron and HS during transit from peat soil to river mouth.

Study of As(III) and As(V) Oxoanion Adsorption onto Single and Mixed Ferrite and Hausmannite Nanomaterials

PubMed Central

Garcia, Sandra; Sardar, Saima; Maldonado, Stephanie; Garcia, Velia; Tamez, C.; Parsons, J. G.

2014-01-01

The removal of arsenic(III) and arsenic(V) from an aqueous solution through adsorption on to Fe3O4, MnFe2O4, 50% Mn substituted Fe3O4, 75% Mn substituted Fe3O4, and Mn3O4 nanomaterials was investigated. Characterization of the nanomaterials using XRD showed only pure phases for Mn3O4, MnFe2O4, and Fe3O4. The 50% and 75% substituted nanomaterials were found to be mixtures of Mn3O4 and Fe3O4. From batch studies the optimum binding pH of arsenic(III) and arsenic(V) to the nanomaterials was determined to be pH 3. The binding capacity for As(III) and As(VI) to the various nanomaterials was determined using Isotherm studies. The binding capacity of Fe3O4 was determined to be 17.1 mg/g for arsenic(III) and 7.0 mg/g for arsenic(V). The substitution of 25% Mn into the Fe3O4 lattice showed a slight increase in the binding capacity for As(III) and As(VI) to 23.8 mg/g and 7.9 mg/g, respectively. The 50% substituted showed the maximum binding capacity of 41.5 mg/g and 13.9 mg/g for arsenic(III) and arsenic(V). The 75% Mn substituted Fe3O4 capacities were 16.7 mg/g for arsenic(III) and 8.2 mg/g for arsenic(V). The binding capacity of the Mn3O4 was determined to be 13.5 mg/g for arsenic(III) and 7.5 mg/g for arsenic(V). In addition, interference studies on the effects of SO2−4, PO3−4, Cl−, and NO−3 investigated. All the interferences had very minimal effects on the As(III) and As(V) binding never fell below 20% even in the presence of 1000 ppm interfering ions. PMID:25097269

The decrease in the IgG-binding capacity of intensively dry heated whey proteins is associated with intense Maillard reaction, structural changes of the proteins and formation of RAGE-ligands.

PubMed

Liu, Fahui; Teodorowicz, Małgorzata; van Boekel, Martinus A J S; Wichers, Harry J; Hettinga, Kasper A

2016-01-01

Heat treatment is the most common way of milk processing, inducing structural changes as well as chemical modifications in milk proteins. These modifications influence the immune-reactivity and allergenicity of milk proteins. This study shows the influence of dry heating on the solubility, particle size, loss of accessible thiol and amino groups, degree of Maillard reaction, IgG-binding capacity and binding to the receptor for advanced glycation end products (RAGE) of thermally treated and glycated whey proteins. A mixture of whey proteins and lactose was dry heated at 130 °C up to 20 min to mimic the baking process in two different water activities, 0.23 to mimic the heating in the dry state and 0.59 for the semi-dry state. The dry heating was accompanied by a loss of soluble proteins and an increase in the size of dissolved aggregates. Most of the Maillard reaction sites were found to be located in the reported conformational epitope area on whey proteins. Therefore the structural changes, including exposure of the SH group, SH-SS exchange, covalent cross-links and the loss of available lysine, subsequently resulted in a decreased IgG-binding capacity (up to 33%). The binding of glycation products to RAGE increased with the heating time, which was correlated with the stage of the Maillard reaction and the decrease in the IgG-binding capacity. The RAGE-binding capacity was higher in samples with a lower water activity (0.23). These results indicate that the intensive dry heating of whey proteins as it occurs during baking may be of importance to the immunological properties of allergens in cow's milk, both due to chemical modifications of the allergens and formation of AGEs.

Oxytocin and vasopressin: distinct receptors in myometrium

DOE Office of Scientific and Technical Information (OSTI.GOV)

Guillon, G.; Balestre, M.N.; Roberts, J.M.

1987-06-01

The binding characteristics of (/sup 3/H)oxytocin (( /sup 3/H)OT) and (/sup 3/H)lysine vasopressin (( /sup 3/H)LVP) to nonpregnant human myometrium were investigated. Binding of both radioligands was saturable, time dependent, and reversible. Whereas (/sup 3/H)OT was found to bind to a single class of sites with high affinity (Kd, 1.5 +/- 0.4 (+/- SEM) nM) and low capacity (maximum binding (Bmax), 34 +/- 6 fmol/mg protein), (/sup 3/H)LVP bound to two classes of sites, one with high affinity (Kd, 2.2 +/- 0.1 nM) and low capacity (Bmax, 198 +/- 7 fmol/mg protein) and another with low affinity (Kd, 655 +/-more » 209 nM) and high capacity (Bmax, 5794 +/- 1616 fmol/mg protein). The binding of the labeled peptides also displayed a marked difference in sensitivity to Mg2+ and guanine nucleotides. These differences in binding characteristics as well as the differences in potency of analogs in competing for (/sup 3/H)OT and (/sup 3/H)LVP binding indicate the presence of distinct receptors for OT and vasopressin in human myometrium. Pharmacological characterization of the high affinity binding sites for (/sup 3/H)LVP indicated that these are of the V1 subtype. Although, as suggested by others, vasopressin and OT can bind to the same sites, the presence of distinct receptors for both peptides provides an explanation for the previously reported difference in myometrial responsiveness to OT and vasopressin.« less

Binding and Utilization of Human Transferrin by Prevotella nigrescens

PubMed Central

Duchesne, Pascale; Grenier, Daniel; Mayrand, Denis

1999-01-01

To survive and multiply within their hosts, pathogens must possess efficient iron-scavenging mechanisms. In the present study, we investigate the capacity of Prevotella nigrescens and Prevotella intermedia to use various sources of iron for growth and characterize the transferrin-binding activity of P. nigrescens. Iron-saturated human transferrin and lactoferrin, but not ferric chloride and the iron-free form of transferrin, could be used as sources of iron by P. nigrescens and P. intermedia. Neither siderophore activity nor ferric reductase activity could be detected in P. nigrescens and P. intermedia. However, both species showed transferrin-binding activity as well as the capacity to proteolytically cleave transferrin. To various extents, all strains of P. nigrescens and P. intermedia tested demonstrated transferrin-binding activity. The activity was heat and protease sensitive. The capacity of P. nigrescens to bind transferrin was decreased when cells were grown in the presence of hemin. Preincubation of bacterial cells with hemin, hemoglobin, lactoferrin, fibrinogen, immunoglobulin G, or laminin did not affect transferrin-binding activity. The transferrin-binding protein could be extracted from the cell surface of P. nigrescens by treatment with a zwitterionic detergent. Subjecting the cell surface extract to affinity chromatography on an agarose-transferrin column revealed that it contained a protein having an estimated molecular mass of 37 kDa and possessing transferrin-binding activity. The transferrin-binding activity of P. nigrescens and P. intermedia may permit the bacteria to obtain iron for survival and growth in periodontal pockets. PMID:9916061

Hupresin Retains Binding Capacity for Butyrylcholinesterase and Acetylcholinesterase after Sanitation with Sodium Hydroxide.

PubMed

Onder, Seda; David, Emilie; Tacal, Ozden; Schopfer, Lawrence M; Lockridge, Oksana

2017-01-01

Hupresin is a new affinity resin that binds butyrylcholinesterase (BChE) in human plasma and acetylcholinesterase (AChE) solubilized from red blood cells (RBC). Hupresin is available from the CHEMFORASE company. BChE in human plasma binds to Hupresin and is released with 0.1 M trimethylammonium bromide (TMA) with full activity and 10-15% purity. BChE immunopurified from plasma by binding to immobilized monoclonal beads has fewer contaminating proteins than the one-step Hupresin-purified BChE. However, when affinity chromatography on Hupresin follows ion exchange chromatography at pH 4.5, BChE is 99% pure. The membrane bound AChE, solubilized from human RBC with 0.6% Triton X-100, binds to Hupresin and remains bound during washing with sodium chloride. Human AChE is not released in significant quantities with non-denaturing solvents, but is recovered in 1% trifluoroacetic acid. The denatured, partially purified AChE is useful for detecting exposure to nerve agents by mass spectrometry. Our goal was to determine whether Hupresin retains binding capacity for BChE and AChE after Hupresin is washed with 0.1 M NaOH. A 2 mL column of Hupresin equilibrated in 20 mM TrisCl pH 7.5 was used in seven consecutive trials to measure binding and recovery of BChE from 100 mL human plasma. Between each trial the Hupresin was washed with 10 column volumes of 0.1 M sodium hydroxide. A similar trial was conducted with red blood cell AChE in 0.6% Triton X-100. It was found that the binding capacity for BChE and AChE was unaffected by washing Hupresin with 0.1 M sodium hydroxide. Hupresin could be washed with sodium hydroxide at least seven times without losing binding capacity.

Adaptation of avian influenza A (H6N1) virus from avian to human receptor-binding preference

PubMed Central

Wang, Fei; Qi, Jianxun; Bi, Yuhai; Zhang, Wei; Wang, Min; Zhang, Baorong; Wang, Ming; Liu, Jinhua; Yan, Jinghua; Shi, Yi; Gao, George F

2015-01-01

The receptor-binding specificity of influenza A viruses is a major determinant for the host tropism of the virus, which enables interspecies transmission. In 2013, the first human case of infection with avian influenza A (H6N1) virus was reported in Taiwan. To gather evidence concerning the epidemic potential of H6 subtype viruses, we performed comprehensive analysis of receptor-binding properties of Taiwan-isolated H6 HAs from 1972 to 2013. We propose that the receptor-binding properties of Taiwan-isolated H6 HAs have undergone three major stages: initially avian receptor-binding preference, secondarily obtaining human receptor-binding capacity, and recently human receptor-binding preference, which has been confirmed by receptor-binding assessment of three representative virus isolates. Mutagenesis work revealed that E190V and G228S substitutions are important to acquire the human receptor-binding capacity, and the P186L substitution could reduce the binding to avian receptor. Further structural analysis revealed how the P186L substitution in the receptor-binding site of HA determines the receptor-binding preference change. We conclude that the human-infecting H6N1 evolved into a human receptor preference. PMID:25940072

Mutational analysis of vaccinia virus E3 protein: the biological functions do not correlate with its biochemical capacity to bind double-stranded RNA.

PubMed

Dueck, Kevin J; Hu, YuanShen Sandy; Chen, Peter; Deschambault, Yvon; Lee, Jocelyn; Varga, Jessie; Cao, Jingxin

2015-05-01

Vaccinia E3 protein has the biochemical capacity of binding to double-stranded RNA (dsRNA). The best characterized biological functions of the E3 protein include its host range function, suppression of cytokine expression, and inhibition of interferon (IFN)-induced antiviral activity. Currently, the role of the dsRNA binding capacity in the biological functions of the E3 protein is not clear. To further understand the mechanism of the E3 protein biological functions, we performed alanine scanning of the entire dsRNA binding domain of the E3 protein to examine the link between its biochemical capacity of dsRNA binding and biological functions. Of the 115 mutants examined, 20 were defective in dsRNA binding. Although the majority of the mutants defective in dsRNA binding also showed defective replication in HeLa cells, nine mutants (I105A, Y125A, E138A, F148A, F159A, K171A, L182A, L183A, and I187/188A) retained the host range function to various degrees. Further examination of a set of representative E3L mutants showed that residues essential for dsRNA binding are not essential for the biological functions of E3 protein, such as inhibition of protein kinase R (PKR) activation, suppression of cytokine expression, and apoptosis. Thus, data described in this communication strongly indicate the E3 protein performs its biological functions via a novel mechanism which does not correlate with its dsRNA binding activity. dsRNAs produced during virus replication are important pathogen-associated molecular patterns (PAMPs) for inducing antiviral immune responses. One of the strategies used by many viruses to counteract such antiviral immune responses is achieved by producing dsRNA binding proteins, such as poxvirus E3 family proteins, influenza virus NS1, and Ebola virus V35 proteins. The most widely accepted model for the biological functions of this class of viral dsRNA binding proteins is that they bind to and sequester viral dsRNA PAMPs; thus, they suppress the related antiviral immune responses. However, no direct experimental data confirm such a model. In this study of vaccinia E3 protein, we found that the biological functions of the E3 protein are not necessarily linked to its biochemical capacity of dsRNA binding. Thus, our data strongly point to a new concept of virus modulation of cellular antiviral responses triggered by dsRNA PAMPs. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Characterization of [(3)H]harmane binding to rat whole brain membranes.

PubMed

Anderson, N J; Robinson, E S J; Husbands, S M; Delagrange, P; Nutt, D J; Hudson, A L

2003-12-01

This study investigates the binding of [(3)H]harmane to rat whole brain homogenates. Saturation studies revealed [(3)H]harmane labels a single, saturable, high-capacity population with high affinity. All the test compounds displaced [(3)H]harmane completely and in an apparently monophasic manner. The displacement profile of the test ligands indicated labeling of MAO-A. Given the high level of MAO-A binding, it is unlikely that a low-capacity I(2) site would be distinguishable from the total [(3)H]harmane population.

Hydrophobic interaction chromatography in dual salt system increases protein binding capacity.

PubMed

Senczuk, Anna M; Klinke, Ralph; Arakawa, Tsutomu; Vedantham, Ganesh; Yigzaw, Yinges

2009-08-01

Hydrophobic interaction chromatography (HIC) uses weakly hydrophobic resins and requires a salting-out salt to promote protein-resin interaction. The salting-out effects increase with protein and salt concentration. Dynamic binding capacity (DBC) is dependent on the binding constant, as well as on the flow characteristics during sample loading. DBC increases with the salt concentration but decreases with increasing flow rate. Dynamic and operational binding capacity have a major raw material cost/processing time impact on commercial scale production of monoclonal antibodies. In order to maximize DBC the highest salt concentration without causing precipitation is used. We report here a novel method to maintain protein solubility while increasing the DBC by using a combination of two salting-out salts (referred to as dual salt). In a series of experiments, we explored the dynamic capacity of a HIC resin (TosoBioscience Butyl 650M) with combinations of salts. Using a model antibody, we developed a system allowing us to increase the dynamic capacity up to twofold using the dual salt system over traditional, single salt system. We also investigated the application of this novel approach to several other proteins and salt combinations, and noted a similar protein solubility and DBC increase. The observed increase in DBC in the dual salt system was maintained at different linear flow rates and did not impact selectivity.

Revised Model of Calcium and Magnesium Binding to the Bacterial Cell Wall

PubMed Central

Thomas, Kieth J.; Rice, Charles V.

2014-01-01

Metals bind to the bacterial cell wall yet the binding mechanisms and affinity constants are not fully understood. The cell wall of gram positive bacteria is characterized by a thick layer of peptidoglycan and anionic teichoic acids anchored in the cytoplasmic membrane (lipoteichoic acid) or covalently bound to the cell wall (wall teichoic acid). The polyphosphate groups of teichoic acid provide one-half of the metal binding sites for calcium and magnesium, contradicting previous reports that calcium binding is 100% dependent on teichoic acid. The remaining binding sites are formed with the carboxyl units of peptidoglycan. In this work we report equilibrium association constants and total metal binding capacities for the interaction of calcium and magnesium ions with the bacterial cell wall. Metal binding is much stronger and previously reported. Curvature of Scatchard plots from the binding data and the resulting two regions of binding affinity suggest the presence of negative cooperative binding, meaning that the binding affinity decreases as more ions become bound to the sample. For Ca2+, Region I has a KA = (1.0 ± 0.2) × 106 M−1 and Region II has a KA = (0.075 ± 0.058) × 106 M−1. For Mg2+, KA1 = (1.5 ± 0.1) × 106 and KA2 = (0.17 ± 0.10) × 106. A binding capacity (η) is reported for both regions. However, since binding is still occurring in Region II, the total binding capacity is denoted by η2, which are 0.70 ± 0.04 µmol/mg and 0.67 ± 0.03 µmol/mg for Ca2+ and Mg2+ respectively. These data contradict the current paradigm of there being a single metal affinity value that is constant over a range of concentrations. We also find that measurement of equilibrium binding constants is highly sample dependent, suggesting a role for diffusion of metals through heterogeneous cell wall fragments. As a result, we are able to reconcile many contradictory theories that describe binding affinity and the binding mode of divalent metal cations. PMID:25315444

Seasonal changes in plasma androgens, glucocorticoids and glucocorticoid-binding proteins in the marsupial sugar glider Petaurus breviceps.

PubMed

Bradley, A J; Stoddart, D M

1992-01-01

An investigation spanning two breeding seasons was carried out to examine endocrine changes associated with reproduction in a wild population of the marsupial sugar glider Petaurus breviceps, a small arboreal gliding possum. Using techniques of equilibrium dialysis and polyacrylamide gel electrophoresis at steady-state conditions, a high-affinity, low-capacity glucocorticoid-binding protein was demonstrated in the plasma of Petaurus breviceps. Equilibrium dialysis at 36 degrees C using cortisol gave a high-affinity binding constant of 95 +/- 5.2 litres/mumol for a presumed corticosteroid-binding globulin (CBG) while the binding constant for the cortisol-albumin interaction was 3.5 +/- 0.4 litres/mmol. There was no difference between the sexes in the affinity of binding of cortisol to CBG; however, the cortisol-binding capacity underwent seasonal variation in both sexes. Progesterone was bound strongly to the presumed CBG while neither oestradiol nor aldosterone appeared to be bound with high affinity to P. breviceps plasma. In the males, peaks in the plasma concentration of testosterone coincided with the July-September breeding season in both years. A significant inverse relationship was shown to exist between the plasma testosterone concentration and the CBG-binding capacity. In both sexes an increase occurred in the plasma concentration of free cortisol during the first breeding season, a pattern which was not repeated in the subsequent breeding season, possibly due to a lower population density in that year.

Modelling the interdependence between the stoichiometry of receptor oligomerization and ligand binding for a coexisting dimer/tetramer receptor system.

PubMed

Rovira, X; Vivó, M; Serra, J; Roche, D; Strange, P G; Giraldo, J

2009-01-01

Many G protein-coupled receptors have been shown to exist as oligomers, but the oligomerization state and the effects of this on receptor function are unclear. For some G protein-coupled receptors, in ligand binding assays, different radioligands provide different maximal binding capacities. Here we have developed mathematical models for co-expressed dimeric and tetrameric species of receptors. We have considered models where the dimers and tetramers are in equilibrium and where they do not interconvert and we have also considered the potential influence of the ligands on the degree of oligomerization. By analogy with agonist efficacy, we have considered ligands that promote, inhibit or have no effect on oligomerization. Cell surface receptor expression and the intrinsic capacity of receptors to oligomerize are quantitative parameters of the equations. The models can account for differences in the maximal binding capacities of radioligands in different preparations of receptors and provide a conceptual framework for simulation and data fitting in complex oligomeric receptor situations.

Dietary fibers from mushroom Sclerotia: 2. In vitro mineral binding capacity under sequential simulated physiological conditions of the human gastrointestinal tract.

PubMed

Wong, Ka-Hing; Cheung, Peter C K

2005-11-30

The in vitro mineral binding capacity of three novel dietary fibers (DFs) prepared from mushroom sclerotia, namely, Pleurotus tuber-regium, Polyporous rhinocerus, and Wolfiporia cocos, to Ca, Mg, Cu, Fe, and Zn under sequential simulated physiological conditions of the human stomach, small intestine, and colon was investigated and compared. Apart from releasing most of their endogenous Ca (ranged from 96.9 to 97.9% removal) and Mg (ranged from 95.9 to 96.7% removal), simulated physiological conditions of the stomach also attenuated the possible adverse binding effect of the three sclerotial DFs to the exogenous minerals by lowering their cation-exchange capacity (ranged from 20.8 to 32.3%) and removing a substantial amount of their potential mineral chelators including protein (ranged from 16.2 to 37.8%) and phytate (ranged from 58.5 to 64.2%). The in vitro mineral binding capacity of the three sclerotial DF under simulated physiological conditions of small intestine was found to be low, especially for Ca (ranged from 4.79 to 5.91% binding) and Mg (ranged from 3.16 to 4.18% binding), and was highly correlated (r > 0.97) with their residual protein contents. Under simulated physiological conditions of the colon with slightly acidic pH (5.80), only bound Ca was readily released (ranged from 34.2 to 72.3% releasing) from the three sclerotial DFs, and their potential enhancing effect on passive Ca absorption in the human large intestine was also discussed.

A streptavidin linker layer that functions after drying.

PubMed

Xia, Nan; Shumaker-Parry, Jennifer S; Zareie, M Hadi; Campbell, Charles T; Castner, David G

2004-04-27

The ability of streptavidin (SA) to simultaneously bind four biotins is often used in linker layers, where a biotinylated molecule is linked to a biotin-functionalized surface via SA. For biosensor and array applications, it is desirable that the SA linker layer be stable to drying and rehydration. In this study it was observed that a significant decrease in binding capacity of a SA layer occurred when that layer was dried. For this study a SA linker layer was constructed by binding SA to a biotin-containing alkylthiolate monolayer (BAT/OEG) self-assembled onto gold. Its stability after drying was investigated using surface plasmon resonance (SPR). Approximately a quarter of the SA layer was removed from the BAT/OEG surface upon drying and rehydration, suggesting disruption of SA-biotin binding when dry. This resulted in the dried SA layer losing approximately 40% of its biotinylated ferritin (BF) binding capacity. Coating the layer with trehalose before drying was found to inhibit the loss of SA from the BAT/OEG surface. SPR showed that the trehalose-protected SA linker layer retained approximately 91% of its original BF binding capacity after drying and rehydration. Atomic force microscopy, which was used to image individual surface-bound SA and BF molecules, qualitatively confirmed these observations.

Lead-binding capacity of calcium pectates with different molecular weight.

PubMed

Khotimchenko, Maksim; Makarova, Ksenia; Khozhaenko, Elena; Kovalev, Valeri

2017-04-01

Nowadays, heavy metal contamination of environment is considered as a serious threat to public health because of toxicity of these pollutants and the lack of effective materials with metal-binding properties. Some biopolymers such as pectins were proposed for removal of metal ions from industrial water disposals. Chemical structure of pectins is quite variable and substantially affects their metal binding properties. In this work, relationship between molecular weight and Pb(II)-binding capacity of calcium pectates was investigated in a batch sorption system. The results showed that all pectate samples are able to form complexes with Pb(II) ions. The effects of contact time, pH of the media and equilibrium metal concentration on metal-binding process were tested in experiments. The equilibrium time min required for uptake of Pb(II) by pectate compounds was found to be 60min. Langmuir and Freundlich models were applied for description of interactions between pectates and metal ions. Binding capacity of low molecular pectate was highest among all the samples tested. Langmuir model was figured out to be the best fit within the whole range of pH values. These results demonstrate that calcium pectate with low molecular weight is more promising agent for elimination of Pb(II) ions from contaminated wastewaters. Copyright © 2017 Elsevier B.V. All rights reserved.

Ontogeny of growth hormone (GH) binding in the domestic turkey: evidence of sexual dimorphism and developmental changes in relationship to plasma GH.

PubMed

Vasilatos-Younken, R; Gray, K S; Bacon, W L; Nestor, K E; Long, D W; Rosenberger, J L

1990-07-01

The post-hatch ontogeny of hepatic GH binding and its relationship to GH plasma profile characteristics in male and female turkeys of slow- (RBC-2) and fast-growing (F; selected from RBC-2) genetic lines were determined. Specific binding of 125I-labelled recombinant chicken GH to crude hepatic membrane preparations (100,000 g pellet) was determined at 2, 4, 8, 14 and 24 weeks of age for both total (occupied plus free; 4 mol MgCl2/l pretreatment) and free (without MgCl2 pretreatment) binding sites. Characteristics of the plasma GH profile were measured at each age by serial blood sampling through indwelling jugular vein catheters. When specific binding to either free or total sites was expressed on a whole organ basis (i.e. hepatic GH-binding capacity/bird), binding increased dramatically (P less than 0.0001) with increasing age over both lines and sexes. Total binding capacity (free plus occupied sites) per bird was greater for females than for males at 24 weeks of age (P less than 0.04), as birds reached sexual maturity, but did not differ between fast- and slow-growing lines at any age. Available binding capacity (free sites) per bird was greater for the faster growing F than RBC-2 line at the older ages when body size was most divergent (14 and 24 weeks of age; P less than 0.01, P less than 0.06 respectively), but did not differ between sexes. Correlation analysis at individual ages revealed a progressive change in the nature of the relationship between hepatic GH binding, plasma GH and somatic growth.(ABSTRACT TRUNCATED AT 250 WORDS)

Serotonin and dopamine transporter binding in children with autism determined by SPECT.

PubMed

Makkonen, Ismo; Riikonen, Raili; Kokki, Hannu; Airaksinen, Mauno M; Kuikka, Jyrki T

2008-08-01

Disturbances in the serotonergic system have been recognized in autism. To investigate the association between serotonin and dopamine transporters and autism, we studied 15 children (14 males, one female; mean age 8 y 8 mo [SD 3 y 10 mo]) with autism and 10 non-autistic comparison children (five males, five females; mean age 9 y 10 mo [SD 2 y 8 mo]) using single-photon emission computed tomography (SPECT) with [123 I] nor-beta-CIT. The children, with autism were studied during light sedation. They showed reduced serotonin transporter (SERT) binding capacity in the medial frontal cortex, midbrain, and temporal lobe areas. However, after correction due to the estimated effect of sedation, the difference remained significant only in the medial frontal cortex area (p=0.002). In the individuals with autism dopamine transporter (DAT) binding did not differ from that of the comparison group. The results indicate that SERT binding capacity is disturbed in autism. The reduction is more evident in adolescence than in earlier childhood. The low SERT binding reported here and the low serotonin synthesis capacity shown elsewhere may indicate maturation of a lesser number of serotonergic nerve terminals in individuals with autism.

From neural oscillations to reasoning ability: Simulating the effect of the theta-to-gamma cycle length ratio on individual scores in a figural analogy test.

PubMed

Chuderski, Adam; Andrelczyk, Krzysztof

2015-02-01

Several existing computational models of working memory (WM) have predicted a positive relationship (later confirmed empirically) between WM capacity and the individual ratio of theta to gamma oscillatory band lengths. These models assume that each gamma cycle represents one WM object (e.g., a binding of its features), whereas the theta cycle integrates such objects into the maintained list. As WM capacity strongly predicts reasoning, it might be expected that this ratio also predicts performance in reasoning tasks. However, no computational model has yet explained how the differences in the theta-to-gamma ratio found among adult individuals might contribute to their scores on a reasoning test. Here, we propose a novel model of how WM capacity constraints figural analogical reasoning, aimed at explaining inter-individual differences in reasoning scores in terms of the characteristics of oscillatory patterns in the brain. In the model, the gamma cycle encodes the bindings between objects/features and the roles they play in the relations processed. Asynchrony between consecutive gamma cycles results from lateral inhibition between oscillating bindings. Computer simulations showed that achieving the highest WM capacity required reaching the optimal level of inhibition. When too strong, this inhibition eliminated some bindings from WM, whereas, when inhibition was too weak, the bindings became unstable and fell apart or became improperly grouped. The model aptly replicated several empirical effects and the distribution of individual scores, as well as the patterns of correlations found in the 100-people sample attempting the same reasoning task. Most importantly, the model's reasoning performance strongly depended on its theta-to-gamma ratio in same way as the performance of human participants depended on their WM capacity. The data suggest that proper regulation of oscillations in the theta and gamma bands may be crucial for both high WM capacity and effective complex cognition. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Capture and separation of l-histidine through optimized zinc-decorated magnetic silica spheres.

PubMed

Cardoso, Vanessa F; Sebastián, Víctor; Silva, Carlos J R; Botelho, Gabriela; Lanceros-Méndez, Senentxu

2017-09-01

Zinc-decorated magnetic silica spheres were developed, optimized and tested for the capture and separation of l-histidine. The magnetic silica spheres were prepared using a simple sol-gel method and show excellent magnetic characteristics, adsorption capacity toward metal ions, and stability in aqueous solution in a wide pH range. The binding capacity of zinc-decorated magnetic silica spheres to histidine proved to be strongly influenced by the morphology, composition and concentration of metal at the surface of the magnetic silica spheres and therefore these parameters should be carefully controlled in order to maximize the performance for protein purification purposes. Optimized zinc-decorated magnetic silica spheres demonstrate a binding capacity to l-histidine of approximately 44mgg -1 at the optimum binding pH buffer. Copyright © 2017 Elsevier B.V. All rights reserved.

Thiacarbocyanine as ligand in dye-affinity chromatography for protein purification. II. Dynamic binding capacity using lysozyme as a model.

PubMed

Boto, R E F; Anyanwu, U; Sousa, F; Almeida, P; Queiroz, J A

2009-09-01

A constant development of dye-affinity chromatography to replace more traditional techniques is verified, with the aim of increasing specificity in the purification of biomolecules. The establishment of a new dye-affinity chromatographic support imposes their complete characterization, namely with relation to the binding capacity for proteins, in order to evaluate its applicability on global purification processes. Following previous studies, the adsorption of lysozyme onto a thiacarbocyanine dye immobilized on beaded cellulose was investigated. The effect of different parameters, such as temperature, ionic strength, pH, protein concentration and flow rate, on the dynamic binding capacity of the support to retain lysozyme was also studied. Increasing the temperature and the lysozyme concentration had a positive effect on the dynamic binding capacity (DBC), whereas increasing the ionic strength and the flow rate resulted in the opposite. It was also discovered that the pH used had an important impact on the lysozyme binding onto the immobilized dye. The maximum DBC value obtained for lysozyme was 8.6 mg/mL, which was achieved at 30 degrees C and pH 9 with a protein concentration of 0.5 mg/mL and a flow rate of 0.05 mL/min. The dissociation constant (K(d)) obtained was 2.61 +/- 0.36 x 10(-5 )m, proving the affinity interaction between the thiacarbocyanine dye ligand and the lysozyme. Copyright (c) 2009 John Wiley & Sons, Ltd.

Development of high-productivity, strong cation-exchange adsorbers for protein capture by graft polymerization from membranes with different pore sizes

PubMed Central

Chenette, Heather C.S.; Robinson, Julie R.; Hobley, Eboni; Husson, Scott M.

2012-01-01

This paper describes the surface modification of macroporous membranes using ATRP (atom transfer radical polymerization) to create cation-exchange adsorbers with high protein binding capacity at high product throughput. The work is motivated by the need for a more economical and rapid capture step in downstream processing of protein therapeutics. Membranes with three reported nominal pore sizes (0.2, 0.45, 1.0 μm) were modified with poly(3-sulfopropyl methacrylate, potassium salt) tentacles, to create a high density of protein binding sites. A special formulation was used in which the monomer was protected by a crown ether to enable surface-initiated ATRP of this cationic polyelectrolyte. Success with modification was supported by chemical analysis using Fourier-transform infrared spectroscopy and indirectly by measurement of pure water flux as a function of polymerization time. Uniformity of modification within the membranes was visualized with confocal laser scanning microscopy. Static and dynamic binding capacities were measured using lysozyme protein to allow comparisons with reported performance data for commercial cation-exchange materials. Dynamic binding capacities were measured for flow rates ranging from 13 to 109 column volumes (CV)/min. Results show that this unique ATRP formulation can be used to fabricate cation-exchange membrane adsorbers with dynamic binding capacities as high as 70 mg/mL at a throughput of 100 CV/min and unprecedented productivity of 300 mg/mL/min. PMID:23175597

Dextran hydrogel coated surface plasmon resonance imaging (SPRi) sensor for sensitive and label-free detection of small molecule drugs

NASA Astrophysics Data System (ADS)

Li, Shaopeng; Yang, Mo; Zhou, Wenfei; Johnston, Trevor G.; Wang, Rui; Zhu, Jinsong

2015-11-01

The label-free and sensitive detection of small molecule drugs on SPRi is still a challenging task, mainly due to the limited surface immobilization capacity of the sensor. In this research, a dextran hydrogel-coated gold sensor chip for SPRi was successfully fabricated via photo-cross-linking for enhanced surface immobilization capacity. The density of the dextran hydrogel was optimized for protein immobilization and sensitive small molecule detection. The protein immobilization capacity of the hydrogel was 10 times greater than a bare gold surface, and 20 times greater than an 11-mercaptoundecanoic acid (MUA) surface. Such a drastic improvement in immobilization capacity allowed the SPRi sensor to detect adequate response signals when probing small molecule binding events. The binding signal of 4 nM liquid-phase biotin to streptavidin immobilized on the dextran surface reached 435 RU, while no response was observed on bare gold or MUA surfaces. The dextran hydrogel-coated SPRi sensor was also applied in a kinetic study of the binding between an immunosuppressive drug (FK506) and its target protein (FKBP12) in a high-throughput microarray format. The measured binding affinity was shown to be consistent with reported literature values, and a detection limit of 0.5 nM was achieved.

Improved purification of immunoglobulin G from plasma by mixed-mode chromatography.

PubMed

Chai, Dong-Sheng; Sun, Yan; Wang, Xiao-Ning; Shi, Qing-Hong

2014-12-01

Efficient loading of immunoglobulin G in mixed-mode chromatography is often a serious bottleneck in the chromatographic purification of immunoglobulin G. In this work, a mixed-mode ligand, 4-(1H-imidazol-1-yl) aniline, was coupled to Sepharose Fast Flow to fabricate AN SepFF adsorbents with ligand densities of 15-64 mmol/L, and the chromatographic performances of these adsorbents were thoroughly investigated to identify a feasible approach to improve immunoglobulin G purification. The results indicate that a critical ligand density exists for immunoglobulin G on the AN SepFF adsorbents. Above the critical ligand density, the adsorbents showed superior selectivity to immunoglobulin G at high salt concentrations, and also exhibited much higher dynamic binding capacities. For immunoglobulin G purification, both the yield and binding capacity increased with adsorbent ligand density along with a decrease in purity. It is difficult to improve the binding capacity, purity, and yield of immunoglobulin G simultaneously in AN SepFF chromatography. By using tandem AN SepFF chromatography, a threefold increase in binding capacity as well as high purity and yield of immunoglobulin G were achieved. Therefore, the tandem chromatography demonstrates that AN SepFF adsorbent is a practical and feasible alternative to MEP HyperCel adsorbents for immunoglobulin G purification. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Influence of binding pH and protein solubility on the dynamic binding capacity in hydrophobic interaction chromatography.

PubMed

Baumann, Pascal; Baumgartner, Kai; Hubbuch, Jürgen

2015-05-29

Hydrophobic interaction chromatography (HIC) is one of the most frequently used purification methods in biopharmaceutical industry. A major drawback of HIC, however, is the rather low dynamic binding capacity (DBC) obtained when compared to e.g. ion exchange chromatography (IEX). The typical purification procedure for HIC includes binding at neutral pH, independently of the proteins nature and isoelectric point. Most approaches to process intensification are based on resin and salt screenings. In this paper a combination of protein solubility data and varying binding pH leads to a clear enhancement of dynamic binding capacity. This is shown for three proteins of acidic, neutral, and alkaline isoelectric points. High-throughput solubility screenings as well as miniaturized and parallelized breakthrough curves on Media Scout RoboColumns (Atoll, Germany) were conducted at pH 3-10 on a fully automated robotic workstation. The screening results show a correlation between the DBC and the operational pH, the protein's isoelectric point and the overall solubility. Also, an inverse relationship of DBC in HIC and the binding kinetics was observed. By changing the operational pH, the DBC could be increased up to 30% compared to the standard purification procedure performed at neutral pH. As structural changes of the protein are reported during HIC processes, the applied samples and the elution fractions were proven not to be irreversibly unfolded. Copyright © 2015 Elsevier B.V. All rights reserved.

Binding of /sup 125/I-hCG to rainbow trout (Salmo gairdneri) testis in vitro. [Human Chorionic Gonadotropin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Schlaghecke, R.

1983-02-01

Homogenates of maturing rainbow trout testes show specific binding sites for /sup 125/I-labeled hCG (. /sup 125/I-labeled hCG). The binding is competitively inhibited by unlabeled hCG and by a hypophyseal extract of rainbow trout. It could be demonstrated that the tissue /sup 125/I-hCG binding specificity is restricted to the gonadal preparation. The trout testis was characterized by determining affinity and capacity from Scatchard plot analysis giving a high constant of dissociation Kd 3.65 x 10(-10)/M and a low binding capacity of 0.88 x 10(-15) M/mg tissue. The test system is markedly dependent on temperature, incubation-time, and pH. The maximum bindingmore » was found at 37 degrees during 2 hr of incubation in a buffer of pH 7.5.« less

Thermochemistry of the specific binding of C12 surfactants to bovine serum albumin.

PubMed

Nielsen, A D; Borch, K; Westh, P

2000-06-15

The specific binding to bovine serum albumin (BSA) of anionic and non-ionic surfactants with C12 acyl chains has been studied by high sensitivity isothermal titration calorimetry. This method proved particularly effective in resolving the binding of anionic surfactants into separate classes of sites with different affinity. For sodium dodecylsulfate (SDS) the measured binding curves could be rationalized as association to two classes (high affinity/low affinity) of sites comprising, respectively, three and six similar (i.e. thermodynamically equivalent), independent sites. Changes in the thermodynamic functions enthalpy, standard free energy, standard entropy and heat capacity could be discerned for each class of binding site, as well as for micelle formation. These data suggest that binding to low affinity sites (in analogy with micelle formation) exhibits energetic parameters; in particular, a large negative change in heat capacity, which is characteristic of hydrophobic interactions. The thermodynamics of high affinity binding, on the other hand, is indicative of other dominant forces; most likely electrostatic interactions. Other anionic ligands investigated (laurate and dodecyl benzylsulfonate) showed a behavior similar to SDS, the most significant difference being the high affinity binding of the alkylbenzyl sulfonate. For this ligand, the thermodynamic data is indicative of a more loosely associated complex than for SDS and laurate. BSA was found to bind one or two of the non-ionic surfactants (NIS) hepta- or penta(ethylene glycol) monododecyl ether (C12EO7 and C12EO5) with binding constants about three orders of magnitude lower than for SDS. Hence, the free energy of the surfactant in the weakly bound BSA-NIS complex is only slightly favored over the micellar state. The binding process is characterized by very large exothermic enthalpy changes (larger than for the charged surfactants) and a large, positive increment in heat capacity. These observations cannot be reconciled with a molecular picture based on simple hydrophobic condensation onto non-polar patches on the protein surface.

Drug Design Relating Amebicides to Inhibition of Protein Synthesis.

DTIC Science & Technology

1977-09-01

A study of the effect of emetine on protein synthesis in E. histolytica was made on log phase amebas as compared to stationary phase amebas ...Sensitivity to emetine was maintained independently of the rate of protein synthesis. Furthermore, both stages of amebas had the same capacity to bind emetine...elongation site. Finally, evidence was obtained that the capacity to bind emetine provides a basis for conferring drug resistance in amebas . A direct

Glycation of whey protein with dextrans of different molar mass: Effect on immunoglobulin E-binding capacity with blood sera obtained from patients with cow milk protein allergy.

PubMed

Xu, Lei; Gong, Yuansheng; Gern, James E; Ikeda, Shinya; Lucey, John A

2018-05-16

A growing concern around the world is the number of people who are suffering from food protein allergies. One potential approach to decrease protein allergenicity is to block IgE-binding epitopes of the protein allergen by attachment of polysaccharides via the Maillard reaction (i.e., glycation). Protein glycation has been extensively studied to modify various functional properties. We wanted to examine whether glycates could reduce IgE binding in patients with cow milk protein allergy and to explore how the size (molar mass; M W ) of the polysaccharide affects this IgE-binding capacity. Glycation was performed using the initial step of the Maillard reaction performed in aqueous solutions. The specific goal of this study was to reduce the IgE-binding capacity of whey protein isolate (WPI) through glycation with dextran (DX). Blood sera were obtained from 8 patients who had been diagnosed with cow milk protein allergy, and a composite sera sample was used for IgE-binding analysis by the ImmunoCap (Phadia, Uppsala, Sweden) method. The WPI was glycated with DX of M W ranging from 1 to 2,000 kDa, and the M W of purified glycates was determined using size-exclusion chromatography coupled with multiangle laser light scattering. The WPI to DX molar ratios in the glycates made from DX that had M W values of 1, 3.5, 10 (G10), 150, 500, and 2,000 kDa were 1:4, 1:3, 1:2, 1:1.5, 1:1, and 1:1, respectively. With the increase in the M W of DX, there was an increase in the M W values of the corresponding glycates but a decrease in the number of bound DX. The WPI-DX glycates had lower whey protein IgE-binding capacity than native WPI, with the lowest IgE-binding capacity obtained in the G10 glycate. The DX binding ratios and morphology results from atomic force microscopy images suggested that glycation of WPI with small-M W DX resulted in extensive protein surface coverage, probably due to the attachment of up to 4 DX molecules per whey protein. The lower IgE binding of the G10 glycate was likely due to greater steric hindrance (or a physical barrier) at the surface of the protein. In summary, our results demonstrate that glycating WPI with DX via Maillard reaction can potentially be used to decrease the allergenicity of whey protein. Copyright © 2018 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Salt-soluble proteins from wheat-derived foodstuffs show lower allergenic potency than those from raw flour.

PubMed

de Gregorio, Marta; Armentia, Alicia; Díaz-Perales, Araceli; Palacín, Arantxa; Dueñas-Laita, Antonio; Martín, Blanca; Salcedo, Gabriel; Sánchez-Monge, Rosa

2009-04-22

Salt-soluble proteins from wheat flour have been described as main allergens associated with both baker's asthma and food allergy. However, most studies have used raw flour as starting material, thus not considering potential changes in allergenic properties induced by the heat treatment and other industrial processing to produce wheat-derived foodstuffs. Salt extracts from different commercial wheat-derived products were obtained and their allergenic properties investigated by IgE-immunodetection, ELISA assays, and skin prick test. The IgE-binding capacity of salt-soluble proteins from commercial breads and cooked pastas was reduced around 50% compared with that of raw flour, the reduction being less dramatic in noncooked pastas and biscuits. Several wheat-derived foodstuffs showed major IgE-binding components of 20 and 35 kDa, identified as avenin-like and globulin proteins, respectively. These proteins, as well as most flour and bread salt-soluble proteins, were hydrolyzed when subjected to simulated gastrointestinal digestion. However, the digested products still exhibited a residual IgE-binding capacity. Therefore, processing of wheat flour to obtain derived foodstuffs decreases the IgE binding-capacity of the major salt-soluble wheat proteins. Moreover, simulated gastric fluid digestion further inactivates some heat-resistant IgE-binding proteins.

Transient chloride binding as a contributory factor to corneal stromal swelling in the ox.

PubMed Central

Hodson, S; Kaila, D; Hammond, S; Rebello, G; al-Omari, Y

1992-01-01

1. Investigations were made of the cation exchange capacity of fresh isolated ox corneal stroma (Q, units: mequiv fixed stromal charge/kg stromal fluid) at pH 7.4 over a variety of stomal hydrations (H, units: kg stromal fluid/kg dry tissue) both above and below the physiological hydration of 3.2, whilst the stromas were immersed in a variety of sodium chloride solutions (range 5-1000 mM). 2. At any particular salt concentration, the product QH (dry tissue exchange capacity, units: mequiv/kg dry tissue) appeared constant, over all the hydrations investigated. 3. Dry tissue exchange capacity (QH) varied, however, when the bathing salt concentration was altered. It varied between 55 mequiv/kg dry tissue (e.g. Q = 17 mequiv at H = 3.2) in 5 mM-NaCl to 240 mequiv/kg dry tissue (e.g. Q = 75 mequiv/l at H = 3.2) in 1000 mM-NaCl. 4. The variation of stromal exchange capacity in NaCl solutions of different concentrations was similar when detected by three independent procedures: stromal gel pressure measurements, intrastromal sodium ion distributions, and intrastromal electrical potentials. 5. Intrastromal chloride ion distributions were anomalous. Total chloride (measured by radio-isotopes) was consistently higher than that predicted by Donnan theory. 6. The data were consistent with Elliott's hypothesis that a fraction of intrastromal chloride ions bind to the corneal stromal matrix and in so doing contribute to the fixed negative charge of the stroma. 7. Our observations may be explained by a model of the cation exchange capacity of ox cornea which has two types of components. On is (at constant pH) invariant, and has a dry tissue exchange capacity of about 50 mequiv/kg dry tissue, and is probably generated by the sulphonic and carboxylic acid groups of the glycosaminoglycans. The other is explained by supposing it to consist of a chloride binding ligand which exhibits first order binding, is half occupied at ambient chloride concentrations of 300 mM, and has a total capacity of 240 mequiv/kg dry tissue. 8. Partial stromal extraction with 4 M-guanidine HCl indicated that the chloride binding ligand is not associated with the collagen molecules in the corneal stromal fibrils. 9. It is suggested that such a stromal chloride ion binding ligand would help to stabilize the hydration and transparency of the living cornea when it is exposed to environments of varying tonicity (such as in river or sea bathing). PMID:1432722

Hydrogen storage in engineered carbon nanospaces.

PubMed

Burress, Jacob; Kraus, Michael; Beckner, Matt; Cepel, Raina; Suppes, Galen; Wexler, Carlos; Pfeifer, Peter

2009-05-20

It is shown how appropriately engineered nanoporous carbons provide materials for reversible hydrogen storage, based on physisorption, with exceptional storage capacities (approximately 80 g H2/kg carbon, approximately 50 g H2/liter carbon, at 50 bar and 77 K). Nanopores generate high storage capacities (a) by having high surface area to volume ratios, and (b) by hosting deep potential wells through overlapping substrate potentials from opposite pore walls, giving rise to a binding energy nearly twice the binding energy in wide pores. Experimental case studies are presented with surface areas as high as 3100 m(2) g(-1), in which 40% of all surface sites reside in pores of width approximately 0.7 nm and binding energy approximately 9 kJ mol(-1), and 60% of sites in pores of width>1.0 nm and binding energy approximately 5 kJ mol(-1). The findings, including the prevalence of just two distinct binding energies, are in excellent agreement with results from molecular dynamics simulations. It is also shown, from statistical mechanical models, that one can experimentally distinguish between the situation in which molecules do (mobile adsorption) and do not (localized adsorption) move parallel to the surface, how such lateral dynamics affects the hydrogen storage capacity, and how the two situations are controlled by the vibrational frequencies of adsorbed hydrogen molecules parallel and perpendicular to the surface: in the samples presented, adsorption is mobile at 293 K, and localized at 77 K. These findings make a strong case for it being possible to significantly increase hydrogen storage capacities in nanoporous carbons by suitable engineering of the nanopore space.

An Intrinsic MicroRNA Timer Regulates Progressive Decline in Shoot Regenerative Capacity in Plants

PubMed Central

Zhang, Tian-Qi; Lian, Heng; Tang, Hongbo; Dolezal, Karel; Zhou, Chuan-Miao; Yu, Sha; Chen, Juan-Hua; Chen, Qi; Liu, Hongtao; Ljung, Karin

2015-01-01

Plant cells are totipotent and competent to regenerate from differentiated organs. It has been shown that two phytohormones, auxin and cytokinin, play critical roles within this process. As in animals, the regenerative capacity declines with age in plants, but the molecular basis for this phenomenon remains elusive. Here, we demonstrate that an age-regulated microRNA, miR156, regulates shoot regenerative capacity. As a plant ages, the gradual increase in miR156-targeted SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) transcription factors leads to the progressive decline in shoot regenerative capacity. In old plants, SPL reduces shoot regenerative capacity by attenuating the cytokinin response through binding with the B-type ARABIDOPSIS RESPONSE REGULATORs, which encode the transcriptional activators in the cytokinin signaling pathway. Consistently, the increased amount of exogenous cytokinin complements the reduced shoot regenerative capacity in old plants. Therefore, the recruitment of age cues in response to cytokinin contributes to shoot regenerative competence. PMID:25649435

Chemical modification of protein A chromatography ligands with polyethylene glycol. I: Effects on IgG adsorption equilibrium, kinetics, and transport.

PubMed

Weinberg, Justin; Zhang, Shaojie; Crews, Gillian; Carta, Giorgio; Przybycien, Todd

2018-04-20

Chemical modification of Protein A (ProA) chromatography ligands with polyethylene glycol (PEGylation) has been proposed as a strategy to increase the process selectivity and resin robustness by providing the ligand with a steric repulsion barrier against non-specific binding. This article comprises a comprehensive study of IgG adsorption and transport in Repligen CaptivA PriMAB resin with PEGylated ProA ligands that are modified using 5.2 and 21.5 kDa PEG chains. We studied the impact of the molecular weight of the PEG as well as the extent of PEGylation for the 5.2 kDa PEG modification. In all cases, PEGylation of ProA ligands decreases the resin average pore size, particle porosity, and static binding capacity for IgG proportional to the volume of conjugated PEG in the resin. Resin batch uptake experiments conducted in bulk via a stirred-tank system and with individual resin particles under confocal laser scanning microscopy suggests that PEGylation introduces heterogeneity into IgG binding kinetics: a fraction of the IgG binding sites are transformed from typical fast association kinetic behavior to slow kinetic behavior. pH gradient elution experiments of an IgG molecule on the modified resins show an increase in IgG elution pH for all modified resins, implying a decrease in IgG-ProA binding affinity on modification. Despite losses in static binding capacity for all resins with PEGylated ligands, the loss of dynamic binding capacity at 10% breakthrough (DBC 10% ) ranged more broadly from almost 0-47% depending on the PEG molecular weight and the extent of PEGylation. Minimal losses in DBC 10% were observed with a low extent of PEGylation with a smaller molecular weight PEG, while higher losses were observed at higher extents of PEGylation and with higher molecular weight PEG due to decreased static binding capacity and increased mass transfer resistance. This work provides insight into the practical implications for resin performance if PEGylation is considered as a strategy for selectivity enhancement in affinity chromatography with macromolecular ligands. Copyright © 2018 Elsevier B.V. All rights reserved.

An evaluation of ferrihydrite- and Metsorb™-DGT techniques for measuring oxyanion species (As, Se, V, P): effective capacity, competition and diffusion coefficients.

PubMed

Price, Helen L; Teasdale, Peter R; Jolley, Dianne F

2013-11-25

This study investigated several knowledge gaps with respect to the diffusive gradients in thin films (DGT) technique for measurement of oxyanions (As(III), As(V), Se(IV), Se(VI), PO4(3-), and V(V)) using the ferrihydrite and Metsorb™ binding layers. Elution efficiencies for each binding layer were higher with 1:20 dilutions, as analytical interferences for ICP-MS were minimised. Diffusion coefficients measured by diffusion cell and by DGT time-series experiments were found to agree well and generally agreed with previously reported values, although a range of diffusion coefficients have been reported for inorganic As and Se species. The relative binding affinity for both ferrihydrite and Metsorb™ was PO4(3-) ≈ As(V)>V(V) ≈ As(III)>Se(IV) > Se(VI) and effective binding capacities were measured in single ion solutions, and spiked synthetic freshwater and seawater, advising practical decisions about DGT monitoring. Under the conditions tested the performance of both ferrihydrite and Metsorb™ binding layers was directly comparable for As(V), As(III) Se(IV), V(V) and PO4(3-) over a deployment spanning ≤ 2 days for both freshwater and seawater. In order to return quantitative data for several analytes we recommend that the DGT method using either ferrihydrite or Metsorb™ be deployed for a maximum of 2 days in marine waters likely to contain high levels of the most strongly adsorbing oxyanions contaminants. The high pH, the competitive ions present in seawater and the identity of co-adsorbing ions affect the capacity of each binding layer for the analytes of interest. In freshwaters, longer deployment times can be considered but the concentration and identity of co-adsorbing ions may impact on quantitative uptake of Se(IV). This study found ferrihydrite-DGT outperformed Metsorb-DGT while previous studies have found the opposite, with variation in binding materials masses used being a likely reason. Clearly, preparation of both binding layers should always be optimised to produce the highest capacity possible, especially for seawater deployments. Crown Copyright © 2013. Published by Elsevier B.V. All rights reserved.

Inflammatory markers following acute fuel oil exposure or bacterial lipopolysaccharide in mallard ducks (Anas platyrhynchos).

PubMed

Lee, Kelly A; Tell, Lisa A; Mohr, F Charles

2012-12-01

Adult mallard ducks (Anas platyrhynchos) were orally dosed with bunker C fuel oil for 5 days, and five different inflammatory markers (haptoglobin, mannan-binding lectin, ceruloplasmin, unsaturated iron-binding capacity, and plasma iron) were measured in blood plasma prior to and 8, 24, 48, and 72 hr following exposure. In order to contrast the response to fuel oil with that of a systemic inflammatory response, an additional five ducks were injected intramuscularly with bacterial lipopolysaccharide (LPS). Oil-treated birds had an inflammatory marker profile that was significantly different from control and LPS-treated birds, showing decreases in mannan-binding lectin-dependent hemolysis and unsaturated iron-binding capacity, but no changes in any of the other inflammatory markers. Birds treated with oil also exhibited increased liver weights, decreased body and splenic weights, and decreased packed cell volume.

A comparative study for Hydrogen storage in metal decorated graphyne nanotubes and graphyne monolayers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lu, Jinlian; Guo, Yanhua; Zhang, Yun

A comparative study for hydrogen storage in metal decorated graphyne nanotubes and graphyne monolayers has been investigated within the framework of first-principle calculations. Our results show that the binding energies of Li, Ca, Sc, Ti on graphyne nanotubes are stronger than that on graphyne monolayers. Such strong binding would prevent the formation of metal clusters on graphyne nanotubes. From the charge transfer and partial density of states, it is found that the curvature effect of nanotubes plays an important role for the strong binding strength of metal on graphyne nanotubes. And the hydrogen storage capacity is 4.82 wt%, 5.08 wt%,more » 4.88 wt%, 4.76 wt% for Li, Ca, Sc, Ti decorated graphyne nanotubes that promise a potential material for storing hydrogen. - Graphical abstract: Metal atoms (Li, Ca, Sc and Ti) can strongly bind to graphyne nanotubes to avoid the formation of metal clusters, and a capacity of Ca@graphyne nanotube is 5.08 wt% which is close to the requirement of DOE in 2015. Twenty-four hydrogen molecules absorb to Ti-decorated graphyne nanotube. - Highlights: • The binding strength for metal on graphyne nanotubes is much stronger than that on γ-graphyne monolayer. • Metal atoms can strongly bind to the curving triangle acetylenes rings to avoid the formation of metal clusters. • A capacity of Ca@graphyne nanotube is 5.08 wt% which is close to the requirement of DOE in 2015.« less

Bio-nanocapsule-based scaffold improves the sensitivity and ligand-binding capacity of mammalian receptors on the sensor chip.

PubMed

Iijima, Masumi; Yoshimoto, Nobuo; Niimi, Tomoaki; Maturana, Andrés D; Kuroda, Shun'ichi

2016-06-01

Mammalian receptors are recognized as target molecules for drug discovery, and chemical libraries have been screened for both potential antagonists and agonists mainly by ligand-binding assays using immobilized receptors. A bio-nanocapsule (BNC) of approximately 30 nm that displays a tandem form of the protein A-derived immunoglobulin G (IgG) Fc-binding Z domains (denoted as ZZ-BNC) has been developed for both clustering and oriented immobilization of IgGs on the solid phase of immunosensors. In this study, human IgG1 Fc-fused vascular endothelial growth factor (VEGF) receptor was immobilized through ZZ-BNC on the sensor chip of quartz crystal microbalance (ZZ-BNC-coating). When compared with direct adsorption and protein A-coating, the sensor chip showed higher sensitivity (∽46- and ∽165-fold, respectively) and larger ligand-binding capacity (∽4- and ∽18-fold, respectively). Furthermore, the number of VEGF molecules bound to its receptor increased from 0.20 (direct adsorption) to 2.06 by ZZ-BNC-coating, strongly suggesting that ZZ-BNC reduced the steric hindrance near ligand recognition sites through oriented immobilization. Similarly, the sensitivity and ligand-binding capacity of leptin and prolactin receptors were both enhanced at a level comparable to that observed for the VEGF receptor. Thus, the combination of ZZ-BNC and Fc-fused receptors could significantly improve the function of ligand-binding assays. Copyright © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Comparing dissolved reactive phosphorus measured by DGT with ferrihydrite and titanium dioxide adsorbents: anionic interferences, adsorbent capacity and deployment time.

PubMed

Panther, Jared G; Teasdale, Peter R; Bennett, William W; Welsh, David T; Zhao, Huijun

2011-07-18

Two adsorbents (Metsorb and ferrihydrite) used in binding layers with the diffusive gradients in a thin film technique were evaluated for the measurement of dissolved reactive phosphorous (DRP) in synthetic and natural waters. Possible interferences were investigated with Cl(-) (up to 1.35 mol L(-1)) and SO(4)(2-) (up to 0.056 mol L(-1)) having no affect on either DGT binding layer, and HCO(3)(-) (up to 5.7 mmol L(-1)) having no effect on Metsorb-DGT, over 4 days. However, HCO(3)(-) interfered with the ferrihydrite-DGT measurement at concentrations typical of many natural waters (≥0.7 mmol L(-1)) after a deployment period of 1-2 days. The capacity of the Metsorb binding phase for DGT response was ∼37,000 ng P, whereas the capacities of a low-mass (17.8 mg of adsorbent per DGT sampler) and high-mass (29.2mg of adsorbent per DGT sampler) ferrihydrite binding phase were substantially lower (∼15,000 ng P and ∼25,000 ng P, low-mass and high-mass, respectively). Increasing the capacity of the ferrihydrite adsorbent allowed the ferrihydrite-DGT to be utilized for up to 3 days before interference by HCO(3)(-) was observed. Seawater deployments demonstrated that even high-capacity ferrihydrite-DGT devices underestimated the DRP concentration by 37%, whereas Metsorb-DGT measurements were accurate. The Metsorb-DGT is superior to the ferrihydrite-DGT for determining DRP over deployment times greater than 1 day and in waters with ≥0.7 mmol L(-1) HCO(3)(-). Based on the experience obtained from this detailed validation process, the authors propose a number of key requirements that need to be considered when developing new DGT binding layers, with testing the performance over longer deployment times being critical. Copyright © 2011 Elsevier B.V. All rights reserved.

The Effect of Hydraulic Loading Rate and Influent Source on the Binding Capacity of Phosphorus Filters

PubMed Central

Herrmann, Inga; Jourak, Amir; Hedström, Annelie; Lundström, T. Staffan; Viklander, Maria

2013-01-01

Sorption by active filter media can be a convenient option for phosphorus (P) removal and recovery from wastewater for on-site treatment systems. There is a need for a robust laboratory method for the investigation of filter materials to enable a reliable estimation of their longevity. The objectives of this study were to (1) investigate and (2) quantify the effect of hydraulic loading rate and influent source (secondary wastewater and synthetic phosphate solution) on P binding capacity determined in laboratory column tests and (3) to study how much time is needed for the P to react with the filter material (reaction time). To study the effects of these factors, a 22 factorial experiment with 11 filter columns was performed. The reaction time was studied in a batch experiment. Both factors significantly (α = 0.05) affected the P binding capacity negatively, but the interaction of the two factors was not significant. Increasing the loading rate from 100 to 1200 L m−2 d−1 decreased P binding capacity from 1.152 to 0.070 g kg−1 for wastewater filters and from 1.382 to 0.300 g kg−1 for phosphate solution filters. At a loading rate of 100 L m−2 d−1, the average P binding capacity of wastewater filters was 1.152 g kg−1 as opposed to 1.382 g kg−1 for phosphate solution filters. Therefore, influent source or hydraulic loading rate should be carefully controlled in the laboratory. When phosphate solution and wastewater were used, the reaction times for the filters to remove P were determined to be 5 and 15 minutes, respectively, suggesting that a short residence time is required. However, breakthrough in this study occurred unexpectedly quickly, implying that more time is needed for the P that has reacted to be physically retained in the filter. PMID:23936313

Feature Binding in Visual Working Memory Evaluated by Type Identification Paradigm

ERIC Educational Resources Information Center

Saiki, Jun; Miyatsuji, Hirofumi

2007-01-01

Memory for feature binding comprises a key ingredient in coherent object representations. Previous studies have been equivocal about human capacity for objects in the visual working memory. To evaluate memory for feature binding, a type identification paradigm was devised and used with a multiple-object permanence tracking task. Using objects…

Binding of environmental carcinogens to asbestos and mineral fibres.

PubMed Central

Harvey, G; Pagé, M; Dumas, L

1984-01-01

A rapid method has been developed for measuring the binding capacity of asbestos and other mineral fibres for environmental carcinogens. Benzo(alpha)pyrene (B(alpha)P), nitrosonornicotine (NNN), and N-acetyl-2-aminofluorene (NAAF) were assayed in the presence of Canadian grade 4T30 chrysotile, chrysotile A, amosite, crocidolite, glass microfibres, glasswool, attapulgite, and titanium dioxide. Chrysotile binds significantly more carcinogens than the other mineral fibres. This binding assay is reproducible with coefficients of variation of less than 8% and 6% respectively for inter and intra assay. The influence of pH was also studied, and there is good correlation between the carcinogen binding and the charge of the tested mineral fibres. The in vitro cytotoxicity on macrophage like cell line P388D1 and the haemolytic activity of various mineral fibres were also measured; a good correlation was found between the binding capacity and the cytotoxicity of tested mineral fibres on P388D1 cells. These results give some explanations for the reported synergism between exposure to asbestos and the smoking habits of workers. PMID:6331497

Binding of carbonyl flavours to canola, pea and wheat proteins using GC/MS approach.

PubMed

Wang, Kun; Arntfield, Susan D

2014-08-15

Interactions of homologous aldehydes (hexanal, heptanal, and octanal) and ketones (2-hexanone, 2-heptanone, and 2-octanone) to salt and alkaline-extracted canola and pea proteins and commercial wheat gluten were studied using GC/MS. Long-chain aldehyde flavours exhibited higher binding affinity, regardless of protein type and isolation method. Salt-extracted canola protein isolates (CPIs) revealed the highest binding capacity to all aldehydes followed by wheat gluten and salt-extracted pea protein isolates (PPIs), while binding of ketone flavours decreased in the order: PPIs>wheat gluten>CPIs. Two aldolisation products, 2-butyl-2-octenal and 2-pentyl-2-nonenal, were detected from the interactions between CPIs with hexanal and heptanal, respectively. Protein thermal behaviour in the presence of these compounds was analysed by differential scanning calorimeter, where decreased ΔH inferred potential conformational changes due to partial denaturation of PPIs. Compared to ketones, aldehyde flavours possessed much higher "unfolding capacity" (lower ΔH), which accounted for their higher binding affinities. Copyright © 2014 Elsevier Ltd. All rights reserved.

Study of the role of bran water binding and the steric hindrance by bran in straight dough bread making.

PubMed

Hemdane, S; Langenaeken, N A; Jacobs, P J; Verspreet, J; Delcour, J A; Courtin, C M

2018-07-01

This study investigates the effect of the physical presence and water binding of wheat bran during bread making, and the possible mechanisms behind this effect. Regular bran, pericarp-enriched bran and synthetic bran-like particles with different water binding capacities and particle sizes were used. Incorporation of regular and pericarp-enriched bran in dough (15% dm) led to a lower oven rise than the control dough. Bread volumes decreased with 11% and 30%, respectively. Dough with synthetic bran, having a low water binding capacity, displayed a near to normal leavening and oven rise and resulted in a bread volume decrease of only 5% compared to the control. Particle size reduction of regular bran and synthetic bran to an average size of 200 µm did not affect final bread quality. Results indicate that water binding by bran affects bread quality the most, whereas steric hindrance by physical presence of bran particles is less determinative. Copyright © 2018 Elsevier Ltd. All rights reserved.

Pharmaceutical-grade albumin: impaired drug-binding capacity in vitro

PubMed Central

Olsen, Harald; Andersen, Anders; Nordbø, Arve; Kongsgaard, Ulf E; Børmer, Ole P

2004-01-01

Background Albumin is the most abundant protein in blood plasma, and due to its ligand binding properties, serves as a circulating depot for endogenous and exogenous (e.g. drugs) compounds. Hence, the unbound drug is the pharmacologically active drug. Commercial human albumin preparations are frequently used during surgery and in critically ill patients. Recent studies have indicated that the use of pharmaceutical-grade albumin is controversial in critically ill patients. In this in vitro study we investigated the drug binding properties of pharmaceutical-grade albumins (Baxter/Immuno, Octapharma, and Pharmacia & Upjohn), native human serum, and commercially available human serum albumin from Sigma Chemical Company. Methods The binding properties of the various albumin solutions were tested in vitro by means of ultrafiltration. Naproxen, warfarin, and digitoxin were used as ligands. HPLC was used to quantitate the total and free drug concentrations. The data were fitted to a model of two classes of binding sites for naproxen and warfarin and one class for digitoxin, using Microsoft Excel and Graphpad Prism. Results The drugs were highly bound to albumin (95–99.5%). The highest affinity (lowest K1) was found with naproxen. Pharmaceutical-grade albumin solutions displayed significantly lower drug-binding capacity compared to native human serum and Sigma albumin. Thus, the free fraction was considerably higher, approximately 40 times for naproxen and 5 and 2 times for warfarin and digitoxin, respectively. The stabilisers caprylic acid and N-acetyl-DL-tryptophan used in the manufacturing procedure seem to be of importance. Adding the stabilisers to human serum and Sigma albumin reduced the binding affinity whereas charcoal treatment of the pharmaceutical-grade albumin from Octapharma almost restored the specific binding capacity. Conclusion This in vitro study demonstrates that the specific binding for warfarin and digitoxin is significantly reduced and for naproxen no longer detectable in pharmaceutical-grade albumin. It further shows that the addition of stabilisers may be of major importance for this effect. PMID:15046641

Potential Functional Byproducts from Guava Purée Processing.

PubMed

Lim, Si Yi; Tham, Paik Yean; Lim, Hilary Yi Ler; Heng, Wooi Shin; Chang, Ying Ping

2018-05-10

The valorization of guava waste requires compositional and functional studies. We tested three byproducts of guava purée processing, namely refiner, siever, and decanter. We analyzed the chemical composition and quantified the prebiotic activity score and selected carbohydrates; we also determined the water holding (WHC), oil holding (OHC), cation exchange capacities, bile acid binding, and glucose dialysis retardation (GDR) of the solid fraction and the antioxidative and α-amylase inhibitory capacities (AIC) of the ethanolic extract. Refiner contained 7.7% lipid, 7.08% protein and a relatively high phytate content; it had a high prebiotic activity score and possessed the highest binding capacity with deoxycholic acid. Siever contained high levels of low molecular weight carbohydrates and total tannin but relatively low crude fiber and cellulose contents. It had the highest binding with chenodeoxycholic acid (74.8%), and exhibited the highest 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging capacity. Decanter was rich in cellulose and had a high prebiotic activity score. The WHC and OHC values of decanter were within a narrow range and also exhibited the highest binding with cholic acid (86.6%), and the highest values of GDR and AIC. The refiner waste could be included in animal feed but requires further processing to reduce the high phytate levels. All three guava byproducts had the potential to be a source of antioxidant dietary fiber (DF), a finding that warrants further in vivo study. To differing extents, the guava byproducts exhibited useful physicochemical binding properties and so possessed the potential for health-promoting activity. These byproducts could also be upgraded to other marketable products so the manufacturers of processed guava might be able to develop their businesses sustainably by making better use of them. © 2018 Institute of Food Technologists®.

High-Affinity Low-Capacity and Low-Affinity High-Capacity N-Acetyl-2-Aminofluorene (AAF) Macromolecular Binding Sites Are Revealed During the Growth Cycle of Adult Rat Hepatocytes in Primary Culture.

PubMed

Koch, Katherine S; Moran, Tom; Shier, W Thomas; Leffert, Hyam L

2018-05-01

Long-term cultures of primary adult rat hepatocytes were used to study the effects of N-acetyl-2-aminofluorene (AAF) on hepatocyte proliferation during the growth cycle; on the initiation of hepatocyte DNA synthesis in quiescent cultures; and, on hepatocyte DNA replication following the initiation of DNA synthesis. Scatchard analyses were used to identify the pharmacologic properties of radiolabeled AAF metabolite binding to hepatocyte macromolecules. Two classes of growth cycle-dependent AAF metabolite binding sites-a high-affinity low-capacity site (designated Site I) and a low-affinity high-capacity site (designated Site II)-associated with two spatially distinct classes of macromolecular targets, were revealed. Based upon radiolabeled AAF metabolite binding to purified hepatocyte genomic DNA or to DNA, RNA, proteins, and lipids from isolated nuclei, Site IDAY 4 targets (KD[APPARENT] ≈ 2-4×10-6 M and BMAX[APPARENT] ≈ 6 pmol/106 cells/24 h) were consistent with genomic DNA; and with AAF metabolized by a nuclear cytochrome P450. Based upon radiolabeled AAF binding to total cellular lysates, Site IIDAY 4 targets (KD[APPARENT] ≈ 1.5×10-3 M and BMAX[APPARENT] ≈ 350 pmol/106 cells/24 h) were consistent with cytoplasmic proteins; and with AAF metabolized by cytoplasmic cytochrome P450s. DNA synthesis was not inhibited by concentrations of AAF that saturated DNA binding in the neighborhood of the Site I KD. Instead, hepatocyte DNA synthesis inhibition required higher concentrations of AAF approaching the Site II KD. These observations raise the possibility that carcinogenic DNA adducts derived from AAF metabolites form below concentrations of AAF that inhibit replicative and repair DNA synthesis.

Construction of a Functional S-Layer Fusion Protein Comprising an Immunoglobulin G-Binding Domain for Development of Specific Adsorbents for Extracorporeal Blood Purification

PubMed Central

Völlenkle, Christine; Weigert, Stefan; Ilk, Nicola; Egelseer, Eva; Weber, Viktoria; Loth, Fritz; Falkenhagen, Dieter; Sleytr, Uwe B.; Sára, Margit

2004-01-01

The chimeric gene encoding a C-terminally-truncated form of the S-layer protein SbpA from Bacillus sphaericus CCM 2177 and two copies of the Fc-binding Z-domain was constructed, cloned, and heterologously expressed in Escherichia coli HMS174(DE3). The Z-domain is a synthetic analogue of the B-domain of protein A, capable of binding the Fc part of immunoglobulin G (IgG). The S-layer fusion protein rSbpA31-1068/ZZ retained the specific properties of the S-layer protein moiety to self-assemble in suspension and to recrystallize on supports precoated with secondary cell wall polymer (SCWP), which is the natural anchoring molecule for the S-layer protein in the bacterial cell wall. Due to the construction principle of the S-layer fusion protein, the ZZ-domains remained exposed on the outermost surface of the protein lattice. The binding capacity of the native or cross-linked monolayer for human IgG was determined by surface plasmon resonance measurements. For batch adsorption experiments, 3-μm-diameter, biocompatible cellulose-based, SCWP-coated microbeads were used for recrystallization of the S-layer fusion protein. In the case of the native monolayer, the binding capacity for human IgG was 5.1 ng/mm2, whereas after cross-linking with dimethyl pimelimidate, 4.4 ng of IgG/mm2 was bound. This corresponded to 78 and 65% of the theoretical saturation capacity of a planar surface for IgGs aligned in the upright position, respectively. Compared to commercial particles used as immunoadsorbents to remove autoantibodies from sera of patients suffering from an autoimmune disease, the IgG binding capacity of the S-layer fusion protein-coated microbeads was at least 20 times higher. For that reason, this novel type of microbeads should find application in the microsphere-based detoxification system. PMID:15006773

Human adenovirus serotypes 4p and 11p are efficiently expressed in cell lines of neural tumour origin.

PubMed

Skog, Johan; Mei, Ya-Fang; Wadell, Göran

2002-06-01

Most currently used adenovirus vectors are based upon adenovirus serotypes 2 and 5 (Ad2 and Ad5), which have limited efficiencies for gene transfer to human neural cells. Both serotypes bind to the known adenovirus receptor, CAR (coxsackievirus and adenovirus receptor), and have restricted cell tropism. The purpose of this study was to find vector candidates that are superior to Ad5 in infecting human neural tumours. Using flow cytometry, the vector candidates Ad4p, Ad11p and Ad17p were compared to the commonly used adenovirus vector Ad5v for their binding capacity to neural cell lines derived from glioblastoma, medulloblastoma and neuroblastoma cell lines. The production of viral structural proteins and the CAR-binding properties of the different serotypes were also assessed in these cells. Computer-based models of the fibre knobs of Ad4p and Ad17 were created based upon the crystallized fibre knob structure of adenoviruses and analysed for putative receptor-interacting regions that differed from the fibre knob of Ad5. The non CAR-binding vector candidate Ad11p showed clearly the best binding capacity to all of the neural cell lines, binding more than 90% of cells of all of the neural cell lines tested, in contrast to 20% or less for the commonly used vector Ad5v. Ad4p and Ad11p were also internalized and produced viral proteins more successfully than Ad5. Ad4p showed a low binding ability but a very efficient capacity for infection in cell culture. Ad17p virions neither bound or efficiently infected any of the neural cell lines studied.

Newborn Jaundice Technologies: Unbound Bilirubin and Bilirubin Binding Capacity In Neonates

PubMed Central

Amin, Sanjiv B.; Lamola, Angelo A.

2011-01-01

Neonatal jaundice (hyperbilirubinemia), extremely common in neonates, can be associated with neurotoxicity. A safe level of bilirubin has not been defined in either premature or term infants. Emerging evidence suggest that the level of unbound (or “free”) bilirubin has a better sensitivity and specificity than total serum bilirubin for bilirubin-induced neurotoxicity. Although recent studies suggest the usefulness of free bilirubin measurements in managing high-risk neonates including premature infants, there currently exists no widely available method to assay the serum free bilirubin concentration. To keep pace with the growing demand, in addition to reevaluation of old methods, several promising new methods are being developed for sensitive, accurate, and rapid measurement of free bilirubin and bilirubin binding capacity. These innovative methods need to be validated before adopting for clinical use. We provide an overview of some promising methods for free bilirubin and binding capacity measurements with the goal to enhance research in this area of active interest and apparent need. PMID:21641486

Estimating Prion Adsorption Capacity of Soil by BioAssay of Subtracted Infectivity from Complex Solutions (BASICS)

PubMed Central

Wyckoff, A. Christy; Lockwood, Krista L.; Meyerett-Reid, Crystal; Michel, Brady A.; Bender, Heather; VerCauteren, Kurt C.; Zabel, Mark D.

2013-01-01

Prions, the infectious agent of scrapie, chronic wasting disease and other transmissible spongiform encephalopathies, are misfolded proteins that are highly stable and resistant to degradation. Prions are known to associate with clay and other soil components, enhancing their persistence and surprisingly, transmissibility. Currently, few detection and quantification methods exist for prions in soil, hindering an understanding of prion persistence and infectivity in the environment. Variability in apparent infectious titers of prions when bound to soil has complicated attempts to quantify the binding capacity of soil for prion infectivity. Here, we quantify the prion adsorption capacity of whole, sandy loam soil (SLS) typically found in CWD endemic areas in Colorado; and purified montmorillonite clay (Mte), previously shown to bind prions, by BioAssay of Subtracted Infectivity in Complex Solutions (BASICS). We incubated prion positive 10% brain homogenate from terminally sick mice infected with the Rocky Mountain Lab strain of mouse-adapted prions (RML) with 10% SLS or Mte. After 24 hours samples were centrifuged five minutes at 200×g and soil-free supernatant was intracerebrally inoculated into prion susceptible indicator mice. We used the number of days post inoculation to clinical disease to calculate the infectious titer remaining in the supernatant, which we subtracted from the starting titer to determine the infectious prion binding capacity of SLS and Mte. BASICS indicated SLS bound and removed ≥ 95% of infectivity. Mte bound and removed lethal doses (99.98%) of prions from inocula, effectively preventing disease in the mice. Our data reveal significant prion-binding capacity of soil and the utility of BASICS to estimate prion loads and investigate persistence and decomposition in the environment. Additionally, since Mte successfully rescued the mice from prion disease, Mte might be used for remediation and decontamination protocols. PMID:23484043

Estimating prion adsorption capacity of soil by BioAssay of Subtracted Infectivity from Complex Solutions (BASICS).

PubMed

Wyckoff, A Christy; Lockwood, Krista L; Meyerett-Reid, Crystal; Michel, Brady A; Bender, Heather; VerCauteren, Kurt C; Zabel, Mark D

2013-01-01

Prions, the infectious agent of scrapie, chronic wasting disease and other transmissible spongiform encephalopathies, are misfolded proteins that are highly stable and resistant to degradation. Prions are known to associate with clay and other soil components, enhancing their persistence and surprisingly, transmissibility. Currently, few detection and quantification methods exist for prions in soil, hindering an understanding of prion persistence and infectivity in the environment. Variability in apparent infectious titers of prions when bound to soil has complicated attempts to quantify the binding capacity of soil for prion infectivity. Here, we quantify the prion adsorption capacity of whole, sandy loam soil (SLS) typically found in CWD endemic areas in Colorado; and purified montmorillonite clay (Mte), previously shown to bind prions, by BioAssay of Subtracted Infectivity in Complex Solutions (BASICS). We incubated prion positive 10% brain homogenate from terminally sick mice infected with the Rocky Mountain Lab strain of mouse-adapted prions (RML) with 10% SLS or Mte. After 24 hours samples were centrifuged five minutes at 200 × g and soil-free supernatant was intracerebrally inoculated into prion susceptible indicator mice. We used the number of days post inoculation to clinical disease to calculate the infectious titer remaining in the supernatant, which we subtracted from the starting titer to determine the infectious prion binding capacity of SLS and Mte. BASICS indicated SLS bound and removed ≥ 95% of infectivity. Mte bound and removed lethal doses (99.98%) of prions from inocula, effectively preventing disease in the mice. Our data reveal significant prion-binding capacity of soil and the utility of BASICS to estimate prion loads and investigate persistence and decomposition in the environment. Additionally, since Mte successfully rescued the mice from prion disease, Mte might be used for remediation and decontamination protocols.

Dissecting the link between the enzymatic activity and the SaPI inducing capacity of the phage 80α dUTPase.

PubMed

Alite, Christian; Humphrey, Suzanne; Donderis, Jordi; Maiques, Elisa; Ciges-Tomas, J Rafael; Penadés, José R; Marina, Alberto

2017-09-11

The trimeric staphylococcal phage-encoded dUTPases (Duts) are signalling molecules that induce the cycle of some Staphylococcal pathogenicity islands (SaPIs) by binding to the SaPI-encoded Stl repressor. To perform this regulatory role, these Duts require an extra motif VI, as well as the Dut conserved motifs IV and V. While the apo form of Dut is required for the interaction with the Stl repressor, usually only those Duts with normal enzymatic activity can induce the SaPI cycle. To understand the link between the enzymatic activities and inducing capacities of the Dut protein, we analysed the structural, biochemical and physiological characteristics of the Dut80α D95E mutant, which loses the SaPI cycle induction capacity despite retaining enzymatic activity. Asp95 is located at the threefold central channel of the trimeric Dut where it chelates a divalent ion. Here, using state-of-the-art techniques, we demonstrate that D95E mutation has an epistatic effect on the motifs involved in Stl binding. Thus, ion binding in the central channel correlates with the capacity of motif V to twist and order in the SaPI-inducing disposition, while the tip of motif VI is disturbed. These alterations in turn reduce the affinity for the Stl repressor and the capacity to induce the SaPI cycle.

Modulation of FadR Binding Capacity for Acyl-CoA Fatty Acids Through Structure-Guided Mutagenesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bacik, John-Paul; Yeager, Chris M.; Twary, Scott N.

FadR is a versatile global regulator in Escherichia coli that controls fatty acid metabolism and thereby modulates the ability of this bacterium to grow using fatty acids or acetate as the sole carbon source. FadR regulates fatty acid metabolism in response to intra-cellular concentrations of acyl-CoA lipids. The ability of FadR to bind acyl-CoA fatty acids is hence of significant interest for the engineering of biosynthetic pathways for the production of lipid-based biofuels and commodity chemicals. Based on the available crystal structure of E. coli bound to myristoyl- CoA, we predicted amino acid positions within the effector binding pocket thatmore » would alter the ability of FadR to bind acyl-CoA fatty acids without affecting DNA binding. We utilized fluorescence polarization to characterize the in-vitro binding properties of wild type and mutant FadR. We found that a Leu102Ala mutant enhanced binding of the effector, likely by increasing the size of the binding pocket for the acyl moiety of the molecule. Conversely, the elimination of the guanidine side chain (Arg213Ala and Arg213Met mutants) of the CoA moiety binding site severely diminished the ability of FadR to bind the acyl-CoA effector. These results demonstrate the ability to fine tune FadR binding capacity. The validation of an efficient method to fully characterize all the binding events involved in the specific activity (effector and DNA operator binding) of FadR has allowed us to increase our understanding of the role of specific amino acids in the binding and recognition of acyl-CoA fatty acids and will greatly facilitate efforts aimed at engineering tunable FadR regulators for synthetic biology.« less

Modulation of FadR Binding Capacity for Acyl-CoA Fatty Acids Through Structure-Guided Mutagenesis

DOE PAGES

Bacik, John-Paul; Yeager, Chris M.; Twary, Scott N.; ...

2015-09-18

FadR is a versatile global regulator in Escherichia coli that controls fatty acid metabolism and thereby modulates the ability of this bacterium to grow using fatty acids or acetate as the sole carbon source. FadR regulates fatty acid metabolism in response to intra-cellular concentrations of acyl-CoA lipids. The ability of FadR to bind acyl-CoA fatty acids is hence of significant interest for the engineering of biosynthetic pathways for the production of lipid-based biofuels and commodity chemicals. Based on the available crystal structure of E. coli bound to myristoyl- CoA, we predicted amino acid positions within the effector binding pocket thatmore » would alter the ability of FadR to bind acyl-CoA fatty acids without affecting DNA binding. We utilized fluorescence polarization to characterize the in-vitro binding properties of wild type and mutant FadR. We found that a Leu102Ala mutant enhanced binding of the effector, likely by increasing the size of the binding pocket for the acyl moiety of the molecule. Conversely, the elimination of the guanidine side chain (Arg213Ala and Arg213Met mutants) of the CoA moiety binding site severely diminished the ability of FadR to bind the acyl-CoA effector. These results demonstrate the ability to fine tune FadR binding capacity. The validation of an efficient method to fully characterize all the binding events involved in the specific activity (effector and DNA operator binding) of FadR has allowed us to increase our understanding of the role of specific amino acids in the binding and recognition of acyl-CoA fatty acids and will greatly facilitate efforts aimed at engineering tunable FadR regulators for synthetic biology.« less

Relating saturation capacity to charge density in strong cation exchangers.

PubMed

Steinebach, Fabian; Coquebert de Neuville, Bertrand; Morbidelli, Massimo

2017-07-21

In this work the relation between physical and chemical resin characteristics and the total amount of adsorbed protein (saturation capacity) for ion-exchange resins is discussed. Eleven different packing materials with a sulfo-functionalization and one multimodal resin were analyzed in terms of their porosity, pore size distribution, ligand density and binding capacity. By specifying the ligand density and binding capacity by the total and accessible surface area, two different groups of resins were identified: Below a ligand density of approx. 2.5μmol/m 2 area the ligand density controls the saturation capacity, while above this limit the accessible surface area becomes the limiting factor. This results in a maximum protein uptake of around 2.5mg/m 2 of accessible surface area. The obtained results allow estimating the saturation capacity from independent resin characteristics like the saturation capacity mainly depends on "library data" such as the accessible and total surface area and the charge density. Hence these results give an insight into the fundamentals of protein adsorption and help to find suitable resins, thus limiting the experimental effort in early process development stages. Copyright © 2017 Elsevier B.V. All rights reserved.

Comparison of the fibronectin-binding ability and antitumor efficacy of various mycobacteria.

PubMed

Hudson, M A; Ritchey, J K; Catalona, W J; Brown, E J; Ratliff, T L

1990-07-01

Although the mechanism by which Bacillus Calmette-Guerin (BCG) exerts an antitumor effect on superficial bladder tumors is not fully understood, recent evidence has implicated binding of BCG organisms to fibronectin (FN) as requisite for this antitumor efficacy. Various substrains of BCG and other mycobacteria were tested in vitro for their relative capacities to bind both matrix and soluble FN. A substrain of Mycobacterium kansasii, designated the "high-binding strain," was found to bind FN more readily (P less than 0.05) in in vitro studies, when compared to commercially available substrains of BCG (Tice, Connaught, and Armand Frappier). The binding by the three commercial strains of BCG to FN in vitro appeared to be equivalent. The high-binding strain was further demonstrated to attach more readily in vivo to the acutely injured murine bladder (P less than 0.005) than the Armand Frappier substrain. Finally, using the MB49 murine bladder tumor model, an enhanced antitumor effect (P less than 0.05) was noted in mice treated with intravesical high-binding strain, in comparison to the Armand Frappier substrain, during five weekly treatments. It appears not only that the commercial substrains of BCG bind FN in an equivalent manner but also that the relative binding capacities of the substrains correlate directly with antitumor activity. A substrain of M. kansasii appears to have been identified which may prove more clinically effective than the currently available strains of BCG.

Kinetic and Thermodynamic Analyses of Interaction between a High-Affinity RNA Aptamer and Its Target Protein.

PubMed

Amano, Ryo; Takada, Kenta; Tanaka, Yoichiro; Nakamura, Yoshikazu; Kawai, Gota; Kozu, Tomoko; Sakamoto, Taiichi

2016-11-15

AML1 (RUNX1) protein is an essential transcription factor involved in the development of hematopoietic cells. Several genetic aberrations that disrupt the function of AML1 have been frequently observed in human leukemia. AML1 contains a DNA-binding domain known as the Runt domain (RD), which recognizes the RD-binding double-stranded DNA element of target genes. In this study, we identified high-affinity RNA aptamers that bind to RD by systematic evolution of ligands by exponential enrichment. The binding assay using surface plasmon resonance indicated that a shortened aptamer retained the ability to bind to RD when 1 M potassium acetate was used. A thermodynamic study using isothermal titration calorimetry (ITC) showed that the aptamer-RD interaction is driven by a large enthalpy change, and its unfavorable entropy change is compensated by a favorable enthalpy change. Furthermore, the binding heat capacity change was identified from the ITC data at various temperatures. The aptamer binding showed a large negative heat capacity change, which suggests that a large apolar surface is buried upon such binding. Thus, we proposed that the aptamer binds to RD with long-range electrostatic force in the early stage of the association and then changes its conformation and recognizes a large surface area of RD. These findings about the biophysics of aptamer binding should be useful for understanding the mechanism of RNA-protein interaction and optimizing and modifying RNA aptamers.

[EFFICIENCY OF HAEMOGLOBIN REGENERATION IN THE NUTRITIONAL FERROPENIC ANAEMIA RECOVERY WITH GOAT MILK-BASED DIETS].

PubMed

Serrano Reina, José Antonio; Nestares Pleguezuelo, Teresa; Muñoz Alférez, Ma José; Díaz Castro, Javier; López Aliaga, Ma Inmaculada

2015-10-01

in spite of the high incidence/prevalence of iron deficiency anemia (IDA) and the beneficial effects derived from the consumption of goat milk, scarce is known about the recovery of the anemia following a balanced diet accompanied by the intake of goat milk of goat. The aim of the current study is to assess, in rats with experimentally induced nutritional iron deficiency anemia, the effects of goat or cow milk-based diets, supplied during 30 days, on the recovery of the anemia and the efficiency of regeneration of the hemoglobin. 40 male Wistar albino rats newly weaned were divided at random in two experimental groups and they were fed ad libitum for 40 days with AIN-93G diet, either with normal iron content (control group, 45 mg/kg diet), or low iron content (anaemic group, 5 mg/kg diet). Samples of blood form the caudal vein were collected for the hematologic control of the anemia. Later, both experimental groups (control and iron deficient) were fed for 30 days with goat or cow milk- based diets. After finishing the experimental period and previous anesthesia the animals were withdrawn by canulation of the abdominal aorta, and the obtained blood was gathered in tubes with EDTA as anticoagulant for the later determination of hematologic parameters and the efficiency of regeneration of the hemoglobin. after the consumption of a diet with low iron content during 40 days, the rats were anaemic, with a concentration of hemoglobin, hematocrit, serum iron, mean corpuscular volume (MCV), serum ferritin and low transferrin (p < 0.001), whereas the levels of platelets and the total iron binding capacity (TIBC) were raised (p < 0.001), findings consistent with the anemia induced experimentally in the animals. The efficiency of regeneration of the hemoglobin was higher in control and anaemic rats fed goat milk-based diet in comparison with those fed cow milk-based diet (p < 0.001) due to, partly, to the major levels of serum iron and hemoglobin, and to the best nutritive utilization of iron in the animals that consumed the goat milk-based diet thanks to the excellent nutritional characteristics of this type of milk. the consumption during 30 days of goat or cow milk-based diets favors the recovery of the iron deficiency anemia, especially with the goat milk, due to the major efficiency of regeneration of the hemoglobin, index that shows the quantity of iron of the diet used for the synthesis of hemoglobin. Therefore, it would be recommendable the consumption of goat milk in the context of a balanced diet in healthy populations and, especially in those at risk of suffering iron deficiency. Copyright AULA MEDICA EDICIONES 2014. Published by AULA MEDICA. All rights reserved.

Effect of chain length on binding of fatty acids to Pluronics in microemulsions.

PubMed

James-Smith, Monica A; Shekhawat, Dushyant; Cheung, Sally; Moudgil, Brij M; Shah, Dinesh O

2008-03-15

We investigated the effect of fatty acid chain length on the binding capacity of drug and fatty acid to Pluronic F127-based microemulsions. This was accomplished by using turbidity experiments. Pluronic-based oil-in-water microemulsions of various compositions were synthesized and titrated to turbidity with concentrated Amitriptyline, an antidepressant drug. Sodium salts of C(8), C(10), or C(12) fatty acid were used in preparation of the microemulsion and the corresponding binding capacities were observed. It has been previously determined that, for microemulsions prepared with sodium caprylate (C(8) fatty acid soap), a maximum of 11 fatty acid molecules bind to the microemulsion per 1 molecule of Pluronic F127 and a maximum of 12 molecules of Amitriptyline bind per molecule of F127. We have found that with increasing the chain length of the fatty acid salt component of the microemulsion, the binding capacity of both the fatty acid and the Amitriptyline to the microemulsion decreases. For sodium salts of C(8), C(10) and C(12) fatty acids, respectively, a maximum of approximately 11, 8.4 and 8.3 molecules of fatty acid molecules bind to 1 Pluronic F127 molecule. We propose that this is due to the decreasing number of free monomers with increasing chain length. As chain length increases, the critical micelle concentration (cmc) decreases, thus leading to fewer monomers. Pluronics are symmetric tri-block copolymers consisting of propylene oxide (PO) and ethylene oxide (EO). The polypropylene oxide block, PPO is sandwiched between two polyethylene oxide (PEO) blocks. The PEO blocks are hydrophilic while PPO is hydrophobic portion in the Pluronic molecule. Due to this structure, we propose that the fatty acid molecules that are in monomeric form most effectively diffuse between the PEO "tails" and bind to the hydrophobic PPO groups.

[Fiber in the diet--certainties and speculation].

PubMed

Peters, P; Peters, K M

1988-06-01

This report defines dietary fibre and summarizes its effects on dental, gastrointestinal and metabolic diseases. A higher intake of dietary fibre is important in prophylaxis of caries, paradentosis, constipation, diverticulosis, colon cancer, diabetes and hypercholesteraemia. An ideal preparation must have the following abilities: It should be coarse, hard and swallowable and without cariogenic sugars in order to prevent dental diseases. It should be a mixture of several kinds of fibre getting water binding capacity and bile acid binding capacity. Mechanical crushing and heatening of fibre are to be avoided. The preparation should not contain phytic acid.

Bilirubin Binding Capacity in the Preterm Neonate

PubMed Central

Amin, Sanjiv B

2016-01-01

SYNOPSIS Total serum/plasma bilirubin (TB), the biochemical measure currently used to evaluate and manage hyperbilirubinemia, is not a useful predictor of bilirubin-induced neurotoxicity in premature infants. Altered bilirubin-albumin binding in premature infants limits the usefulness of TB in premature infants. In this article, bilirubin-albumin binding, a modifying factor for bilirubin-induced neurotoxicity, in premature infants is reviewed. PMID:27235205

Affinity binding of antibodies to supermacroporous cryogel adsorbents with immobilized protein A for removal of anthrax toxin protective antigen.

PubMed

Ingavle, Ganesh C; Baillie, Les W J; Zheng, Yishan; Lis, Elzbieta K; Savina, Irina N; Howell, Carol A; Mikhalovsky, Sergey V; Sandeman, Susan R

2015-05-01

Polymeric cryogels are efficient carriers for the immobilization of biomolecules because of their unique macroporous structure, permeability, mechanical stability and different surface chemical functionalities. The aim of the study was to demonstrate the potential use of macroporous monolithic cryogels for biotoxin removal using anthrax toxin protective antigen (PA), the central cell-binding component of the anthrax exotoxins, and covalent immobilization of monoclonal antibodies. The affinity ligand (protein A) was chemically coupled to the reactive hydroxyl and epoxy-derivatized monolithic cryogels and the binding efficiencies of protein A, monoclonal antibodies to the cryogel column were determined. Our results show differences in the binding capacity of protein A as well as monoclonal antibodies to the cryogel adsorbents caused by ligand concentrations, physical properties and morphology of surface matrices. The cytotoxicity potential of the cryogels was determined by an in vitro viability assay using V79 lung fibroblast as a model cell and the results reveal that the cryogels are non-cytotoxic. Finally, the adsorptive capacities of PA from phosphate buffered saline (PBS) were evaluated towards a non-glycosylated, plant-derived human monoclonal antibody (PANG) and a glycosylated human monoclonal antibody (Valortim(®)), both of which were covalently attached via protein A immobilization. Optimal binding capacities of 108 and 117 mg/g of antibody to the adsorbent were observed for PANG attached poly(acrylamide-allyl glycidyl ether) [poly(AAm-AGE)] and Valortim(®) attached poly(AAm-AGE) cryogels, respectively, This indicated that glycosylation status of Valortim(®) antibody could significantly increase (8%) its binding capacity relative to the PANG antibody on poly(AAm-AGE)-protien-A column (p < 0.05). The amounts of PA which remained in the solution after passing PA spiked PBS through PANG or Valortim bound poly(AAm-AGE) cryogel were significantly (p < 0.05) decreased relative to the amount of PA remained in the solution after passing through unmodified as well as protein A modified poly(AAm-AGE) cryogel columns, indicates efficient PA removal from spiked PBS over 60 min of circulation. The high adsorption capacity towards anthrax toxin PA of the cryogel adsorbents indicated potential application of these materials for treatment of Bacillus anthracis infection. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Studies with an immobilized metal affinity chromatography cassette system involving binuclear triazacyclononane-derived ligands: automation of batch adsorption measurements with tagged recombinant proteins.

PubMed

Petzold, Martin; Coghlan, Campbell J; Hearn, Milton T W

2014-07-18

This study describes the determination of the adsorption isotherms and binding kinetics of tagged recombinant proteins using a recently developed IMAC cassette system and employing automated robotic liquid handling procedures for IMAC resin screening. These results confirm that these new IMAC resins, generated from a variety of different metal-charged binuclear 1,4,7-triaza-cyclononane (tacn) ligands, interact with recombinant proteins containing a novel N-terminal metal binding tag, NT1A, with static binding capacities similar to those obtained with conventional hexa-His tagged proteins, but with significantly increased association constants. In addition, higher kinetic binding rates were observed with these new IMAC systems, an attribute that can be positively exploited to increase process productivity. The results from this investigation demonstrate that enhancements in binding capacities and affinities were achieved with these new IMAC resins and chosen NT1A tagged protein. Further, differences in the binding performances of the bis(tacn) xylenyl-bridged ligands were consistent with the distance between the metal binding centres of the two tacn moieties, the flexibility of the ligand and the potential contribution from the aromatic ring of the xylenyl group to undergo π/π stacking interactions with the tagged proteins. Copyright © 2014 Elsevier B.V. All rights reserved.

Characterization of the Mel1c melatoninergic receptor in platypus (Ornithorhynchus anatinus)

PubMed Central

Gautier, Célia; Guenin, Sophie-Penelope; Riest-Fery, Isabelle; Perry, Tahlia Jade; Legros, Céline; Nosjean, Olivier; Simonneaux, Valerie; Grützner, Frank

2018-01-01

Melatonin is a neurohormone produced in both animals and plants. It binds at least three G-protein-coupled receptors: MT1 and MT2, and Mel1cGPR. Mammalian GPR50 evolved from the reptilian/avian Mel1c and lost its capacity to bind melatonin in all the therian mammal species that have been tested. In order to determine if binding is lost in the oldest surviving mammalian lineage of monotremes we investigated whether the melatonin receptor has the ability to bind melatonin in the platypus (Ornithorhynchus anatinus), and evaluated its pharmacological profile. Sequence and phylogenetic analysis showed that platypus has in fact retained the ancestral Mel1c and has the capacity to bind melatonin similar to other mammalian melatonin receptors (MT1 and MT2), with an affinity in the 1 nM range. We also investigated the binding of a set of melatoninergic ligands used previously to characterize the molecular pharmacology of the melatonin receptors from sheep, rats, mice, and humans and found that the general profiles of these compounds make Mel1c resemble human MT1 more than MT2. This work shows that the loss of GPR50 binding evolved after the divergence of monotremes less than 190MYA in therian mammals. PMID:29529033

Characterization of the Mel1c melatoninergic receptor in platypus (Ornithorhynchus anatinus).

PubMed

Gautier, Célia; Guenin, Sophie-Penelope; Riest-Fery, Isabelle; Perry, Tahlia Jade; Legros, Céline; Nosjean, Olivier; Simonneaux, Valerie; Grützner, Frank; Boutin, Jean A

2018-01-01

Melatonin is a neurohormone produced in both animals and plants. It binds at least three G-protein-coupled receptors: MT1 and MT2, and Mel1cGPR. Mammalian GPR50 evolved from the reptilian/avian Mel1c and lost its capacity to bind melatonin in all the therian mammal species that have been tested. In order to determine if binding is lost in the oldest surviving mammalian lineage of monotremes we investigated whether the melatonin receptor has the ability to bind melatonin in the platypus (Ornithorhynchus anatinus), and evaluated its pharmacological profile. Sequence and phylogenetic analysis showed that platypus has in fact retained the ancestral Mel1c and has the capacity to bind melatonin similar to other mammalian melatonin receptors (MT1 and MT2), with an affinity in the 1 nM range. We also investigated the binding of a set of melatoninergic ligands used previously to characterize the molecular pharmacology of the melatonin receptors from sheep, rats, mice, and humans and found that the general profiles of these compounds make Mel1c resemble human MT1 more than MT2. This work shows that the loss of GPR50 binding evolved after the divergence of monotremes less than 190MYA in therian mammals.

Down-modulation of receptors for phorbol ester tumor promoter in primary epidermal cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Solanki, V.; Slaga, T.J.

1982-01-01

The specific (20-/sup 3/H)phorbol 12,13-dibutyrate ((/sup 3/H)PDBu) binding to intact epidermal cells displayed the phenomenon of down-modulation, i.e., the specific binding of (/sup 3/H)PDBu to its receptors on primary epidermal cells reached a maximum within 1 h and steadily declined thereafter. The apparent down-modulation of radiolabel resulted from a partial loss in the total number of receptors; the affinity of receptors for the ligand was essentially unchanged. A number of agents such as chloroquine, methylamine, or arginine which are known to prevent clustering, down-modulation, and/or internalization of several hormone receptors did not affect the down-modulation of phorbol ester receptors. Furthermore,more » cycloheximide had no effect either on down-modulation or on the binding capacity of cells. The surface binding capacity of down-modulated cells following a 90-min incubation with unlabeled ligand was almost returned to normal within 1 h. The effect of the antidepressant drug chlorpromazine, which is known to interact with calmodulin, on (/sup 3/H)PDBu binding was also investigated. Our data indicate that the effect of chlorpromazine on (/sup 3/H)PDBu binding is probably unrelated to its calmodulin-binding activity.« less

Ca sup 2+ binding capacity of cytoplasmic proteins from rod photoreceptors is mainly due to arrestin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huppertz, B.; Weyand, I.; Bauer, P.J.

1990-06-05

Arrestin (also called S-antigen or 48-kDa protein) binds to photoexcited and phosphorylated rhodopsin and, thereby, blocks competitively the activation of transducin. Using Ca{sup 2+} titration in the presence of the indicator arsenazo III and {sup 45}Ca{sup 2+} autoradiography, we show that arrestin is a Ca2(+)-binding protein. The Ca{sup 2+} binding capacity of arresting-containing protein extracts from bovine rod outer segments is about twice as high as that of arrestin-depleted extracts. The difference in the Ca{sup 2+} binding of arrestin-containing and arrestin-depleted protein extracts was attributed to arrestin. Both, these difference-measurements of protein extracts and the measurements of purified arrestin yieldmore » dissociation constants for the Ca{sup 2+} binding of arrestin between 2 and 4 microM. The titration curves are consistent with a molar ratio of one Ca{sup 2+} binding site per arrestin. No Ca{sup 2+} binding in the micromolar range was found in extracts containing mainly transducin and cGMP-phosphodiesterase. Since arrestin is one of the most abundant proteins in rod photoreceptors occurring presumably up to millimolar concentrations in rod outer segments, we suggest that aside from its function to prevent the activation of transducin, arrestin acts probably as an intracellular Ca{sup 2+} buffer.« less

Decreasing methylation of pectin caused by nitric oxide leads to higher aluminium binding in cell walls and greater aluminium sensitivity of wheat roots

PubMed Central

Sun, Chengliang; Lu, Lingli; Yu, Yan; Liu, Lijuan; Hu, Yan; Ye, Yiquan; Jin, Chongwei; Lin, Xianyong

2016-01-01

Nitric oxide (NO) is an important bioactive molecule involved in cell wall metabolism, which has been recognized as a major target of aluminium (Al) toxicity. We have investigated the effects of Al-induced NO production on cell wall composition and the subsequent Al-binding capacity in roots of an Al-sensitive cultivar of wheat (Triticum aestivum L. cv. Yang-5). We found that Al exposure induced NO accumulation in the root tips. Eliminating NO production with an NO scavenger (cPTIO) significantly alleviated the Al-induced inhibition of root growth and thus reduced Al accumulation. Elimination of NO, however, did not significantly affect malate efflux or rhizosphere pH changes under Al exposure. Levels of cell wall polysaccharides (pectin, hemicelluloses 1, and hemicelluloses 2) and pectin methylesterase activity, as well as pectin demethylation in the root apex, significantly increased under Al treatment. Exogenous cPTIO application significantly decreased pectin methylesterase activity and increased the degree of methylation of pectin in the root cell wall, thus decreasing the Al-binding capacity of pectin. These results suggest that the Al-induced enhanced production of NO decreases cell wall pectin methylation, thus increasing the Al-binding capacity of pectin and negatively regulating Al tolerance in wheat. PMID:26663393

Metal binding characterization and conformational studies using Raman microscopy of resin-bound poly(aspartic acid).

PubMed

Stair, Jacqueline L; Holcombe, James A

2007-03-01

The metal binding capacities, conditional stability constants, and secondary structure of immobilized polyaspartic acid (PLAsp) (n = 6, 20, and 30) on TentaGel resin were determined when binding Mg2+, Co2+, Cd2+, and Ni2+. Metal binding to the synthesized peptides was evaluated using breakthrough curves from a packed microcolumn and flame atomic absorption spectrophotometry (FAAS) detection. The metal capacities reached values of 590, 2160, and 3710 mumol of metal/g of resin for the 6-mer, 20-mer, and 30-mer, respectively, and this resulted in 2-3 residues per metal for all peptides and metals tested. Surprisingly, the concentrated environment of the resin along with the spatial distribution of attachment groups allowed for most residues to participate in metal binding regardless of the peptide length. Conditional stability constants calculated using single metal binding isotherms indicated that binding strength decreased as the chain length increased on the resin. Raman microscopy on single beads was used to determine PLAsp secondary structure, and all peptides were of a mixed conformation (i.e., beta-sheets, alpha-helices, random chain, etc.) during neutral conditioning and metal binding. Uniquely, the longer 20-mer and 30-mer peptides showed a distinct change from a mixed conformation to beta-sheets and alpha-helices during metal release with acid. This study confirms that metal release by longer immobilized peptides is often assisted by a conformational change, which easily spoils the binding cavity, while shorter peptides may release metal primarily by H+ displacement.

Bilirubin Binding Capacity in the Preterm Neonate.

PubMed

Amin, Sanjiv B

2016-06-01

Total serum/plasma bilirubin (TB), the biochemical measure currently used to evaluate and manage hyperbilirubinemia, is not a useful predictor of bilirubin-induced neurotoxicity in premature infants. Altered bilirubin-albumin binding in premature infants limits the usefulness of TB in premature infants. In this article, bilirubin-albumin binding, a modifying factor for bilirubin-induced neurotoxicity, in premature infants is reviewed. Copyright © 2016 Elsevier Inc. All rights reserved.

Novel Peptide with Specific Calcium-Binding Capacity from Schizochytrium sp. Protein Hydrolysates and Calcium Bioavailability in Caco-2 Cells

PubMed Central

Cai, Xixi; Lin, Jiaping; Wang, Shaoyun

2016-01-01

Peptide-calcium can probably be a suitable supplement to improve calcium absorption in the human body. In this study, a specific peptide Phe-Tyr (FY) with calcium-binding capacity was purified from Schizochytrium sp. protein hydrolysates through gel filtration chromatography and reversed phase HPLC. The calcium-binding capacity of FY reached 128.77 ± 2.57 μg/mg. Results of ultraviolet spectroscopy, fluorescence spectroscopy, and infrared spectroscopy showed that carboxyl groups, amino groups, and amido groups were the major chelating sites. FY-Ca exhibited excellent thermal stability and solubility, which were beneficial to be absorbed and transported in the basic intestinal tract of the human body. Moreover, the calcium bioavailability in Caco-2 cells showed that FY-Ca could enhance calcium uptake efficiency by more than three times when compared with CaCl2, and protect calcium ions against dietary inhibitors, such as tannic acid, oxalate, phytate, and Zn2+. Our findings further the progress of algae-based peptide-calcium, suggesting that FY-Ca has the potential to be developed as functionally nutraceutical additives. PMID:28036002

In vitro digestibility of goat milk and kefir with a new standardised static digestion method (INFOGEST cost action) and bioactivities of the resultant peptides.

PubMed

Nehir El, Sedef; Karakaya, Sibel; Simsek, Sebnem; Dupont, Didier; Menfaatli, Esra; Eker, Alper Tolga

2015-07-01

The hydrolysis degrees of goat milk and kefir during simulated gastrointestinal digestion and some bioactivities of the resulting peptides after fermentation and digestion were studied. A static in vitro digestion method by the COST FA1005 Action INFOGEST was used and goat milk and kefir were partially hydrolyzed during the gastric phase and had above 80% hydrolysis after duodenal digestion. There were no differences between the digestibility of goat milk and kefir (p > 0.05). Goat milk and kefir displayed about 7-fold antioxidant activity after digestion (p < 0.05). Fermentation showed no effect on the calcium-binding capacity of the samples (p > 0.05), however, after in vitro digestion calcium-binding capacity of the goat milk and kefir increased 2 and 5 fold, respectively (p < 0.05). Digested goat milk and kefir showed a higher dose-dependent inhibitory effect on α-amylase compared to undigested samples (p < 0.05). α-Glucosidase inhibitory activities and in vitro bile acid-binding capacities of the samples were not determined at the studied concentrations.

[3H]-nitrendipine binding in membranes obtained from hypoxic and reoxygenated heart.

PubMed

Matucci, R; Bennardini, F; Sciammarella, M L; Baccaro, C; Stendardi, I; Franconi, F; Giotti, A

1987-04-01

We compared the binding properties of [3H]-nitrendipine in heart membranes from normal guinea-pig heart and from hypoxic or hypoxic and reoxygenated heart. The [3H]-nitrendipine binds a single class of high capacity (Bmax 667.2 +/- 105.2) with high affinity (KD 0.14 +/- 0.02) binding sites. By contrast, in membranes of hypoxic and reoxygenated heart the Bmax decreases significantly while it remains unaffected during hypoxia. Xanthinoxidase activity is increased in hypoxic-reoxygenated hearts.

Techno-functional properties and in vitro bile acid-binding capacities of tamarillo (Solanum betaceum Cav.) hydrocolloids.

PubMed

Gannasin, Sri Puvanesvari; Adzahan, Noranizan Mohd; Mustafa, Shuhaimi; Muhammad, Kharidah

2016-04-01

Hydrocolloids were extracted from seed mucilage and the pulp fractions from red tamarillo (Solanum betaceum Cav.) mesocarp, and characterisation of their techno-functional properties and in vitro bile acid-binding capacities was performed. The seed mucilage hydrocolloids that were extracted, using either 1% citric acid (THC) or water (THW), had a good foaming capacity (32-36%), whereas the pulp hydrocolloids that were extracted, using 72% ethanol (THE) or 20mM HEPES buffer (THH), had no foaming capacity. The pulp hydrocolloid, however, possessed high oil-holding and water-holding capacities in the range of 3.3-3.6 g oil/g dry sample and 25-27 g water/g dry sample, respectively. This enabled the pulp hydrocolloid to entrap more bile acids (35-38% at a hydrocolloid concentration of 2%) in its gelatinous network in comparison to commercial oat fibre and other hydrocolloids studied. The exceptional emulsifying properties (80-96%) of both hydrocolloids suggest their potential applications as food emulsifiers and bile acid binders. Copyright © 2015 Elsevier Ltd. All rights reserved.

Binding of Radioactive Benzylpenicillin to Sporulating Bacillus Cultures: Chemistry and Fluctuations in Specific Binding Capacity

PubMed Central

Lawrence, Paul J.; Rogolsky, Marvin; Hanh, Vo Thi

1971-01-01

The chemistry of the binding of 14C-benzylpenicillin to sporulating cultures of Bacillus megaterium and B. subtilis is similar to that in a 4-hr vegetative culture of Staphylococcus aureus. Unlabeled penicillins prevent the binding of 14C-benzylpenicillin, but benzylpenicilloic acid and benzylpenilloic acid do not. Bound antibiotic can be removed from cells with neutral hydroxylamine at 25 C. Sporulating cultures display two intervals of enhanced binding, whereas binding to stationaryphase S. aureus cells remains constant. The first period of increased binding activity occurs during formation of the spore septum or cell wall primordium development, and the second coincides with cortex biosynthesis. PMID:4942758

Comprehensive meta-analysis of Signal Transducers and Activators of Transcription (STAT) genomic binding patterns discerns cell-specific cis-regulatory modules

PubMed Central

2013-01-01

Background Cytokine-activated transcription factors from the STAT (Signal Transducers and Activators of Transcription) family control common and context-specific genetic programs. It is not clear to what extent cell-specific features determine the binding capacity of seven STAT members and to what degree they share genetic targets. Molecular insight into the biology of STATs was gained from a meta-analysis of 29 available ChIP-seq data sets covering genome-wide occupancy of STATs 1, 3, 4, 5A, 5B and 6 in several cell types. Results We determined that the genomic binding capacity of STATs is primarily defined by the cell type and to a lesser extent by individual family members. For example, the overlap of shared binding sites between STATs 3 and 5 in T cells is greater than that between STAT5 in T cells and non-T cells. Even for the top 1,000 highly enriched STAT binding sites, ~15% of STAT5 binding sites in mouse female liver are shared by other STATs in different cell types while in T cells ~90% of STAT5 binding sites are co-occupied by STAT3, STAT4 and STAT6. In addition, we identified 116 cis-regulatory modules (CRM), which are recognized by all STAT members across cell types defining a common JAK-STAT signature. Lastly, in liver STAT5 binding significantly coincides with binding of the cell-specific transcription factors HNF4A, FOXA1 and FOXA2 and is associated with cell-type specific gene transcription. Conclusions Our results suggest that genomic binding of STATs is primarily determined by the cell type and further specificity is achieved in part by juxtaposed binding of cell-specific transcription factors. PMID:23324445

Studies on the interactions between purified bovine caseins and alkaline-earth-metalions

PubMed Central

Dickson, I. R.; Perkins, D. J.

1971-01-01

1. Alkaline-earth-metal cations at low concentrations form soluble complexes with bovine caseins. The relative order of binding capacities is: Mg2+>Ca2+>Ba2+>Sr2+. 2. The cations interact with both free ionized carboxyl groups of aspartic acid and glutamic acid and with monoester phosphate groups covalently bound to serine and threonine; at low concentrations of the cations interactions are predominantly with the phosphate groups. 3. The order of binding capacities for purified components of the casein complex is: αs1-casein>β-casein>κ-casein. PMID:5166590

Solubility and binding properties of PEGylated lysozyme derivatives with increasing molecular weight on hydrophobic-interaction chromatographic resins.

PubMed

Müller, Egbert; Josic, Djuro; Schröder, Tim; Moosmann, Anna

2010-07-09

Dynamic binding capacities and resolution of PEGylated lysozyme derivatives with varying molecular weights of poly (ethylene) glycol (PEG) with 5 kDa, 10 kDa and 30 kDa for HIC resins and columns are presented. To find the optimal range for the operating conditions, solubility studies were performed by high-throughput analyses in a 96-well plate format, and optimal salt concentrations and pH values were determined. The solubility of PEG-proteins was strongly influenced by the length of the PEG moiety. Large differences in the solubilities of PEGylated lysozymes in two different salts, ammonium sulfate and sodium chloride were found. Solubility of PEGylated lysozyme derivatives in ammonium sulfate decreases with increased length of attached PEG chains. In sodium chloride all PEGylated lysozyme derivatives are fully soluble in a concentration range between 0.1 mg protein/ml and 10 mg protein/ml. The binding capacities for PEGylated lysozyme to HIC resins are dependent on the salt type and molecular weight of the PEG polymer. In both salt solutions, ammonium sulfate and sodium chloride, the highest binding capacity of the resin was found for 5 kDa PEGylated lysozyme. For both native lysozyme and 30 kDa mono-PEGylated lysozyme the binding capacities were lower. In separation experiments on a TSKgel Butyl-NPR hydrophobic-interaction column with ammonium sulfate as mobile phase, the elution order was: native lysozyme, 5 kDa mono-PEGylated lysozyme and oligo-PEGylated lysozyme. This elution order was found to be reversed when sodium chloride was used. Furthermore, the resolution of the three mono-PEGylated forms was not possible with this column and ammonium sulfate as mobile phase. In 4 M sodium chloride a resolution of all PEGylated lysozyme forms was achieved. A tentative explanation for these phenomena can be the increased solvation of the PEG polymers in sodium chloride which changes the usual attractive hydrophobic forces in ammonium sulfate to more repulsive hydration forces in this hydrotrophic salt.

Surface display of metal binding domain derived from PbrR on Escherichia coli specifically increases lead(II) adsorption.

PubMed

Hui, Chang-Ye; Guo, Yan; Yang, Xue-Qin; Zhang, Wen; Huang, Xian-Qing

2018-05-01

To improve the Pb 2+ biosorption capacity of the potential E. coli biosorbent, a putative Pb 2+ binding domain (PbBD) derived from PbrR was efficiently displayed on to the E. coli cell surface. The PbBD was obtained by truncating the N-terminal DNA-binding domain and C-terminal redundant amino acid residues of the Pb 2+ -sensing transcriptional factor PbrR. Whole-cell sorbents were constructed with the full-length PbrR and PbBD of PbrR genetically engineered onto the surface of E. coli cells using Lpp-OmpA as the anchor. Followed by a 1.71-fold higher display of PbBD than PbrR, the presence of PbBD on the surface of E. coli cells enabled a 1.92-fold higher Pb 2+ biosorption than that found in PbrR-displayed cells. Specific Pb 2+ binding via PbBD was the same as Pb 2+ binding via the full-length PbrR, with no observable decline even in the presence of Zn 2+ and Cd 2+ . Since surface-engineered E. coli cells with PbBD increased the Pb 2+ binding capacity and did not affect the adsorption selectivity, this suggests that surface display of the metal binding domain derived from MerR-like proteins may be used for the bioremediation of specific toxic heavy metals.

Rigidification of the autolysis loop enhances Na[superscript +] binding to thrombin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pozzi, Nicola; Chen, Raymond; Chen, Zhiwei

2011-09-20

Binding of Na{sup +} to thrombin ensures high activity toward physiological substrates and optimizes the procoagulant and prothrombotic roles of the enzyme in vivo. Under physiological conditions of pH and temperature, the binding affinity of Na{sup +} is weak due to large heat capacity and enthalpy changes associated with binding, and the K{sub d} = 80 mM ensures only 64% saturation of the site at the concentration of Na{sup +} in the blood (140 mM). Residues controlling Na{sup +} binding and activation have been identified. Yet, attempts to improve the interaction of Na{sup +} with thrombin and possibly increase catalyticmore » activity under physiological conditions have so far been unsuccessful. Here we report how replacement of the flexible autolysis loop of human thrombin with the homologous rigid domain of the murine enzyme results in a drastic (up to 10-fold) increase in Na{sup +} affinity and a significant improvement in the catalytic activity of the enzyme. Rigidification of the autolysis loop abolishes the heat capacity change associated with Na{sup +} binding observed in the wild-type and also increases the stability of thrombin. These findings have general relevance to protein engineering studies of clotting proteases and trypsin-like enzymes.« less

Sorption of Cr(III) and Cr(VI) to High and Low Pressure Synthetic Nano-Magnetite (Fe3O4)Particles

PubMed Central

Parsons, Jason G.; Hernandez, Jeffrey; Gonzalez, Christina M.; Gardea-Torresdey, J. L.

2014-01-01

The binding of Cr(III) and Cr(VI) to synthetic nano-magnetie particles synthesized under open vessel conditions and a microwave assisted hydrothermal synthesis techniques was investigated. Batch studies showed that the binding of both the Cr(III) and Cr(VI) bound to the nano-materials in a pH dependent manner. The Cr(III) maximized at binding at pH 4 and 100% binding. Similarly, the Cr(VI) ions showed a maximum binding of 100% at pH 4. The data from the time dependency studies showed for the most part the majority of the binding occurred within the first 5 minutes of contact with the nanomaterial and remained constant thereafter. In addition, the effects of the possible interferences were investigated which showed some effects on the binding of both Cr(III) and Cr(VI). However, the interferences never completely eliminated the chromium binding. Isotherm studies conducted at room temperature showed the microwave synthesized nanomaterials had a binding capacity of 1208 ± 43.9 mg/g and 555 ± 10.5 mg/g for Cr(VI) and Cr(III), respectively. However, the microwave assisted synthesized nanomaterials had capacities of 1705 ± 14.5 and 555± 10.5 mg/g for Cr(VI) and Cr(III), respectively. XANES studies showed the Cr(VI) was reduced to Cr(III), and the Cr(III) remained as Cr(III). In addition, the XANES studies indicated that the chromium remained coordinated in an octahedral arrangement of oxygen atoms. PMID:25097452

Nanoporous sorbent material as an oral phosphate binder and for aqueous phosphate, chromate, and arsenate removal

PubMed Central

Sangvanich, Thanapon; Ngamcherdtrakul, Worapol; Lee, Richard; Morry, Jingga; Castro, David; Fryxell, Glen E.; Yantasee, Wassana

2014-01-01

Phosphate removal is both biologically and environmentally important. Biologically, hyperphosphatemia is a critical condition in end-stage chronic kidney disease patients. Patients with hyperphosphatemia are treated long-term with oral phosphate binders to prevent phosphate absorption to the body by capturing phosphate in the gastrointestinal (GI) tract followed by fecal excretion. Environmentally, phosphate levels in natural water resources must be regulated according to limits set forth by the US Environmental Protection Agency. By utilizing nanotechnology and ligand design, we developed a new material to overcome limitations of traditional sorbent materials such as low phosphate binding capacity, slow binding kinetics, and negative interference by other anions. A phosphate binder based on iron-ethylenediamine on nanoporous silica (Fe-EDA-SAMMS) has been optimized for substrates and Fe(III) deposition methods. The Fe-EDA-SAMMS material had a 4-fold increase in phosphate binding capacity and a broader operating pH window compared to other reports. The material had a faster phosphate binding rate and was significantly less affected by other anions than Sevelamer HCl, the gold standard oral phosphate binder, and AG® 1-X8, a commercially available anion exchanger. It had less cytotoxicity to Caco-2 cells than lanthanum carbonate, another prescribed oral phosphate binder. The Fe-EDA-SAMMS also had high capacity for arsenate and chromate, two of the most toxic anions in natural water. PMID:25554735

Melanin-Based Coatings as Lead-Binding Agents

PubMed Central

Sono, Karin; Lye, Diane; Moore, Christine A.; Boyd, W. Christopher; Gorlin, Thomas A.; Belitsky, Jason M.

2012-01-01

Interactions between metal ions and different forms of melanin play significant roles in melanin biochemistry. The binding properties of natural melanin and related synthetic materials can be exploited for nonbiological applications, potentially including water purification. A method for investigating metal ion-melanin interactions on solid support is described, with lead as the initial target. 2.5 cm discs of the hydrophobic polymer PVDF were coated with synthetic eumelanin from the tyrosinase-catalyzed polymerization of L-dopa, and with melanin extracted from human hair. Lead (Pb2+) binding was quantified by atomic absorption spectroscopy (flame mode), and the data was well fit by the Langmuir model. Langmuir affinities ranged from 3.4 · 103 to 2.2 · 104 M−1. At the maximum capacity observed, the synthetic eumelanin coating bound ~9% of its mass in lead. Binding of copper (Cu2+), zinc (Zn2+), and cadmium (Cd2+) to the synthetic-eumelanin-coated discs was also investigated. Under the conditions tested, the Langmuir affinities for Zn2+, Cd2+, and Cu2+ were 35%, 53%, and 77%, respectively, of the Langmuir affinity for Pb2+. The synthetic-eumelanin-coated discs have a slightly higher capacity for Cu2+ on a per mole basis than for Pb2+, and lower capacities for Cd2+ and Zn2+. The system described can be used to address biological questions and potentially be applied toward melanin-based water purification. PMID:22611345

Important role of N108 residue in binding of bovine foamy virus transactivator Tas to viral promoters.

PubMed

Bing, Tiejun; Zhang, Suzhen; Liu, Xiaojuan; Liang, Zhibin; Shao, Peng; Zhang, Song; Qiao, Wentao; Tan, Juan

2016-06-30

Bovine foamy virus (BFV) encodes the transactivator BTas, which enhances viral gene transcription by binding to the long terminal repeat promoter and the internal promoter. In this study, we investigated the different replication capacities of two similar BFV full-length DNA clones, pBS-BFV-Y and pBS-BFV-B. Here, functional analysis of several chimeric clones revealed a major role for the C-terminal region of the viral genome in causing this difference. Furthermore, BTas-B, which is located in this C-terminal region, exhibited a 20-fold higher transactivation activity than BTas-Y. Sequence alignment showed that these two sequences differ only at amino acid 108, with BTas-B containing N108 and BTas-Y containing D108 at this position. Results of mutagenesis studies demonstrated that residue N108 is important for BTas binding to viral promoters. In addition, the N108D mutation in pBS-BFV-B reduced the viral replication capacity by about 1.5-fold. Our results suggest that residue N108 is important for BTas binding to BFV promoters and has a major role in BFV replication. These findings not only advances our understanding of the transactivation mechanism of BTas, but they also highlight the importance of certain sequence polymorphisms in modulating the replication capacity of isolated BFV clones.

Characterization of product capture resin during microbial cultivations.

PubMed

Frykman, Scott; Tsuruta, Hiroko; Galazzo, Jorge; Licari, Peter

2006-06-01

Various bioactive small molecules produced by microbial cultivation are degraded in the culture broth or may repress the formation of additional product. The inclusion of hydrophobic adsorber resin beads to capture these products in situ and remove them from the culture broth can reduce or prevent this degradation and repression. These product capture beads are often subjected to a dynamic and stressful microenvironment for a long cultivation time, affecting their physical structure and performance. Impact and collision forces can result in the fracturing of these beads into smaller pieces, which are difficult to recover at the end of a cultivation run. Various contaminating compounds may also bind in a non-specific manner to these beads, reducing the binding capacity of the resin for the product of interest (fouling). This study characterizes resin bead binding capacity (to monitor bead fouling), and resin bead volume distributions (to monitor bead fracture) for an XAD-16 adsorber resin used to capture epothilone produced during myxobacterial cultivations. Resin fouling was found to reduce the product binding capacity of the adsorber resin by 25-50%. Additionally, the degree of resin bead fracture was found to be dependent on the cultivation length and the impeller rotation rate. Microbial cultivations and harvesting processes should be designed in such a way to minimize bead fragmentation and fouling during cultivation to maximize the amount of resin and associated product harvested at the end of a run.

The effect of prolonged intrauterine hyperinsulinemia on iron utilization in fetal sheep.

PubMed

Georgieff, M K; Widness, J A; Mills, M M; Stonestreet, B S

1989-11-01

Newborn infants of poorly controlled insulin-dependent diabetic mothers demonstrate a redistribution of iron from serum and tissue stores into red blood cells. These changes may be due to increases in iron utilization during augmented Hb synthesis, which compensates for chronic intrauterine hypoxemia induced by prolonged fetal hyperinsulinemia. We tested this hypothesis by measuring plasma iron, total iron-binding capacity, percent iron-binding capacity saturation (total iron-binding capacity saturation), Hb concentration, total red cell Hb, and total red cell iron in the arterial blood of 11 chronically instrumented fetal sheep after 7-12 d of infusion with 15 U/day of insulin (n = 5) or placebo (n = 6). The insulin-infused fetal sheep had higher mean +/- SD plasma insulin concentrations (448 +/- 507 versus 11 +/- 8 mU/L; p less than 0.001) and lower arterial oxygen saturations (38 +/- 7 versus 54 +/- 9%; p less than 0.02). The insulin-infused group had a lower mean plasma iron concentration (20.8 +/- 10.9 versus 42.1 +/- 14.7 microM/L; p less than 0.02) and total iron-binding capacity saturation (36 +/- 20 versus 64 +/- 22%; p less than 0.02) and a higher total red cell Hb (45.4 +/- 8.7 versus 32.6 +/- 8.8 g; p less than 0.02) and total red cell iron content (154 +/- 29 versus 111 +/- 29 mg; p less than 0.02) when compared with the placebo group. Seven to 12 d of intrauterine hyperinsulinemia decreases serum iron and increases total red cell iron, most likely by stimulating increased Hb synthesis in response to low arterial oxygen saturation.(ABSTRACT TRUNCATED AT 250 WORDS)

Avian Nanostructured Tissues as Models for New Defensive Coatings and Photonic Crystal Fibers

DTIC Science & Technology

2012-03-31

promiscuous binding capacity of chitin , the chemical backbone of the arthropod cuticle (Kumar 2000). This polysaccharide binds many proteins and other...properties. The greater refractive index contrast between light and dark layers afforded by chitin may allow Arthropoda to attain brighter and more 71

The Thermal Stabilization of Vaccines Against Agents of Bioterrorism

DTIC Science & Technology

2005-09-01

to determine (1) whether rPA in the formulation buffer in the absence of excipients binds to Alhydrogel®and (2) the binding capacity . The aluminum...botulinum toxin (Allergan), A ricin vaccine (DOR Biopharma ) and a vaccine against Norwalk virus (Ligocyte) were also initiated and are in various

A CE-FL based method for real-time detection of in-capillary self-assembly of the nanoconjugates of polycysteine ligand and quantum dots.

PubMed

Wang, Jianhao; Zhu, Zhilan; Qiu, Lin; Wang, Jianpeng; Wang, Xiang; Xiao, Qicai; Xia, Jiang; Liu, Li; Liu, Xiaoqian; Feng, Wei; Wang, Jinmei; Miao, Peng; Gao, Liqian

2018-07-06

Small molecules with free thiol groups always show high binding affinity to quantum dots (QDs). However, it is still highly challenging to detect the binding capacity between thiol-containing molecules and QDs inside a capillary. To conquer this limitation, a capillary electrophoresis with fluorescence detection (CE-FL) based assay was proposed and established to investigate the binding capacity between QDs and a poly-thiolated peptide (ATTO 590-DDSSGGCCPGCC, ATTO-C4). Interestingly, the results showed that interval time had a great influence on QDs and ATTO-C4 self-assembly, which can be attributed to longer interval time benefitting the binding of QDs to ATTO-C4. The stability assays on ATTO-C4-QD assembly indicated that high concentration of imidazole or GSH had a high capability of competing with the bound ATTO-C4, evidenced by dramatically dropping of S 625 /S 565 ratio from 0.78 to 0.30 or 0.29. Therefore, all these results above suggested that this novel CE-FL based detection assay could be successfully applied to the binding studies between QDs and thiol-containing biomolecules.

Magnetic, core-shell structured and surface molecularly imprinted polymers for the rapid and selective recognition of salicylic acid from aqueous solutions

NASA Astrophysics Data System (ADS)

Zhang, Zulei; Niu, Dechao; Li, Yongsheng; Shi, Jianlin

2018-03-01

In this work, a novel kind of magnetic, core-shell structured and surface molecularly imprinted polymers (MMIPs) for the recognition of salicylic acid (SA) was facilely synthesized through a surface imprinting and sol-gel polymerization approach. The as-synthesized MMIPs exhibit uniform core-shell structure and favorable magnetic properties with a saturation magnetization of 22.8 emu g-1. The binding experiments demonstrated that MMIPs possessed high binding and specific recognition capacity, as well as fast binding kinetics and phase separation rate. The maximum binding capacity of MMIPs is around 36.8 mg g-1, nearly 6 times that of the magnetic non-imprinted polymers (MNIPs). Moreover, the selectivity experiments show that all the relative selectivity coefficients towards SA over its structure analogs are higher than 18, further indicating the markedly enhanced binding selectivity of MMIPs. Furthermore, the MMIPs were successfully applied for the determination of SA in environmental water samples with the recovery rates ranging from 94.0 to 108.0 %. This strategy may provide a versatile approach for the fabrication of well-defined molecularly imprinted polymers on nanomaterials for the analysis of complicated matrixes.

A CE-FL based method for real-time detection of in-capillary self-assembly of the nanoconjugates of polycysteine ligand and quantum dots

NASA Astrophysics Data System (ADS)

Wang, Jianhao; Zhu, Zhilan; Qiu, Lin; Wang, Jianpeng; Wang, Xiang; Xiao, Qicai; Xia, Jiang; Liu, Li; Liu, Xiaoqian; Feng, Wei; Wang, Jinmei; Miao, Peng; Gao, Liqian

2018-07-01

Small molecules with free thiol groups always show high binding affinity to quantum dots (QDs). However, it is still highly challenging to detect the binding capacity between thiol-containing molecules and QDs inside a capillary. To conquer this limitation, a capillary electrophoresis with fluorescence detection (CE-FL) based assay was proposed and established to investigate the binding capacity between QDs and a poly-thiolated peptide (ATTO 590-DDSSGGCCPGCC, ATTO-C4). Interestingly, the results showed that interval time had a great influence on QDs and ATTO-C4 self-assembly, which can be attributed to longer interval time benefitting the binding of QDs to ATTO-C4. The stability assays on ATTO-C4-QD assembly indicated that high concentration of imidazole or GSH had a high capability of competing with the bound ATTO-C4, evidenced by dramatically dropping of S 625/S 565 ratio from 0.78 to 0.30 or 0.29. Therefore, all these results above suggested that this novel CE-FL based detection assay could be successfully applied to the binding studies between QDs and thiol-containing biomolecules.

LH-RH binding to purified pituitary plasma membranes: absence of adenylate cyclase activation.

PubMed

Clayton, R N; Shakespear, R A; Marshall, J C

1978-06-01

Purified bovine pituitary plasma membranes possess two specific LH-RH binding sites. The high affinity site (2.5 X 10(9) l/mol) has low capacity (9 X 10(-15) mol/mg membrane protein) while the low affinity site 6.1 X 10(5) l/mol) has a much higher capacity (1.1 X 10(-10) mol/mg). Specific LH-RH binding to plasma membranes is increased 8.5-fold during purification from homogenate whilst adenylate cyclase activity is enriched 7--8-fold. Distribution of specific LH-RH binding to sucrose density gradient interface fractions parallels that of adenylate cyclase activity. Mg2+ and Ca2+ inhibit specific [125I]LH-RH binding at micromolar concentrations. Synthetic LH-RH, up to 250 microgram/ml, failed to stimulate adenylase cyclase activity of the purified bovine membranes. Using a crude 10,800 g rat pituitary membrane preparation, LH-RH similarly failed to activate adenylate cyclase even in the presence of guanyl nucleotides. These data confirm the presence of LH-RH receptor sites on pituitary plasma membranes and suggest that LH-RH-induced gonadotrophin release may be mediated by mechanisms other than activation of adenylate cyclase.

Gold Binding by Native and Chemically Modified Hops Biomasses

PubMed Central

López, M. Laura; Peralta-Videa, J. R.; de la Rosa, G.; Armendáriz, V.; Herrera, I.; Troiani, H.; Henning, J.

2005-01-01

Heavy metals from mining, smelting operations and other industrial processing facilities pollute wastewaters worldwide. Extraction of metals from industrial effluents has been widely studied due to the economic advantages and the relative ease of technical implementation. Consequently, the search for new and improved methodologies for the recovery of gold has increased. In this particular research, the use of cone hops biomass (Humulus lupulus) was investigated as a new option for gold recovery. The results showed that the gold binding to native hops biomass was pH dependent from pH 2 to pH 6, with a maximum percentage binding at pH 3. Time dependency studies demonstrated that Au(III) binding to native and modified cone hops biomasses was found to be time independent at pH 2 while at pH 5, it was time dependent. Capacity experiments demonstrated that at pH 2, esterified hops biomass bound 33.4 mg Au/g of biomass, while native and hydrolyzed hops biomasses bound 28.2 and 12.0 mg Au/g of biomass, respectively. However, at pH 5 the binding capacities were 38.9, 37.8 and 11.4 mg of Au per gram of native, esterified and hydrolyzed hops biomasses, respectively. PMID:18365087

Testing the Underlying Chemical Principles of the Biotic Ligand Model (BLM) to Marine Copper Systems: Measuring Copper Speciation Using Fluorescence Quenching.

PubMed

Tait, Tara N; McGeer, James C; Smith, D Scott

2018-01-01

Speciation of copper in marine systems strongly influences the ability of copper to cause toxicity. Natural organic matter (NOM) contains many binding sites which provides a protective effect on copper toxicity. The purpose of this study was to characterize copper binding with NOM using fluorescence quenching techniques. Fluorescence quenching of NOM with copper was performed on nine sea water samples. The resulting stability constants and binding capacities were consistent with literature values of marine NOM, showing strong binding with [Formula: see text] values from 7.64 to 10.2 and binding capacities ranging from 15 to 3110 nmol mg [Formula: see text] Free copper concentrations estimated at total dissolved copper concentrations corresponding to previously published rotifer effect concentrations, in the same nine samples, were statistically the same as the range of free copper calculated for the effect concentration in NOM-free artificial seawater. These data confirms the applicability of fluorescence spectroscopy techniques for NOM and copper speciation characterization in sea water and demonstrates that such measured speciation is consistent with the chemical principles underlying the biotic ligand model approach for bioavailability-based metals risk assessment.

Removal of cyanotoxins from surface water resources using reusable molecularly imprinted polymer adsorbents.

PubMed

Krupadam, Reddithota J; Patel, Govind P; Balasubramanian, Rajasekhar

2012-06-01

Microcystins (MCs; cyclic heptapeptides) are produced by freshwater cyanobacteria and cause public health concern in potable water supplies. There are more than 60 types of MCs identified to date, of which MC-LR is the most common found worldwide. For MC-LR, the WHO has established a threshold value of 1 μg L(-1) for drinking water. The present MCs removal methods such as coagulation, flocculation, adsorption, and filtration showed low efficiency for removing dissolved MC fraction from surface waters to the stipulated limit prescribed by WHO based on MC health impacts. The search for cost-effective and efficient removal method is still warranted for remediation of dissolved MC-LR-contaminated water resources. Molecularly imprinted polymer (MIP) adsorbent has been prepared using non-covalent imprinting approach. Using MC-LR as a template, itaconic acid as a functional monomer, and ethylene glycol dimethacrylate as a cross-linking monomer, a MIP has been synthesized. Computer simulations were used to design effective binding sites for MC-LR binding in aqueous solutions. Batch binding adsorption assay was followed to determine binding capacity of MIP under the influence of environmental parameters such as total dissolved solids and pH. The adsorptive removal of MC-LR from lake water has been investigated using MIPs. The MIP showed excellent adsorption potential toward MC-LR in aqueous solutions with a binding capacity of 3.64 μg mg(-1) which is about 60% and 70% more than the commercially used powdered activated carbon (PAC) and resin XAD, respectively. Environmental parameters such as total organic carbon (represented as chemical oxygen demand (COD)) and total dissolved solids (TDS) showed no significant interference up to 300 mg L(-1) for MC-LR removal from lake water samples. It was found that the binding sites on PAC and XAD have more affinity toward COD and TDS than the MC-LR. Further, the adsorption capacity of the MIP was evaluated rigorously by its repeated contact with fresh lake water, and it was found that the adsorption capacity of the MIP did not change even after seven adsorption/desorption cycles. The contaminated water of MC-LR (1.0 μg L(-1)) of 3,640 L could be treated by 1 g of MIP with an estimated cost of US $1.5. The adsorption capacity of the MIP is 40% more than commercially used PAC and resins and also the polymer showed reusable potential which is one of the important criteria in selection of cyanotoxins remediation methods.

Nanofiber Ion-Exchange Membranes for the Rapid Uptake and Recovery of Heavy Metals from Water

PubMed Central

Chitpong, Nithinart; Husson, Scott M.

2016-01-01

An evaluation of the performance of polyelectrolyte-modified nanofiber membranes was undertaken to determine their efficacy in the rapid uptake and recovery of heavy metals from impaired waters. The membranes were prepared by grafting poly(acrylic acid) (PAA) and poly(itaconic acid) (PIA) to cellulose nanofiber mats. Performance measurements quantified the dynamic ion-exchange capacity for cadmium (Cd), productivity, and recovery of Cd(II) from the membranes by regeneration. The dynamic binding capacities of Cd(II) on both types of nanofiber membrane were independent of the linear flow velocity, with a residence time of as low as 2 s. Analysis of breakthrough curves indicated that the mass flow rate increased rapidly at constant applied pressure after membranes approached equilibrium load capacity for Cd(II), apparently due to a collapse of the polymer chains on the membrane surface, leading to an increased porosity. This mechanism is supported by hydrodynamic radius (Rh) measurements for PAA and PIA obtained from dynamic light scattering, which show that Rh values decrease upon Cd(II) binding. Volumetric productivity was high for the nanofiber membranes, and reached 0.55 mg Cd/g/min. The use of ethylenediaminetetraacetic acid as regeneration reagent was effective in fully recovering Cd(II) from the membranes. Ion-exchange capacities were constant over five cycles of binding-regeneration. PMID:27999394

Nanofiber Ion-Exchange Membranes for the Rapid Uptake and Recovery of Heavy Metals from Water.

PubMed

Chitpong, Nithinart; Husson, Scott M

2016-12-20

An evaluation of the performance of polyelectrolyte-modified nanofiber membranes was undertaken to determine their efficacy in the rapid uptake and recovery of heavy metals from impaired waters. The membranes were prepared by grafting poly(acrylic acid) (PAA) and poly(itaconic acid) (PIA) to cellulose nanofiber mats. Performance measurements quantified the dynamic ion-exchange capacity for cadmium (Cd), productivity, and recovery of Cd(II) from the membranes by regeneration. The dynamic binding capacities of Cd(II) on both types of nanofiber membrane were independent of the linear flow velocity, with a residence time of as low as 2 s. Analysis of breakthrough curves indicated that the mass flow rate increased rapidly at constant applied pressure after membranes approached equilibrium load capacity for Cd(II), apparently due to a collapse of the polymer chains on the membrane surface, leading to an increased porosity. This mechanism is supported by hydrodynamic radius (R h ) measurements for PAA and PIA obtained from dynamic light scattering, which show that R h values decrease upon Cd(II) binding. Volumetric productivity was high for the nanofiber membranes, and reached 0.55 mg Cd/g/min. The use of ethylenediaminetetraacetic acid as regeneration reagent was effective in fully recovering Cd(II) from the membranes. Ion-exchange capacities were constant over five cycles of binding-regeneration.

The minor house dust mite allergen Der p 13 is a fatty acid-binding protein and an activator of a TLR2-mediated innate immune response.

PubMed

Satitsuksanoa, P; Kennedy, M; Gilis, D; Le Mignon, M; Suratannon, N; Soh, W T; Wongpiyabovorn, J; Chatchatee, P; Vangveravong, M; Rerkpattanapipat, T; Sangasapaviliya, A; Piboonpocanun, S; Nony, E; Ruxrungtham, K; Jacquet, A

2016-10-01

The house dust mite (HDM) allergen Der p 13 could be a lipid-binding protein able to activate key innate signaling pathways in the initiation of the allergic response. We investigated the IgE reactivity of recombinant Der p 13 (rDer p 13), its lipid-binding activities, and its capacity to stimulate airway epithelium cells. Purified rDer p 13 was characterized by mass spectrometry, circular dichroism, fluorescence-based lipid-binding assays, and in silico structural prediction. IgE-binding activity and allergenic potential of Der p 13 were examined by ELISA, basophil degranulation assays, and in vitro airway epithelial cell activation assays. Protein modeling and biophysical analysis indicated that Der p 13 adopts a β-barrel structure with a predominately apolar pocket representing a potential binding site for hydrophobic ligands. Fluorescent lipid-binding assays confirmed that the protein is highly selective for ligands and that it binds a fatty acid with a dissociation constant typical of lipid transporter proteins. The low IgE-binding frequency (7%, n = 224) in Thai HDM-allergic patients as well as the limited propensity to activate basophil degranulation classifies Der p 13 as a minor HDM allergen. Nevertheless, the protein with its presumptively associated lipid(s) triggered the production of IL-8 and GM-CSF in respiratory epithelial cells through a TLR2-, MyD88-, NF-kB-, and MAPK-dependent signaling pathway. Although a minor allergen, Der p 13 may, through its lipid-binding capacity, play a role in the initiation of the HDM-allergic response through TLR2 activation. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Molecular Evolution of the Oxygen-Binding Hemerythrin Domain

PubMed Central

Alvarez-Carreño, Claudia; Becerra, Arturo; Lazcano, Antonio

2016-01-01

Background The evolution of oxygenic photosynthesis during Precambrian times entailed the diversification of strategies minimizing reactive oxygen species-associated damage. Four families of oxygen-carrier proteins (hemoglobin, hemerythrin and the two non-homologous families of arthropodan and molluscan hemocyanins) are known to have evolved independently the capacity to bind oxygen reversibly, providing cells with strategies to cope with the evolutionary pressure of oxygen accumulation. Oxygen-binding hemerythrin was first studied in marine invertebrates but further research has made it clear that it is present in the three domains of life, strongly suggesting that its origin predated the emergence of eukaryotes. Results Oxygen-binding hemerythrins are a monophyletic sub-group of the hemerythrin/HHE (histidine, histidine, glutamic acid) cation-binding domain. Oxygen-binding hemerythrin homologs were unambiguously identified in 367/2236 bacterial, 21/150 archaeal and 4/135 eukaryotic genomes. Overall, oxygen-binding hemerythrin homologues were found in the same proportion as single-domain and as long protein sequences. The associated functions of protein domains in long hemerythrin sequences can be classified in three major groups: signal transduction, phosphorelay response regulation, and protein binding. This suggests that in many organisms the reversible oxygen-binding capacity was incorporated in signaling pathways. A maximum-likelihood tree of oxygen-binding hemerythrin homologues revealed a complex evolutionary history in which lateral gene transfer, duplications and gene losses appear to have played an important role. Conclusions Hemerythrin is an ancient protein domain with a complex evolutionary history. The distinctive iron-binding coordination site of oxygen-binding hemerythrins evolved first in prokaryotes, very likely prior to the divergence of Firmicutes and Proteobacteria, and spread into many bacterial, archaeal and eukaryotic species. The later evolution of the oxygen-binding hemerythrin domain in both prokaryotes and eukaryotes led to a wide variety of functions, ranging from protection against oxidative damage in anaerobic and microaerophilic organisms, to oxygen supplying to particular enzymes and pathways in aerobic and facultative species. PMID:27336621

Sex Differences in Serotonin 1 Receptor Binding in Rat Brain

NASA Astrophysics Data System (ADS)

Fischette, Christine T.; Biegon, Anat; McEwen, Bruce S.

1983-10-01

Male and female rats exhibit sex differences in binding by serotonin 1 receptors in discrete areas of the brain, some of which have been implicated in the control of ovulation and of gonadotropin release. The sex-specific changes in binding, which occur in response to the same hormonal (estrogenic) stimulus, are due to changes in the number of binding sites. Castration alone also affects the number of binding sites in certain areas. The results lead to the conclusion that peripheral hormones modulate binding by serotonin 1 receptors. The status of the serotonin receptor system may affect the reproductive capacity of an organism and may be related to sex-linked emotional disturbances in humans.

Point mutation increases a form of the NK1 receptor with high affinity for neurokinin A and B and septide

PubMed Central

Ciucci, Alessandra; Palma, Carla; Manzini, Stefano; Werge, Thomas M

1998-01-01

The binding modalities of substance P and neurokinin A on the wild type and Gly166 to-Cys mutant NK1 receptors expressed on CHO cells were investigated in homologous and heterologous binding experiments using both radiolabelled substance P and neurokinin A.On the wild type NK1 receptor NKA displaces radiolabelled substance P with very low apparent affinity, despite its high-affinity binding constant (determined in homologous binding experiments). The Gly166 to-Cys substitution in the NK1 tachykinin receptor greatly enhances the apparent affinity of neurokinin A in competition for radiolabelled substance P, but it does not change the binding constant of neurokinin A. The mutation, thereby, eliminates the discrepancy between the low apparent affinity and the high binding constant of neurokinin A.On the wild type receptor the binding capacity of neurokinin A is significantly smaller than that of substance P. In contrast, the two tachykinins bind to approximately the same number of sites on the mutant receptor.Simultaneous mass action law analysis of binding data in which multiple radioligands were employed in parallel demonstrated that a one-site model was unable to accommodate all the experimental data, whereas a two-site model provided a dramatically better description.These two receptor-sites display equally high affinity for substance P, while neurokinin A strongly discriminates between a high and a low affinity component. The binding affinities of neurokinin A are not affected by the mutation, which instead specifically alters the distribution between receptor sites in favour of a high affinity neurokinin A binding form.The low apparent affinity and binding capacity of neurokinin A on the wild type receptor results from neurokinin A binding with high affinity only to a fraction of the sites labelled by substance P. The mutation increases the proportion of this site, and consequently enhances the apparent affinity and binding capacity of neurokinin A.The binding modalities of septide-like ligands (i.e. neurokinin B, SP(6-11), SP-methyl ester) are affected similarly to neurokinin A and are better resolved into two sites. The mutation leaves the affinity of these ligands for the two receptor forms unchanged, but increases the fraction of high-affinity sites. On the other hand, the binding of non-peptide and peptide antagonists (SR140.333 and FK888) behaved similarly to substance P with a single high affinity site that is unaffected by the mutation.These findings may suggest that the NK1 receptor exists in two different forms with similar affinity for substance P and NK1 antagonists, but with a high and a low affinity for neurokinin A and septide-like ligands. Hence, the Gly166 in the NK1 receptor would seem to control the distribution between a pan-reactive form and a substance P-selective form of the receptor. PMID:9786514

Effect of chromatographic conditions and plasmid DNA size on the dynamic binding capacity of a monolithic support.

PubMed

Bicho, Diana; Sousa, Ângela; Sousa, Fani; Queiroz, João; Tomaz, Cãndida

2014-09-01

DNA therapies are becoming recognized alternatives for the treatment and prevention of severe pathologies. Although most current trials have used plasmids <10 kbp, in the future larger plasmids would be required. The purpose of this work was to study the chromatographic behavior of nongrafted carbonyldiimidazole monolithic disks using plasmids with different sizes under hydrophobic conditions. Thereunto, the purification of several plasmids was performed. Higher size plasmids needed lower ammonium sulfate concentration, due to the greater number of interactions between the plasmids and monolith. The dynamic binding capacity experiments for the different plasmids revealed a lower capacity for bigger plasmids. It was also verified that the increase of salt concentration from 2.5 to 3 M of ammonium sulfate increased the capacity. At the highest salt concentration, a slight improvement in the capacity using lower flow rate was observed, possibly due to compaction of plasmid molecules and its better organization on the monolith channels. Finally, a low pH also had a positive effect on the capacity. So, this monolithic support proved to be appropriate to purify the supercoiled isoform of different plasmids with different sizes, providing a valuable instrument as a purification technique. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Rapid incremental methods for the determination of serum iron and iron-binding capacity

PubMed Central

Beale, R. N.; Bostrom, J. O.; Taylor, R. F.

1961-01-01

Rapid methods depending on differential absorptiometry are described for the determination of the transferrin iron content and the latent iron-binding capacity of blood serum. Each determination requires as little as 0·5 ml. serum. The methods are well adapted for routine use in the `average' laboratory. Three or four sera may be completely analysed in 30 minutes. All operations are carried out in the cells or tubes used for the colorimetric measurements, no precipitation or heating being employed at any stage. Critical investigations of the reliability of the methods are attempted and ranges of normal values are included. PMID:13866116

Origin of low sodium capacity in graphite and generally weak substrate binding of Na and Mg among alkali and alkaline earth metals.

PubMed

Liu, Yuanyue; Merinov, Boris V; Goddard, William A

2016-04-05

It is well known that graphite has a low capacity for Na but a high capacity for other alkali metals. The growing interest in alternative cation batteries beyond Li makes it particularly important to elucidate the origin of this behavior, which is not well understood. In examining this question, we find a quite general phenomenon: among the alkali and alkaline earth metals, Na and Mg generally have the weakest chemical binding to a given substrate, compared with the other elements in the same column of the periodic table. We demonstrate this with quantum mechanics calculations for a wide range of substrate materials (not limited to C) covering a variety of structures and chemical compositions. The phenomenon arises from the competition between trends in the ionization energy and the ion-substrate coupling, down the columns of the periodic table. Consequently, the cathodic voltage for Na and Mg is expected to be lower than those for other metals in the same column. This generality provides a basis for analyzing the binding of alkali and alkaline earth metal atoms over a broad range of systems.

Xyloglucan-cellulose interaction depends on the sidechains and molecular weight of xyloglucan.

PubMed

Lima, Denis U; Loh, Watson; Buckeridge, Marcos S

2004-05-01

Recent papers have brought evidence against the hypothesis that the fucosyl branching of primary wall xyloglucans (Xg) are responsible for their higher capacity of binding to cellulose. Reinforcement of this questioning has been obtained in this work where we show that the binding capacity was improved when the molecular weight (MW) of the Xg polymers is decreased by enzymatic hydrolysis. Moreover, the enthalpy changes associated with the adsorption process between Xg and cellulose is similar for Xgs with similar MW (but differing in the fine structure such as the presence/absence of fucose). On the basis of these results, we suggest that the fine structure and MW of Xg determines the energy and amount of binding to cellulose, respectively. Thus, the occurrence of different fine structural domains of Xg (e.g. the presence of fucose and the distribution of galactoses) might have several different functions in the wall. Besides the structural function in primary wall, these results might have impact on the packing features of storage Xg in seed cotyledons, since the MW and absence of fucose could also be associated with the self-association capacity. Copyright 2004 Elsevier SAS

Differential inhibition of [3H]-oxotremorine-M and [3H]-quinuclinidyl benzilate binding to muscarinic receptors in rat brain membranes with acetylcholinesterase inhibitors.

PubMed

Lockhart, B; Closier, M; Howard, K; Steward, C; Lestage, P

2001-04-01

The potential interaction of acetylcholinesterase inhibitors with cholinergic receptors may play a significant role in the therapeutic and/or side-effects associated with this class of compound. In the present study, the capacity of acetylcholinesterase inhibitors to interact with muscarinic receptors was assessed by their ability to displace both [3H]-oxotremorine-M and [3H]-quinuclinidyl benzilate binding in rat brain membranes. The [3H]-quinuclinidyl benzilate/[3H]-oxotremorine-M affinity ratios permitted predictions to be made of either the antagonist or agonist properties of the different compounds. A series of compounds, representative of the principal classes of acetylcholinesterase inhibitors, displaced [3H]-oxotremorine-M binding with high-to-moderate potency (ambenonium>neostigmine=pyridostigmine=tacrine>physostigmine> edrophonium=galanthamine>desoxypeganine) whereas only ambenonium and tacrine displaced [3H]-quinuclinidyl benzilate binding. Inhibitors such as desoxypeganine, parathion and gramine demonstrated negligible inhibition of the binding of both radioligands. Scatchard plots constructed from the inhibition of [3H]-oxotremorine-M binding in the absence and presence of different inhibitors showed an unaltered Bmax and a reduced affinity constant, indicative of potential competitive or allosteric mechanisms. The capacity of acetylcholinesterase inhibitors, with the exception of tacrine and ambenonium, to displace bound [3H]-oxotremorine-M in preference to [3H]quinuclinidyl benzilate predicts that the former compounds could act as potential agonists at muscarinic receptors. Moreover, the rank order for potency in inhibiting acetylcholinesterase (ambenonium>neostigmine=physostigmine =tacrine>pyridostigmine=edrophonium=galanthamine >desoxypeganine>parathion>gramine) indicated that the most effective inhibitors of acetylcholinesterase also displaced [3H]-oxotremorine-M to the greatest extent. The capacity of these inhibitors to displace [3H]-oxotremorine-M binding preclude their utilisation for the prevention of acetylcholine catabolism in rat brain membranes, the latter being required to estimate the binding of acetylcholine to [3H]-oxotremorine-M-labelled muscarinic receptors. However, fasciculin-2, a potent peptide inhibitor of acetylcholinesterase (IC50 24 nM), did prevent catabolism of acetylcholine in rat brain membranes with an atypical inhibition isotherm of [3H]-oxotremorine-M binding, thus permitting an estimation of the "global affinity" of acetylcholine (Ki 85 nM) for [3H]-oxotremorine-M-labelled muscarinic receptors in rat brain.

Thermodynamics of T cell receptor – peptide/MHC interactions: progress and opportunities

PubMed Central

Armstrong, Kathryn M.; Insaidoo, Francis K.; Baker, Brian M.

2013-01-01

αβ T cell receptors (TCR) recognize peptide antigens presented by class I or class II major histocompatibility complex molecules (pMHC). Here we review the use of thermodynamic measurements in the study of TCR-pMHC interactions, with attention to the diversity in binding thermodynamics and how this is related to the variation in TCR-pMHC interfaces. We show that there is no enthalpic or entropic signature for TCR binding; rather, enthalpy and entropy changes vary in a compensatory manner that reflects a narrow free energy window for the interactions that have been characterized. Binding enthalpy and entropy changes do not correlate with structural features such as buried surface area or the number of hydrogen bonds within TCR-pMHC interfaces, possibly reflecting the myriad of contributors to binding thermodynamics, but likely also reflecting a reliance on van’t Hoff over calorimetric measurements and the unaccounted influence of equilibria linked to binding. TCR-pMHC binding heat capacity changes likewise vary considerably. In some cases the heat capacity changes are consistent with conformational differences between bound and free receptors, but there is little data indicating these conformational differences represent the need to organize commonly disordered CDR loops. In this regard, we discuss how thermodynamics may provide additional insight into conformational changes occurring upon TCR binding. Finally, we highlight opportunities for the further use of thermodynamic measurements in the study of TCR-pMHC interactions, not only for understanding TCR binding in general, but for understanding specifics of individual interactions and the engineering of T cell receptors with desired molecular recognition properties. PMID:18496839

Development of gastrointestinal surface. VIII. Lectin identification of carbohydrate differences

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pang, K.Y.; Bresson, J.L.; Walker, W.A.

Binding of microvillus membranes (MVM) from newborn and adult rats by concanavalin A (Con A), Ulex europaeus (UEA I), Dolichos bifluorus (DBA), and Triticum vulgaris (WGA) was examined to determine the availability of carbohydrate-containing sites for these lectins on the intestinal surface during development. Consistent patterns of differences in the reaction of MVM with these lectins were found. Con A and UEA had much higher reactivities to MVM of adult than newborn rats. /sup 125/I-labeled-UEA gel overlay experiments revealed the abundance of UEA-binding sites in MVM of adult rat in contrast to the two binding sites in MVM of amore » newborn rat. DBA bound only to MVM of the adults, and very few binding sites were found in immature MVM. In contrast to these lectins, WGA binding was much higher in MVM of the newborns and decreased with maturation. Additional experiments on the age dependence of UEA and DBA reactivities revealed that the most striking changes occur in animals from 2 to 2 wk of age. In MVM from 2-wk-old rats, there were only 13.9% and < 0.2% of the adult binding capacities for UEA and DBA, respectively. By the time the animals were 4 wk old, the binding capacity for UEA had attained close to the level of the adults, whereas for DBA it reached 71.3% of the adult value. These results provide definite evidence of changes in the intestinal surface during perinatal development.« less

Extracellular polymeric substances facilitate the biosorption of phenanthrene on cyanobacteria Microcystis aeruginosa.

PubMed

Bai, Leilei; Xu, Huacheng; Wang, Changhui; Deng, Jiancai; Jiang, Helong

2016-11-01

Phytoplankton-derived extracellular polymeric substances (EPS) are of vital importance for the biogeochemical cycles of hydrophobic organic pollutants in lake ecosystems. In this study, roles of loosely-bound EPS (LB-EPS) and tightly bound EPS (TB-EPS) in biosorption of phenanthrene (PHE) on a typical cyanobacteria Microcystis aeruginosa were investigated. The results showed that the biosorption of PHE on M. aeruginosa cell varied lasted 24 h, while the binding of PHE to LB-EPS and TB-EPS reached equilibrium within less than 2 h. The equilibrium biosorption capacities of M. aeruginosa cell, LB-EPS and TB-EPS were 6.78, 12.31, and 9.47 μg mg(-1), respectively, indicating that the binding of PHE to EPS was a considerable process involved in biosorption. Fluorescence quenching titration revealed that increasing temperature induced more binding sites in EPS for PHE and the binding process was driven by electrostatic force and hydrophobic interactions. Interestingly, dynamic and static quenching processes occurred simultaneously for the binding of PHE to protein-like substances in EPS, whereas the binding of PHE to humic-like substances belonged to static quenching. The relatively higher contents of proteins in LB-EPS produced a stronger binding capacity of PHE. Overall, the interactions between hydrophobic organic pollutants and cyanobacterial EPS are favorable to the bioaccumulation of hydrophobic organic pollutants in cyanobacteria and facilitate the regulatory function of cyanobacterial biomass as a biological pump. Copyright © 2016 Elsevier Ltd. All rights reserved.

High affinity binding of 125I-neurotensin to dispersed cells from chicken liver and brain.

PubMed

Mitra, S P; Carraway, R E

1997-01-01

Dispersed cells from chicken brain and liver were found to possess cell surface binding sites for 125I-neurotensin (125I-NT). Scatchard analyses indicated the presence of high affinity (K4, 25-80 pM) and low affinity (Kd, 250-450 pM) components in adult tissues. Binding capacity was reduced 25-40% by incubation with pertussis toxin. Ontogenetic studies indicated that NT receptor capacity increased approximately 20-fold from the embryonic stage to adult. Cross-linking of 125I-NT to intact cells labeled one major band (52 kDa, > or = 90%) and two minor bands (40 and 90 kDa, < or = 10%) which could represent distinct NT-receptors or one receptor partly degraded or cross-linked to G-protein(s). The binding of 125I-NT to dispersed cells was enhanced by reduction with dithoithreitol and suppressed by alkylation with N-ethyl-maleimide (NEM), maleimidocaproic acid (MCA) and p-chloromercuribenzenesulfonate (PCMBS). Since MCA and PCMBS do not permeate cells, this suggests that the sulfhydryl group(s) critical to binding are located within the NT receptor itself. Preincubation of cells with NT prior to treatment with NEM diminished its inhibitory effect, suggesting that the critical SH-group(s) were within the NT binding pocket or were protected by an allosteric effect. These results suggest that one or more of the nine cysteine residues in the NT receptor is involved in the NT binding reaction.

Endocrine changes of Paralichthys olivaceus after oral administration with exogenous growth hormone

NASA Astrophysics Data System (ADS)

Liu, Zong-Zhu; Xu, De-Wu; Wang, Yong; Xu, Yong-Li; Zhang, Pei-Jun

2000-12-01

Recombinant salmon growth hormone contained in yeast was given for 5 months to flounder in its diet. Both free and total specific binding sites for the growth hormone were examined in liver membranes of control and treated fish. The association constants of both free and total specific binding sites were of the same order (1 nM-1), and no significant difference was found between any two groups in the capacity of their free binding sites. The capacity of total binding sites in the liver of treated fish increased significantly compared with that of control. Insulin-like growth factor I (IGF-I) levels in the plasma of treated fish increased by 22.61% (P<0.05), compared with that of control. While the T4 levels in plasma did not increase significantly (from 1.35±0.91 ng/ml to 2.29±1.13 ng/ml), T3 levels were elevated significantly (from 1.78±1.14 ng/ml to 4.87±1.22 ng/ml, P<0.01), as compared with that of control.

Heat Capacity Changes and Disorder-to-Order Transitions in Allosteric Activation.

PubMed

Cressman, William J; Beckett, Dorothy

2016-01-19

Allosteric coupling in proteins is ubiquitous but incompletely understood, particularly in systems characterized by coupling over large distances. Binding of the allosteric effector, bio-5'-AMP, to the Escherichia coli biotin protein ligase, BirA, enhances the protein's dimerization free energy by -4 kcal/mol. Previous studies revealed that disorder-to-order transitions at the effector binding and dimerization sites, which are separated by 33 Å, are integral to functional coupling. Perturbations to the transition at the ligand binding site alter both ligand binding and coupled dimerization. Alanine substitutions in four loops on the dimerization surface yield a range of energetic effects on dimerization. A glycine to alanine substitution at position 142 in one of these loops results in a complete loss of allosteric coupling, disruption of the disorder-to-order transitions at both functional sites, and a decreased affinity for the effector. In this work, allosteric communication between the effector binding and dimerization surfaces in BirA was further investigated by performing isothermal titration calorimetry measurements on nine proteins with alanine substitutions in three dimerization surface loops. In contrast to BirAG142A, at 20 °C all variants bind to bio-5'-AMP with free energies indistinguishable from that measured for wild-type BirA. However, the majority of the variants exhibit altered heat capacity changes for effector binding. Moreover, the ΔCp values correlate with the dimerization free energies of the effector-bound proteins. These thermodynamic results, combined with structural information, indicate that allosteric activation of the BirA monomer involves formation of a network of intramolecular interactions on the dimerization surface in response to bio-5'-AMP binding at the distant effector binding site.

Multilayer affinity adsorption of albumin on polymer brushes modified membranes in a continuous-flow system.

PubMed

Hu, Meng-Xin; Li, Xiang; Li, Ji-Nian; Huang, Jing-Jing; Ren, Ge-Rui

2018-02-23

Polymer brushes modified surfaces have been widely used for protein immobilization and isolation. Modification of membranes with polymer brushes increases the surface concentration of affinity ligands used for protein binding. Albumin is one of the transporting proteins and shows a high affinity to bile acids. In this work, the modified membranes with cholic acid-containing polymer brushes can be facilely prepared by the immobilization of cholic acid on the poly(2-hydroxyethyl methacrylate) grafted microporous polypropylene membranes (MPPMs) for affinity adsorption of albumin. ATR/FT-IR and X-ray photoelectron spectroscopy were used to characterize the chemical composition of the modified membranes. Water contact angle measurements were used to analyze the hydrophilic/hydrophobic properties of the membrane surface. The modified MPPMs show a high affinity to albumin and have little non-specific adsorption of hemoglobin. The dynamic binding capacity of albumin in the continous-flow system increases with the cycle number and feed rate as the binding degree of cholic acid is moderate. The highest binding capacity of affinity membranes is about 52.49 g/m 2 membrane, which is about 24 times more than the monolayer binding capacity. These results reveal proteins could be captured in multilayers by the polymer brushes containing affinity ligands similar to the polymer brushes containing ion-exchange groups, which open up the potential of the polymer brushes containing affinity ligands in protein or another components separation. And the cholic acid containing polymer brushes modified membranes has the promising potential for albumin separation and purification rapidly from serum or fermented solution in medical diagnosis and bioseparation. Copyright © 2018 Elsevier B.V. All rights reserved.

Thermodynamics of Interaction between Some Cellulose Ethers and SDS by Titration Microcalorimetry.

PubMed

Singh; Nilsson

1999-05-01

The interaction between certain nonionic cellulose ethers (ethyl hydroxyethyl cellulose and hydroxypropyl methyl cellulose) and sodium dodecyl sulphate (SDS) has been investigated using isothermal titration microcalorimetry at temperatures between 25-50 degrees C. The observed heat flow curves have been interpreted in terms of a plausible mechanism of the interaction of the substituent groups with SDS monomers and clusters. The data have been related to changes occuring in the system at the macro- and microscopic levels with the addition of surfactants and with temperature. The process consists predominantly of polymer-surfactant interactions initially and surfactant-surfactant interactions at the later stages. A phenomenological model of the cooperative interaction (adsorption) process has been derived, and earlier published equilibrium binding data have been used to recover binding constants and Gibbs energy changes for this process. The adsorption enthalpies and entropies have been recovered along with the heat capacity change. The enthalpic cost of confining the nonpolar regions of the polymers in surfactant clusters is high, but the entropy gain from release of hydration shell water molecules as well as increased freedom of movement of these nonpolar regions in the clusters gives the process a strong entropic driving force. The process is entropy-driven initially and converts to being both enthalpy and entropy-driven at high SDS concentrations. An enthalpy-entropy compensation behavior is seen. Strongly negative heat capacity changes have been obtained resulting from the transfer of nonpolar groups from aqueous into nonpolar environments, as well as a reduction of conformational domains that the chains can populate. Changes in these two components cause the heat capacity change to become less negative at the higher binding levels. The system can be classified as exhibiting nonclassical hydrophobic binding at the later stages of binding. Copyright 1999 Academic Press.

Distribution and Kinetics of Lipoprotein-Bound Lipoteichoic Acid

PubMed Central

Levels, Johannes H. M.; Abraham, Philip R.; van Barreveld, Erik P.; Meijers, Joost C. M.; van Deventer, Sander J. H.

2003-01-01

Lipoteichoic acid (LTA), a major cell wall component of gram-positive bacteria, is an amphipathic anionic glycolipid with structural similarities to lipopolysaccharide (LPS) from gram-negative bacteria. LTA has been implicated as one of the primary immunostimulatory components that may trigger the systemic inflammatory response syndrome. Plasma lipoproteins have been shown to sequester LPS, which results in attenuation of the host response to infection, but little is known about the LTA binding characteristics of plasma lipid particles. In this study, we have examined the LTA binding capacities and association kinetics of the major lipoprotein classes under simulated physiological conditions in human whole blood (ex vivo) by using biologically active, fluorescently labeled LTA and high-performance gel permeation chromatography. The average distribution of an LTA preparation from Staphylococcus aureus in whole blood from 10 human volunteers revealed that >95% of the LTA was associated with total plasma lipoproteins in the following proportions: high-density lipoprotein (HDL), 68% ± 10%; low-density lipoprotein (LDL), 28% ± 8%; and very low density lipoprotein (VLDL), 4% ± 5%. The saturation capacity of lipoproteins for LTA was in excess of 150 μg/ml. The LTA distribution was temperature dependent, with an optimal binding between 22 and 37°C. The binding of LTA by lipoproteins was essentially complete within 10 min and was followed by a subsequent redistribution from HDL and VLDL to LDL. We conclude that HDL has the highest binding capacity for LTA and propose that the loading and redistribution of LTA among plasma lipoproteins is a specific process that closely resembles that previously described for LPS (J. H. M. Levels, P. R. Abraham, A. van den Ende, and S. J. H. van Deventer, Infect. Immun. 68:2821-2828, 2001). PMID:12761109

Glucocorticoid receptor ligand binding in monocytic cells using a microplate assay.

PubMed

Jansen, J; Uitdehaag, B; Koper, J W; van Den Berg, T K

1999-01-01

Glucocorticoids have profound effects on macrophage function and are widely used as anti-inflammatory drugs. Glucocorticoids receptor (GR) ligand binding capacity is a major determinant of cellular glucocorticoid sensitivity. The number and affinity of GR can be measured in a whole cell binding assay using (3)H-dexamethasone. Here, we describe a rapid and simple microplate assay for GR measurement using the human promonocytic cell line THP-1. Copyright 2000 S. Karger AG, Basel.

Comparative studies on drug binding to the purified and pharmaceutical-grade human serum albumins: Bridging between basic research and clinical applications of albumin.

PubMed

Ashrafi-Kooshk, Mohammad Reza; Ebrahimi, Farangis; Ranjbar, Samira; Ghobadi, Sirous; Moradi, Nastaran; Khodarahmi, Reza

2015-09-01

Human serum albumin (HSA), the most abundant protein in blood plasma, is a monomeric multidomain protein that possesses an extraordinary capacity for binding, so that serves as a circulating depot for endogenous and exogenous compounds. During the heat sterilization process, the structure of pharmaceutical-grade HSA may change and some of its activities may be lost. In this study, to provide deeper insight on this issue, we investigated drug-binding and some physicochemical properties of purified albumin (PA) and pharmaceutical-grade albumin (PGA) using two known drugs (indomethacin and ibuprofen). PGA displayed significantly lower drug binding capacity compared to PA. Analysis of the quenching and thermodynamic parameters indicated that intermolecular interactions between the drugs and the proteins are different from each other. Surface hydrophobicity as well as the stability of PGA decreased compared to PA, also surface hydrophobicity of PA and PGA increased upon drugs binding. Also, kinetic analysis of pseudo-esterase activities indicated that Km and Vmax parameters for PGA enzymatic activity are more and less than those of PA, respectively. This in vitro study demonstrates that the specific drug binding of PGA is significantly reduced. Such studies can act as connecting bridge between basic research discoveries and clinical applications. Copyright © 2015 The International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.

The Glycine Synaptic Receptor: Evidence That Strychnine Binding Is Associated with the Ionic Conductance Mechanism

PubMed Central

Young, Anne B.; Snyder, Solomon H.

1974-01-01

The ability of a series of anions to inhibit [3H]strychnine binding to spinal cord synaptic membranes correlates closely with their neurophysiologic capacity to reverse inhibitory postsynaptic potentials in the mammalian spinal cord. Seven neurophysiologically active anions are also effective inhibitors of [3H]strychnine binding with mean effective doses ranging from 160 to 620 mM. Seven other anions that are ineffective neurophysiologically also fail to alter strychnine binding. Chloride inhibits strychnine binding in a noncompetitive fashion. Hill plots of the displacement of [3H]strychnine by chloride give coefficients of 2.3-2.7. The inhibition of strychnine binding by these anions suggests that strychnine binding is closely associated with the ionic conductance mechanism for chloride in the glycine receptor. PMID:4372600

Binding of corroded ions to human saliva.

PubMed

Mueller, H J

1985-05-01

Employing equilibrium dialysis, the binding abilities of Cu, Al, Co and Cr ions from corroded Cu-Al and Co-Cr dental casting alloys towards human saliva and two of its gel chromatographic fractions were determined. Results indicate that both Cu and Co bind to human saliva i.e. 0.045 and 0.027 mg/mg protein, respectively. Besides possessing the largest binding ability, Cu also possessed the largest binding capacity. The saturation of Cu binding was not reached up to the limit of 0.35 mg protein/ml employed in the tests, while Co reached full saturation at about 0.2 mg protein/ml. Chromium showed absolutely no binding to human saliva while Al ions did not pass through the dialysis membranes. Compared to the binding with solutions that were synthetically made up to contain added salivary-type proteins, it is shown that the binding to human saliva is about 1 order of magnitude larger, at least for Cu ions.

Prion removal effect of a specific affinity ligand introduced into the manufacturing process of the pharmaceutical quality solvent/detergent (S/D)-treated plasma OctaplasLG.

PubMed

Neisser-Svae, A; Bailey, A; Gregori, L; Heger, A; Jordan, S; Behizad, M; Reichl, H; Römisch, J; Svae, T-E

2009-10-01

A new chromatographic step for the selective binding of abnormal prion protein (PrP(Sc)) was developed, and optimization for PrP(Sc) capture was achieved by binding to an affinity ligand attached to synthetic resin particles. This step was implemented into the manufacturing process of the solvent/detergent (S/D)-treated biopharmaceutical quality plasma Octaplas to further improve the safety margin in terms of risk for variant Creutzfeldt-Jakob disease (vCJD) transmission. Intermediates and Octaplas final container material, spiked with hamster brain-derived PrP(Sc)-containing fractions, were used for experiments to establish the feasibility of introducing this novel chromatography step. The binding capacity per millilitre of ligand gel was determined under the selected manufacturing conditions. In addition, the specificity of the ligand gel to bind PrP(Sc) from human sources was investigated. A validated Western blot test was used for the identification and quantification of PrP(Sc). A reduction factor of > or = 3.0 log(10) could be demonstrated by Western blotting, utilizing the relevant Octaplas matrix from manufacturing. In this particular cell-free plasma solution, the PrP(Sc) binding capacity of the selected gel was very high (> or = 6 log(10) ID(50)/ml, equivalent to roughly 10 log(10) ID(50)/column at manufacturing scale). The gel binds specifically PrP(Sc) from both animal (hamster and mouse) and human (sporadic and variant CJD) sources. This new single-use, disposable PrP(Sc)-harvesting gel ensures a very high capacity in terms of removing the pathogenic agent causing vCJD from the new generation OctaplasLG, in the event that prions can be found in plasma from donors incubating the disease and thereby contaminating the raw material plasma used for manufacturing.

Tea Dietary Fiber Improves Serum and Hepatic Lipid Profiles in Mice Fed a High Cholesterol Diet.

PubMed

Guo, Wenxin; Shu, Yang; Yang, Xiaoping

2016-06-01

Tea dietary fiber (TDF) was prepared from tea residues and modified to get cellulose-modified TDF (CTDF) by cellulase or micronized TDF (MTDF) by ultrafine grinding. The in vitro lipid-binding capacities of the three fibers and their effects on serum and hepatic lipid profiles in mice fed a high cholesterol diet were evaluated. The results showed that the three fibers had excellent lipid-binding capacities, and the cholesterol- and sodium cholate-binding capacities of CTDF and MTDF were significantly higher than those of TDF. Animal studies showed that, compared to model control, the three fibers significantly decreased mice average daily gain, gain: feed, and liver index, reduced total cholesterol (TC), triglyceride, and low density lipoprotein-cholesterol of serum and liver, increased serum and hepatic high density lipoprotein-cholesterol to TC ratio, and promoted the excretion of fecal lipids, and they also significantly increased the activities of superoxide dismutase and glutathione peroxidase of serum and liver, and decreased lipid peroxidation; moreover, the effects of CTDF and MTDF were better than that of TDF. It was concluded that the three fibers could improve serum and hepatic lipid profiles in mice fed a high cholesterol diet and the mechanism of action might be due to the promotion of fecal excretion of lipids through their lipid-binding ability and the inhibition of lipid peroxidation. These findings suggest that tea dietary fiber has the potential to be used as a functional ingredient to control cardiovascular disease.

Effects of saw palmetto extract on micturition reflex of rats and its autonomic receptor binding activity.

PubMed

Oki, Tomomi; Suzuki, Mayumi; Nishioka, Yasuhiko; Yasuda, Akio; Umegaki, Keizo; Yamada, Shizuo

2005-04-01

We examined the effects of saw palmetto extract (SPE) on the rat micturition reflex and on autonomic receptors in the lower urinary tract. The effect of SPE was examined on cystometrograms of anesthetized rats induced by intravesical infusion of saline or 0.1% acetic acid. SHR/NDmc-cp (cp/cp) rats received repeat oral administration of SPE and nighttime urodynamic function was determined. The autonomic receptor binding activity of SPE in the rat bladder and prostate was examined by radioligand binding assay. Intraduodenal administration of SPE (60 mg/kg) in anesthetized rat cystometry caused a significant increase in the micturition interval, micturition volume and bladder capacity during intravesical saline infusion. Also, similar administration of SPE at doses of 12 and 20 mg/kg significantly reversed the shortened micturition interval as well as the decreased micturition volume and bladder capacity due to 0.1% acetic acid infusion in a dose dependent manner. In conscious SHR/NDmc-cp (cp/cp) rats repeat oral administration of SPE (6 mg/kg daily) constantly increased the micturition interval and concomitantly decreased voiding frequency. SPE inhibited specific binding of [H]NMS ([N-methyl-H]scopolamine methyl chloride) (bladder) and [H]prazosin (prostate) with IC50 values of 46.1 and 183 microg/ml, respectively. SPE significantly alleviates urodynamic symptoms in hyperactive rat bladders by increasing bladder capacity and subsequently prolonging the micturition interval. Our data may support the clinical efficacy of SPE for the treatment of lower urinary tract symptoms.

Paradoxical roles of hydrogen in electrochemical performance of graphene: High rate capacity and atomistic origins

DOE PAGES

Ye, Jianchao C.; Ong, Mitchell T.; Heo, Tae Wook; ...

2015-11-05

Atomic hydrogen exists ubiquitously in graphene materials made by chemical methods. Yet determining the effect of hydrogen on the electrochemical performance of graphene remains a significant challenge. Here we report the experimental observations of high rate capacity in hydrogen-treated 3-dimensional (3D) graphene nanofoam electrodes for lithium ion batteries. Structural and electronic characterization suggests that defect sites and hydrogen play synergistic roles in disrupting sp 2 graphene to facilitate fast lithium transport and reversible surface binding, as evidenced by the fast charge-transfer kinetics and increased capacitive contribution in hydrogen-treated 3D graphene. In concert with experiments, multiscale calculations reveal that defect complexesmore » in graphene are prerequisite for low-temperature hydrogenation, and that the hydrogenation of defective or functionalized sites at strained domain boundaries plays a beneficial role in improving rate capacity by opening gaps to facilitate easier Li penetration. Additional reversible capacity is provided by enhanced lithium binding near hydrogen-terminated edge sites. Furthermore, these findings provide qualitative insights in helping the design of graphene-based materials for high-power electrodes.« less

Universal roles of hydrogen in electrochemical performance of graphene: high rate capacity and atomistic origins

PubMed Central

Ye, Jianchao; Ong, Mitchell T.; Heo, Tae Wook; Campbell, Patrick G.; Worsley, Marcus A.; Liu, Yuanyue; Shin, Swanee J.; Charnvanichborikarn, Supakit; Matthews, Manyalibo J.; Bagge-Hansen, Michael; Lee, Jonathan R.I.; Wood, Brandon C.; Wang, Y. Morris

2015-01-01

Atomic hydrogen exists ubiquitously in graphene materials made by chemical methods. Yet determining the effect of hydrogen on the electrochemical performance of graphene remains a significant challenge. Here we report the experimental observations of high rate capacity in hydrogen-treated 3-dimensional (3D) graphene nanofoam electrodes for lithium ion batteries. Structural and electronic characterization suggests that defect sites and hydrogen play synergistic roles in disrupting sp2 graphene to facilitate fast lithium transport and reversible surface binding, as evidenced by the fast charge-transfer kinetics and increased capacitive contribution in hydrogen-treated 3D graphene. In concert with experiments, multiscale calculations reveal that defect complexes in graphene are prerequisite for low-temperature hydrogenation, and that the hydrogenation of defective or functionalized sites at strained domain boundaries plays a beneficial role in improving rate capacity by opening gaps to facilitate easier Li penetration. Additional reversible capacity is provided by enhanced lithium binding near hydrogen-terminated edge sites. These findings provide qualitative insights in helping the design of graphene-based materials for high-power electrodes. PMID:26536830

Gold Binding by Native and Chemically Modified Hops Biomasses

DOE PAGES

López, M. Laura; Gardea-Torresdey, J. L.; Peralta-Videa, J. R.; ...

2005-01-01

Heavy metals from mining, smelting operations and other industrial processing facilities pollute wastewaters worldwide. Extraction of metals from industrial effluents has been widely studied due to the economic advantages and the relative ease of technical implementation. Consequently, the search for new and improved methodologies for the recovery of gold has increased. In this particular research, the use of cone hops biomass ( Humulus lupulus ) was investigated as a new option for gold recovery. The results showed that the gold binding to native hops biomass was pH dependent from pH 2 to pH 6, with a maximum percentage binding atmore » pH 3. Time dependency studies demonstrated that Au(III) binding to native and modified cone hops biomasses was found to be time independent at pH 2 while at pH 5, it was time dependent. Capacity experiments demonstrated that at pH 2, esterified hops biomass bound 33.4 mg Au/g of biomass, while native and hydrolyzed hops biomasses bound 28.2 and 12.0 mg Au/g of biomass, respectively. However, at pH 5 the binding capacities were 38.9, 37.8 and 11.4 mg of Au per gram of native, esterified and hydrolyzed hops biomasses, respectively.« less

The in vitro interactions between serum lipoproteins and proteoglycans of the neointima of rabbit aorta after a single balloon catheter injury.

PubMed

Alavi, M Z; Richardson, M; Moore, S

1989-02-01

The effect of injury-induced alterations in the aortic neointimal proteoglycans on their binding with homologous serum lipoproteins was examined. Proteoglycans of the aortic intimal-medial tissues of rabbits that had undergone denudation with a balloon catheter 12 weeks earlier were isolated after homogenization of the tissues in 0.33 M sucrose, ultracentrifugation and subsequently by gel-exclusion chromatography. Lipoproteins from the plasma of healthy donors were prepared by sequential, ultracentrifugal floatation after density adjustment with KBr. To study the interactions, aliquots of electrophoretically pure very low-density lipoproteins (VLDL, d less than 1.006 g/ml), low-density lipoproteins (LDL, d = 1.019-1.063 g/ml), or high-density lipoproteins (HDL, d = 1.210 g/ml) were incubated with proteoglycans in the presence of Ca++ and Mg++ at 4 C. The amount of cholesterol found in the resulting pellet was measured as a marker of the binding capacity of the proteoglycans. Among lipoprotein fractions both VLDL and LDL showed strong binding with proteoglycans, whereas no appreciable binding was observed when incubation experiments were done with HDL. There were significant differences in the lipoprotein binding capacity of proteoglycan of control and injured animals, indicating that injury induced changes in proteoglycan composition exert profound influences on their ionic interactions.

Exercise-induced TBC1D1 Ser237 phosphorylation and 14-3-3 protein binding capacity in human skeletal muscle.

PubMed

Frøsig, Christian; Pehmøller, Christian; Birk, Jesper B; Richter, Erik A; Wojtaszewski, Jørgen F P

2010-11-15

TBC1D1 is a Rab-GTPase activating protein involved in regulation of GLUT4 translocation in skeletal muscle. We here evaluated exercise-induced regulation of TBC1D1 Ser237 phosphorylation and 14-3-3 protein binding capacity in human skeletal muscle. In separate experiments healthy men performed all-out cycle exercise lasting either 30 s, 2  min or 20  min. After all exercise protocols, TBC1D1 Ser237 phosphorylation increased (∼70-230%, P < 0.005), with the greatest response observed after 20  min of cycling. Interestingly, capacity of TBC1D1 to bind 14-3-3 protein showed a similar pattern of regulation, increasing 60-250% (P < 0.001). Furthermore, recombinant 5AMP-activated protein kinase (AMPK) induced both Ser237 phosphorylation and 14-3-3 binding properties on human TBC1D1 when evaluated in vitro. To further characterize the role of AMPK as an upstream kinase regulating TBC1D1, extensor digitorum longus muscle (EDL) from whole body α1 or α2 AMPK knock-out and wild-type mice were stimulated to contract in vitro. In wild-type and α1 knock-out mice, contractions resulted in a similar ∼100% increase (P < 0.001) in Ser237 phosphorylation. Interestingly, muscle of α2 knock-out mice were characterized by reduced protein content of TBC1D1 (∼50%, P < 0.001) as well as in basal and contraction-stimulated (∼60%, P < 0.001) Ser237 phosphorylation, even after correction for the reduced TBC1D1 protein content. This study shows that TBC1D1 is Ser237 phosphorylated and 14-3-3 protein binding capacity is increased in response to exercise in human skeletal muscle. Furthermore, we show that the catalytic α2 AMPK subunit is the main (but probably not the only) donor of AMPK activity regulating TBC1D1 Ser237 phosphorylation in mouse EDL muscle.

Specific binding of magnetic nanoparticle probes to platelets in whole blood detected by magnetorelaxometry

NASA Astrophysics Data System (ADS)

Eberbeck, Dietmar; Wiekhorst, Frank; Steinhoff, Uwe; Schwarz, Kay Oliver; Kummrow, Andreas; Kammel, Martin; Neukammer, Jörg; Trahms, Lutz

2009-05-01

The binding of monoclonal antibodies labelled with magnetic nanoparticles to CD61 surface proteins expressed by platelets in whole blood samples was measured by magnetorelaxometry. This technique is sensitive to immobilization of the magnetic labels upon binding. Control experiments with previous saturation of the epitopes on the platelet surfaces demonstrated the specificity of the binding. The kinetics of the antibody antigen reaction is accessible with a temporal resolution of 12 s. The minimal detectable platelet concentration is about 2000 μL -1 (sample volume 150 μL). The proportionality of the magnetic relaxation amplitude to the number of bound labels allows a quantification of the antibody binding capacity.

Staphylococcal surface display of metal-binding polyhistidyl peptides

DOE Office of Scientific and Technical Information (OSTI.GOV)

Samuelson, P.; Wernerus, H.; Svedberg, M.

2000-03-01

Recombinant Staphylococcus xylosus and Staphylococcus carnosus strains were generated with surface-exposed chimeric proteins containing polyhistidyl peptides designed for binding to divalent metal ions. Surface accessibility of the chimeric surface proteins was demonstrated and the chimeric surface proteins were found to be functional in terms of metal binding, since the recombinant staphylococcal cells were shown to have gained Ni{sup 2+}- and Cd{sup 2+}-binding capacity, suggesting that such bacteria could find use in bioremediation of heavy metals. This is, to their knowledge, the first time that recombinant, surface-exposed metal-binding peptides have been expressed on gram-positive bacteria. Potential environmental or biosensor applications formore » such recombinant staphylococci as biosorbents are discussed.« less

The effect of vegetarian diet on selected essential nutrients in children.

PubMed

Laskowska-Klita, Teresa; Chełchowska, Magdalena; Ambroszkiewicz, Jadwiga; Gajewska, Joanna; Klemarczyk, Witold

2011-01-01

Vegetarian diets are considered to promote health and reduce the risk of some chronic diseases. It is also known that restriction or exclusion of animal foods may result in low intake of essential nutrients. The aim of the presented study was to assess the intake and serum status of vitamin B12, folate, vitamins A, E and D, as well as concentrations of homocysteine, total antioxidant status and iron balance in Polish vegetarian children. The study included 50 children, aged 5-11 who had been referred to the Institute of Mother and Child for dietary consultation. From those, 32 were vegetarians (aged 6.5±4.2 years) and 18 omnivores (aged 7.9±2.7 years). Dietary constituents were analyzed using the nutritional programme Dietetyk2®. Folate and vitamin B12 were determined with a chemiluminescence immunoassay, total homocysteine with a fluorescence polarization immunoassay and TAS (total antioxidant status) by colorimetric method. Vitamin A and E in serum were determined by the high-pressure liquid chromatography method (HPLC) and vitamin D by immunoenzymatic assay (ELISA). Concentrations of iron, ferritin, transferrin and total iron-binding capacity (TIBC) in serum were determined by commercially available kits. In vegetarian children daily intake of vitamin B12 (1.6 ěg) was in the recommended range, that of folate (195 ěg) and vitamin A (1245 ěg) higher, but vitamin E slightly lower (6.6 ěg) and three-fold lower vitamin D (1.1 ěg) than references allowance. Serum concentrations of vitamin B12 (548 pg/ml), folate (12.8 ng/ml), vitamin A (1.2 ěmol/L), vitamin E (15.6 ěmol/l) were within physiological range, but that of vitamin D (13.7 ěg/L) was only half of the lowest limit of the reference value. In vegetarian children in comparison to omnivorous similar levels of homocysteine (6.13 ěmol/L vs 5.45 ěmol/L) and vitamin A (1,17 ěmol/L vs 1.32 ěmol/L) were observed. Lower (p<0.05) values of vitamin E (15.6 ěmol/L vs 18.4 ěmol/L) and TAS (1.21 mmol/L vs 1.30 mmol/L; p<0.0001) were found. Concentrations of iron markers were in physiological range. Obtained results indicated that intakes of vitamin B12 and folic acid from vegetarian diets are sufficient to maintain serum concentrations of both homocysteine and iron in the range observed in omnivorous children. High consumption of vitamin A and low vitamin E only slightly affected their serum values. Significantly lower concentration of serum vitamin E in vegetarian children in comparison to nonvegetarians may be reflected with statistically significant lowering of total antioxidant status. Insufficient intake of vitamin D and its low serum concentration should be under close monitoring in vegetarian children. In order to prevent vitamin D deficiency appropriate age-dependent supplementation should be considered.

Fundamental considerations in ski binding analysis.

PubMed

Mote, C D; Hull, M L

1976-01-01

1. The static adjustment of a ski binding by hand or by available machines is only an adjustment and is neither a static nor a dynamic evaluation of the binding design. Bindings of different design with identical static adjustments will perform differently in environments in which the forces are static or dynamic. 2. The concept of binding release force is a useful measure of binding adjustment, but it is inappropriate as a criterion for binding evaluation. First, it does not direct attention toward the injury causing mechanism, strain, or displacement in the leg. Second, it is only part of the evaluation in dynamic problems. 3. The binding release decision in present bindings is displacement controlled. The relative displacement of the boot and ski is the system variable. For any specified relative displacement the binding force can be any of an infinite number of possibilities determined by the loading path. 4. The response of the leg-ski system to external impulses applied to the ski is independent of the boot-ski relative motion as long as the boot recenters quickly in the binding. Response is dependent upon the external impulse plus system inertia, damping and stiffness. 5. When tested under half sinusoidal forces applied to a test ski, all bindings will demonstrate static and impulse loading regions. In the static region the force drives the binding to a relative release displacement. In the impulse region the initial velocity of the ski drives the binding to a release displacement. 6. The transition between the static and impulse loading regions is determined by the binding's capacity to store and dissipate energy along the principal loading path. Increased energy capacity necessitates larger external impulses to produce release. 7. In all bindings examined to date, the transmitted leg displacement or strain at release under static loading exceeds leg strain under dynamic or impact loading. Because static loading is responsible for many injuries, a skier should be able to release his bindings in every mode by simply pulling or twisting his foot outward. If that cannot be done without injury, the skier has identified for himself one type of fall that will result in injury. 8. And lastly, a little advice from Ben Franklin--"Carelessness does more harm than a want of knowledge."

Structural Reorganization and the Cooperative Binding of Single-stranded Telomere DNA in Sterkiella nova*

PubMed Central

Buczek, Pawel; Horvath, Martin P.

2009-01-01

In Sterkiella nova, α and β telomere proteins bind cooperatively with single-stranded DNA to form a ternary α·β·DNA complex. Association of telomere protein subunits is DNA-dependent, and α-β association enhances DNA affinity. To further understand the molecular basis for binding cooperativity, we characterized several possible stepwise assembly pathways using isothermal titration calorimetry. In one path, α and DNA first form a stable α·DNA complex followed by addition of β in a second step. Binding energy accumulates with nearly equal free energy of association for each of these steps. Heat capacity is nonetheless dramatically different with ΔCp = −305 ± 3 cal mol−1 K−1 for α binding with DNA and ΔCp = −2010 ± 20 cal mol−1 K−1 for addition of β to complete the α·β·DNA complex. By examining alternate routes including titration of single-stranded DNA with a preformed α·β complex, a significant portion of binding energy and heat capacity could be assigned to structural reorganization involving protein-protein interactions and repositioning of the DNA. Structural reorganization probably affords a mechanism to regulate high affinity binding of telomere single-stranded DNA with important implications for telomere biology. Regulation of telomere complex dissociation is thought to involve post-translational modifications in the lysine-rich C-terminal portion of β. We observed no difference in binding energetics or crystal structure when comparing complexes prepared with full-length β or a C-terminally truncated form, supporting interesting parallels between the intrinsically disordered regions of histones and this portion of β. PMID:17082188

Potential of goat probiotic to bind mutagens.

PubMed

Apás, Ana Lidia; González, Silvia Nelina; Arena, Mario Eduardo

2014-08-01

The mutagen binding ability of the goat probiotics (Lactobacillus reuteri DDL 19, Lactobacillus alimentarius DDL 48, Enterococcus faecium DDE 39, and Bifidobacterium bifidum DDBA) was evaluated. The oral administration of these probiotics reduced fecal mutagens and intestinal cancer markers in goats. Secondly, the effects of probiotics against the mutagenesis induced by sodium azide (SA), and Benzopyrene (B[α]P) by performing the modified Ames test using Salmonella typhimurium TA 100 was investigated. The capacity to bind benzopyrene and the stability of the bacterial-mutagen complex was analyzed by HPLC. The dismutagenic potential against both mutagens was proportional to probiotic concentration. Results showed that probiotic antimutagenic capacity against SA was ranging from 13 to 78%. The mixture of four goat probiotics (MGP) displayed higher antimutagenic activity against SA than any individual strains at the same cell concentration. This study shows that the highest diminution of mutagenicity in presence of B[α]P (74%) was observed in presence of MGP. The antimutagenic activity of nearly all the individual probiotic and the MGP were in concordance with the B[α]P binding determined by HPLC. According to our results, the B[α]P binding to probiotic was irreversible still after being washed with DMSO solution. The stability of the toxic compounds-bacterial cell binding is a key consideration when probiotic antimutagenic property is evaluated. MGP exhibits the ability to bind and detoxify potent mutagens, and this property can be useful in supplemented foods for goats since it can lead to the removal of potent mutagens and protect and enhance ruminal health and hence food safety of consumers. Copyright © 2014 Elsevier Ltd. All rights reserved.

Functional interaction of CCAAT/enhancer-binding-protein-α basic region mutants with E2F transcription factors and DNA.

PubMed

Kowenz-Leutz, Elisabeth; Schuetz, Anja; Liu, Qingbin; Knoblich, Maria; Heinemann, Udo; Leutz, Achim

2016-07-01

The transcription factor CCAAT/enhancer-binding protein α (C/EBPα) regulates cell cycle arrest and terminal differentiation of neutrophils and adipocytes. Mutations in the basic leucine zipper domain (bZip) of C/EBPα are associated with acute myeloid leukemia. A widely used murine transforming C/EBPα basic region mutant (BRM2) entails two bZip point mutations (I294A/R297A). BRM2 has been discordantly described as defective for DNA binding or defective for interaction with E2F. We have separated the two BRM2 mutations to shed light on the intertwined reciprocity between C/EBPα-E2F-DNA interactions. Both, C/EBPα I294A and R297A retain transactivation capacity and interaction with E2F-DP. The C/EBPα R297A mutation destabilized DNA binding, whereas the C/EBPα I294A mutation enhanced binding to DNA. The C/EBPα R297A mutant, like BRM2, displayed enhanced interaction with E2F-DP but failed to repress E2F-dependent transactivation although both mutants were readily suppressed by E2F1 for transcription through C/EBP cis-regulatory sites. In contrast, the DNA binding enhanced C/EBPα I294A mutant displayed increased repression of E2F-DP mediated transactivation and resisted E2F-DP mediated repression. Thus, the efficient repression of E2F dependent S-phase genes and the activation of differentiation genes reside in the balanced DNA binding capacity of C/EBPα. Copyright © 2016 Elsevier B.V. All rights reserved.

Colorectal mucus binds DC-SIGN and inhibits HIV-1 trans-infection of CD4+ T-lymphocytes.

PubMed

Stax, Martijn J; Mouser, Emily E I M; van Montfort, Thijs; Sanders, Rogier W; de Vries, Henry J C; Dekker, Henk L; Herrera, Carolina; Speijer, Dave; Pollakis, Georgios; Paxton, William A

2015-01-01

Bodily secretions, including breast milk and semen, contain factors that modulate HIV-1 infection. Since anal intercourse caries one of the highest risks for HIV-1 transmission, our aim was to determine whether colorectal mucus (CM) also contains factors interfering with HIV-1 infection and replication. CM from a number of individuals was collected and tested for the capacity to bind DC-SIGN and inhibit HIV-1 cis- or trans-infection of CD4+ T-lymphocytes. To this end, a DC-SIGN binding ELISA, a gp140 trimer competition ELISA and HIV-1 capture/ transfer assays were utilized. Subsequently we aimed to identify the DC-SIGN binding component through biochemical characterization and mass spectrometry analysis. CM was shown to bind DC-SIGN and competes with HIV-1 gp140 trimer for binding. Pre-incubation of Raji-DC-SIGN cells or immature dendritic cells (iDCs) with CM potently inhibits DC-SIGN mediated trans-infection of CD4+ T-lymphocytes with CCR5 and CXCR4 using HIV-1 strains, while no effect on direct infection is observed. Preliminary biochemical characterization demonstrates that the component seems to be large (>100kDa), heat and proteinase K resistant, binds in a α1-3 mannose independent manner and is highly variant between individuals. Immunoprecipitation using DC-SIGN-Fc coated agarose beads followed by mass spectrometry indicated lactoferrin (fragments) and its receptor (intelectin-1) as candidates. Using ELISA we showed that lactoferrin levels within CM correlate with DC-SIGN binding capacity. In conclusion, CM can bind the C-type lectin DC-SIGN and block HIV-1 trans-infection of both CCR5 and CXCR4 using HIV-1 strains. Furthermore, our data indicate that lactoferrin is a DC-SIGN binding component of CM. These results indicate that CM has the potential to interfere with pathogen transmission and modulate immune responses at the colorectal mucosa.

Generation of therapeutic protein variants with the human serum albumin binding capacity via site-specific fatty acid conjugation.

PubMed

Cho, Jinhwan; Lim, Sung In; Yang, Byung Seop; Hahn, Young S; Kwon, Inchan

2017-12-21

Extension of the serum half-life is an important issue in developing new therapeutic proteins and expanding applications of existing therapeutic proteins. Conjugation of fatty acid, a natural human serum albumin ligand, to a therapeutic protein/peptide was developed as a technique to extend the serum half-life in vivo by taking advantages of unusually long serum half-life of human serum albumin (HSA). However, for broad applications of fatty acid-conjugation, several issues should be addressed, including a poor solubility of fatty acid and a substantial loss in the therapeutic activity. Therefore, herein we systematically investigate the conditions and components in conjugation of fatty acid to a therapeutic protein resulting in the HSA binding capacity without compromising therapeutic activities. By examining the crystal structure and performing dye conjugation assay, two sites (W160 and D112) of urate oxidase (Uox), a model therapeutic protein, were selected as sites for fatty acid-conjugation. Combination of site-specific incorporation of a clickable p-azido-L-phenylalanine to Uox and strain-promoted azide-alkyne cycloaddition allowed the conjugation of fatty acid (palmitic acid analog) to Uox with the HSA binding capacity and retained enzyme activity. Deoxycholic acid, a strong detergent, greatly enhanced the conjugation yield likely due to the enhanced solubility of palmitic acid analog.

Antibacterial Activity of Silver Nanoparticle-Loaded Soft Contact Lens Materials: The Effect of Monomer Composition.

PubMed

Shayani Rad, Maryam; Khameneh, Bahman; Sabeti, Zahra; Mohajeri, Seyed Ahmad; Fazly Bazzaz, Bibi Sedigheh

2016-10-01

In the present work, the effect of monomer composition on silver nanoparticles' (SNPs) binding capacity of hydrogels was investigated and their antibacterial efficacy was evaluated. Three series of poly-hydroxyethyl methacrylate (HEMA) hydrogels were prepared using methacrylic acid (MAA), methacrylamide (MAAM), and 4-vinylpyridine (4VP) as co-monomers, and ethylene glycol dimethacrylate (EGDMA) as cross-linker. SNPs binding capacity of hydrogels was evaluated in different concentrations (2, 10, and 20 ppm). In vitro antibacterial activity of SNP-loaded hydrogels was studied against Pseudomonas aeruginosa (P. aeruginosa) isolated from patients' eyes. Then, inhibitory effect of hydrogels in biofilm formation was evaluated in the presence of Staphylococcus epidermidis (S. epidermidis) (DSMZ 3270). Our data indicated that poly(HEMA-co-MAA-co-EGDMA) had superior binding affinity for SNPs in comparison with other hydrogels. All SNP-loaded hydrogels demonstrated excellent antimicrobial effects at all times against P. aeruginosa and S. epidermidis after soaking in 10 and 20 ppm SNP suspensions. Scanning electron microscope (SEM) images revealed excellent inhibitory effect of SNPs against biofilm formation on the surface of the hydrogels. This study indicated the effect of monomer compositions in SNP loading capacity of poly(HEMA) hydrogels and antibacterial efficacy of SNP-loaded hydrogels against P. aeruginosa and S. epidermidis, but further in vivo evaluation is necessary.

Arginine homopeptides for plasmid DNA purification using monolithic supports.

PubMed

Cardoso, Sara; Sousa, Ângela; Queiroz, João A; Azzoni, Adriano R; Sousa, Fani

2018-06-15

Purification of plasmid DNA targeting therapeutic applications still presents many challenges, namely on supports and specific ligand development. Monolithic supports have emerged as interesting approaches for purifying pDNA due to its excellent mass transfer properties and higher binding capacity values. Moreover, arginine ligands were already described to establish specific and preferential interactions with pDNA. Additionally, some studies revealed the ability of arginine based cationic peptides to condense plasmid DNA, which increased lengthening can result in strongest interactions with higher binding capacities for chromatographic purposes of large molecules such as pDNA. In this work, arginine homopeptides were immobilized in monolithic supports and their performance was evaluated and compared with a single arginine monolithic column regarding supercoiled (sc) plasmid DNA purification. Specific interactions of arginine based peptides with several nucleic acids present in a clarified Escherichia coli lysate sample showed potential for the sc pDNA purification. Effectively, the immobilization of the arginine homopeptides became more functional compared with the single arginine amino acid, showing higher binding capacities, which was also reflected in the intensity of the interactions. The combination of structural versatilities of monoliths with the specificity of arginine peptides raised as a promising strategy for sc pDNA purification. Copyright © 2018 Elsevier B.V. All rights reserved.

The effect of microwave on the interaction of flavour compounds with G-actin from grass carp (Catenopharyngodon idella).

PubMed

Lou, Xiaowei; Yang, Qiuli; Sun, Yangying; Pan, Daodong; Cao, Jinxuan

2017-09-01

In order to investigate the influence of non-thermal effects of microwaves on the flavour of fish and meat products, the G-actin of grass carp in ice baths was exposed to different microwave powers (0, 100, 300 or 500 W); the surface hydrophobicity, sulfhydryl contents, secondary structures and adsorption capacity of G-actin to ketones were determined. As microwave power increased from 0 to 300 W, the surface hydrophobicity, total and reactive sulfhydryls increased; α-helix, β-sheet and random coil fractions turned into β-turn fractions. As microwave power increased from 300 to 500 W, however, hydrophobicity and sulfhydryl contents decreased; β-turn and random coil fractions turned into α-helix and β-sheet fractions. The tendencies of adsorbed capacity of ketones were similar to hydrophobicity and sulfhydryl contents. The increased adsorbing of ketones could be attributed to the unfolding of secondary structures by revealing new binding sites, including thiol groups and hydrophobic binding sites. The decreased binding capacity was related to the refolding and aggregation of protein. The results suggested that microwave powers had obvious effects on the flavour retention and proteins structures in muscle foods. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

Chemiluminescence analysis of antioxidant capacity for serum albumin isolated from healthy or uremic volunteers.

PubMed

Huang, Chih-Yang; Liou, Show-Yih; Kuo, Wei-Wen; Wu, Hsi-Chin; Chang, Yen-Lin; Chen, Tung-Sheng

2016-12-01

Regular hemodialysis treatment induces an elevation in oxidative stress in patients with end-stage renal failure, resulting in oxidative damage of the most abundant serum protein, albumin. Oxidation of serum albumin causes depletion of albumin reactive thiols, leading to oxidative modification of serum albumin. The aim of this study was to screen the antioxidant capacity of albumins isolated from uremic patients (HD-ALB) or healthy volunteers (N-ALB). From high-performance liquid chromatography spectra, we observed that one uremic solute binds to HD-ALB via the formation of disulfide bonds between HD-ALB and the uremic solute. Furthermore, we found using chemiluminescent analysis that the antioxidant capacities for N-ALB to scavenge reactive oxygen species including singlet oxygen, hypochlorite and hydrogen peroxide were higher than HD-ALB. Our results suggest that protein-bound uremic solute binds to albumin via formation of disulfide bonds, resulting in the depletion of albumin reactive thiols. The depletion of albumin reactive thiols leads to a reduced antioxidant capacity of HD-ALB, implying postmodification of albumin. This situation may reduce the antioxidant capacity of albumin and increase oxidative stress, resulting in increase in complications related to oxidative damage in uremic patients. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Assessment of the nickel-albumin binding assay for diagnosis of acute coronary syndrome.

PubMed

da Silva, Sandra Huber; Pereira, Renata da Silva; Hausen, Bruna dos Santos; Signor, Cristiane; Gomes, Patrícia; de Campos, Marli Matiko Anraku; Moresco, Rafael Noal

2011-03-01

Myocardial ischemia may alter the metal binding capacity of circulating serum albumin. Thus, the aim of this study was to describe an automated method to measure ischemia-induced alterations in the binding capacity of serum albumin for exogenous nickel, and to evaluate the diagnostic characteristics of this assay for the assessment of acute coronary syndrome (ACS) in patients presenting to the emergency room (ER) with acute chest pain. We assessed the concentrations of cardiac troponin I (cTnI), serum albumin, ischemia-modified albumin (IMA) measured by the cobalt-albumin binding assay (CABA), and by an automated nickel-albumin binding assay (NABA) in the following groups: ACS (n=63) and non-ischemic chest pain (NICP, n=26). Biochemical markers were determined in blood samples obtained from patients within 3 h of ER admission. cTnI, CABA and NABA concentrations were higher in ACS group in comparison to the NICP group. A significant correlation between NABA and CABA was observed (r=0.5387, p<0.001). Areas under the curve for CABA and NABA were 0.7289 and 0.7582, respectively. Both CABA and NABA have the ability to discriminate patients with ACS. However, NABA has a slightly higher ability to discriminate ACS compared with CABA. Patients with ACS have reduced nickel binding to human serum albumin, and NABA may have an important role as an early marker of myocardial ischemia, particularly in patients presenting to the ER with acute chest pain.

The role of living/controlled radical polymerization in the formation of improved imprinted polymers.

PubMed

Salian, Vishal D; Vaughan, Asa D; Byrne, Mark E

2012-06-01

In this work, living/controlled radical polymerization (LRP) is compared with conventional free radical polymerization in the creation of highly and weakly cross-linked imprinted poly(methacrylic acid-co-ethylene glycol dimethacrylate) networks. It elucidates, for the first time, the effect of LRP on the chain level and begins to explain why the efficiency of the imprinting process is improved using LRP. Imprinted polymers produced via LRP exhibited significantly higher template affinity and capacity compared with polymers prepared using conventional methods. The use of LRP in the creation of highly cross-linked imprinted polymers resulted in a fourfold increase in binding capacity without a decrease in affinity; whereas weakly cross-linked gels demonstrated a nearly threefold increase in binding capacity at equivalent affinity when LRP was used. In addition, by adjusting the double bond conversion, we can choose to increase either the capacity or the affinity in highly cross-linked imprinted polymers, thus allowing the creation of imprinted polymers with tailorable binding parameters. Using free radical polymerization in the creation of polymer chains, as the template-monomer ratio increased, the average molecular weight of the polymer chains decreased despite a slight increase in the double bond conversion. Thus, the polymer chains formed were shorter but greater in number. Using LRP neutralized the effect of the template. The addition of chain transfer agent resulted in slow, uniform, simultaneous chain growth, resulting in the formation of longer more monodisperse chains. Reaction analysis revealed that propagation time was extended threefold in the formation of highly cross-linked polymers when LRP techniques were used. This delayed the transition to the diffusion-controlled stage of the reaction, which in turn led to the observed enhanced binding properties, decreased polydispersity in the chains, and a more homogeneous macromolecular architecture. Copyright © 2012 John Wiley & Sons, Ltd.

Dispersion of single-wall carbon nanotubes with supramolecular Congo red - properties of the complexes and mechanism of the interaction.

PubMed

Jagusiak, Anna; Piekarska, Barbara; Pańczyk, Tomasz; Jemioła-Rzemińska, Małgorzata; Bielańska, Elżbieta; Stopa, Barbara; Zemanek, Grzegorz; Rybarska, Janina; Roterman, Irena; Konieczny, Leszek

2017-01-01

A method of dispersion of single-wall carbon nanotubes (SWNTs) in aqueous media using Congo red (CR) is proposed. Nanotubes covered with CR constitute the high capacity system that provides the possibility of binding and targeted delivery of different drugs, which can intercalate into the supramolecular, ribbon-like CR structure. The study revealed the presence of strong interactions between CR and the surface of SWNTs. The aim of the study was to explain the mechanism of this interaction. The interaction of CR and carbon nanotubes was studied using spectral analysis of the SWNT-CR complex, dynamic light scattering (DLS), differential scanning calorimetry (DSC) and microscopic methods: atomic force microscopy (AFM), transmission (TEM), scanning (SEM) and optical microscopy. The results indicate that the binding of supramolecular CR structures to the surface of the nanotubes is based on the "face to face stacking". CR molecules attached directly to the surface of the nanotubes can bind further, parallel-oriented molecules and form supramolecular and protruding structures. This explains the high CR binding capacity of carbon nanotubes. The presented system - containing SWNTs covered with CR - offers a wide range of biomedical applications.

Effect of geometry and scale for axial and radial flow membrane chromatography-Experimental study of bovin serum albumin adsorption.

PubMed

Teepakorn, Chalore; Fiaty, Koffi; Charcosset, Catherine

2015-07-17

During the last 10 years, membrane chromatography (MC) has been increasingly reported for biomolecule purification at both small and large scales. Although, several axial and radial flow MC devices are commercialized, the effect of the device dimensions on the adsorption performance has not been fully investigated. In this study, axial and radial flow anion ion-exchange MC devices were used for bovine serum albumin (BSA) adsorption. For both axial and radial flow, three devices at different scales were compared, two having similar diameter and two similar bed height. The pressure drop and the flow distribution using acetone as a non-binding solute were measured, as well as BSA breakthrough curves at different flow rates and BSA loading concentrations. For all devices, it was observed that the flow rate had no effect on the breakthrough curve, which confirms the advantage of MC to be used at high flow rates. In addition, the BSA binding capacity increased with increasing BSA concentration, which suggests that it could be preferable to work with concentrated solutions rather than with very dilute solutions, when using buffer at high phosphate concentration. For both axial and radial flow, the bed height had a negative impact on the binding capacity, as the lowest binding capacities per membrane volume were obtained with the devices having the highest bed height. Radial flow MC has potential at large-scale applications, as a short bed thickness can be combined with a large inlet surface area. Copyright © 2015 Elsevier B.V. All rights reserved.

Influence of processing on the allergenic properties of pistachio nut assessed in vitro.

PubMed

Noorbakhsh, Reihaneh; Mortazavi, Seyed Ali; Sankian, Mojtaba; Shahidi, Fakhri; Maleki, Soheila J; Nasiraii, Leila Roozbeh; Falak, Reza; Sima, Hamid Reza; Varasteh, AbdolReza

2010-09-22

Pistachio (Pistacia vera) is a tree nut that has been reported to cause IgE-mediated allergic reactions. This study was undertaken to investigate the distinctions between different cultivars of pistachio nut and the influence of different processing on the IgE-binding capacity of whole pistachio protein extracts. The influence of different processes on allergenicity was investigated using competitive inhibition ELISA and Western blotting assays. The Western blotting results of extracts from pistachio cultivars showed no marked difference among them. The IgE-binding capacity was significantly lower for the protein extract prepared from steam-roasted than from raw and dry-roasted pistachio nuts. The results of sensory evaluation analysis and hedonic rating proved no significant differences in color, taste, flavor, and overall quality of raw, roasted, and steam-roasted pistachio nut treatments. The most significant finding of the present study was the successful reduction of IgE-binding by pistachio extracts using steam-roast processing without any significant changes in sensory quality of product.

Transition-metal dispersion on carbon-doped boron nitride nanostructures: Applications for high-capacity hydrogen storage

NASA Astrophysics Data System (ADS)

Chen, Ming; Zhao, Yu-Jun; Liao, Ji-Hai; Yang, Xiao-Bao

2012-07-01

Using density-functional theory calculations, we investigated the adsorption of transition-metal (TM) atoms (TM = Sc, Ti, V, Cr, Mn, Fe, Co, and Ni) on carbon doped hexagonal boron nitride (BN) sheet and the corresponding cage (B12N12). With carbon substitution of nitrogen, Sc, V, Cr, and Mn atoms were energetically favorable to be dispersed on the BN nanostructures without clustering or the formation of TM dimers, due to the strong binding between TM atoms and substrate, which contains the half-filled levels above the valence bands maximum. The carbon doped BN nanostructures with dispersed Sc could store up to five and six H2, respectively, with the average binding energy of 0.3 ˜ 0.4 eV, indicating the possibility of fabricating hydrogen storage media with high capacity. We also demonstrated that the geometrical effect is important for the hydrogen storage, leading to a modulation of the charge distributions of d levels, which dominates the binding between H2 and TM atoms.

Protection of Dentate Hilar Cells from Prolonged Stimulation by Intracellular Calcium Chelation

NASA Astrophysics Data System (ADS)

Scharfman, Helen E.; Schwartzkroin, Philip A.

1989-10-01

Prolonged afferent stimulation of the rat dentate gyrus in vivo leads to degeneration only of those cells that lack immunoreactivity for the calcium binding proteins parvalbumin and calbindin. In order to test the hypothesis that calcium binding proteins protect against the effects of prolonged stimulation, intracellular recordings were made in hippocampal slices from cells that lack immunoreactivity for calcium binding proteins. Calcium binding protein--negative cells showed electrophysiological signs of deterioration during prolonged stimulation; cells containing calcium binding protein did not. When neurons without calcium binding proteins were impaled with microelectrodes containing the calcium chelator BAPTA, and BAPTA was allowed to diffuse into the cells, these cells showed no deterioration. These results indicate that, in a complex tissue of the central nervous system, an activity-induced increase in intracellular calcium can trigger processes leading to cell deterioration, and that increasing the calcium binding capacity of a cell decreases its vulnerability to damage.

High-capacity thermo-responsive magnetic molecularly imprinted polymers for selective extraction of curcuminoids.

PubMed

You, Qingping; Zhang, Yuping; Zhang, Qingwen; Guo, Junfang; Huang, Weihua; Shi, Shuyun; Chen, Xiaoqin

2014-08-08

Thermo-responsive magnetic molecularly imprinted polymers (TMMIPs) for selective recognition of curcuminoids with high capacity and selectivity have firstly been developed. The resulting TMMIPs were characterized by TEM, FT-IR, TGA, VSM and UV, which indicated that TMMIPs showed thermo-responsiveness [lower critical solution temperature (LCST) at 33.71°C] and rapid magnetic separation (5s). The polymerization, adsorption and release conditions were optimized in detail to obtain the highest binding capacity, selectivity and release ratio. We found that the adopted thermo-responsive monomer [N-isopropylacrylamide (NIPAm)] could be considered not only as inert polymer backbone for thermo-responsiveness but also as functional co-monomers combination with basic monomer (4-VP) for more specific binding sites when ethanol was added in binding solution. The maximum adsorption capacity with highest selectivity of curcumin was 440.3μg/g (1.93 times that on MMIPs with no thermosensitivity) at 45°C (above LCST) in 20% (v/v) ethanol solution on shrunk TMMIPs, and the maximum release proportion was about 98% at 20°C (below LCST) in methanol-acetic acid (9/1, v/v) solution on swelled TMMIPs. The adsorption process between curcumin and TMMIPs followed Langumuir adsorption isotherm and pseudo-first-order reaction kinetics. The prepared TMMIPs also showed high reproducibility (RSD<6% for batch-to-batch evaluation) and stability (only 7% decrease after five cycles). Subsequently, the TMMIPs were successfully applied for selective extraction of curcuminoids from complex natural product, Curcuma longa. Copyright © 2014 Elsevier B.V. All rights reserved.

On the limited recognition of inorganic surfaces by short peptides compared with antibodies.

PubMed

Artzy-Schnirman, Arbel; Abu-Shah, Enas; Dishon, Matan; Soifer, Hadas; Sivan, Yotam; Reiter, Yoram; Benhar, Itai; Sivan, Uri

2014-06-01

The vast potential applications of biomolecules that bind inorganic surfaces led mostly to the isolation of short peptides that target selectively specific materials. The demonstrated differential affinity toward certain surfaces created the impression that the recognition capacity of short peptides may match that of rigid biomolecules. In the following, we challenge this view by comparing the capacity of antibody molecules to discriminate between the (100) and (111A) facets of a gallium arsenide semiconductor crystal with the capacity of short peptides to do the same. Applying selection from several peptide and single chain phage display libraries, we find a number of antibody molecules that bind preferentially a given crystal facet but fail to isolate, in dozens of attempts, a single peptide capable of such recognition. The experiments underscore the importance of rigidity to the recognition of inorganic flat targets and therefore set limitations on potential applications of short peptides in biomimetics. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd.

Identification and characterization of a Fc receptor activity on the Toxoplasma gondii tachyzoite.

PubMed

Vercammen, M; el Bouhdidi, A; Ben Messaoud, A; de Meuter, F; Bazin, H; Dubremetz, J F; Carlier, Y

1998-01-01

The Immunoglobulin (Ig) binding capacity of Toxoplasma gondii tachyzoites was investigated using fluorescence flow-cytometry analysis. Polyclonal mouse, human and rat immunoglobulins without specific anti-Toxoplasma activity bound to parasites in a concentration-dependent manner, saturating them at circulating serum concentrations. The immunoglobulin class and subclass specificity of binding was investigated using irrelevant monoclonal antibodies. IgM, IgA and IgG reacted with the parasite membrane. The attachment of mouse IgM to the parasite surface was hampered by mouse IgG1, IgG2a, IgG2b and IgG3. The binding of mouse IgG was proportionally reduced with increasing concentrations of mouse monoclonal IgM. The binding of murine immunoglobulin was diminished when in presence of human IgG. Purified Fc- but not Fab portions of immunoglobulins, fixed to parasites. Using labelled calibrated beads, the Ig binding capacity of parasites was estimated to be 6900 +/- 500 sites per tachyzoite. The Kd of the T. gondii Fc Receptor (FcR) activity was determined at 1.4 +/- 0.1 microM (mean +/- SEM). Such FcR activity was reduced by phospholipase C, trypsin and pronase treatment of the parasites. These data show a low affinity FcR activity on T. gondii tachyzoites which recognizes Ig of different species and isotypes and is likely supported by a glycosyl-phosphatidylinositol (GPI)-anchored surface protein of the parasite.

Krill Oil-In-Water Emulsion Protects against Lipopolysaccharide-Induced Proinflammatory Activation of Macrophages In Vitro.

PubMed

Bonaterra, Gabriel A; Driscoll, David; Schwarzbach, Hans; Kinscherf, Ralf

2017-03-15

Parenteral nutrition is often a mandatory therapeutic strategy for cases of septicemia. Likewise, therapeutic application of anti-oxidants, anti-inflammatory therapy, and endotoxin lowering, by removal or inactivation, might be beneficial to ameliorate the systemic inflammatory response during the acute phases of critical illness. Concerning anti-inflammatory properties in this setting, omega-3 fatty acids of marine origin have been frequently described. This study investigated the anti-inflammatory and LPS-inactivating properties of krill oil (KO)-in-water emulsion in human macrophages in vitro. Differentiated THP-1 macrophages were activated using specific ultrapure-LPS that binds only on the toll-like receptor 4 (TLR4) in order to determine the inhibitory properties of the KO emulsion on the LPS-binding capacity, and the subsequent release of TNF-α. KO emulsion inhibited the macrophage binding of LPS to the TLR4 by 50% (at 12.5 µg/mL) and 75% (at 25 µg/mL), whereas, at 50 µg/mL, completely abolished the LPS binding. Moreover, KO (12.5 µg/mL, 25 µg/mL, or 50 µg/mL) also inhibited (30%, 40%, or 75%, respectively) the TNF-α release after activation with 0.01 µg/mL LPS in comparison with LPS treatment alone. KO emulsion influences the LPS-induced pro-inflammatory activation of macrophages, possibly due to inactivation of the LPS binding capacity.

Variable ligand- and receptor-binding hot spots in key strains of influenza neuraminidase

PubMed Central

Votapka, Lane; Demir, Özlem; Swift, Robert V; Walker, Ross C; Amaro, Rommie E

2012-01-01

Influenza A continues to be a major public health concern due to its ability to cause epidemic and pandemic disease outbreaks in humans. Computational investigations of structural dynamics of the major influenza glycoproteins, especially the neuraminidase (NA) enzyme, are able to provide key insights beyond what is currently accessible with standard experimental techniques. In particular, all-atom molecular dynamics simulations reveal the varying degrees of flexibility for such enzymes. Here we present an analysis of the relative flexibility of the ligand- and receptor-binding area of three key strains of influenza A: highly pathogenic H5N1, the 2009 pandemic H1N1, and a human N2 strain. Through computational solvent mapping, we investigate the various ligand- and receptor-binding “hot spots” that exist on the surface of NA which interacts with both sialic acid receptors on the host cells and antiviral drugs. This analysis suggests that the variable cavities found in the different strains and their corresponding capacities to bind ligand functional groups may play an important role in the ability of NA to form competent reaction encounter complexes with other species of interest, including antiviral drugs, sialic acid receptors on the host cell surface, and the hemagglutinin protein. Such considerations may be especially useful for the prediction of how such complexes form and with what binding capacity. PMID:22872804

Switching on RNA Silencing Suppressor Activity by Restoring Argonaute Binding to a Viral Protein

PubMed Central

Szabó, Edit Z.; Manczinger, Máté; Göblös, Anikó; Kemény, Lajos

2012-01-01

We found that Sweet potato feathery mottle virus (SPFMV) P1, a close homologue of Sweet potato mild mottle virus P1, did not have any silencing suppressor activity. Remodeling the Argonaute (AGO) binding domain of SPFMV P1 by the introduction of two additional WG/GW motifs converted it to a silencing suppressor with AGO binding capacity. To our knowledge, this is the first instance of the transformation of a viral protein of unknown function to a functional silencing suppressor. PMID:22623784

Drifting from Slow to "D'oh!": Working Memory Capacity and Mind Wandering Predict Extreme Reaction Times and Executive Control Errors

ERIC Educational Resources Information Center

McVay, Jennifer C.; Kane, Michael J.

2012-01-01

A combined experimental, individual-differences, and thought-sampling study tested the predictions of executive attention (e.g., Engle & Kane, 2004) and coordinative binding (e.g., Oberauer, Suss, Wilhelm, & Sander, 2007) theories of working memory capacity (WMC). We assessed 288 subjects' WMC and their performance and mind-wandering rates…

Mutations in type 3 reovirus that determine binding to sialic acid are contained in the fibrous tail domain of viral attachment protein sigma1.

PubMed

Chappell, J D; Gunn, V L; Wetzel, J D; Baer, G S; Dermody, T S

1997-03-01

The reovirus attachment protein, sigma1, determines numerous aspects of reovirus-induced disease, including viral virulence, pathways of spread, and tropism for certain types of cells in the central nervous system. The sigma1 protein projects from the virion surface and consists of two distinct morphologic domains, a virion-distal globular domain known as the head and an elongated fibrous domain, termed the tail, which is anchored into the virion capsid. To better understand structure-function relationships of sigma1 protein, we conducted experiments to identify sequences in sigma1 important for viral binding to sialic acid, a component of the receptor for type 3 reovirus. Three serotype 3 reovirus strains incapable of binding sialylated receptors were adapted to growth in murine erythroleukemia (MEL) cells, in which sialic acid is essential for reovirus infectivity. MEL-adapted (MA) mutant viruses isolated by serial passage in MEL cells acquired the capacity to bind sialic acid-containing receptors and demonstrated a dependence on sialic acid for infection of MEL cells. Analysis of reassortant viruses isolated from crosses of an MA mutant virus and a reovirus strain that does not bind sialic acid indicated that the sigma1 protein is solely responsible for efficient growth of MA mutant viruses in MEL cells. The deduced sigma1 amino acid sequences of the MA mutant viruses revealed that each strain contains a substitution within a short region of sequence in the sigma1 tail predicted to form beta-sheet. These studies identify specific sequences that determine the capacity of reovirus to bind sialylated receptors and suggest a location for a sialic acid-binding domain. Furthermore, the results support a model in which type 3 sigma1 protein contains discrete receptor binding domains, one in the head and another in the tail that binds sialic acid.

Comparison of functional assays used in the clinical development of a placental malaria vaccine.

PubMed

Pehrson, Caroline; Heno, Kristine K; Adams, Yvonne; Resende, Mafalda; Mathiesen, Line; Soegaard, Max; de Jongh, Willem A; Theander, Thor G; Salanti, Ali; Nielsen, Morten A

2017-01-23

Malaria in pregnancy is associated with significant morbidity in pregnant women and their offspring. Plasmodium falciparum infected erythrocytes (IE) express VAR2CSA that mediates binding to chondroitin sulphate A (CSA) in the placenta. Two VAR2CSA-based vaccines for placental malaria are in clinical development. The purpose of this study was to evaluate the robustness and comparability of binding inhibition assays used in the clinical development of placental malaria vaccines. The ability of sera from animals immunised with different VAR2CSA constructs to inhibit IE binding to CSA was investigated in three in vitro assays using 96-well plates, petri dishes, capillary flow and an ex vivo placental perfusion assay. The inter-assay variation was not uniform between assays and ranged from above ten-fold in the flow assay to two-fold in the perfusion assay. The intra-assay variation was highest in the petri dish assay. A positive correlation between IE binding avidity and the level of binding after antibody inhibition in the petri dish assay indicate that high avidity IE binding is more difficult to inhibit. The highest binding inhibition sensitivity was found in the 96-well and petri dish assays compared to the flow and perfusion assays where binding inhibition required higher antibody titers. The inhibitory capacity of antibodies is not easily translated between assays and the high sensitivity of the 96-well and petri dish assays stresses the need for comparing serial dilutions of serum. Furthermore, IE binding avidity must be in the same range when comparing data from different days. There was an overall concordance in the capacity of antibody-mediated inhibition, when comparing the in vitro assays with the perfusion assay, which more closely represents in vivo conditions. Importantly the ID1-ID2a protein in a liposomal formulation, currently in a phase I trial, effectively induced antibodies that inhibited IE adhesion in placental tissue. Copyright © 2016. Published by Elsevier Ltd.

DNA mutagenic activity and capacity for HIV-1 restriction of the cytidine deaminase APOBEC3G depend on whether DNA or RNA binds to tyrosine 315

PubMed Central

Polevoda, Bogdan; Joseph, Rebecca; Friedman, Alan E.; Bennett, Ryan P.; Greiner, Rebecca; De Zoysa, Thareendra; Stewart, Ryan A.; Smith, Harold C.

2017-01-01

APOBEC3G (A3G) belongs to the AID/APOBEC protein family of cytidine deaminases (CDA) that bind to nucleic acids. A3G mutates the HIV genome by deamination of dC to dU, leading to accumulation of virus-inactivating mutations. Binding to cellular RNAs inhibits A3G binding to substrate single-stranded (ss) DNA and CDA activity. Bulk RNA and substrate ssDNA bind to the same three A3G tryptic peptides (amino acids 181–194, 314–320, and 345–374) that form parts of a continuously exposed protein surface extending from the catalytic domain in the C terminus of A3G to its N terminus. We show here that the A3G tyrosines 181 and 315 directly cross-linked ssDNA. Binding experiments showed that a Y315A mutation alone significantly reduced A3G binding to both ssDNA and RNA, whereas Y181A and Y182A mutations only moderately affected A3G nucleic acid binding. Consistent with these findings, the Y315A mutant exhibited little to no deaminase activity in an Escherichia coli DNA mutator reporter, whereas Y181A and Y182A mutants retained ∼50% of wild-type A3G activity. The Y315A mutant also showed a markedly reduced ability to assemble into viral particles and had reduced antiviral activity. In uninfected cells, the impaired RNA-binding capacity of Y315A was evident by a shift of A3G from high-molecular-mass ribonucleoprotein complexes to low-molecular-mass complexes. We conclude that Tyr-315 is essential for coordinating ssDNA interaction with or entry to the deaminase domain and hypothesize that RNA bound to Tyr-315 may be sufficient to competitively inhibit ssDNA deaminase-dependent antiviral activity. PMID:28381554

Evaluation of the endotoxin binding efficiency of clay minerals using the Limulus Amebocyte lysate test: an in vitro study

PubMed Central

2014-01-01

Endotoxins are part of the cell wall of Gram-negative bacteria. They are potent immune stimulators and can lead to death if present in high concentrations. Feed additives, which bind endotoxins in the gastrointestinal tract of animals, could help to prevent their negative impact. The objective of our study was to determine the potential of a bentonite (Bentonite 1), a sodium bentonite (Bentonite 2), a chemically treated smectite (Organoclay 1) and a modified attapulgite (Organoclay 2) to bind endotoxins in vitro. Polymyxin B served as positive control. The kinetic chromogenic Limulus Amebocyte lysate test was adapted to measure endotoxin activity. Firstly, a single sorption experiment (10 endotoxin units/mL (EU/mL)) was performed. Polymyxin B and organoclays showed 100% binding efficiency. Secondly, the adsorption efficiency of sorbents in aqueous solution with increasing endotoxin concentrations (2,450 – 51,700 EU/mL) was investigated. Organoclay 1 (0.1%) showed a good binding efficiency in aqueous solution (average 81%), whereas Bentonite 1 (0.1%) obtained a lower binding efficiency (21-54%). The following absorbent capacities were calculated in highest endotoxin concentration: 5.59 mg/g (Organoclay 1) > 3.97 mg/g (Polymyxin B) > 2.58mg/g (Organoclay 2) > 1.55 mg/g (Bentonite 1) > 1.23 mg/g (Bentonite 2). Thirdly, a sorption experiment in artificial intestinal fluid was conducted. Especially for organoclays, which are known to be unspecific adsorbents, the endotoxin binding capacity was significantly reduced. In contrast, Bentonite 1 showed comparable results in artificial intestinal fluid and aqueous solution. Based on the results of this in vitro study, the effect of promising clay minerals will be investigated in in vivo trials. PMID:24383578

Insulin binding and glycolytic activity in erythrocytes from dialyzed and nondialyzed uremic patients.

PubMed

Weisinger, J R; Contreras, N E; Cajias, J; Bellorin-Font, E; Amair, P; Guitierrez, L; Sylva, V; Paz-Martínez, V

1988-01-01

Insulin resistance in uremia has been attributed to impaired hormone-receptor binding or to postbinding defects. Oral glucose tolerance tests, insulin binding, and in vitro glycolytic activity were studied in purified red blood cells from normal control subjects (C) and from uremic patients belonging to three groups: nondialyzed (U), on chronic hemodialysis (HD), and on continuous ambulatory peritoneal dialysis (CAPD). Glucose intolerance and hyperinsulinemia were demonstrated in all groups of patients. Maximal specific binding of 125I-insulin to erythrocytes, kinetically derived receptor numbers per cell, and affinity constants for insulin binding did not differ between control and patient groups. No correlation was found between the degree of glucose intolerance and insulin binding parameters. Basal lactate production by erythrocytes incubated in vitro was significantly higher in U and HD patients than in C, whereas CAPD patients did not differ from C in this respect. Addition of 1 mM dibutyryl-cAMP and 0.5 mM isobutyl-methyl-xanthine during incubation of erythrocytes caused an increase in the rate of lactate production that was similar in magnitude in the U, HD and C groups, whereas cells from CAPD subjects showed a significantly larger absolute response to these compounds after 1 h of incubation. There was no evidence of impairment of glycolytic capacity in red blood cells from uremic patients. In addition, no correlation was found between the degree of glucose intolerance and basal or stimulated lactate production by erythrocytes. Our results obtained in human erythrocytes suggest that the insulin resistance observed in uremia does not involve a defect in hormone binding or in the intracellular capacity to utilize glucose through glycolysis.

Contrasting effects of photochemical and microbial degradation on Cu(II) binding with fluorescent DOM from different origins.

PubMed

Xu, Huacheng; Guan, Dong-Xing; Zou, Li; Lin, Hui; Guo, Laodong

2018-08-01

Effects of photochemical and microbial degradation on variations in composition and molecular-size of dissolved organic matter (DOM) from different sources (algal and soil) and the subsequent influence on Cu(II) binding were investigated using UV-Vis, fluorescence excitation-emission matrices coupled with parallel factor analysis, flow field-flow fractionation (FlFFF), and metal titration. The degradation processes resulted in an initial rapid decline in the bulk dissolved organic carbon and chromophoric and fluorescent DOM components, followed by a small or little decrease. Specifically, photochemical reaction decreased the aromaticity, humification and apparent molecular weights of all DOM samples, whereas a reverse trend was observed during microbial degradation. The FlFFF fractograms revealed that coagulation of both protein- and humic-like DOM induced an increase in molecular weights for algal-DOM, while the molecular weight enhancement for allochthonous soil samples was mainly attributed to the self-assembly of humic-like components. The Cu(II) binding capacity of algal-derived humic-like and fulvic-like DOM consistently increased during photo- and bio-degradation, while the soil-derived DOM exhibited a slight decline in Cu(II) binding capacity during photo-degradation but a substantial increase during microbial degradation, indicating source- and degradation-dependent metal binding heterogeneities. Pearson correlation analysis demonstrated that the Cu(II) binding potential was mostly related with aromaticity and molecular size for allochthonous soil-derived DOM, but was regulated by both DOM properties and specific degradation processes for autochthonous algal-derived DOM. This study highlighted the coupling role of inherent DOM properties and external environmental processes in regulating metal binding, and provided new insights into metal-DOM interactions and the behavior and fate of DOM-bound metals in aquatic environments. Copyright © 2018 Elsevier Ltd. All rights reserved.

In vitro bile acid binding of mustard greens, kale, broccoli, cabbage and green bell pepper improves with sautéing compared with raw or other methods of preparation.

USDA-ARS?s Scientific Manuscript database

Bile acid binding capacity has been related to cholesterol-lowering potential of foods and food fractions. Lowered recirculating bile acids results in utilization of cholesterol to synthesize bile acid and reduced fat absorption. Secondary bile acids have been associated with increased risk of can...

Changes in protease activity and Cry3Aa toxin binding in the Colorado potato beetle: implications for insect resistance to Bacillus thuringiensis toxins

Treesearch

Olga Loseva; Mohamed Ibrahim; Mehmet Candas; C. Noah Koller; Leah S. Bauer; Lee A. Jr. Bulla

2002-01-01

Widespread commercial use of Bacillus thuringiensis Cry toxins to control pest insects has increased the likelihood for development of insect resistance to this entomopathogen. In this study, we investigated protease activity profiles and toxin-binding capacities in the midgut of a strain of Colorado potato beetle (CPB) that has developed resistance...

Pectin-like carbohydrates in the green alga Micrasterias characterized by cytochemical analysis and energy filtering TEM.

PubMed

Eder, M; Lütz-Meindl, U

2008-08-01

Pectins are the major matrix polysaccharides of plant cell walls and are important for controlling growth, wall porosity and regulation of the ionic environment in plant cells. Pectic epitopes recognized by the monoclonal antibodies JIM5, JIM7 and 2F4 could be localized in the primary wall during development of the green alga Micrasterias. As the degree of pectin esterification determines the calcium-binding capacity and thus the physical properties of the cell wall, chemical and enzymatic in situ de-esterification was performed. This resulted in displacement of epitopes recognized by JIM5, JIM7 and 2F4, respectively, in changes in the intensity of the antibody labelling as visualized in CLSM. In addition, calcium-binding capacities of cell walls and components of the secretory apparatus were determined in transmission electron microscopy by electron energy loss spectroscopy and electron spectroscopic imaging. These analyses revealed that pectic polysaccharides are transported to the cell wall in a de-esterified form. At the primary wall, pectins get methyl-esterified at the inner side, thus allowing flexibility of the wall. At the outer side of the wall they become again de-esterified and bind high amounts of calcium which leads to cell wall stiffening. Mucilage vesicles possess the highest calcium-binding capacity of all structures observed in Micrasterias, indicating that the pectic polysaccharides of mucilage are secreted in a de-esterified, compact form. When mucilage is excreted through the cell wall, it loses its ability to bind calcium. The esterification of pectins involved is obviously required for swelling of mucilage by water uptake, which generates the motive force for orientation of this unicellular organism in respect to light. Incubation of Micrasterias in pectin methylesterase (PME), which de-esterifies pectic polymers in higher plants, resulted in growth inhibition, cell shape malformation and primary wall thickening. A PME-like enzyme could be found in Micrasterias by PME activity assays.

Human immunodeficiency virus type 1 Tat binds to Candida albicans, inducing hyphae but augmenting phagocytosis in vitro

PubMed Central

Gruber, Andreas; Lell, Claudia P; Speth, Cornelia; Stoiber, Heribert; Lass-Flörl, Cornelia; Sonneborn, Anja; Ernst, Joachim F; Dierich, Manfred P; Würzner, Reinhard

2001-01-01

Tat, the human immunodeficiency virus type 1 (HIV-1) transactivating protein, binds through its RGD-motif to human integrin receptors. Candida albicans, the commonest cause of mucosal candidiasis in subjects infected with HIV-1, also possesses RGD-binding capacity. The present study reveals that Tat binds to C. albicans but not to C. tropicalis. Tat binding was markedly reduced by laminin and to a lesser extent by a complement C3 peptide containing the RGD motif, but not by a control peptide. The outgrowth of C. albicans was accelerated following binding of Tat, but phagocytosis of opsonized C. albicans was also increased after Tat binding. Thus, Tat binding promotes fungal virulence by inducing hyphae but may also reduce it by augmenting phagocytosis. The net effect of Tat in vivo is difficult to judge but in view of the many disease-promoting effects of Tat we propose that accelerating the formation of hyphae dominates over the augmentation of phagocytosis. PMID:11899432

High voltage and high specific capacity dual intercalating electrode Li-ion batteries

NASA Technical Reports Server (NTRS)

Blanco, Mario (Inventor); West, William C. (Inventor)

2010-01-01

The present invention provides high capacity and high voltage Li-ion batteries that have a carbonaceous cathode and a nonaqueous electrolyte solution comprising LiF salt and an anion receptor that binds the fluoride ion. The batteries can comprise dual intercalating electrode Li ion batteries. Methods of the present invention use a cathode and electrode pair, wherein each of the electrodes reversibly intercalate ions provided by a LiF salt to make a high voltage and high specific capacity dual intercalating electrode Li-ion battery. The present methods and systems provide high-capacity batteries particularly useful in powering devices where minimizing battery mass is important.

Nuclear magnetic resonance-based model of a TF1/HmU-DNA complex.

PubMed

Silva, M V; Pasternack, L B; Kearns, D R

1997-12-15

Transcription factor 1 (TF1), a type II DNA-binding protein encoded by the Bacillus subtilis bacteriophage SPO1, has the capacity for sequence-selective DNA binding and a preference for 5-hydroxymethyl-2'-deoxyuridine (HmU)-containing DNA. In NMR studies of the TF1/HmU-DNA complex, intermolecular NOEs indicate that the flexible beta-ribbon and C-terminal alpha-helix are involved in the DNA-binding site of TF1, placing it in the beta-sheet category of DNA-binding proteins proposed to bind by wrapping two beta-ribbon "arms" around the DNA. Intermolecular and intramolecular NOEs were used to generate an energy-minimized model of the protein-DNA complex in which both DNA bending and protein structure changes are evident.

Comparative analyses of the thermodynamic RNA binding signatures of different types of RNA recognition motifs

PubMed Central

Cléry, Antoine; Allain, Frédéric H-T

2017-01-01

Abstract RNA recognition motifs (RRMs) are structurally versatile domains important in regulation of alternative splicing. Structural mechanisms of sequence-specific recognition of single-stranded RNAs (ssRNAs) by RRMs are well understood. The thermodynamic strategies are however unclear. Therefore, we utilized microcalorimetry and semi-empirical analyses to comparatively analyze the cognate ssRNA binding thermodynamics of four different RRM domains, each with a different RNA binding mode. The different binding modes are: canonical binding to the β-sheet surface; canonical binding with involvement of N- and C-termini; binding to conserved loops; and binding to an α-helix. Our results identify enthalpy as the sole and general force driving association at physiological temperatures. Also, networks of weak interactions are a general feature regulating stability of the different RRM–ssRNA complexes. In agreement, non-polyelectrolyte effects contributed between ∼75 and 90% of the overall free energy of binding in the considered complexes. The various RNA binding modes also displayed enormous heat capacity differences, that upon dissection revealed large differential changes in hydration, conformations and dynamics upon binding RNA. Altogether, different modes employed by RRMs to bind cognate ssRNAs utilize various thermodynamics strategies during the association process. PMID:28334819

Multimodal charge-induction chromatography for antibody purification.

PubMed

Tong, Hong-Fei; Lin, Dong-Qiang; Chu, Wen-Ning; Zhang, Qi-Lei; Gao, Dong; Wang, Rong-Zhu; Yao, Shan-Jing

2016-01-15

Hydrophobic charge-induction chromatography (HCIC) has advantages of high capacity, salt-tolerance and convenient pH-controlled elution. However, the binding specificity might be improved with multimodal molecular interactions. New ligand W-ABI that combining tryptophan and 5-amino-benzimidazole was designed with the concept of mutimodal charge-induction chromatography (MCIC). The indole and benzimidazole groups of the ligand could provide orientated mutimodal binding to target IgG under neutral pH, while the imidazole groups could induce the electrostatic repulsion forces for efficient elution under acidic pH. W-ABI ligand was coupled successfully onto agarose gel, and IgG adsorption behaviors were investigated. High affinity to IgG was found with the saturated adsorption capacity of 70.4 mg/ml at pH 7, and the flow rate of mobile phase showed little impact on the dynamic binding capacity. In addition, efficient elution could be achieved at mild acidic pH with high recovery. Two separation cases (IgG separation from albumin containing feedstock and monoclonal antibody purification from cell culture supernatant) were verified with high purity and recovery. In general, MCIC with the specially-designed ligand is an expanding of HCIC with improved adsorption selectivity, which would be a potential alternative to Protein A-based capture for the cost-effective purification of antibodies. Copyright © 2015 Elsevier B.V. All rights reserved.

Stabilization of glucocorticoid receptors in isolated rat hepatocytes by radioprotectants

DOE Office of Scientific and Technical Information (OSTI.GOV)

Karle, J.M.; Ridder, W.E.; Wright, N.

1986-05-01

Previous work has shown that glucocorticoid receptors in rat liver homogenate can be stabilized by the addition of MoO/sub 4/ plus the sulfhydryl-containing compounds dithiothreitol and WR 1065. The latter is the dephosphorylated, principal metabolite of the radioprotectant WR 2721 (or S-2-(3-aminopropylamino)ethanesphosphorothioic acid). The current work results from applying this knowledge to intact rat hepatocytes. Cells were isolated by collagenase perfusion and incubated in supplemented minimum essential medium at 37/sup 0/C with various concentrations of WR 2721, WR 1065, or vehicle. Samples of these cell suspensions were analyzed at various times for steroid binding capacity by incubating homogenates (27,000 xmore » g supernates) with 50 nM /sup 3/H-triamcinolone acetonide in the presence or absence of excess unlabelled dexamethasone. Concentrations of 10 mM WR 2721 provided marked preservation of the binding capacity (>85% of the initial value at 5 hours) compared to control at <20% by 2 hours. WR 2721 at 1 mM preserved >60% of the binding capacity. WR 1065 at 10 mM provided no such protection. This is consistent with the observation that WR 1065 does not pass cell membranes. The authors propose that supplying reducing equivalents to intracellular components such as the glucocorticoid receptor may be one mechanism of the radioprotection afforded by WR 2721.« less

High-capacity composite adsorbents for nucleic acids.

PubMed

Tiainen, Peter; Rokebul Anower, M; Larsson, Per-Olof

2011-08-05

Cytopore™ is a bead-shaped, macroporous and easily compressible cellulose-based anion-exchange material intended for cultivation of anchor-dependent animal cells. Reticulated vitreous carbon (RVC) is a strong, non-compressible, high voidage (97%) matrix material that can be cut to desired geometrical shapes. Cytopore and RVC were combined to cylindrical composites (25 mm × 10 mm) fitted inside chromatography columns. The composite combined the advantageous properties of both its constituents, making it suitable for column chromatography. The composite could withstand very high flow rates without compaction of the bed (>25 column volumes/min; 4000 cm h(-1)). Chromatography runs with tracers showed a low HETP value (0.3mm), suggesting that pore flow was in operation. The dynamic binding capacities (10% breakthrough) per gram of dry weight Cytopore were determined for several compounds including DNA and RNA and were found to be 240-370 mg/g. The composite was used to isolate pUC 18-type plasmids from a cleared alkaline lysate in a good yield. Confocal microscopy studies showed that plasmids were bound not only to the surface of the Cytopore material but also within the matrix walls, thus offering an explanation to the very high binding capacities observed. The concept of using a composite prepared from a mechanically weak, high-binding material and a strong scaffold material may be applied to other systems as well. Copyright © 2011 Elsevier B.V. All rights reserved.

Characterization of large-pore polymeric supports for use in perfusion biochromatography.

PubMed

Whitney, D; McCoy, M; Gordon, N; Afeyan, N

1998-05-22

Perfusion chromatography is uniquely characterized by the flow of a portion of the column eluent directly through the resin in the packed bed. The benefits of this phenomenon and some of the properties of perfusive resins have been described before, and can be summarized as enhanced mass transport to interior binding sites. Here we extend the understanding of this phenomenon by comparing resins with different pore size distributions. Resins are chosen to give approximately the same specific pore volumes (as shown in the characterization section) but the varying contribution of large pores is used to control the amount of liquid flowing through the beads. POROS R1 has the largest contribution of throughpores, and therefore the greatest intraparticle flow. POROS R2 has a lower contribution of throughpores, and a higher surface area coming from a greater population of diffusive pores, but still shows significant mass transport enhancements relative to a purely diffusive control. Oligo R3 is dominated by a high population of diffusive pores, and is used comparatively as a non-perfusive resin. Although the pore size distribution can be engineered to control mass transport rates, the resulting surface area is not the only means by which binding capacity can be controlled. Surface coatings are employed to increase binding capacity without fundamentally altering the mass transport properties. Models are used to describe the amount of flow transecting the beads, and comparisons of coated resins to uncoated (polystyrene) resins leads to the conclusion that these coatings do not obstruct the throughpore structures. This is an important conclusion since the binding capacity of the coated product, in some cases, is shown to be over 10-fold higher than the precursor polystyrene scaffold (i.e., POROS R1 or POROS R2).

Purification, biochemical, and immunological characterisation of a major food allergen: different immunoglobulin E recognition of the apo- and calcium-bound forms of carp parvalbumin

PubMed Central

Bugajska-Schrette..., A; Grote, M; Vangelista, L; Valent, P; Sperr, W; Rumpold, H; Pastore, A; Reichelt, R; Valenta, R; Spitzauer, S

2000-01-01

BACKGROUND—Almost 4% of the population suffer from food allergy which is an adverse reaction to food with an underlying immunological mechanism.
AIMS—To characterise one of the most frequent IgE defined food allergens, fish parvalbumin.
METHODS—Tissue and subcellular distribution of carp parvalbumin was analysed by immunogold electron microscopy and cell fractionation. Parvalbumin was purified to homogeneity, analysed by mass spectrometry and circular dichroism (CD) spectroscopy, and its allergenic activity was analysed by IgE binding and basophil histamine release tests.
RESULTS—The isoelectric point (pI) 4.7 form of carp parvalbumin, a three EF-hand calcium-binding protein, was purified to homogeneity. CD analysis revealed a remarkable stability and refolding capacity of calcium-bound parvalbumin. This may explain why parvalbumin, despite cooking and exposure to the gastrointestinal tract, can sensitise patients. Purified parvalbumin reacted with IgE of more than 95% of individuals allergic to fish, induced dose-dependent basophil histamine release and contained, on average, 83% of the IgE epitopes present in other fish species. Calcium depletion reduced the IgE binding capacity of parvalbumin which, according to CD analysis, may be due to conformation-dependent IgE recognition.
CONCLUSIONS—Purified carp parvalbumin represents an important cross reactive food allergen. It can be used for in vitro and in vivo diagnosis of fish-induced food allergy. Our finding that the apo-form of parvalbumin had a greatly reduced IgE binding capacity indicates that this form may be a candidate for safe immunotherapy of fish-related food allergy.


Keywords: food allergy; parvalbumin; circular dichroism; epitopes; antibodies; immunochemistry PMID:10764710

A hypoallergenic variant of the major birch pollen allergen shows distinct characteristics in antigen processing and T-cell activation.

PubMed

Kitzmüller, C; Wallner, M; Deifl, S; Mutschlechner, S; Walterskirchen, C; Zlabinger, G J; Ferreira, F; Bohle, B

2012-11-01

BM4 is a novel genetically engineered variant of the major birch pollen allergen Bet v 1 that lacks the typical Bet v 1-like fold and displays negligible IgE-binding but strong T cell-activating capacity. The aim of this study was to elucidate possible differences between BM4 and Bet v 1 in internalization, antigen processing, and presentation. Proliferative responses to BM4 and Bet v 1 of peripheral blood mononuclear cells and Bet v 1-specific T-cell clones were compared. Fluorescently labeled BM4 and Bet v 1 were used to study surface binding, endocytosis, and intracellular degradation by monocyte-derived DC (mdDC). Both proteins were digested by endolysosomal extracts of mdDC. BM4- and Bet v 1-pulsed mdDC were employed to assess the kinetics of activation of Bet v 1-specific T-cell clones and the polarization of naïve T cells. BM4 displayed a significantly stronger T cell-activating capacity than Bet v 1. Furthermore, BM4 showed increased surface binding and internalization as well as faster endolysosomal degradation compared with Bet v 1. BM4-pulsed mdDC induced enhanced proliferative responses at earlier time-points in Bet v 1-specific T-cell clones and promoted less IL-5 production in T cells than Bet v 1-pulsed mdDC. The loss of the Bet v 1-fold changes the protein's interaction with the human immune system at the level of antigen-presenting cells resulting in altered T-cell responses. By combining low IgE-binding with strong and modulating T cell-activating capacity, BM4 represents a highly interesting candidate for specific immunotherapy of birch pollen allergy. © 2012 John Wiley & Sons A/S.

Effect of transfection and co-incubation of bovine sperm with exogenous DNA on sperm quality and functional parameters for its use in sperm-mediated gene transfer.

PubMed

Arias, María Elena; Sánchez-Villalba, Esther; Delgado, Andrea; Felmer, Ricardo

2017-02-01

Sperm-mediated gene transfer (SMGT) is based on the capacity of sperm to bind exogenous DNA and transfer it into the oocyte during fertilization. In bovines, the progress of this technology has been slow due to the poor reproducibility and efficiency of the production of transgenic embryos. The aim of the present study was to evaluate the effects of different sperm transfection systems on the quality and functional parameters of sperm. Additionally, the ability of sperm to bind and incorporate exogenous DNA was assessed. These analyses were carried out by flow cytometry and confocal fluorescence microscopy, and motility parameters were also evaluated by computer-assisted sperm analysis (CASA). Transfection was carried out using complexes of plasmid DNA with Lipofectamine, SuperFect and TurboFect for 0.5, 1, 2 or 4 h. The results showed that all of the transfection treatments promoted sperm binding and incorporation of exogenous DNA, similar to sperm incorporation of DNA alone, without affecting the viability. Nevertheless, the treatments and incubation times significantly affected the motility parameters, although no effect on the integrity of DNA or the levels of reactive oxygen species (ROS) was observed. Additionally, we observed that transfection using SuperFect and TurboFect negatively affected the acrosome integrity, and TurboFect affected the mitochondrial membrane potential of sperm. In conclusion, we demonstrated binding and incorporation of exogenous DNA by sperm after transfection and confirmed the capacity of sperm to spontaneously incorporate exogenous DNA. These findings will allow the establishment of the most appropriate method [intracytoplasmic sperm injection (ICSI) or in vitro fertilization (IVF)] of generating transgenic embryos via SMGT based on the fertilization capacity of transfected sperm.

Soluble PD-L1 with PD-1-binding capacity exists in the plasma of patients with non-small cell lung cancer.

PubMed

Takeuchi, Masahiro; Doi, Tomomitsu; Obayashi, Kunie; Hirai, Ayako; Yoneda, Kazue; Tanaka, Fumihiro; Iwai, Yoshiko

2018-04-01

PD-L1 is one of the important immune checkpoint molecules that can be targeted by cancer immunotherapies. PD-L1 has a soluble form (sPD-L1) and a membrane-bound form (mPD-L1). Conventional enzyme-linked immunosorbent assay (ELISA) systems can detect sPD-L1 using anti-PD-L1 capture antibody through the antigen-antibody reaction, but cannot evaluate the quality and function of sPD-L1. In this study, we developed a novel ELISA system for the detection and quantification of sPD-L1 with PD-1-binding capacity (bsPD-L1). To capture bsPD-L1 through the ligand-receptor reaction, the anti-PD-L1 capture antibody in the conventional ELISA was replaced with PD-1-Ig fusion protein in the new ELISA. The new ELISA could detect bsPD-L1 in 29 out of 75 plasma samples from patients with non-small cell lung cancer (NSCLC), with higher sensitivity and frequency than the conventional ELISA. The western blot analysis showed that sPD-L1 in the plasma was glycosylated. Treatment of the samples with glycosidase reduced the absorbance determined by the new ELISA but had no effect on the absorbance determined by the conventional ELISA. These results suggest that glycosylation of sPD-L1 is important for its binding to the immobilized PD-1 in the new ELISA. Our new ELISA system may be useful for the evaluation of functional sPD-L1 with PD-1-binding capacity in cancer patients. Copyright © 2018 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

Identical linkage and cooperativity of oxygen and carbon monoxide binding to Octopus dofleini hemocyanin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Connelly, P.R.; Gill, S.J.; Miller, K.I.

1989-02-21

Employment of high-precision thin-layer methods has enabled detailed functional characterization of oxygen and carbon monoxide binding for (1) the fully assembled form with 70 binding sites and (2) the isolated chains with 7 binding sites of octopus dofleini hemocyanin. The striking difference in the cooperativities of the two ligands for the assembled decamer is revealed through an examination of the binding capacities and the partition coefficient, determined as functions of the activities of both ligands. A global analysis of the data sets supported by a two-state allosteric model assuming an allosteric unit of 7. Higher level allosteric interactions were notmore » indicated. This contrasts to results obtained for arthropod hemocyanins. Oxygen and carbon monoxide experiments performed on the isolated subunit chain confirmed the presence of functional heterogeneity reported previously. The analysis shows two types of binding sites in the ratio of 4:3.« less

Clinical relevance of drug binding to plasma proteins

NASA Astrophysics Data System (ADS)

Ascenzi, Paolo; Fanali, Gabriella; Fasano, Mauro; Pallottini, Valentina; Trezza, Viviana

2014-12-01

Binding to plasma proteins highly influences drug efficacy, distribution, and disposition. Serum albumin, the most abundant protein in plasma, is a monomeric multi-domain macromolecule that displays an extraordinary ligand binding capacity, providing a depot and carrier for many endogenous and exogenous compounds, such as fatty acids and most acidic drugs. α-1-Acid glycoprotein, the second main plasma protein, is a glycoprotein physiologically involved in the acute phase reaction and is the main carrier for basic and neutral drugs. High- and low-density lipoproteins play a limited role in drug binding and are natural drug delivery system only for few lipophilic drugs or lipid-based formulations. Several factors influence drug binding to plasma proteins, such as pathological conditions, concurrent administration of drugs, sex, and age. Any of these factors, in turn, influences drug efficacy and toxicity. Here, biochemical, biomedical, and biotechnological aspects of drug binding to plasma proteins are reviewed.

What is working memory capacity, and how can we measure it?

PubMed Central

Wilhelm, Oliver; Hildebrandt, Andrea; Oberauer, Klaus

2013-01-01

A latent variable study examined whether different classes of working-memory tasks measure the same general construct of working-memory capacity (WMC). Data from 270 subjects were used to examine the relationship between Binding, Updating, Recall-N-back, and Complex Span tasks, and the relations of WMC with secondary memory measures, indicators of cognitive control from two response-conflict paradigms (Simon task and Eriksen flanker task), and fluid intelligence. Confirmatory factor analyses support the concept of a general WMC factor. Results from structural-equation modeling show negligible relations of WMC with response-conflict resolution, and very strong relations of WMC with secondary memory and fluid intelligence. The findings support the hypothesis that individual differences in WMC reflect the ability to build, maintain and update arbitrary bindings. PMID:23898309

Enhancement of anion-exchange chromatography of DNA using compaction agents

NASA Technical Reports Server (NTRS)

Murphy, Jason C.; Fox, George E.; Willson, Richard C.

2003-01-01

The use of adsorptive chromatography for preparative nucleic acid separations is often limited by low capacity. The possibility that the adsorbent surface area sterically accessible to nucleic acid molecules could be increased by reducing their radius of gyration with compaction agents has been investigated. The equilibrium adsorption capacity of Q Sepharose anion-exchange matrix for plasmid DNA at 600 mM NaCl was enhanced by up to ca. 40% in the presence of 2.5 mM spermine. In addition, compaction agent selectivity has been demonstrated. Spermine, for example, enhances the adsorption of both plasmid and genomic DNA, spermidine enhances binding only of plasmid, and hexamine cobalt enhances only the binding of genomic DNA. Compaction may be generally useful for enhancing adsorptive separations of nucleic acids.

Preparation of well-defined erythromycin imprinted non-woven fabrics via radiation-induced RAFT-mediated grafting

NASA Astrophysics Data System (ADS)

Söylemez, Meshude Akbulut; Barsbay, Murat; Güven, Olgun

2018-01-01

Radiation-induced RAFT polymerization technique was applied to synthesize well-defined molecularly imprinted polymers (MIPs) of erythromycin (ERY). Methacrylic acid (MAA) was grafted onto porous polyethylene (PE)/polypropylene (PP) nonwoven fabrics, under γ-irradiation by employing 2-pheny-2-propyl benzodithioate as the RAFT agent and ethylene glycol dimethacrylate (EGDMA) as the crosslinker. MAA/erythromycin ratios of 2/1, 4/1, 6/1 were tested to optimize the synthesis of MIPs. The highest binding capacity was encountered at a MAA/ERY ratio of 4/1. Non-imprinted polymers (NIPs) were also synthesized in the absence of ERY. The MIPs synthesized by RAFT method presented a better binding capacity compared to those prepared by conventional method where no RAFT agent was employed.

Facile synthesis of zwitterionic polymer-coated core-shell magnetic nanoparticles for highly specific capture of N-linked glycopeptides

NASA Astrophysics Data System (ADS)

Chen, Yajing; Xiong, Zhichao; Zhang, Lingyi; Zhao, Jiaying; Zhang, Quanqing; Peng, Li; Zhang, Weibing; Ye, Mingliang; Zou, Hanfa

2015-02-01

Highly selective and efficient capture of glycosylated proteins and peptides from complex biological samples is of profound significance for the discovery of disease biomarkers in biological systems. Recently, hydrophilic interaction liquid chromatography (HILIC)-based functional materials have been extensively utilized for glycopeptide enrichment. However, the low amount of immobilized hydrophilic groups on the affinity material has limited its specificity, detection sensitivity and binding capacity in the capture of glycopeptides. Herein, a novel affinity material was synthesized to improve the binding capacity and detection sensitivity for glycopeptides by coating a poly(2-(methacryloyloxy)ethyl)-dimethyl-(3-sulfopropyl) ammonium hydroxide (PMSA) shell onto Fe3O4@SiO2 nanoparticles, taking advantage of reflux-precipitation polymerization for the first time (denoted as Fe3O4@SiO2@PMSA). The thick polymer shell endows the nanoparticles with excellent hydrophilic property and several functional groups on the polymer chains. The resulting Fe3O4@SiO2@PMSA demonstrated an outstanding ability for glycopeptide enrichment with high selectivity, extremely high detection sensitivity (0.1 fmol), large binding capacity (100 mg g-1), high enrichment recovery (above 73.6%) and rapid magnetic separation. Furthermore, in the analysis of real complicated biological samples, 905 unique N-glycosylation sites from 458 N-glycosylated proteins were reliably identified in three replicate analyses of a 65 μg protein sample extracted from mouse liver, showing the great potential of Fe3O4@SiO2@PMSA in the detection and identification of low-abundance N-linked glycopeptides in biological samples.Highly selective and efficient capture of glycosylated proteins and peptides from complex biological samples is of profound significance for the discovery of disease biomarkers in biological systems. Recently, hydrophilic interaction liquid chromatography (HILIC)-based functional materials have been extensively utilized for glycopeptide enrichment. However, the low amount of immobilized hydrophilic groups on the affinity material has limited its specificity, detection sensitivity and binding capacity in the capture of glycopeptides. Herein, a novel affinity material was synthesized to improve the binding capacity and detection sensitivity for glycopeptides by coating a poly(2-(methacryloyloxy)ethyl)-dimethyl-(3-sulfopropyl) ammonium hydroxide (PMSA) shell onto Fe3O4@SiO2 nanoparticles, taking advantage of reflux-precipitation polymerization for the first time (denoted as Fe3O4@SiO2@PMSA). The thick polymer shell endows the nanoparticles with excellent hydrophilic property and several functional groups on the polymer chains. The resulting Fe3O4@SiO2@PMSA demonstrated an outstanding ability for glycopeptide enrichment with high selectivity, extremely high detection sensitivity (0.1 fmol), large binding capacity (100 mg g-1), high enrichment recovery (above 73.6%) and rapid magnetic separation. Furthermore, in the analysis of real complicated biological samples, 905 unique N-glycosylation sites from 458 N-glycosylated proteins were reliably identified in three replicate analyses of a 65 μg protein sample extracted from mouse liver, showing the great potential of Fe3O4@SiO2@PMSA in the detection and identification of low-abundance N-linked glycopeptides in biological samples. Electronic supplementary information (ESI) available. See DOI: 10.1039/c4nr05955g

Phospholipid epitopes for mouse antibodies against bromelain-treated mouse erythrocytes.

PubMed Central

Kawaguchi, S

1987-01-01

The reactivity of mouse antibodies against bromelain-treated mouse erythrocytes (BrMRBC) with phospholipid epitopes was assessed by ELISA, using four clones of monoclonal anti-BrMRBC antibodies that had idiotypes distinct from one another. The four antibodies could bind to low-density lipoproteins (LDL) from human and chicken, but not to LDL from mouse and rat. As to liposomes of natural phospholipids, all the clones reacted with liposomes of phosphatidylcholine, and some of them could react with liposomes of sphingomyelin, phosphatidylglycerol, phosphatidylic acid or cardiolipin. For liposomes of synthetic phosphatidylcholine with different fatty acids, the length of carbon chains and the number of unsaturated carbon chains of the fatty acids markedly affected the binding of each monoclonal antibody to the liposomes. The addition of dicetyl phosphate or stearylamine to phosphatidylcholine liposomes changed the reactivity of the liposomes. These results support the view that mouse anti-BrMRBC antibodies can recognize appropriately spaced phosphorylcholine residues on the surface of phospholipid liposomes, LDL and cells. The four clones had similar capacities for binding to LDL as well as to BrMRBC, but they had obviously different capacities for binding to phospholipid liposomes; the epitopes on phospholipid liposomes used in the present study were not so perfect as to react well with every anti-BrMRBC antibody. PMID:2443446

Enhanced binding by dextran-grafting to Protein A affinity chromatographic media.

PubMed

Zhao, Lan; Zhu, Kai; Huang, Yongdong; Li, Qiang; Li, Xiunan; Zhang, Rongyue; Su, Zhiguo; Wang, Qibao; Ma, Guanghui

2017-04-01

Dextran-grafted Protein A affinity chromatographic medium was prepared by grafting dextran to agarose-based matrix, followed by epoxy-activation and Protein A coupling site-directed to sulfhydryl groups of cysteine molecules. An enhancement of both the binding performance and the stability was achieved for this dextran-grafted Protein A chromatographic medium. Its dynamic binding capacity was 61 mg immunoglobulin G/mL suction-dried gel, increased by 24% compared with that of the non-grafted medium. The binding capacity of dextran-grafted medium decreased about 7% after 40 cleaning-in-place cycles, much lower than that of the non-grafted medium as decreased about 15%. Confocal laser scanning microscopy results showed that immunoglobulin G was bound to both the outside and the inside of dextran-grafted medium faster than that of non-grafted one. Atomic force microscopy showed that this dextran-grafted Protein A medium had much rougher surface with a vertical coordinate range of ±80 nm, while that of non-grafted one was ±10 nm. Grafted dextran provided a more stereo surface morphology and immunoglobulin G molecules were more easily to be bound. This high-performance dextran-grafted Protein A affinity chromatographic medium has promising applications in large-scale antibody purification. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Eating habits modulate short term memory and epigenetical regulation of brain derived neurotrophic factor in hippocampus of low- and high running capacity rats.

PubMed

Torma, Ferenc; Bori, Zoltan; Koltai, Erika; Felszeghy, Klara; Vacz, Gabriella; Koch, Lauren; Britton, Steven; Boldogh, Istvan; Radak, Zsolt

2014-08-01

Exercise capacity and dietary restriction (DR) are linked to improved quality of life, including enhanced brain function and neuro-protection. Brain derived neurotrophic factor (BDNF) is one of the key proteins involved in the beneficial effects of exercise on brain. Low capacity runner (LCR) and high capacity runner (HCR) rats were subjected to DR in order to investigate the regulation of BDNF. HCR-DR rats out-performed other groups in a passive avoidance test. BDNF content increased significantly in the hippocampus of HCR-DR groups compared to control groups (p<0.05). The acetylation of H3 increased significantly only in the LCR-DR group. However, chip-assay revealed that the specific binding between acetylated histone H3 and BNDF promoter was increased in both LCR-DR and HCR-DR groups. In spite of these increases in binding, at the transcriptional level only, the LCR-DR group showed an increase in BDNF mRNA content. Additionally, DR also induced the activity of cAMP response element-binding protein (CREB), while the content of SIRT1 was not altered. Peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) was elevated in HCR-DR groups. But, based on the levels of nuclear respiratory factor-1 and cytocrome c oxidase, it appears that DR did not cause mitochondrial biogenesis. The data suggest that DR-mediated induction of BDNF levels includes chromatin remodeling. Moreover, DR does not induce mitochondrial biogenesis in the hippocampus of LCR/HCR rats. DR results in different responses to a passive avoidance test, and BDNF regulation in LCR and HCR rats. Copyright © 2014 Elsevier Inc. All rights reserved.

Polysulfide Binding to Several Nanoscale Magnéli Phases Synthesized in Carbon for Long-Life Lithium-Sulfur Battery Cathodes.

PubMed

Zubair, Usman; Amici, Julia; Francia, Carlotta; McNulty, David; Bodoardo, Silvia; O'Dwyer, Colm

2018-06-11

In Li-S batteries, it is important to ensure efficient reversible conversion of sulfur to lithium polysulfide (LiPS). Shuttling effects caused by LiPS dissolution can lead to reduced performance and cycle life. Although carbon materials rely on physical trapping of polysulfides, polar oxide surfaces can chemically bind LiPS to improve the stability of sulfur cathodes. We show a simple synthetic method that allows high sulfur loading into mesoporous carbon preloaded with spatially localized nanoparticles of several Magnéli-phase titanium oxide (Ti n O 2n-1 ). This material simultaneously suppresses polysulfide shuttling phenomena by chemically binding Li polysulfides onto several Magnéli-phase surfaces in a single cathode and ensures physical confinement of sulfur and LiPS. The synergy between chemical immobilization of significant quantities of LiPS at the surface of several Ti n O 2n-1 phases and physical entrapment results in coulombically efficient high-rate cathodes with long cycle life and high capacity. These cathodes function efficiently at low electrolyte-to-sulfur ratios to provide high gravimetric and volumetric capacities in comparison with their highly porous carbon counterparts. Assembled coin cells have an initial discharge capacity of 1100 mAh g -1 at 0.1C and maintain a reversible capacity of 520 mAh g -1 at 0.2C for more than 500 cycles. Even at 1C, the cell loses only 0.06 % per cycle for 1000 cycles with a coulombic efficiency close to 99 %. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

The roles of protein disulphide isomerase family A, member 3 (ERp57) and surface thiol/disulphide exchange in human spermatozoa-zona pellucida binding.

PubMed

Wong, Chi-Wai; Lam, Kevin K W; Lee, Cheuk-Lun; Yeung, William S B; Zhao, Wei E; Ho, Pak-Chung; Ou, Jian-Ping; Chiu, Philip C N

2017-04-01

Are multimeric sperm plasma membrane protein complexes, ERp57 and sperm surface thiol content involved in human spermatozoa-zona pellucida (ZP) interaction? ERp57 is a component of a multimeric spermatozoa-ZP receptor complex involved in regulation of human spermatozoa-ZP binding via up-regulation of sperm surface thiol content. A spermatozoon acquires its fertilization capacity within the female reproductive tract by capacitation. Spermatozoa-ZP receptor is suggested to be a composite structure that is assembled into a functional complex during capacitation. Sperm surface thiol content is elevated during capacitation. ERp57 is a protein disulphide isomerase that modulates the thiol-disulphide status of proteins. The binding ability and components of protein complexes in extracted membrane protein fractions of spermatozoa were studied. The roles of capacitation, thiol-disulphide reagent treatments and ERp57 on sperm functions and sperm surface thiol content were assessed. Spermatozoa were obtained from semen samples from normozoospermic men. Human oocytes were obtained from an assisted reproduction programme. Blue native polyacrylamide gel electrophoresis, western ligand blotting and mass spectrometry were used to identify the components of solubilized ZP/ZP3-binding complexes. The localization and expression of sperm surface thiol and ERp57 were studied by immunostaining and sperm surface protein biotinylation followed by western blotting. Sperm functions were assessed by standard assays. Several ZP-binding complexes were isolated from the cell membrane of capacitated spermatozoa. ERp57 was a component of one of these complexes. Capacitation significantly increased the sperm surface thiol content, acrosomal thiol distribution and ERp57 expression on sperm surface. Sperm surface thiol and ERp57 immunoreactivity were localized to the acrosomal region of spermatozoa, a region responsible for ZP-binding. Up-regulation of the surface thiol content or ERp57 surface expression in vitro stimulated ZP-binding capacity of human spermatozoa. Blocking of ERp57 function by specific antibody or inhibitors against ERp57 reduced the surface thiol content and ZP-binding capacity of human spermatozoa. N/A. The mechanisms by which up-regulation of surface thiol content stimulates spermatozoa-ZP binding have not been depicted. Thiol-disulphide exchange is a crucial event in capacitation. ERp57 modulates the event and the subsequent fertilization process. Modulation of the surface thiol content of the spermatozoa of subfertile men may help to increase fertilization rate in assisted reproduction. This work was supported by The Hong Kong Research Grant Council Grant HKU764611 and HKU764512M to P.C.N.C. The authors have no competing interests. © The Author 2017. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com

Theme: Innovative Curricula.

ERIC Educational Resources Information Center

Coffey, David M.; And Others

1994-01-01

Includes "Risking the Future" (Coffey); "Breaking Tradition" (Paynton); "Sustainable Farm Plan Activity" (Vahoviak et al.); "Curriculum Integration and Ornamental Horticulture" (Clark); "Ties That Bind" (Barden et al.); "Building Capacity for an Innovative Elementary Agriscience…

Krill Oil-In-Water Emulsion Protects against Lipopolysaccharide-Induced Proinflammatory Activation of Macrophages In Vitro

PubMed Central

Bonaterra, Gabriel A.; Driscoll, David; Schwarzbach, Hans; Kinscherf, Ralf

2017-01-01

Background: Parenteral nutrition is often a mandatory therapeutic strategy for cases of septicemia. Likewise, therapeutic application of anti-oxidants, anti-inflammatory therapy, and endotoxin lowering, by removal or inactivation, might be beneficial to ameliorate the systemic inflammatory response during the acute phases of critical illness. Concerning anti-inflammatory properties in this setting, omega-3 fatty acids of marine origin have been frequently described. This study investigated the anti-inflammatory and LPS-inactivating properties of krill oil (KO)-in-water emulsion in human macrophages in vitro. Materials and Methods: Differentiated THP-1 macrophages were activated using specific ultrapure-LPS that binds only on the toll-like receptor 4 (TLR4) in order to determine the inhibitory properties of the KO emulsion on the LPS-binding capacity, and the subsequent release of TNF-α. Results: KO emulsion inhibited the macrophage binding of LPS to the TLR4 by 50% (at 12.5 µg/mL) and 75% (at 25 µg/mL), whereas, at 50 µg/mL, completely abolished the LPS binding. Moreover, KO (12.5 µg/mL, 25 µg/mL, or 50 µg/mL) also inhibited (30%, 40%, or 75%, respectively) the TNF-α release after activation with 0.01 µg/mL LPS in comparison with LPS treatment alone. Conclusion: KO emulsion influences the LPS-induced pro-inflammatory activation of macrophages, possibly due to inactivation of the LPS binding capacity. PMID:28294970

Multiple sup 3 H-oxytocin binding sites in rat myometrial plasma membranes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Crankshaw, D.; Gaspar, V.; Pliska, V.

1990-01-01

The affinity spectrum method has been used to analyse binding isotherms for {sup 3}H-oxytocin to rat myometrial plasma membranes. Three populations of binding sites with dissociation constants (Kd) of 0.6-1.5 x 10(-9), 0.4-1.0 x 10(-7) and 7 x 10(-6) mol/l were identified and their existence verified by cluster analysis based on similarities between Kd, binding capacity and Hill coefficient. When experimental values were compared to theoretical curves constructed using the estimated binding parameters, good fits were obtained. Binding parameters obtained by this method were not influenced by the presence of GTP gamma S (guanosine-5'-O-3-thiotriphosphate) in the incubation medium. The bindingmore » parameters agree reasonably well with those found in uterine cells, they support the existence of a medium affinity site and may allow for an explanation of some of the discrepancies between binding and response in this system.« less

Bead mediated separation of microparticles in droplets.

PubMed

Wang, Sida; Sung, Ki-Joo; Lin, Xiaoxia Nina; Burns, Mark A

2017-01-01

Exchange of components such as particles and cells in droplets is important and highly desired in droplet microfluidic assays, and many current technologies use electrical or magnetic fields to accomplish this process. Bead-based microfluidic techniques offer an alternative approach that uses the bead's solid surface to immobilize targets like particles or biological material. In this paper, we demonstrate a bead-based technique for exchanging droplet content by separating fluorescent microparticles in a microfluidic device. The device uses posts to filter surface-functionalized beads from a droplet and re-capture the filtered beads in a new droplet. With post spacing of 7 μm, beads above 10 μm had 100% capture efficiency. We demonstrate the efficacy of this system using targeted particles that bind onto the functionalized beads and are, therefore, transferred from one solution to another in the device. Binding capacity tests performed in the bulk phase showed an average binding capacity of 5 particles to each bead. The microfluidic device successfully separated the targeted particles from the non-targeted particles with up to 98% purity and 100% yield.

Bead mediated separation of microparticles in droplets

PubMed Central

Sung, Ki-Joo; Lin, Xiaoxia Nina; Burns, Mark A.

2017-01-01

Exchange of components such as particles and cells in droplets is important and highly desired in droplet microfluidic assays, and many current technologies use electrical or magnetic fields to accomplish this process. Bead-based microfluidic techniques offer an alternative approach that uses the bead’s solid surface to immobilize targets like particles or biological material. In this paper, we demonstrate a bead-based technique for exchanging droplet content by separating fluorescent microparticles in a microfluidic device. The device uses posts to filter surface-functionalized beads from a droplet and re-capture the filtered beads in a new droplet. With post spacing of 7 μm, beads above 10 μm had 100% capture efficiency. We demonstrate the efficacy of this system using targeted particles that bind onto the functionalized beads and are, therefore, transferred from one solution to another in the device. Binding capacity tests performed in the bulk phase showed an average binding capacity of 5 particles to each bead. The microfluidic device successfully separated the targeted particles from the non-targeted particles with up to 98% purity and 100% yield. PMID:28282412

In vitro evaluation of whole faba bean and its seed coat as a potential source of functional food components.

PubMed

Çalışkantürk Karataş, Selen; Günay, Demet; Sayar, Sedat

2017-09-01

In vitro studies were conducted to evaluate the particular nutritional benefits of whole faba bean seed (WFB) and fava bean seed coat (FBSC). Total dietary fiber contents of WFB and FBSC were 27.5% and 82.3%, respectively. FBSC were contained much higher total phenolic substances, condensed tannins, and total antioxidant activity than WFB. Bile acid (BA)-binding capacities of in vitro digested samples and nutritionally important products produced by in vitro fermentation of digestion residues were also studied. The BA-binding capacities of WFB and FBSC were 1.94 and 37.50μmol/100mg, respectively. Total BA bound by FBSC was even higher than the positive standard cholestyramine. Lignin and other constituents of the Klason residue were found to influence BA-binding properties. Moreover, the extent of the in vitro fermentation process showed that, fermentability of FBSC residue was significantly lower than that of WFB residue. Overall, faba bean, especially its seed coat, has great potential as a functional food. Copyright © 2017 Elsevier Ltd. All rights reserved.

Exploring the binding energy profiles of full agonists, partial agonists, and antagonists of the α7 nicotinic acetylcholine receptor.

PubMed

Tabassum, Nargis; Ma, Qianyun; Wu, Guanzhao; Jiang, Tao; Yu, Rilei

2017-09-01

Nicotinic acetylcholine receptors (nAChRs) belong to the Cys-loop receptor family and are important drug targets for the treatment of neurological diseases. However, the precise determinants of the binding efficacies of ligands for these receptors are unclear. Therefore, in this study, the binding energy profiles of various ligands (full agonists, partial agonists, and antagonists) were quantified by docking those ligands with structural ensembles of the α7 nAChR exhibiting different degrees of C-loop closure. This approximate treatment of interactions suggested that full agonists, partial agonists, and antagonists of the α7 nAChR possess distinctive binding energy profiles. Results from docking revealed that ligand binding efficacy may be related to the capacity of the ligand to stabilize conformational states with a closed C loop.

Soluble Fiber with High Water-Binding Capacity, Swelling Capacity, and Fermentability Reduces Food Intake by Promoting Satiety Rather Than Satiation in Rats.

PubMed

Tan, Chengquan; Wei, Hongkui; Zhao, Xichen; Xu, Chuanhui; Zhou, Yuanfei; Peng, Jian

2016-10-02

To understand whether soluble fiber (SF) with high water-binding capacity (WBC), swelling capacity (SC) and fermentability reduces food intake and whether it does so by promoting satiety or satiation or both, we investigated the effects of different SFs with these properties on the food intake in rats. Thirty-two male Sprague-Dawley rats were randomized to four equal groups and fed the control diet or diet containing 2% konjac flour (KF), pregelatinized waxy maize starch (PWMS) plus guar gum (PG), and PWMS starch plus xanthan gum (PX) for three weeks, with the measured values of SF, WBC, and SC in the four diets following the order of PG > KF > PX > control. Food intake, body weight, meal pattern, behavioral satiety sequence, and short-chain fatty acids (SCFAs) in cecal content were evaluated. KF and PG groups reduced the food intake, mainly due to the decreased feeding behavior and increased satiety, as indicated by decreased meal numbers and increased inter-meal intervals. Additionally, KF and PG groups increased concentrations of acetate acid, propionate acid, and SCFAs in the cecal contents. Our results indicate that SF with high WBC, SC, and fermentability reduces food intake-probably by promoting a feeling of satiety in rats to decrease their feeding behavior.

Soluble Fiber with High Water-Binding Capacity, Swelling Capacity, and Fermentability Reduces Food Intake by Promoting Satiety Rather Than Satiation in Rats

PubMed Central

Tan, Chengquan; Wei, Hongkui; Zhao, Xichen; Xu, Chuanhui; Zhou, Yuanfei; Peng, Jian

2016-01-01

To understand whether soluble fiber (SF) with high water-binding capacity (WBC), swelling capacity (SC) and fermentability reduces food intake and whether it does so by promoting satiety or satiation or both, we investigated the effects of different SFs with these properties on the food intake in rats. Thirty-two male Sprague-Dawley rats were randomized to four equal groups and fed the control diet or diet containing 2% konjac flour (KF), pregelatinized waxy maize starch (PWMS) plus guar gum (PG), and PWMS starch plus xanthan gum (PX) for three weeks, with the measured values of SF, WBC, and SC in the four diets following the order of PG > KF > PX > control. Food intake, body weight, meal pattern, behavioral satiety sequence, and short-chain fatty acids (SCFAs) in cecal content were evaluated. KF and PG groups reduced the food intake, mainly due to the decreased feeding behavior and increased satiety, as indicated by decreased meal numbers and increased inter-meal intervals. Additionally, KF and PG groups increased concentrations of acetate acid, propionate acid, and SCFAs in the cecal contents. Our results indicate that SF with high WBC, SC, and fermentability reduces food intake—probably by promoting a feeling of satiety in rats to decrease their feeding behavior. PMID:27706095

Metal-functionalized silicene for efficient hydrogen storage.

PubMed

Hussain, Tanveer; Chakraborty, Sudip; Ahuja, Rajeev

2013-10-21

First-principles calculations based on density functional theory are used to investigate the electronic structure along with the stability, bonding mechanism, band gap, and charge transfer of metal-functionalized silicene to envisage its hydrogen-storage capacity. Various metal atoms including Li, Na, K, Be, Mg, and Ca are doped into the most stable configuration of silicene. The corresponding binding energies and charge-transfer mechanisms are discussed from the perspective of hydrogen-storage compatibility. The Li and Na metal dopants are found to be ideally suitable, not only for strong metal-to-substrate binding and uniform distribution over the substrate, but also for the high-capacity storage of hydrogen. The stabilities of both Li- and Na-functionalized silicene are also confirmed through molecular dynamics simulations. It is found that both of the alkali metals, Li(+) and Na(+), can adsorb five hydrogen molecules, attaining reasonably high storage capacities of 7.75 and 6.9 wt %, respectively, with average adsorption energies within the range suitable for practical hydrogen-storage applications. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Engineering Copper Carboxylate Functionalities on Water Stable Metal–Organic Frameworks for Enhancement of Ammonia Removal Capacities

DOE Office of Scientific and Technical Information (OSTI.GOV)

Joshi, Jayraj N.; Garcia-Gutierrez, Erika Y.; Moran, Colton M.

Functionalization of copper carboxylate groups on a series of UiO-66 metal organic framework (MOF) analogues and their corresponding impact on humid and dry ammonia adsorption behavior were studied. Relative locations of possible carboxylic acid binding sites for copper on the MOF analogues were varied on ligand and missing linker defect sites. Materials after copper incorporation exhibited increased water vapor and ammonia affinity during isothermal adsorption and breakthrough experiments, respectively. The introduction of copper markedly increased ammonia adsorption capacities for all adsorbents possessing carboxyl binding sites. In particular, the new MOF UiO-66-(COOCu)2 displayed the highest ammonia breakthrough capacities of 6.38 andmore » 6.84 mmol g–1 under dry and humid conditions, respectively, while retaining crystallinity and porosity. Relative carboxylic acid site locations were also found to impact sorbent stability, as missing linker defect functionalized materials degraded under humid conditions after copper incorporation. Postsynthetic metal insertion provides a method for adding sites that are analogous to open metal sites while maintaining good structural stability.« less

Preparation and evaluation of magnetic core-shell mesoporous molecularly imprinted polymers for selective adsorption of tetrabromobisphenol S.

PubMed

Wang, Xuemei; Huang, Pengfei; Ma, Xiaomin; Wang, Huan; Lu, Xiaoquan; Du, Xinzhen

2017-05-01

Novel magnetic mesoporous molecularly imprinted polymers (MMIPs) with core-shell structure were prepared by simple surface molecular imprinting polymerization using tetrabromobisphenol-S (TBBPS) as the template. The MMIPs-TBBPS were characterized by fourier-transform infrared spectrometry (FTIR), scanning electron microscopy (SEM), transmission electron microscopy (TEM), N 2 adsorption-desorption transmission, and vibrating sample magnetometry. The resultant MMIPs-TBBPS were successfully applied magnetic solid-phase extraction (MSPE) coupled with HPLC determination of TBBPS in spiked real water samples with recoveries of 77.8-88.9%. The adsorption experiments showed that the binding capacity of MMIPs-TBBPS to TBBPS and six structural analogs were significantly higher than that of the magnetic nonimprinted polymers (MNIPs). Meanwhile, the MMIPs-TBBPS possessed rapid binding affinity, excellent magnetic response, specific selectivity and high adsorption capacity toward TBBPS with a maximum adsorption capacity of 1626.8µgg -1 . The analytical results indicate that the MMIPs-TBBPS are promising materials for selective separation and fast enrichment of TBBPS from complicated enviromental samples. Copyright © 2017 Elsevier B.V. All rights reserved.

Metal binding stoichiometry and isotherm choice in biosorption

DOE Office of Scientific and Technical Information (OSTI.GOV)

Schiewer, S.; Wong, M.H.

1999-11-01

Seaweeds that possess a high metal binding capacity may be used as biosorbents for the removal of toxic heavy metals from wastewater. The binding of Cu and Ni by three brown algae (Sargassum, Colpomenia, Petalonia) and one green alga (Ulva) was investigated at pH 4.0 and pH 3.0. The greater binding strength of Cu is reflected in a binding constant that is about 10 times as high as that of Ni. The extent of metal binding followed the order Petalonia {approximately} Sargassum > Colpomenia > Ulva. This was caused by a decreasing number of binding sites and by much lowermore » metal binding constants for Ulva as compared to the brown algae. Three different stoichiometric assumptions are compared for describing the metal binding, which assume either that each metal ion M binds to one binding site B forming a BM complex or that a divalent metal ion M binds to two monovalent sites B forming BM{sub 0.5} or B{sub 2}M complexes, respectively. Stoichiometry plots are proposed as tools to discern the relevant binding stoichiometry. The pH effect in metal binding and the change in proton binding were well predicted for the B{sub 2}M or BM{sub 0.5} stoichiometries with the former being better for Cu and the latter preferable for Ni. Overall, the BM{sub 0.5} model is recommended because it avoids iterations.« less

Total iron binding capacity

MedlinePlus

... ed. Philadelphia, PA: Elsevier Saunders; 2014:chap 12. Review Date 2/11/2016 Updated by: Todd Gersten, ... Oncology, Florida Cancer Specialists & Research Institute, Wellington, FL. Review provided by VeriMed Healthcare Network. Also reviewed by ...

Binding of human serum proteins to titanium dioxide particles in vitro.

PubMed

Zaqout, Mazen S K; Sumizawa, Tomoyuki; Igisu, Hideki; Higashi, Toshiaki; Myojo, Toshihiko

2011-01-01

To determine the capacity of human serum proteins to bind to titanium dioxide (TiO(2)) particles of different polymorphs and sizes. TiO(2) particles were mixed with diluted human serum, purified human serum albumin (HSA) or purified human serum gamma-globulin (HGG) solutions. After incubation at 37°C for 1 h, the particles were sedimented by centrifugation, and proteins in the supernatant, as well as those bound to the particles, were analyzed. The total protein concentration in the supernatant was lowered by TiO(2), whereas the albumin/globulin ratio was elevated by the particles. Incubation with TiO(2) also lowered the immunoglobulin, pre-albumin, beta2-microglobulin, ceruloplasmin and retinol-binding protein levels, but not ferritin levels, in the supernatant. After sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), proteins in the supernatant, especially HGG, were observed to decrease, while those released from the particles (after adding 1% SDS and heating) increased, depending on the dose of TiO(2). Purified HGG and HSA were also bound to TiO(2), although the former appeared to have a higher affinity. All the proteins tested showed the highest binding potency to the amorphous particles (<50 nm) and the lowest to the rutile particles (<5,000 nm), while binding to anatase particles was intermediate. The affinity to the larger anatase was higher than that to smaller anatase particles in most cases. Human serum proteins, including the two major components, HSA and HGG, are bound by TiO(2) particles. The polymorph of the particles seems to be important for determining the binding capacity of the particles and it may affect distribution of the particles in the body.

Improvement of impaired albumin binding capacity in acute-on-chronic liver failure by albumin dialysis.

PubMed

Klammt, Sebastian; Mitzner, Steffen R; Stange, Jan; Loock, Jan; Heemann, Uwe; Emmrich, Jörg; Reisinger, Emil C; Schmidt, Reinhard

2008-09-01

Extracorporeal albumin dialysis (ECAD) enables the elimination of albumin bound substances and is used as artificial liver support system. Albumin binding function for the benzodiazepine binding site specific marker Dansylsarcosine was estimated in plasma samples of 22 patients with cirrhosis and hyperbilirubinaemia (ECAD: n = 12; control: n = 10) during a period of 30 days in a randomized controlled clinical ECAD trial. Albumin Binding Capacity (ABiC) at baseline was reduced to 31.8% (median; range 24%-74%) and correlated to the severity of liver disease. Within two weeks a significant improvement of ABiC and a reduction of the albumin bound markers bilirubin and bile acids were observed in the ECAD group. During single treatments a significant decrease of albumin bound substances (bilirubin and bile acids) as well as an increase in ABiC was observed. In the control group, baseline ABiC was significantly lower in patients who died during study period (34.2% vs. 41.7%; P < 0.028), whereas no significant differences were observed for CHILD, coagulation factors, albumin, bile acids nor bilirubin. At baseline 13 patients had a severely impaired ABiC (<40%), improvement of ABiC was more frequent in the ECAD group (5/6) than in the SMT group (2/7). Reduced albumin binding function is present in decompensated liver failure and is related to severity and 30 day survival. ABiC can be improved by ECAD. The beneficial effect of this treatment may be related to the improvement of albumin binding function more than to the elimination of specific substances. Characterization of albumin function by the ABiC test may help to evaluate different liver support systems and other therapeutic measures.

Probiotic preparation has the capacity to hydrolyze proteins responsible for wheat allergy.

PubMed

De Angelis, Maria; Rizzello, Carlo G; Scala, Enrico; De Simone, Claudio; Farris, Giovanni A; Turrini, Francesco; Gobbetti, Marco

2007-01-01

This study was aimed at showing the capacity of probiotic VSL#3 to hydrolyze wheat flour allergens. Hydrolysis was investigated either by the use of baker's yeast bread treated with digestive enzymes and VSL#3, an experimental design that mimicked the activity of probiotics during gut colonization, or by the use of VSL#3 as a starter for dough fermentation, an experimental design that mimicked the predigestion of wheat flour proteins during food processing. Albumins, globulins, and gliadins extracted from wheat flour and chemically acidified and started dough and total proteins extracted from breads were analyzed by immunoblotting with pooled sera from patients with an allergy to wheat. Hydrolysis of wheat flour proteins was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and two-dimensional electrophoresis (2DE). Mass spectrometry matrix-assisted laser desorption and ionization-time of flight was used to identify some immunoglobulin E (IgE)-binding proteins. As shown by immunoblotting with sera from allergic patients, several IgE-binding proteins persisted after treatment of baker's yeast bread by pepsin and pancreatin. The signal of all these IgE-binding proteins disappeared after further treatment by VSL#3. As shown by SDS-PAGE and related immunoblotting and 2DE analyses, when VSL#3 was used as a starter for bread making, it caused a marked degradation of wheat proteins, including some IgE-binding proteins such as the putative transcription factor APFI and wheat alpha-amylase inhibitors. Indeed, the IgE-binding profile of the bread manufactured by VSL#3 was largely different from that of baker's yeast bread. The IgE-binding proteins that persisted in the bread made with VSL#3 were completely degraded by pepsin and pancreatin.

Thermodynamics of the GTP-GDP-operated Conformational Switch of Selenocysteine-specific Translation Factor SelB*

PubMed Central

Paleskava, Alena; Konevega, Andrey L.; Rodnina, Marina V.

2012-01-01

SelB is a specialized translation factor that binds GTP and GDP and delivers selenocysteyl-tRNA (Sec-tRNASec) to the ribosome. By analogy to elongation factor Tu (EF-Tu), SelB is expected to control the delivery and release of Sec-tRNASec to the ribosome by the structural switch between GTP- and GDP-bound conformations. However, crystal structures of SelB suggested a similar domain arrangement in the apo form and GDP- and GTP-bound forms of the factor, raising the question of how SelB can fulfill its delivery function. Here, we studied the thermodynamics of guanine nucleotide binding to SelB by isothermal titration calorimetry in the temperature range between 10 and 25 °C using GTP, GDP, and two nonhydrolyzable GTP analogs, guanosine 5′-O-(γ-thio)triphosphate (GTPγS) and guanosine 5′-(β,γ-imido)-triphosphate (GDPNP). The binding of SelB to either guanine nucleotide is characterized by a large heat capacity change (−621, −467, −235, and −275 cal × mol−1 × K−1, with GTP, GTPγS, GDPNP, and GDP, respectively), associated with compensatory changes in binding entropy and enthalpy. Changes in heat capacity indicate a large decrease of the solvent-accessible surface area in SelB, amounting to 43 or 32 amino acids buried upon binding of GTP or GTPγS, respectively, and 15–19 amino acids upon binding GDP or GDPNP. The similarity of the GTP and GDP forms in the crystal structures can be attributed to the use of GDPNP, which appears to induce a structure of SelB that is more similar to the GDP than to the GTP-bound form. PMID:22740700

Thermodynamics of the GTP-GDP-operated conformational switch of selenocysteine-specific translation factor SelB.

PubMed

Paleskava, Alena; Konevega, Andrey L; Rodnina, Marina V

2012-08-10

SelB is a specialized translation factor that binds GTP and GDP and delivers selenocysteyl-tRNA (Sec-tRNA(Sec)) to the ribosome. By analogy to elongation factor Tu (EF-Tu), SelB is expected to control the delivery and release of Sec-tRNA(Sec) to the ribosome by the structural switch between GTP- and GDP-bound conformations. However, crystal structures of SelB suggested a similar domain arrangement in the apo form and GDP- and GTP-bound forms of the factor, raising the question of how SelB can fulfill its delivery function. Here, we studied the thermodynamics of guanine nucleotide binding to SelB by isothermal titration calorimetry in the temperature range between 10 and 25 °C using GTP, GDP, and two nonhydrolyzable GTP analogs, guanosine 5'-O-(γ-thio)triphosphate (GTPγS) and guanosine 5'-(β,γ-imido)-triphosphate (GDPNP). The binding of SelB to either guanine nucleotide is characterized by a large heat capacity change (-621, -467, -235, and -275 cal × mol(-1) × K(-1), with GTP, GTPγS, GDPNP, and GDP, respectively), associated with compensatory changes in binding entropy and enthalpy. Changes in heat capacity indicate a large decrease of the solvent-accessible surface area in SelB, amounting to 43 or 32 amino acids buried upon binding of GTP or GTPγS, respectively, and 15-19 amino acids upon binding GDP or GDPNP. The similarity of the GTP and GDP forms in the crystal structures can be attributed to the use of GDPNP, which appears to induce a structure of SelB that is more similar to the GDP than to the GTP-bound form.

Specific ganglioside binding to receptor sites on T lymphocytes that couple to ganglioside-induced decrease of CD4 expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Morrison, W.J.; Offner, H.; Vandenbark, A.A.

1989-01-01

The binding of different gangliosides to rat T-helper lymphocytes was characterized under conditions that decrease CD4 expression on different mammalian T-helper lymphoctyes. Saturation binding by monosialylated ({sub 3}H)-GM{sub 1} to rat T-lymphocytes was time- and temperature-dependent, had a dissociation constant (K{sub D}) of 2.2 {plus minus} 1.4 {mu}M and a binding capacity near 2 fmoles/cell. Competitive inhibition of ({sup 3}H)- GM{sub 1} binding demonstrated a structural-activity related to the number of unconstrained sialic acid moieties on GM{sub 1}-congeneric gangliosides. A comparison between the results of these binding studies and gangliosides-induced decrease of CD4 expression demonstrated that every aspect of ({supmore » 3}H)-GM{sub 1} binding concurs with ganglioside modulation of CD4 expression. It is concluded that the specific decrease of CD4 expression induced by pretreatment with gangliosides involves the initial process of gangliosides binding to specific sites on CD4{sup {double dagger}} T-helper lymphocytes.« less

The contribution of perceptual factors and training on varying audiovisual integration capacity.

PubMed

Wilbiks, Jonathan M P; Dyson, Benjamin J

2018-06-01

The suggestion that the capacity of audiovisual integration has an upper limit of 1 was challenged in 4 experiments using perceptual factors and training to enhance the binding of auditory and visual information. Participants were required to note a number of specific visual dot locations that changed in polarity when a critical auditory stimulus was presented, under relatively fast (200-ms stimulus onset asynchrony [SOA]) and slow (700-ms SOA) rates of presentation. In Experiment 1, transient cross-modal congruency between the brightness of polarity change and pitch of the auditory tone was manipulated. In Experiment 2, sustained chunking was enabled on certain trials by connecting varying dot locations with vertices. In Experiment 3, training was employed to determine if capacity would increase through repeated experience with an intermediate presentation rate (450 ms). Estimates of audiovisual integration capacity (K) were larger than 1 during cross-modal congruency at slow presentation rates (Experiment 1), during perceptual chunking at slow and fast presentation rates (Experiment 2), and, during an intermediate presentation rate posttraining (Experiment 3). Finally, Experiment 4 showed a linear increase in K using SOAs ranging from 100 to 600 ms, suggestive of quantitative rather than qualitative changes in the mechanisms in audiovisual integration as a function of presentation rate. The data compromise the suggestion that the capacity of audiovisual integration is limited to 1 and suggest that the ability to bind sounds to sights is contingent on individual and environmental factors. (PsycINFO Database Record (c) 2018 APA, all rights reserved).

Lead biosorption of probiotic bacteria: effects of the intestinal content from laying hens.

PubMed

Xing, Sicheng; Wang, Jie; Liang, Juan Boo; Jahromi, Mohammad Faseleh; Zhu, Cui; Shokryazdan, Parisa; Laudadio, Vito; Tufarelli, Vincenzo; Liao, Xindi

2017-05-01

This study investigated the effects and the possible mechanisms of intestinal content (IC) from laying hens on in vitro lead (Pb 2+ ) biosorption of four probiotic bacterial strains (Bifidobacterium longum BB79, Lactobacillus paracasei Kgl6, Lactobacillus pentosus ITA23, and Lactobacillus acidipiscis ITA44). The total Pb 2+ removal capacity of the four probiotic strains, with and without capsule polysaccharides (CPSs), increased in the presence of IC compared to the control (without IC). SEM imaging revealed certain unidentified particles from the IC adhered on the surface of bacterial cells sorted out using flow cytometry. Follow-up experiment showed an overall trend of increase in the Pb 2+ removal capacity of the sorted bacteria, but statistically significant for L. pentosus ITA23 and B. longum BB79 after incubation with IC, particularly with the suspended solid portion of the IC. In addition, the Fourier transform infrared spectrophotometer data showed that functional groups such as C-H, O-H, C=O, and C-O-C which possibly associated with Pb 2+ binding were mainly presented in the suspended solid portion of IC. Putting the above together, we postulated that the enhanced Pb 2+ binding capacity the probiotic bacteria incubated in IC is due to the adherence of the yet to be identified particles which could much exist in suspended solid portion of IC containing negatively charged functional groups which bind with the positive Pb 2+ ions.

Treatment of meats with ionising radiations. VIII.—pH, water-binding capacity and proteolysis of irradiated raw beef and pork during storage, and the ATP-ase activity of irradiated rabbit muscle

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lawrie, R. A.; Sharp, J. G.; Bendall, J. R.

1961-11-01

The immediate effects of 5-Mrad ionizing radiation on beef and pork longissimus dorsi muscles were an increase in pH, a decrease in water-holding capacity, in increment in gel-volume for a given pH rise, and in soluble protein, and increased resistance to low- and high-speed homogenization. The indications of cross-binding induced by irradiation were supported by studies of isolated myofibrils from rabbit psoas muscle. Irradiation markedly reduced the syneresis (18 deg , mu = 0.04) and the swelling (0 deg , mu = 0.25) induced by ATP and, to a lesser extent, over-all fibrillar ATP-ase activity (the initial fast phase beingmore » depressed more than the slower second phase of the reaction). On storage (at -20 deg +37 deg pH and water-binding capacity increased generally with increase of temperature. Changes in pH occurred earlier with pork and to a greater extent than with beef. In sterile beef longissimus dorsi (irradiated or unirradiated) there was a decrease in soluble protein during storage for 60-90 days at 37- (indicating denaturation) and lncreases in TCA-soluble nitrogen and tyrosine (indicating proteolysis, which was more marked in unirradiated samples). The absence of soluble hydroxyproline and the presence of clearly marked cross- striations indicated that the autolysis must have involved sarcoplasmic and not fibrillar or connective tissue protein.« less

Anemia, Iron Deficiency and Iodine Deficiency among Nepalese School Children.

PubMed

Khatiwada, Saroj; Lamsal, Madhab; Gelal, Basanta; Gautam, Sharad; Nepal, Ashwini Kumar; Brodie, David; Baral, Nirmal

2016-07-01

To assess iodine and iron nutritional status among Nepalese school children. A cross-sectional, community based study was conducted in the two districts, Ilam (hilly region) and Udayapur (plain region) of eastern Nepal. A total of 759 school children aged 6-13 y from different schools within the study areas were randomly enrolled. A total of 759 urine samples and 316 blood samples were collected. Blood hemoglobin level, serum iron, total iron binding capacity and urinary iodine concentration was measured. Percentage of transferrin saturation was calculated using serum iron and total iron binding capacity values. The mean level of hemoglobin, serum iron, total iron binding capacity, transferrin saturation and median urinary iodine excretion were 12.29 ± 1.85 g/dl, 70.45 ± 34.46 μg/dl, 386.48 ± 62.48 μg/dl, 19.94 ± 12.07 % and 274.67 μg/L respectively. Anemia, iron deficiency and iodine deficiency (urinary iodine excretion <100 μg/L) were present in 34.5 %, 43.4 % and 12.6 % children respectively. Insufficient urinary iodine excretion (urinary iodine excretion <100 μg/L) was common in anemic and iron deficient children. Iron deficiency and anemia are common in Nepalese children, whereas, iodine nutrition is more than adequate. Low urinary iodine excretion was common in iron deficiency and anemia.

Parietal-Occipital Interactions Underlying Control- and Representation-Related Processes in Working Memory for Nonspatial Visual Features.

PubMed

Gosseries, Olivia; Yu, Qing; LaRocque, Joshua J; Starrett, Michael J; Rose, Nathan S; Cowan, Nelson; Postle, Bradley R

2018-05-02

Although the manipulation of load is popular in visual working memory research, many studies confound general attentional demands with context binding by drawing memoranda from the same stimulus category. In this fMRI study of human observers (both sexes), we created high- versus low-binding conditions, while holding load constant, by comparing trials requiring memory for the direction of motion of one random dot kinematogram (RDK; 1M trials) versus for three RDKs (3M), or versus one RDK and two color patches (1M2C). Memory precision was highest for 1M trials and comparable for 3M and 1M2C trials. And although delay-period activity in occipital cortex did not differ between the three conditions, returning to baseline for all three, multivariate pattern analysis decoding of a remembered RDK from occipital cortex was also highest for 1M trials and comparable for 3M and 1M2C trials. Delay-period activity in intraparietal sulcus (IPS), although elevated for all three conditions, displayed more sensitivity to demands on context binding than to load per se. The 1M-to-3M increase in IPS signal predicted the 1M-to-3M declines in both behavioral and neural estimates of working memory precision. These effects strengthened along a caudal-to-rostral gradient, from IPS0 to IPS5. Context binding-independent load sensitivity was observed when analyses were lateralized and extended into PFC, with trend-level effects evident in left IPS and strong effects in left lateral PFC. These findings illustrate how visual working memory capacity limitations arise from multiple factors that each recruit dissociable brain systems. SIGNIFICANCE STATEMENT Visual working memory capacity predicts performance on a wide array of cognitive and real-world outcomes. At least two theoretically distinct factors are proposed to influence visual working memory capacity limitations: an amodal attentional resource that must be shared across remembered items; and the demands on context binding. We unconfounded these two factors by varying load with items drawn from the same stimulus category ("high demands on context binding") versus items drawn from different stimulus categories ("low demands on context binding"). The results provide evidence for the dissociability, and the neural bases, of these two theorized factors, and they specify that the functions of intraparietal sulcus may relate more strongly to the control of representations than to the general allocation of attention. Copyright © 2018 the authors 0270-6474/18/384357-10$15.00/0.

Capacity for cooperative binding of thyroid hormone (T3) receptor dimers defines wild type T3 response elements.

PubMed

Brent, G A; Williams, G R; Harney, J W; Forman, B M; Samuels, H H; Moore, D D; Larsen, P R

1992-04-01

Thyroid hormone response elements (T3REs) have been identified in a variety of promoters including those directing expression of rat GH (rGH), alpha-myosin heavy chain (rMHC), and malic enzyme (rME). A detailed biochemical and genetic analysis of the rGH element has shown that it consists of three hexamers related to the consensus [(A/G)GGT(C/A)A]. We have extended this analysis to the rMHC and rME elements. Binding of highly purified thyroid hormone receptor (T3R) to T3REs was determined using the gel shift assay, and thyroid hormone (T3) induction was measured in transient tranfections. We show that the wild type version of each of the three elements binds T3R dimers cooperatively. Mutational analysis of the rMHC and rME elements identified domains important for binding T3R dimers and allowed a direct determination of the relationship between T3R binding and function. In each element two hexamers are required for dimer binding, and mutations that interfere with dimer formation significantly reduce T3 induction. Similar to the rGH element, the rMHC T3RE contains three hexameric domains arranged as a direct repeat followed by an inverted copy, although the third domain is weaker than in rGH. All three are required for full function and T3R binding. The rME T3RE is a two-hexamer direct repeat T3RE, which also binds T3R monomer and dimer. Across a series of mutant elements, there was a strong correlation between dimer binding in vitro and function in vivo for rMHC (r = 0.99, P less than 0.01) and rME (r = 0.67, P less than 0.05) T3REs. Our results demonstrate a similar pattern of T3R dimer binding to a diverse array of hexameric sequences and arrangements in three wild type T3REs. Addition of nuclear protein enhanced T3R binding but did not alter the specificity of binding to wild type or mutant elements. Binding of purified T3R to T3REs was highly correlated with function, both with and without the addition of nuclear protein. T3R dimer formation is the common feature which defines the capacity of these elements to confer T3 induction.

Binding mechanism and electrochemical properties of M13 phage-sulfur composite.

PubMed

Dong, Dexian; Zhang, Yongguang; Sutaria, Sanjana; Konarov, Aishuak; Chen, Pu

2013-01-01

Self-assembly of nanostructured materials has been proven a powerful technique in material design and synthesis. By phage display screening, M13 phage was found to strongly bind sulfur particles. Fourier transform infrared and X-ray photoelectron spectroscopy measurements indicated that the strong sulfur-binding ability of M13 phage derives from newly generated S-O and C-S bonds. Using this phage assembled sulfur composite in a lithium battery, the first discharge capacity reached 1117 mAh g(-1), which is more than twice that of the sulfur only cathode. Besides, the negative polysulfide shuttle effect in a lithium-sulfur battery was significantly suppressed.

Binding Mechanism and Electrochemical Properties of M13 Phage-Sulfur Composite

PubMed Central

Dong, Dexian; Zhang, Yongguang; Sutaria, Sanjana; Konarov, Aishuak; Chen, Pu

2013-01-01

Self-assembly of nanostructured materials has been proven a powerful technique in material design and synthesis. By phage display screening, M13 phage was found to strongly bind sulfur particles. Fourier transform infrared and X-ray photoelectron spectroscopy measurements indicated that the strong sulfur-binding ability of M13 phage derives from newly generated S-O and C-S bonds. Using this phage assembled sulfur composite in a lithium battery, the first discharge capacity reached 1117 mAh g-1, which is more than twice that of the sulfur only cathode. Besides, the negative polysulfide shuttle effect in a lithium-sulfur battery was significantly suppressed. PMID:24324560

Heterogeneity of binding of muscarinic receptor antagonists in rat brain homogenates

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, J.H.; el-Fakahany, E.E.

1985-06-01

The binding properties of (-)-(/sup 3/H)quinuclidinyl benzilate and (/sup 3/H) N-methylscopolamine to muscarinic acetylcholine receptors have been investigated in rat brain homogenates. The binding of both antagonists demonstrated high affinity and saturability. Analysis of the binding data resulted in linear Scatchard plots. However, (-)-(/sup 3/H)quinuclidinyl benzilate showed a significantly higher maximal binding capacity than that of (/sup 3/H)N-methylscopolamine. Displacement of both ligands with several muscarinic receptor antagonists resulted in competition curves in accordance with the law of mass-action for quinuclidinyl benzilate, atropine and scopolamine. A similar profile was found for the quaternary ammonium analogs of atropine and scopolamine when (/supmore » 3/H)N-methylscopolamine was used to label the receptors. However, when these hydrophilic antagonists were used to displace (-)-(/sup 3/H) quinuclidinyl benzilate binding, they showed interaction with high- and low-affinity binding sites. On the other hand, the nonclassical muscarinic receptor antagonist, pirenzepine, was able to displace both ligands from two binding sites. The present data are discussed in terms of the relationship of this anomalous heterogenity of binding of these hydrophilic muscarinic receptor antagonists and the proposed M1 and M2 receptor subtypes.« less

Binding of [51Cr]ethylenediaminetetraacetate to proteins of human plasma.

PubMed Central

Babiker, M M

1986-01-01

Binding of [51Cr]EDTA to human plasma proteins was investigated using chemical and chromatographic techniques of separation of the proteins and protein fractions. Total plasma proteins isolated with ethanol retained 12.95 +/- 0.46% of the initial plasma activity. Proteins separated by other precipitants retained about 16% of the initial radioactivity. Globulins exhibited the highest binding capacity for [51Cr]EDTA and retained about 11.7% of the initial plasma activity following chromatographic separation. This value represents about 70% of the radioactivity bound by the total proteins of the plasma. gamma-Globulins contributed most of the binding attributed to the globulins and retained about 8.7% of the initial [51Cr]EDTA activity. The repeatedly reported underestimation of the renal glomerular filtration rate when estimated as the clearance of [51Cr]EDTA could be adequately accounted for by the extent of binding of this marker to the plasma proteins. PMID:2427701

Increased lectin binding capacity of trophoblastic cells of late day 5 rat blastocysts.

PubMed Central

Stein, B A; Shaw, T J; Turner, V F; Murphy, C R

1994-01-01

The binding of lectins to the trophoblast of rat blastocysts has been studied using quantitative ultrastructural cytochemistry. Rat blastocysts from early, mid and late d 5 of gestation were stained using biotinylated lectins (Phytolacca americana [Phy am], fucose binding protein [FBP] and soybean agglutinin [SBA]) and a sensitive avidin-ferritin cytochemical method. Electron micrographs of ferritin particles along the membrane were processed to produce images for which grey scale levels could be established and the ferritin particles automatically counted. The ferritin:membrane ratio was then calculated. Increased binding with Phy am (which detects short chain oligosaccharides) was found after midday of d 5, i.e. after hatching. Binding of FBP and SBA did not alter during the period studied. The increased concentration of oligosaccharides on the blastocyst surface membrane after hatching may have important implications for blastocyst attachment to the endometrium. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:7649802

Non-B-DNA structures on the interferon-beta promoter?

PubMed

Robbe, K; Bonnefoy, E

1998-01-01

The high mobility group (HMG) I protein intervenes as an essential factor during the virus induced expression of the interferon-beta (IFN-beta) gene. It is a non-histone chromatine associated protein that has the dual capacity of binding to a non-B-DNA structure such as cruciform-DNA as well as to AT rich B-DNA sequences. In this work we compare the binding affinity of HMGI for a synthetic cruciform-DNA to its binding affinity for the HMGI-binding-site present in the positive regulatory domain II (PRDII) of the IFN-beta promoter. Using gel retardation experiments, we show that HMGI protein binds with at least ten times more affinity to the synthetic cruciform-DNA structure than to the PRDII B-DNA sequence. DNA hairpin sequences are present in both the human and the murine PRDII-DNAs. We discuss in this work the presence of, yet putative, non-B-DNA structures in the IFN-beta promoter.

LHRH-pituitary plasma membrane binding: the presence of specific binding sites in other tissues.

PubMed

Marshall, J C; Shakespear, R A; Odell, W D

1976-11-01

Two specific binding sites for LHRH are present on plasma membranes prepared from rat and bovine anterior pituitary glands. One site is of high affinity (K = 2X108 1/MOL) and the second is of lower affinity (8-5X105 1/mol) and much greater capacity. Studies on membrane fractions prepared from other tissues showed the presence of a single specific site for LHRH. The kinetics and specificity of this site were similar to those of the lower affinity pituitary receptor. These results indicate that only pituitary membranes possess the higher affinity binding site and suggest that the low affinity site is not of physiological importance in the regulation of gonadotrophin secretion. After dissociation from membranes of non-pituitary tissues 125I-LHRH rebound to pituitary membrane preparations. Thus receptor binding per se does not result in degradation of LHRH and the function of these peripheral receptors remains obscure.

Sulfated Glycopeptide Nanostructures for Multipotent Protein Activation

PubMed Central

Lee, Sungsoo S.; Fyrner, Timmy; Chen, Feng; Álvarez, Zaida; Sleep, Eduard; Chun, Danielle S.; Weiner, Joseph A.; Cook, Ralph W.; Freshman, Ryan D.; Schallmo, Michael S.; Katchko, Karina M.; Schneider, Andrew D.; Smith, Justin T.; Yun, Chawon; Singh, Gurmit; Hashmi, Sohaib Z.; McClendon, Mark T.; Yu, Zhilin; Stock, Stuart R.; Hsu, Wellington K.; Hsu, Erin L.; Stupp, Samuel I.

2017-01-01

Biological systems have evolved to utilize numerous proteins with capacity to bind polysaccharides for the purpose of optimizing their function. A well-known subset of these proteins with binding domains for the highly diverse sulfated polysaccharides are important growth factors involved in biological development and tissue repair. We report here on supramolecular sulfated glycopeptide nanostructures, which display a trisulfated monosaccharide on their surfaces and bind five critical proteins with very different polysaccharide binding domains. Binding does not disrupt the filamentous shape of the nanostructures or their internal β-sheet backbone, but must involve accessible adaptive configurations to interact with such different proteins. The glycopeptide nanostructures amplified signaling of bone morphogenetic protein 2 significantly more than the natural sulfated polysaccharide heparin, and promoted regeneration of bone in the spine with a protein dose that is 100-fold lower than expected. These super-bioactive nanostructures may enable many therapies in the horizon involving proteins. PMID:28650443

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tiberi, M.; Magnan, J.

The binding characteristics of selective and nonselective opioids have been studied in whole guinea pig spinal cord, using a computer fitting method to analyze the data obtained from saturation and competition studies. The delineation of specific binding sites labeled by the mu-selective opioid (3H)D-Ala2,MePhe4,Gly-ol5-enkephalin (Kd = 2.58 nM, R = 4.52 pmol/g of tissue) and by the delta-selective opioid (3H)D-Pen2, D-Pen5-enkephalin (Kd = 2.02 nM, R = 1.47 pmol/g of tissue) suggests the presence of mu and delta-receptors in the spinal cord tissue. The presence of kappa receptors was probed by the kappa-selective opioid (3H)U69593 (Kd = 3.31 nM, Rmore » = 2.00 pmol/g of tissue). The pharmacological characterization of the sites labeled by (3H)U69593 confirms the assumption that this ligand discriminates kappa receptors in guinea pig spinal cord. The benzomorphan (3H)ethylketazocine labels a population of receptors with one homogeneous affinity state (Kd = 0.65 nM, R = 7.39 pmol/g of tissue). The total binding capacity of this ligand was not different from the sum of the binding capacities of mu, delta-, and kappa-selective ligands. Under mu- and delta-suppressed conditions, (3H)ethylketazocine still binds to receptors with one homogeneous affinity state (Kd = 0.45 nM, R = 1.69 pmol/g of tissue). Competition studies performed against the binding of (3H)ethylketazocine under these experimental conditions reveal that the pharmacological profile of the radiolabeled receptors is similar to the profile of the kappa receptors labeled with (3H)U69593. Saturation studies using the nonselective opioid (3H)bremazocine demonstrate that this ligand binds to spinal cord membranes with heterogeneous affinities (Kd1 = 0.28 nM, R1 = 7.91 pmol/g of tissue; Kd2 = 3.24 nM, R2 = 11.2 pmol/g of tissue).« less

Intrasteric inhibition mediates the interaction of the I/LWEQ module proteins Talin1, Talin2, Hip1, and Hip12 with actin.

PubMed

Senetar, Melissa A; Foster, Stanley J; McCann, Richard O

2004-12-14

The I/LWEQ module superfamily is a class of actin-binding proteins that contains a conserved C-terminal actin-binding element known as the I/LWEQ module. I/LWEQ module proteins include the metazoan talins, the cellular slime mold talin homologues TalA and TalB, fungal Sla2p, and the metazoan Sla2 homologues Hip1 and Hip12 (Hip1R). These proteins possess a similar modular organization that includes an I/LWEQ module at their C-termini and either a FERM domain or an ENTH domain at their N-termini. As a result of this modular organization, I/LWEQ module proteins may serve as linkers between cellular compartments, such as the plasma membrane and the endocytic machinery, and the actin cytoskeleton. Previous studies have shown that I/LWEQ module proteins bind to F-actin. In this report, we have determined the affinity of the I/LWEQ module proteins Talin1, Talin2, huntingtin interacting protein-1 (Hip1), and the Hip1-related protein (Hip1R/Hip12) for F-actin and identified a conserved structural element that interferes with the actin binding capacity of these proteins. Our data support the hypothesis that the actin-binding determinants in native talin and other I/LWEQ module proteins are cryptic and indicate that the actin binding capacities of Talin1, Talin2, Hip1, and Hip12 are regulated by intrasteric occlusion of primary actin-binding determinants within the I/LWEQ module. We have also found that the I/LWEQ module contains a dimerization motif and stabilizes actin filaments against depolymerization. This activity may contribute to the function of talin in cell adhesion and the roles of Hip1, Hip12 (Hip1R), and Sla2p in endocytosis.

Bratislava Symposium on Saccharides (7th) Programme and Abstracts

DTIC Science & Technology

1994-09-01

that of cellulose (1). Althoug the binding capacity of cellulose microfibrils is dependent on the sace of the binding un of the kmfbrul& xyloglucans...are not only party embedded in but are also parly free between microfibrils . suggesting cross-link to cellulose microfibuils (2). Xyloglucan...desediftcoli Y.-C.-M a 12. KoIlkovd B., Hricovfrni M., Sirmoutti R.: 43C NMR study of solid-stal, reaction of cellulose with lIgnin monomers 13. Joniak D

Application of a 1.48-microm diode laser for bisecting oocytes into two identical hemizonae for the hemizona assay.

PubMed

Edenfeld, J; Schöpper, B; Sturm, R; Diedrich, K; Al-Hasani, S

2002-04-01

Laser systems are very promising new technical tools in assisted reproduction. It was investigated if laser radiation can replace the mechanical cutting procedure via micromanipulator in the hemizona assay (HZA), a commonly used bioassay to determine the sperm-zona pellucida binding capacity. An oocyte was bisected precisely into two identical hemizonae with approximately 20 laser pulses (pulse length 30 msec) using a 1.48-microm diode laser. Compared with the conventional method using microscalpels for zona bisection, laser treated hemizonae showed equivalent sperm-binding and within the two groups there was no detectable difference between matching hemizonae in their capacity for tight sperm-binding. To evaluate whether laser radiation affects the outcome of the HZA when effects of certain substances are investigated, the spermatozoa were preincubated with human follicular fluid (hFF), which inhibits the binding of spermatozoa to zona pellucida in vitro. Supplementation with follicular fluid exerted an inhibitory effect in both groups. The hemizona index (HZI) showed no statistical differences between the two methods. Therefore, the 1.48-microm diode laser is a suitable new instrument for generating equally sized hemizonae. There is no use for holding pipettes and microscalpels, on the contrary, for performing the HZA the laser is a precise, very quick and easy to use new working tool.

A Specific Peptide with Calcium-Binding Capacity from Defatted Schizochytrium sp. Protein Hydrolysates and the Molecular Properties.

PubMed

Cai, Xixi; Yang, Qian; Lin, Jiaping; Fu, Nanyan; Wang, Shaoyun

2017-03-29

Marine microorganisms have been proposed as a new kind of protein source. Efforts are needed in order to transform the protein-rich biological wastes left after lipid extraction into value-added bio-products. Thus, the utilization of protein recovered from defatted Schizochytrium sp. by-products presents an opportunity. A specific peptide Tyr-Leu (YL) with calcium-binding capacity was purified from defatted Schizochytrium sp. protein hydrolysates through gel filtration chromatography and RP-HPLC. The calcium-binding activity of YL reached 126.34 ± 3.40 μg/mg. The calcium-binding mechanism was investigated through ultraviolet, fluorescence and infrared spectroscopy. The results showed that calcium ions could form dative bonds with carboxyl oxygen atoms and amino nitrogen atoms as well as the nitrogen and oxygen atoms of amide bonds. YL-Ca exhibited excellent thermal stability and solubility, which was beneficial for its absorption and transport in the basic intestinal tract of the human body. Moreover, the cellular uptake of calcium in Caco-2 cells showed that YL-Ca could enhance calcium uptake efficiency and protect calcium ions against precipitation caused by dietary inhibitors such as tannic acid, oxalate, phytate and metal ions. The findings indicate that the by-product of Schizochytrium sp. is a promising source for making peptide-calcium bio-products as algae-based functional supplements for human beings.

Characteristic of theophylline imprinted monolithic column and its application for determination of xanthine derivatives caffeine and theophylline in green tea.

PubMed

Sun, Han-wen; Qiao, Feng-xia; Liu, Guang-yu

2006-11-17

Theophylline imprinted monolithic columns were designed and prepared for rapid separation of a homologous series of xanthine derivatives, caffeine, and theophylline by an in situ thermal-initiated copolymerization technique. Caffeine and theophylline were fully separated both under isocratic and gradient elutions on this kind of monolithic molecularly imprinted polymers (MIP) column. The broad peak showed in isocratic elution could be improved in gradient elution. Some chromatographic conditions such as mobile phase composition, flow rate, and the temperature on the retention times were investigated. Hydrogen bonding interaction and hydrophobic interaction played an important role in the retention and separation. The binding capacity was evaluated by static adsorption and Scatchard analysis, which showed that the dissociation constant (KD) and the maximum binding capacity (Qmax) were 1.50 mol/L, and 236 micromol/g for high affinity binding site, and 7.97 mol/L and 785 micromol/g for lower affinity binding site, respectively. Thermodynamic data (DeltaDeltaH and DeltaDeltaS) obtained by Van't Hoff plots revealed an enthalpy-controlled separation. The morphological characteristics of monolithic MIP were investigated by scanning electron microscope, which showed that both mesopores and macropores were formed in the monolith. The present monolithic MIP column was successfully applied for the quantitative determination of caffeine and theophylline in different kinds of green tea.

The juxtamembrane domain of the E-cadherin cytoplasmic tail contributes to its interaction with Myosin VI

PubMed Central

Mangold, Sabine; Norwood, Suzanne J.; Yap, Alpha S.; Collins, Brett M.

2012-01-01

We recently identified the atypical myosin, Myosin VI, as a component of epithelial cell-cell junctions that interacts with E-cadherin. Recombinant proteins bearing the cargo-binding domain of Myosin VI (Myo VI-CBD) or the cytoplasmic tail of E-cadherin can interact directly with one another. In this report we further investigate the molecular requirements of the interaction between Myo VI-CBD and E-cadherin combining truncation mutation analysis with in vitro binding assays. We report that a short (28 amino acid) juxtamembrane region of the cadherin cytoplasmic tail is sufficient to bind Myo VI-CBD. However, central regions of the cadherin tail adjacent to the juxtamembrane sequence also display binding activity for Myo VI-CBD. It is therefore possible that the cadherin tail bears two binding sites for Myosin VI, or an extended binding site that includes the juxtamembrane region. Nevertheless, our biochemical data highlight the capacity for the juxtamembrane region to interact with functionally-significant cytoplasmic proteins. PMID:23007415

Isolation and preliminary characterization of a Cd-binding protein from Tenebrio molitor (Coleoptera).

PubMed

Pedersen, S A; Kristiansen, E; Andersen, R A; Zachariassen, K E

2007-04-01

The effect of cadmium (Cd) exposure on Cd-binding ligands was investigated for the first time in a beetle (Coleoptera), using the mealworm Tenebrio molitor (L) as a model species. Exposure to Cd resulted in an approximate doubling of the Cd-binding capacity of the protein extracts from whole animals. Analysis showed that the increase was mainly explained by the induction of a Cd-binding protein of 7134.5 Da, with non-metallothionein characteristics. Amino acid analysis and de novo sequencing revealed that the protein has an unusually high content of the acidic amino acids aspartic and glutamic acid that may explain how this protein can bind Cd even without cysteine residues. Similarities in the amino acid composition suggest it to belong to a group of little studied proteins often referred to as "Cd-binding proteins without high cysteine content". This is the first report on isolation and peptide sequence determination of such a protein from a coleopteran.

Understanding how the complex molecular architecture of mannan-degrading hydrolases contributes to plant cell wall degradation.

PubMed

Zhang, Xiaoyang; Rogowski, Artur; Zhao, Lei; Hahn, Michael G; Avci, Utku; Knox, J Paul; Gilbert, Harry J

2014-01-24

Microbial degradation of plant cell walls is a central component of the carbon cycle and is of increasing importance in environmentally significant industries. Plant cell wall-degrading enzymes have a complex molecular architecture consisting of catalytic modules and, frequently, multiple non-catalytic carbohydrate binding modules (CBMs). It is currently unclear whether the specificities of the CBMs or the topology of the catalytic modules are the primary drivers for the specificity of these enzymes against plant cell walls. Here, we have evaluated the relationship between CBM specificity and their capacity to enhance the activity of GH5 and GH26 mannanases and CE2 esterases against intact plant cell walls. The data show that cellulose and mannan binding CBMs have the greatest impact on the removal of mannan from tobacco and Physcomitrella cell walls, respectively. Although the action of the GH5 mannanase was independent of the context of mannan in tobacco cell walls, a significant proportion of the polysaccharide was inaccessible to the GH26 enzyme. The recalcitrant mannan, however, was fully accessible to the GH26 mannanase appended to a cellulose binding CBM. Although CE2 esterases display similar specificities against acetylated substrates in vitro, only CjCE2C was active against acetylated mannan in Physcomitrella. Appending a mannan binding CBM27 to CjCE2C potentiated its activity against Physcomitrella walls, whereas a xylan binding CBM reduced the capacity of esterases to deacetylate xylan in tobacco walls. This work provides insight into the biological significance for the complex array of hydrolytic enzymes expressed by plant cell wall-degrading microorganisms.

Importance of adhesins in the recurrence of pharyngeal infections caused by Streptococcus pyogenes.

PubMed

Wozniak, Aniela; Scioscia, Natalia; Geoffroy, Enrique; Ponce, Iván; García, Patricia

2017-04-01

Pharyngo-amygdalitis is the most common infection caused by Streptococcus pyogenes (S. pyogenes). Reinfection with strains of different M types commonly occurs. However, a second infection with a strain of the same M type can still occur and is referred to as recurrence. We aimed to assess whether recurrence of S. pyogenes could be associated to erythromycin resistance, biofilm formation or surface adhesins like fibronectin-binding proteins and pilus proteins, both located in the fibronectin-binding, collagen-binding, T-antigen (FCT) region. We analyed clinical isolates of S. pyogenes obtained from children with multiple positive cultures of throat swabs. We analysed potential associations between M types, clonal patterns, biofilm production and FCT types with their capacity of producing a recurrent infection. We genetically defined recurrence as an infection with the same M type (same strain) and reinfection as an infection with a different M type. No differences were observed between recurrent and reinfection isolates in relation to erythromycin resistance, presence and number of domains of prtF1 gene, and biofilm formation capacity; the only significant difference was the higher frequency of FCT-4 type among recurrent isolates. However, when all the factors that could contribute to recurrence (erythromycin resistance, biofilm production, presence of prtF1 gene and FCT-4 type) were analysed together, we observed that recurrent isolates have a higher number of factors than reinfection isolates. Recurrence seems not to be associated with biofilm formation. However, pili and fibronectin-binding proteins could be associated with recurrence because FCT-4 isolates which harbour two fibronectin-binding proteins are more frequent among recurrent isolates.

Easy preparation of dietary fiber with the high water-holding capacity from food sources.

PubMed

Yamazaki, Eiji; Murakami, Kazumi; Kurita, Osamu

2005-03-01

Dietary fibers were prepared as alkali- and acid-insoluble fractions with chemical phosphorylation from Tossa jute (Corchorus olitorius), defatted soybean (Glycine max), and Shiitake (Lentinula edodes). The dietary fiber fractions treated with alkaline solution containing sodium metaphosphate had the lower protein content and higher total dietary fiber content than those of the preparations without phosphorylation. Alkaline extraction followed by phosphorylation led to a 1.5-fold increase in the water holding capacity of dietary fiber compared with no phosphorylation, whereas the binding capacity to bile acids of dietary fiber was almost the same. The alkali- and acid-insoluble extraction with phosphorylation provided an efficient preparation of water-insoluble dietary fiber with high-water holding capacity from various food sources.

Behind the scenes of vitamin D binding protein: more than vitamin D binding.

PubMed

Delanghe, Joris R; Speeckaert, Reinhart; Speeckaert, Marijn M

2015-10-01

Although being discovered in 1959, the number of published papers in recent years reveals that vitamin D binding protein (DBP), a member of the albuminoid superfamily, is a hot research topic. Besides the three major phenotypes (DBP1F, DBP1S and DBP2), more than 120 unique variants have been described of this polymorphic protein. The presence of DBP has been demonstrated in different body fluids (serum, urine, breast milk, ascitic fluid, cerebrospinal fluid, saliva and seminal fluid) and organs (brain, heart, lungs, kidneys, placenta, spleen, testes and uterus). Although the major function is binding, solubilization and transport of vitamin D and its metabolites, the name of this glycoprotein hides numerous other important biological functions. In this review, we will focus on the analytical aspects of the determination of DBP and discuss in detail the multifunctional capacity [actin scavenging, binding of fatty acids, chemotaxis, binding of endotoxins, influence on T cell response and influence of vitamin D binding protein-macrophage activating factor (DBP-MAF) on bone metabolism and cancer] of this abundant plasma protein. Copyright © 2015 Elsevier Ltd. All rights reserved.

Peptide-functionalized iron oxide magnetic nanoparticle for gold mining

NASA Astrophysics Data System (ADS)

Shen, Wei-Zheng; Cetinel, Sibel; Sharma, Kumakshi; Borujeny, Elham Rafie; Montemagno, Carlo

2017-02-01

Here, we present our work on preparing a novel nanomaterial composed of inorganic binding peptides and magnetic nanoparticles for inorganic mining. Two previously selected and well-characterized gold-binding peptides from cell surface display, AuBP1 and AuBP2, were exploited. This nanomaterial (AuBP-MNP) was designed to fulfill the following two significant functions: the surface conjugated gold-binding peptide will recognize and selectively bind to gold, while the magnetic nano-sized core will respond and migrate according to the applied external magnetic field. This will allow the smart nanomaterial to mine an individual material (gold) from a pool of mixture, without excessive solvent extraction, filtration, and concentration steps. The working efficiency of AuBP-MNP was determined by showing a dramatic reduction of gold nanoparticle colloid concentration, monitored by spectroscopy. The binding kinetics of AuBP-MNP onto the gold surface was determined using surface plasmon resonance (SPR) spectroscopy, which exhibits around 100 times higher binding kinetics than peptides alone. The binding capacity of AuBP-MNP was demonstrated by a bench-top mining test with gold microparticles.

Identification of gold sensing peptide by integrative proteomics and a bacterial two-component system

NASA Astrophysics Data System (ADS)

Ng, I.-Son; Yu, You-Jin; Yi, Ying-Chen; Tan, Shih-I.; Huang, Bo-Chuan; Han, Yin-Lung

2017-12-01

The proteomics strategy was utilized to analyze and identify the gold adsorption proteins from Tepidimonas fonticaldi AT-A2, due to its outstanding performance in gold-binding and recovery. The results showed that three small proteins, including histidine biosynthesis protein (HisIE), iron donor protein (CyaY) and hypothetical protein_65aa, have a higher ability to adsorb gold ions because of the negatively charged domains or metal binding sites. On the other hand, the Salmonella PmrA/PmrB two-component system first replaces the iron (III)-binding motif using the peptide sequence from hypothetical protein_65aa, and this is then used to reveal the sensing and responsiveness to gold metal ions, which is totally different from the performance of traditional gold binding peptide (GBP) on the crystals on the surface of gold (111). We have successfully demonstrated an integrative proteomics and bacterial two-component system to explore the novel gold binding peptide. Finally, the heterologous over-expression of gold binding peptide by E. coli and the equilibrium of binding capacity for Au(III) have been conducted.

Protein F, a fibronectin-binding protein, is an adhesin of the group A streptococcus Streptococcus pyogenes.

PubMed

Hanski, E; Caparon, M

1992-07-01

Binding to fibronectin has been suggested to play an important role in adherence of the group A streptococcus Streptococcus pyrogenes to host epithelial cells; however, the identity of the streptococcal fibronectin receptor has been elusive. Here we demonstrate that the fibronectin-binding property of S. pyogenes is mediated by protein F, a bacterial surface protein that binds fibronectin at high affinity. The gene encoding protein F (prtF) produced a functional fibronectin-binding protein in Escherichia coli. Insertional mutagenesis of the cloned gene generated a mutation that resulted in the loss of fibronectin-binding activity. When this mutation was introduced into the S. pyrogenes chromosome by homologous recombination with the wild-type allele, the resulting strains no longer produced protein F and lost their ability to bind fibronectin. The mutation could be complemented by prtF introduced on a plasmid. Mutants lacking protein F had a much lower capacity to adhere to respiratory epithelial cells. These results demonstrate that protein F is an important adhesin of S. pyogenes.

Functionality of the STNV translational enhancer domain correlates with affinity for two wheat germ factors

PubMed Central

Lipzig, Rosalinde van; Montagu, Marc Van; Cornelissen, Marc; Meulewaeter, Frank

2001-01-01

The satellite tobacco necrosis virus RNA is uncapped and requires a 3′ translational enhancer domain (TED) for translation. Both in the wheat germ extract and in tobacco, TED stimulates in cis translation of heterologous, uncapped RNAs. In this study we investigated to what extent translation stimulation by TED depends on binding to wheat germ factors. We show that in vitro TED binds at least seven wheat germ proteins. Translation and crosslinking assays, to which TED or TED derivatives with reduced functionality were included as competitor, showed that TED function correlates with binding to a 28 kDa protein (p28). One particular condition of competition revealed that p28 binding is not obligatory for TED function. Under this condition, a 30 kDa protein (p30) binds to TED. Importantly, affinity of p30 correlates with functionality of TED. These results strongly suggest that TED has the capacity to stimulate translation by recruiting the translational machinery either via binding to p28 or via binding to p30. PMID:11222757

Correlation study between sperm concentration, hyaluronic acid-binding capacity and sperm aneuploidy in Hungarian patients.

PubMed

Mokánszki, Attila; Molnár, Zsuzsanna; Ujfalusi, Anikó; Balogh, Erzsébet; Bazsáné, Zsuzsa Kassai; Varga, Attila; Jakab, Attila; Oláh, Éva

2012-12-01

Infertile men with low sperm concentration and/or less motile spermatozoa have an increased risk of producing aneuploid spermatozoa. Selecting spermatozoa by hyaluronic acid (HA) binding may reduce genetic risks such as chromosomal rearrangements and numerical aberrations. Fluorescence in-situ hybridization (FISH) has been used to evaluate the presence of aneuploidies. This study examined spermatozoa of 10 oligozoospermic, 9 asthenozoospermic, 9 oligoasthenozoospermic and 17 normozoospermic men by HA binding and FISH. Mean percentage of HA-bound spermatozoa in the normozoospermic group was 81%, which was significantly higher than in the oligozoospermic (P<0.001), asthenozoospermic (P<0.001) and oligoasthenozoospermic (P<0.001) groups. Disomy of sex chromosomes (P=0.014) and chromosome 17 (P=0.0019), diploidy (P=0.03) and estimated numerical chromosome aberrations (P=0.004) were significantly higher in the oligoasthenozoospermic group compared with the other groups. There were statistically significant relationships (P<0.001) between sperm concentration and HA binding (r=0.658), between sperm concentration and estimated numerical chromosome aberrations (r=-0.668) and between HA binding and estimated numerical chromosome aberrations (r=-0.682). HA binding and aneuploidy studies of spermatozoa in individual cases allow prediction of reproductive prognosis and provision of appropriate genetic counselling. Infertile men with normal karyotypes and low sperm concentrations and/or less motile spermatozoa have significantly increased risks of producing aneuploid (diminished mature) spermatozoa. Selecting spermatozoa by hyaluronic acid (HA) binding, based on a binding between sperm receptors for zona pellucida and HA, may reduce the potential genetic risks such as chromosomal rearrangements and numerical aberrations. In the present study we examined sperm samples of 45 men with different sperm parameters by HA-binding assay and fluorescence in-situ hybridization (FISH). Mean percentage of HA-bound spermatozoa in the normozoospermic group was significantly higher than the oligozoospermic, the asthenozoospermic and the oligoasthenozoospermic groups. Using FISH, disomy of sex chromosomes and chromosome 17, diploidy and estimated numerical chromosome aberration frequencies were significantly higher in the oligoasthenozoospermic group compared with the three other groups. A significant positive correlation was found between the sperm concentration and the HA-binding capacity, and significant negative correlations between the sperm concentration and the estimated numerical chromosomes aberrations as well as between the HA-binding ability and the estimated numerical chromosome aberrations were identified. We conclude that HA-binding assay and sperm aneuploidy study using FISH may help to predict the reproductive ability of selected infertile male patients and to provide appropriate genetic counselling. Crown Copyright © 2012. Published by Elsevier Ltd. All rights reserved.

Thermodynamics of Aryl-Dihydroxyphenyl-Thiadiazole Binding to Human Hsp90

PubMed Central

Kazlauskas, Egidijus; Petrikaitė, Vilma; Michailovienė, Vilma; Revuckienė, Jurgita; Matulienė, Jurgita; Grinius, Leonas; Matulis, Daumantas

2012-01-01

The design of specific inhibitors against the Hsp90 chaperone and other enzyme relies on the detailed and correct understanding of both the thermodynamics of inhibitor binding and the structural features of the protein-inhibitor complex. Here we present a detailed thermodynamic study of binding of aryl-dihydroxyphenyl-thiadiazole inhibitor series to recombinant human Hsp90 alpha isozyme. The inhibitors are highly potent, with the intrinsic Kd approximately equal to 1 nM as determined by isothermal titration calorimetry (ITC) and thermal shift assay (TSA). Dissection of protonation contributions yielded the intrinsic thermodynamic parameters of binding, such as enthalpy, entropy, Gibbs free energy, and the heat capacity. The differences in binding thermodynamic parameters between the series of inhibitors revealed contributions of the functional groups, thus providing insight into molecular reasons for improved or diminished binding efficiency. The inhibitor binding to Hsp90 alpha primarily depended on a large favorable enthalpic contribution combined with the smaller favorable entropic contribution, thus suggesting that their binding was both enthalpically and entropically optimized. The enthalpy-entropy compensation phenomenon was highly evident when comparing the inhibitor binding enthalpies and entropies. This study illustrates how detailed thermodynamic analysis helps to understand energetic reasons for the binding efficiency and develop more potent inhibitors that could be applied for therapeutic use as Hsp90 inhibitors. PMID:22655030

Adrenocortical nuclear progesterone-binding protein: Identification by photoaffinity labeling and evidence for deoxyribonucleic acid binding and stimulation by adrenocorticotropin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Demura, T.; Driscoll, W.J.; Lee, Y.C.

1991-01-01

Nuclei of the guinea pig adrenal cortex contain a protein that specifically binds progesterone and that, biochemically, is clearly distinct from the classical progesterone receptor. The adrenocortical nuclear progesterone-binding protein has now been purified more than 2000-fold by steroid-affinity chromatography with a 75% yield. The purified protein preparation demonstrated three major bands on sodium dodecyl sulfate-polyacrylamide gel of 79K, 74K, and 50K. To determine which of the three might represent the progesterone-binding protein, steroid photoaffinity labeling was performed which resulted in the specific and exclusive labeling of a 50K band. Thus, the adrenocortical nuclear progesterone-binding protein appears to be distinctmore » from the classical progesterone receptor not only biochemically, but also on the basis of molecular size. To test whether the adrenocortical nuclear progesterone-binding protein can be hormonally stimulated, guinea pigs were treated with ACTH. The chronic administration of ACTH caused a 4- to 6-fold increase in the specific progesterone binding capacity without a change in the binding affinity. There appeared to be no significant difference in nuclear progesterone binding between the zona fasciculata and zona reticularis. This finding suggests a mediating role for the progesterone-binding protein in ACTH action. In addition, the nuclear progesterone-binding protein bound to nonspecific DNA sequences, further suggesting a possible transcriptional regulatory role.« less

Protein adsorption to poly(ethylenimine)-modified Sepharose FF: I. a critical ionic capacity for drastically enhanced capacity and uptake kinetics.

PubMed

Yu, Lin-Ling; Tao, Shi-Peng; Dong, Xiao-Yan; Sun, Yan

2013-08-30

To explore the details of protein uptake to polymer-grafted ion exchangers, Sepharose FF was modified with poly(ethylenimine) (PEI) to prepare anion exchanger of 10 different ionic capacities (ICs, 100-1220mmol/L). Adsorption equilibria and kinetics of bovine serum albumin (BSA) were then studied. It is found that ionic capacity, i.e., the coupling density of PEI, had significant effect on both adsorption capacity (qm) and effective protein diffusivity (De). With increasing ionic capacity, the qm value increased rapidly at IC<260mmol/L and then increased slowly till reaching a plateau at IC=600mmol/L. In the IC range of 100-600mmol/L, however, the De values kept at a low level (De/D0<0.07); it first decreased from 0.05±0.01 at IC=100mmol/L to 0.01±0.01 at IC=260mmol/L and then increased to 0.06±0.01 at IC=600mmol/L. Thereafter, sharp increases of the qm and De values [36% (from 201 to 273mg/mL) and 670% (from 0.06±0.01 to 0.49±0.04), respectively] were observed in the narrow range of IC from 600 to 740mmol/L. Finally, at IC>740mmol/L, the qm value decreased significantly while the De value increased moderately with increasing the IC. The results indicate that PEI chains played an important role in protein adsorption and transport. In brief, there was a critical IC (cIC) or PEI chain density, above which protein adsorption and transport behaviors changed drastically. The cIC was identified to be about 600mmol/L. Estimation of PEI grafting-layer thickness suggests that PEI chains formed an extended three-dimensional grafting-layer at IC>cIC, which provided high flexibility as well as accessibility of the chains for protein binding. Therefore, at IC>cIC, the adjacent PEI chains became close and flexible enough, leading to facilitated transport of adsorbed protein molecules by the interactions of neighboring chains mediated by the bound molecules. It is regarded as "chain delivery" effect. At the same time, improved accessibility of binding sites led the significant increase of binding capacity. The decrease of qm value at IC>740mmol/L is considered due to the decrease of effective porosity. The research has thus provided new insight into protein adsorption and transport in polymer-grafted ion-exchange media. Copyright © 2013 Elsevier B.V. All rights reserved.

Peanut flour aggregation with polyphenolic extracts derived from peanut skin inhibits IgE binding capacity and attenuates RBL-2H3 cells degranulation via MAPK signaling pathway.

PubMed

Bansode, Rishipal R; Plundrich, Nathalie J; Randolph, Priscilla D; Lila, Mary Ann; Williams, Leonard L

2018-10-15

This study investigates the anti-allergic properties of peanut skin polyphenols (PSP)-enriched peanut (PN) protein aggregates. PSP was blended with PN flour at concentrations of 5, 10, 15, 30, and 40% (w/w). Rat basophil leukemia cells (RBL-2H3) were sensitized with either anti-DNP-IgE or PN-allergic plasma followed by co-exposure to unmodified PN flour (control) or PSP-PN protein aggregates and Ca 2+ ionophore, ionomycin. Immunoblotting and staining were performed to measure the IgE binding capacity of PSP-PN aggregates. Results showed that 30% PSP-PN aggregate significantly reduced β-hexosaminidase and histamine levels by 54.2% and 49.2%, respectively compared with control. Immunoblotting results revealed 40% PSP-PN aggregates significantly decreased IgE binding by 19%. The phosphorylation of p44/42 MAPK was significantly reduced while phosphorylation of p38 MAPK and SAPK/JNK increased upon PSP-PN protein aggregate exposure to the cells. Our results show that aggregation of PSP to PN proteins reduces allergic response by inhibiting Ca 2+ -induced MAPK-dependent cell degranulation. Copyright © 2018 Elsevier Ltd. All rights reserved.

Treatment of metal-contaminated wastewater: a comparison of low-cost biosorbents.

PubMed

Akunwa, N K; Muhammad, M N; Akunna, J C

2014-12-15

This study aimed to identify some optimum adsorption conditions for the use of low-cost adsorbent, seaweed (Ascophyllum nodosum), sawdust and reed plant (Phragmites australis) root, in the treatment of metal contaminated wastewater for the removal of cadmium, chromium and lead. The effect of pH on the absorption capacity of each of these biosorbents was found to be significant and dependent on the metal being removed. Post-adsorption FTIR analysis showed significant binding activities at the nitro NO groups site in all biosorbents, especially for lead. Competitive metal binding was found to have possibly affected the adsorption capacity for chromium by A. nodosum more than it affected sawdust and P. australis root. Adsorption is believed to take place mainly by ion exchange particularly at low pH values. P. australis root exhibited the highest adsorption for chromium at pH 2, cadmium at pH 10 and lead at pH 7. A. nodosum seaweed species demonstrated the highest adsorption capacity of the three biosorbents used in the study, for cadmium at pH 7 and for lead at pH 2. Sawdust proved to be an efficient biosorbent for lead removal only at pH 7 and 10. No significant effect of temperature on adsorption capacity was observed, particularly for cadmium and lead removal. Copyright © 2014 Elsevier Ltd. All rights reserved.

New functionalized IRMOF-10 with strong affinity for methanol: A simulation study

NASA Astrophysics Data System (ADS)

Liu, Zewei; Zhang, Kai; Wu, Ying; Xi, Hongxia

2018-05-01

Grand Canonical Monte Carlo (GCMC) method simulation combined with density functional theory (DFT) calculation were used to investigate the methanol adsorption in IRMOF-10, with nitrogen and metal-doping functionalizations in order to understand the underlying performance of MOFs in methanol adsorption. New doped IRMOF-10s (M-2N-IRMOF-10, M = Be, Mg, Ca, Sr, Ba) were theoretically constructed by binding nitrogen atoms of organic linkers in N-doping IRMOF-10 (2N-IRMOF-10) with various metal atoms. 2N-IRMOF-10 shows only a little higher methanol capacity in the measured pressure range. However, M-2N-IRMOF-10s (especially Be-2N-IRMOF-10) demonstrate much higher methanol capacity due to the stronger interaction between the induced Be atoms and methanol molecules. Furthermore, the obtained results can be attributed to the new adsorption sites created by metal-doping, as revealed by the more exothermic binding energies (BEs) on Be-sites (-160.8 kJ/mol) than Zn-sites (-19.4 kJ/mol). According to the simulation results, it can be concluded that functionalized IRMOF-10 are capable of enhancing the adsorption capacity of methanol at pressure from 0 to 12 kPa at 298 K. This study provides a new functionalized method to effectively enhance methanol adsorption capacity of MOFs, which might extend the application of MOFs on methanol adsorption in the near future.

Effects of dietary fibers with high water-binding capacity and swelling capacity on gastrointestinal functions, food intake and body weight in male rats.

PubMed

Tan, Chengquan; Wei, Hongkui; Zhao, Xichen; Xu, Chuanhui; Peng, Jian

2017-01-01

Objective : The aim of this study was to investigate the effects of supplementation of dietary soluble fibers with high water-binding capacity (WBC) and swelling capacity (SC) on gastrointestinal tract mass, physicochemical properties of digesta, gastrointestinal mean retention time (MRT), body weight, and food intake in male rats. Methods : Thirty-two male Sprague-Dawley rats were randomized to four equal groups and fed the control diet or diet containing 2% konjac flour (KF), pregelatinized waxy maize starch plus guar gum (PWMS+GG), andPWMS plus xanthan gum (PWMS+XG) for three weeks. Results : WBC and SC of diets followed the order of PWMS+GG > KF > PWMS + XG > control. PWMS+GG and KF groups had a lower average daily food intake than the control group, but all the groups showed no difference in final body weightand the weight gain rate. The high WBC and SC of the PWMS+GG and KF groupsled to an increase of WBC and SC in the stomach digesta, and a gain of the cecal digesta weight, due to increased cecal moisture content. Conclusion : The inclusion of the novel fiber, PWMS+GG, in the diet of male rats appears to facilitate the modulation of WBC and SC of stomach digesta and the reduction of food intake.

Effects of Iron Supplementation and Activity on Serum Iron Depletion and Hemoglobin Levels in Female Athletes

ERIC Educational Resources Information Center

Cooter, G. Rankin; Mowbray, Kathy W.

1978-01-01

Research revealed that a four-month basketball training program did not significantly alter serum iron, total iron binding capacity, hemoglobin, and percent saturation levels in female basketball athletes. (JD)

Mercaptoheterocyclic ligands grafted on a poly(ethylene vinyl alcohol) membrane for the purification of immunoglobulin G in a salt independent thiophilic chromatography.

PubMed

Coffinier, Yannick; Vijayalakshmi, Mookambeswaran A

2004-08-25

In this study, we attempted a limited combinatorial approach for designing affinity ligands based on mercaptoheterocyclic components. The template, divinyl sulfone structure (DVS), which was grafted on poly(ethylene vinyl alcohol) (PEVA) hollow fiber membrane, has served for the tethering of different heterocyclic compounds as pyridine, imidazole, purine and pyrimidine rings. Their ability to adsorb specifically IgG in a salt independent manner out of pure IgG solution, mixture of IgG/albumin and human plasma was demonstrated. Mercapto methyl imidazole (MMI) has shown the best adsorption of IgG in terms of binding capacity. No subclass discrimination was observed on all tested ligands except for mercapto methyl pyrimidine where the major IgG subclass adsorbed was IgG3. MMI gave an IgG binding capacity of 100 microg/cm2 of hollow fiber membrane surface area.

Ferrocene-Promoted Long-Cycle Lithium-Sulfur Batteries.

PubMed

Mi, Yingying; Liu, Wen; Yang, Ke R; Jiang, Jianbing; Fan, Qi; Weng, Zhe; Zhong, Yiren; Wu, Zishan; Brudvig, Gary W; Batista, Victor S; Zhou, Henghui; Wang, Hailiang

2016-11-14

Confining lithium polysulfide intermediates is one of the most effective ways to alleviate the capacity fade of sulfur-cathode materials in lithium-sulfur (Li-S) batteries. To develop long-cycle Li-S batteries, there is an urgent need for material structures with effective polysulfide binding capability and well-defined surface sites; thereby improving cycling stability and allowing study of molecular-level interactions. This challenge was addressed by introducing an organometallic molecular compound, ferrocene, as a new polysulfide-confining agent. With ferrocene molecules covalently anchored on graphene oxide, sulfur electrode materials with capacity decay as low as 0.014 % per cycle were realized, among the best of cycling stabilities reported to date. With combined spectroscopic studies and theoretical calculations, it was determined that effective polysulfide binding originates from favorable cation-π interactions between Li + of lithium polysulfides and the negatively charged cyclopentadienyl ligands of ferrocene. © 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Exopolysaccharide from Ganoderma applanatum as a Promising Bioactive Compound with Cytostatic and Antibacterial Properties

PubMed Central

Osińska-Jaroszuk, Monika; Błachowicz, Adriana; Wydrych, Jerzy; Polak, Jolanta; Jarosz-Wilkołazka, Anna; Kandefer-Szerszeń, Martyna

2014-01-01

A new exopolysaccharide preparation isolated from stationary cultures of the white rot fungus Ganoderma applanatum (GpEPS) was tested in terms of its bioactive properties including its cytotoxic and immunostimulatory effect. The results indicate that the tested GpEPS (at concentrations above 22.85 µg/mL and 228.5 µg/mL) may exhibit selective activity against tumor cells (cell lines SiHa) and stimulate production of TNF-α THP-1-derived macrophages at the level of 752.17 pg/mL. The GpEPS showed antibacterial properties against Staphyloccoccus aureus and a toxic effect against Vibrio fischeri cells (82.8% cell damage). High cholesterol-binding capacity and triglycerides-binding capacity (57.9% and 41.6% after 24 h of incubation with the tested substances, resp.) were also detected for the investigated samples of GpEPS. PMID:25114920

Fusion proteins of flagellin and the major birch pollen allergen Bet v 1 show enhanced immunogenicity, reduced allergenicity, and intrinsic adjuvanticity.

PubMed

Kitzmüller, Claudia; Kalser, Julia; Mutschlechner, Sonja; Hauser, Michael; Zlabinger, Gerhard J; Ferreira, Fatima; Bohle, Barbara

2018-01-01

Recombinant fusion proteins of flagellin and antigens have been demonstrated to induce strong innate and adaptive immune responses. Such fusion proteins can enhance the efficacy of allergen-specific immunotherapy. We sought to characterize different fusion proteins of flagellin and the major birch pollen allergen Bet v 1 for suitability as allergy vaccines. A truncated version of flagellin (NtCFlg) was genetically fused to the N- or C-terminus of Bet v 1. Toll-like receptor (TLR) 5 binding was assessed with HEK293 cells expressing TLR5. Upregulation of CD40, CD80, CD83, and CD86 on monocyte-derived dendritic cells from allergic patients was analyzed by using flow cytometry. The T cell-stimulatory capacity of the fusion proteins was assessed with naive and Bet v 1-specific T cells. IgE binding was tested in inhibition ELISAs and basophil activation tests. Mice were immunized with the fusion proteins in the absence and presence of aluminum hydroxide. Cellular and antibody responses were monitored. Murine antibodies were tested for blocking capacity in basophil activation tests. Both fusion proteins matured monocyte-derived dendritic cells through TLR5. Compared with Bet v 1, the fusion proteins showed stronger T cell-stimulatory and reduced IgE-binding capacity and induced murine Bet v 1-specific antibodies in the absence of aluminum hydroxide. However, only antibodies induced by means of immunization with NtCFlg fused to the C-terminus of Bet v 1 inhibited binding of patients' IgE antibodies to Bet v 1. Bet v 1-flagellin fusion proteins show enhanced immunogenicity, reduced allergenicity, and intrinsic adjuvanticity and thus represent promising vaccines for birch pollen allergen-specific immunotherapy. However, the sequential order of allergen and adjuvant within a fusion protein determines its immunologic characteristics. Copyright © 2017 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Sulfur cathodes with hydrogen reduced titanium dioxide inverse opal structure.

PubMed

Liang, Zheng; Zheng, Guangyuan; Li, Weiyang; Seh, Zhi Wei; Yao, Hongbin; Yan, Kai; Kong, Desheng; Cui, Yi

2014-05-27

Sulfur is a cathode material for lithium-ion batteries with a high specific capacity of 1675 mAh/g. The rapid capacity fading, however, presents a significant challenge for the practical application of sulfur cathodes. Two major approaches that have been developed to improve the sulfur cathode performance include (a) fabricating nanostructured conductive matrix to physically encapsulate sulfur and (b) engineering chemical modification to enhance binding with polysulfides and, thus, to reduce their dissolution. Here, we report a three-dimensional (3D) electrode structure to achieve both sulfur physical encapsulation and polysulfides binding simultaneously. The electrode is based on hydrogen reduced TiO2 with an inverse opal structure that is highly conductive and robust toward electrochemical cycling. The relatively enclosed 3D structure provides an ideal architecture for sulfur and polysulfides confinement. The openings at the top surface allow sulfur infusion into the inverse opal structure. In addition, chemical tuning of the TiO2 composition through hydrogen reduction was shown to enhance the specific capacity and cyclability of the cathode. With such TiO2 encapsulated sulfur structure, the sulfur cathode could deliver a high specific capacity of ∼1100 mAh/g in the beginning, with a reversible capacity of ∼890 mAh/g after 200 cycles of charge/discharge at a C/5 rate. The Coulombic efficiency was also maintained at around 99.5% during cycling. The results showed that inverse opal structure of hydrogen reduced TiO2 represents an effective strategy in improving lithium sulfur batteries performance.

Exploring the effect of oxygen-containing functional groups on the water-holding capacity of lignite.

PubMed

Liu, Jie; Jiang, Xiangang; Cao, Yu; Zhang, Chen; Zhao, Guangyao; Zhao, Maoshuang; Feng, Li

2018-05-07

Graphene oxide with different degrees of oxidation was prepared and selected as a model compound of lignite to study quantitatively, using both experiment and theoretical calculation methods, the effect on water-holding capacity of oxygen-containing functional groups. The experimental results showed that graphite can be oxidized, and forms epoxy groups most easily, followed by hydroxyl and carboxyl groups. The prepared graphene oxide forms a membrane-state as a single layer structure, with an irregular surface. The water-holding capacity of lignite increased with the content of oxygen-containing functional groups. The influence on the configuration of water molecule clusters and binding energy of water molecules of different oxygen-containing functional groups was calculated by density functional theory. The calculation results indicated that the configuration of water molecule clusters was totally changed by oxygen-containing functional groups. The order of binding energy produced by oxygen-containing functional groups and water molecules was as follows: carboxyl > edge phenol hydroxyl >epoxy group. Finally, it can be concluded that the potential to form more hydrogen bonds is the key factor influencing the interaction energy between model compounds and water molecules.

Inorganic nanoparticles for transfection of mammalian cells and removal of viruses from aqueous solutions.

PubMed

Link, Nils; Brunner, Tobias J; Dreesen, Imke A J; Stark, Wendelin J; Fussenegger, Martin

2007-12-01

Owing to their small size, synthetic nanoparticles show unprecedented biophysical and biochemical properties which may foster novel advances in life-science research. Using flame-spray synthesis technology we have produced non-coated aluminum-, calcium-, cerium-, and zirconium-derived inorganic metal oxide nanoparticles which not only exhibit high affinity for nucleic acids, but can sequester such compounds from aqueous solution. This non-covalent DNA-binding capacity was successfully used to transiently transfect a variety of mammalian cells including human, reaching transfection efficiencies which compared favorably with classic calcium phosphate precipitation (CaP) procedures and lipofection. In this straightforward protocol, transfection was enabled by simply mixing nanoparticles with DNA in solution prior to addition to the target cell population. Transiently transfected cells showed higher production levels of the human secreted glycoprotein SEAP compared to isogenic populations transfected with established technologies. Inorganic metal oxide nanoparticles also showed a high binding capacity to human-pathogenic viruses including adenovirus, adeno-associated virus and human immunodeficiency virus type 1 and were able to clear these pathogens from aqueous solutions. The DNA transfection and viral clearance capacities of inorganic metal oxide nanoparticles may provide cost-effective biopharmaceutical manufacturing and water treatment in developing countries.

A comparison of Helicobacter pylori and non-Helicobacter pylori Helicobacter spp. Binding to canine gastric mucosa with defined gastric glycophenotype.

PubMed

Amorim, Irina; Freitas, Daniela P; Magalhães, Ana; Faria, Fátima; Lopes, Célia; Faustino, Augusto M; Smet, Annemieke; Haesebrouck, Freddy; Reis, Celso A; Gärtner, Fátima

2014-08-01

The gastric mucosa of dogs is often colonized by non-Helicobacter pylori helicobacters (NHPH), while H. pylori is the predominant gastric Helicobacter species in humans. The colonization of the human gastric mucosa by H. pylori is highly dependent on the recognition of host glycan receptors. Our goal was to define the canine gastric mucosa glycophenotype and to evaluate the capacity of different gastric Helicobacter species to adhere to the canine gastric mucosa. The glycosylation profile in body and antral compartments of the canine gastric mucosa, with focus on the expression of histo-blood group antigens was evaluated. The in vitro binding capacity of FITC-labeled H. pylori and NHPH to the canine gastric mucosa was assessed in cases representative of the canine glycosylation pattern. The canine gastric mucosa lacks expression of type 1 Lewis antigens and presents a broad expression of type 2 structures and A antigen, both in the surface and glandular epithelium. Regarding the canine antral mucosa, H. heilmannii s.s. presented the highest adhesion score whereas in the body region the SabA-positive H. pylori strain was the strain that adhered more. The canine gastric mucosa showed a glycosylation profile different from the human gastric mucosa suggesting that alternative glycan receptors may be involved in Helicobacter spp. binding. Helicobacter pylori and NHPH strains differ in their ability to adhere to canine gastric mucosa. Among the NHPH, H. heilmannii s.s. presented the highest adhesion capacity in agreement with its reported colonization of the canine stomach. © 2014 John Wiley & Sons Ltd.

Fish protein hydrolysates: application in deep-fried food and food safety analysis.

PubMed

He, Shan; Franco, Christopher; Zhang, Wei

2015-01-01

Four different processes (enzymatic, microwave-intensified enzymatic, chemical, and microwave-intensified chemical) were used to produce fish protein hydrolysates (FPH) from Yellowtail Kingfish for food applications. In this study, the production yield and oil-binding capacity of FPH produced from different processes were evaluated. Microwave intensification significantly increased the production yields of enzymatic process from 42% to 63%. It also increased the production yields of chemical process from 87% to 98%. The chemical process and microwave-intensified chemical process produced the FPH with low oil-binding capacity (8.66 g oil/g FPH and 6.25 g oil/g FPH), whereas the microwave-intensified enzymatic process produced FPH with the highest oil-binding capacity (16.4 g oil/g FPH). The FPH from the 4 processes were applied in the formulation of deep-fried battered fish and deep-fried fish cakes. The fat uptake of deep-fried battered fish can be reduced significantly from about 7% to about 4.5% by replacing 1% (w/w) batter powder with FPH, and the fat uptake of deep-fried fish cakes can be significantly reduced from about 11% to about 1% by replacing 1% (w/w) fish mince with FPH. Food safety tests of the FPH produced by these processes demonstrated that the maximum proportion of FPH that can be safely used in food formulation is 10%, due to its high content of histamine. This study demonstrates the value of FPH to the food industry and bridges the theoretical studies with the commercial applications of FPH. © 2015 Institute of Food Technologists®

Characterization of the cation-binding capacity of a potassium-adsorption filter used in red blood cell transfusion.

PubMed

Suzuki, Takao; Muto, Shigeaki; Miyata, Yukio; Maeda, Takao; Odate, Takayuki; Shimanaka, Kimio; Kusano, Eiji

2015-06-01

A K(+) -adsorption filter was developed to exchange K(+) in the supernatant of stored irradiated red blood cells with Na(+) . To date, however, the filter's adsorption capacity for K(+) has not been fully evaluated. Therefore, we characterized the cation-binding capacity of this filter. Artificial solutions containing various cations were continuously passed through the filter in 30 mL of sodium polystyrene sulfonate at 10 mL/min using an infusion pump at room temperature. The cation concentrations were measured before and during filtration. When a single solution containing K(+) , Li(+) , H(+) , Mg(2+) , Ca(2+) , or Al(3+) was continuously passed through the filter, the filter adsorbed K(+) and the other cations in exchange for Na(+) in direct proportion to the valence number. The order of affinity for cation adsorption to the filter was Ca(2+) >Mg(2+) >K(+) >H(+) >Li(+) . In K(+) -saturated conditions, the filter also adsorbed Na(+) . After complete adsorption of these cations on the filter, their concentration in the effluent increased in a sigmoidal manner over time. Cations that were bound to the filter were released if a second cation was passed through the filter, despite the different affinities of the two cations. The ability of the filter to bind cations, especially K(+) , should be helpful when it is used for red blood cell transfusion at the bedside. The filter may also be useful to gain a better understanding of the pharmacological properties of sodium polystyrene sulfonate. © 2015 The Authors. Therapeutic Apheresis and Dialysis © 2015 International Society for Apheresis.

Inhibition of CYP6B1-mediated detoxification of xanthotoxin by plant allelochemicals in the black swallowtail (Papilio polyxenes).

PubMed

Wen, Zhimou; Berenbaum, May R; Schuler, Mary A

2006-03-01

The structural and biosynthetic diversity of allelochemicals in plants is thought to arise from selection for additive toxicity as a consequence of toxin mixture or for enhanced toxicity as a result of synergism. In order to understand how insects cope with this type of plant defense, we tested the effects of some allelochemicals in host plants of the black swallowtail Papilio polyxenes on the xanthotoxin-metabolic activity of CYP6B1, the principal enzyme responsible for the detoxification of furanocoumarins in this caterpillar. Additionally, the effects of some synthetic compounds not normally encountered by P. polyxenes on CYP6B1 were tested. These studies demonstrate that the integrity of furanocoumarin structure is important for competitive binding to the active site of CYP6B1, even though the carbonyl group on the pyranone ring apparently does not affect its inhibitory capacity, as in the case of furanochromones. Angular furanocoumarins are generally less phototoxic to many organisms than linear furanocoumarins due to their reduced capacity for cross-linking DNA strands, yet they are more toxic than linear furanocoumarins to black swallowtail larvae. This enhanced toxicity in vivo may be due to the ability of angular furanocoumarins to bind to the active site of CYP6B1 without being rapidly metabolized. This binding reduces the availability of CYP6B1 to metabolize other linear furanocoumarins. The structure-activity relationships for methylenedioxyphenyl compounds, flavonoids, imidazole, and imidazole derivatives are also discussed in light of their capacity to inhibit the xanthotoxin-metabolic activity of CYP6B1.

Facile synthesis of zwitterionic polymer-coated core-shell magnetic nanoparticles for highly specific capture of N-linked glycopeptides.

PubMed

Chen, Yajing; Xiong, Zhichao; Zhang, Lingyi; Zhao, Jiaying; Zhang, Quanqing; Peng, Li; Zhang, Weibing; Ye, Mingliang; Zou, Hanfa

2015-02-21

Highly selective and efficient capture of glycosylated proteins and peptides from complex biological samples is of profound significance for the discovery of disease biomarkers in biological systems. Recently, hydrophilic interaction liquid chromatography (HILIC)-based functional materials have been extensively utilized for glycopeptide enrichment. However, the low amount of immobilized hydrophilic groups on the affinity material has limited its specificity, detection sensitivity and binding capacity in the capture of glycopeptides. Herein, a novel affinity material was synthesized to improve the binding capacity and detection sensitivity for glycopeptides by coating a poly(2-(methacryloyloxy)ethyl)-dimethyl-(3-sulfopropyl) ammonium hydroxide (PMSA) shell onto Fe3O4@SiO2 nanoparticles, taking advantage of reflux-precipitation polymerization for the first time (denoted as Fe3O4@SiO2@PMSA). The thick polymer shell endows the nanoparticles with excellent hydrophilic property and several functional groups on the polymer chains. The resulting Fe3O4@SiO2@PMSA demonstrated an outstanding ability for glycopeptide enrichment with high selectivity, extremely high detection sensitivity (0.1 fmol), large binding capacity (100 mg g(-1)), high enrichment recovery (above 73.6%) and rapid magnetic separation. Furthermore, in the analysis of real complicated biological samples, 905 unique N-glycosylation sites from 458 N-glycosylated proteins were reliably identified in three replicate analyses of a 65 μg protein sample extracted from mouse liver, showing the great potential of Fe3O4@SiO2@PMSA in the detection and identification of low-abundance N-linked glycopeptides in biological samples.

Physicochemical properties and biological activities of DEAE-derivatized Sphingomonas gellan.

PubMed

Yoo, Sang-Ho; Lee, Kyung Hee; Lee, Ji-Soo; Cha, Jaeho; Park, Cheon Seok; Lee, Hyeon Gyu

2005-08-10

Physicochemical characteristics and biological activities of Sphingomonas gellan (S-gellan) were investigated. The S-gellan weight fractions of Glc and GlcUA were 0.45 and 0.25, respectively, and the molar ratio of Glc:Rha:GlcUA was approximately 4:2:3. The S-gellan was chemically derivatized with diethylaminoethyl chloride-HCl (DEAE-HCl), and the resulting modified S-gellan contained both positive and negative charges. The elemental and IR analyses were conducted to confirm the successful incorporation of DEAE groups into S-gellan. A large increase in nitrogen fraction was observed from the derivatized S-gellan by elemental analysis. The IR absorption bands induced by C-H, C-N, and C-O-C stretching were noticeable at 2950, 1310-1380, and 1000-1150 cm(-1), respectively, resulting from the DEAE substitution. The characteristic CH3 and CH2 peaks originated from the DEAE group were detected in the 1H NMR spectrum of the derivatized S-gellan as well. The solubility of native S-gellan was improved almost twice from 40% to 75% after DEAE derivatization, while water holding capacity (WHC) drastically decreased from 10026% to 245%. Oil binding capacity (OBC) of S-gellan also significantly dropped from 1528% to 331% after the derivatization. The bile acid binding capacity of S-gellan was indirectly determined by measuring the holding capability of cholic acid inside the dialysis membrane (MWCO 12,000-14,000 Da). Once S-gellan was DEAE derivatized, there was substantial increase in the cholic acid retardation index (CRI). Up to 9 h of dialysis, the derivatized S-gellan released 29.3% less of cholic acid compared to the control group that did not contain S-gellan. From these results of the improved water solubility and stronger bile acid binding capacity, it would be suggested that the DEAE-derivatized S-gellan has more advantages than gellan itself for functional food applications.

[Analysis of the binding capacity of the benzodiazepine site of gabaa receptor in mice C57BL/6 and BALB/C pretreated with anxiolytics].

PubMed

Iarkova, M A

2011-01-01

The level of specific 3H-flunitrazepam binding in synaptosomal membranes of C57BL/6 and BALB/c mice brain underwent to the stress of different types has been studied. Mild stress (Elevated Plus Maze) was shown to induce the decrease of benzodiazepine binding in BALB/c mice only, while the strong one (Exposure to a predator) was revealed to cause this decrease in both strains. Behavioral effects of different non-benzodiazepine drugs possessing anxiolytic properties (Afobazol, Ladasten and Noopept) was accompanied with the normalization of the level of benzodiazepine reception, reduced by the stress of both modalities.

Effect of molecular parameters on the binding of phenoxyacetic acid derivatives to albumins.

PubMed

Cserháti, T; Forgács, E; Deyl, Z; Miksík, I

2001-03-25

The interaction of 12 phenoxyacetic acid derivatives with human and serum albumin as well as with egg albumin was studied by charge-transfer reversed-phase (RP) thin-layer chromatography (TLC) and the relative strength of interaction was calculated. Each phenoxyacetic acid derivative interacted with human and bovine serum albumins whereas no interaction was observed with egg albumin. Stepwise regression analysis proved that the lipophilicity of the derivatives exert a significant impact on their capacity to bind to serum albumins. This result supports the hypothesis that the binding of phenoxyacetic acid derivatives to albumins may involve hydrophobic forces occurring between the corresponding apolar substructures of these derivatives and the amino acid side chains.

Identification of a boron nitride nanosphere-binding peptide for the intracellular delivery of CpG oligodeoxynucleotides

NASA Astrophysics Data System (ADS)

Zhang, Huijie; Yamazaki, Tomohiko; Zhi, Chunyi; Hanagata, Nobutaka

2012-09-01

CpG oligonucleotides (CpG ODNs) interact with Toll-like receptor 9 (TLR9), which results in the induction of immunostimulatory cytokines. We delivered CpG ODNs intracellularly using boron nitride nanospheres (BNNS). To enhance the loading capacity of CpG ODNs on BNNS, we used a phage display technique to identify a 12-amino acid peptide designated as BP7, with specific affinity for BNNS, and used it as a linker to load CpG ODNs on BNNS. The tyrosine residue (Y) at the eighth position from the N-terminus played a crucial role in the affinity of BP7 to BNNS. BNNS that bound BP7 (BNNS-BP7) were taken up by cells and showed no cytotoxicity, and CpG ODNs were successfully crosslinked with BP7 to create BP7-CpG ODN conjugates. Using BP7 as a linker, the loading efficiency of CpG ODNs on BNNS increased 5-fold compared to the direct binding of CpG ODNs to BNNS. Furthermore, the BP7-CpG ODN conjugate-loaded BNNS had a greater capacity to induce interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) production from peripheral blood mononuclear cells (PBMCs) than that of CpG ODNs directly loaded on BNNS. The higher amount of cytokine induction by BP7-CpG ODN conjugate-loaded BNNS may be attributed to a higher loading capacity and stronger binding to BNNS of the linker BP7. The greater functionality of BP7-conjugated CpG ODNs on BNNS expands the potential of BNNS for drug delivery applications.CpG oligonucleotides (CpG ODNs) interact with Toll-like receptor 9 (TLR9), which results in the induction of immunostimulatory cytokines. We delivered CpG ODNs intracellularly using boron nitride nanospheres (BNNS). To enhance the loading capacity of CpG ODNs on BNNS, we used a phage display technique to identify a 12-amino acid peptide designated as BP7, with specific affinity for BNNS, and used it as a linker to load CpG ODNs on BNNS. The tyrosine residue (Y) at the eighth position from the N-terminus played a crucial role in the affinity of BP7 to BNNS. BNNS that bound BP7 (BNNS-BP7) were taken up by cells and showed no cytotoxicity, and CpG ODNs were successfully crosslinked with BP7 to create BP7-CpG ODN conjugates. Using BP7 as a linker, the loading efficiency of CpG ODNs on BNNS increased 5-fold compared to the direct binding of CpG ODNs to BNNS. Furthermore, the BP7-CpG ODN conjugate-loaded BNNS had a greater capacity to induce interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) production from peripheral blood mononuclear cells (PBMCs) than that of CpG ODNs directly loaded on BNNS. The higher amount of cytokine induction by BP7-CpG ODN conjugate-loaded BNNS may be attributed to a higher loading capacity and stronger binding to BNNS of the linker BP7. The greater functionality of BP7-conjugated CpG ODNs on BNNS expands the potential of BNNS for drug delivery applications. Electronic supplementary information (ESI) available. See DOI: 10.1039/c2nr31189e

Diminished hepatic growth hormone receptor binding in sex-linked dwarf broiler and leghorn chickens.

PubMed

Leung, F C; Styles, W J; Rosenblum, C I; Lilburn, M S; Marsh, J A

1987-02-01

Hepatic growth hormone (GH) receptor binding was compared in normal and sex-linked dwarfs (SLD) from both Hubbard and Cornell strain chickens. At 6, 8, and 20 weeks of age, hepatic GH receptor binding in the Hubbard SLD chickens was significantly lower than that of normal fast-growing birds. At 20 weeks of age, only 2 of 22 SLD chickens in the Hubbard broiler strain showed positive binding at a high enough level to allow for Scatchard analysis. The affinity constants and binding capacities of these two SLD chickens were numerically (but not significantly) lower than those of the normal fast-growing birds. We further examined hepatic GH receptor binding in two closely related White Leghorn strains of chickens that have been maintained as closed breeding populations for many years. We observed no detectable hepatic GH binding in the Cornell SLD chickens (N = 20), as compared to the normal-growing control strain (K strain). In both SLD strains, pretreatment with 4 M MgCl2 did not enhance GH binding, suggesting that there was no endogenous GH binding to the receptor. Based on these data, we suggest that the lack, or greatly reduced number, of GH receptors may be a major contributing factor to the dwarfism observed in these strains.

Critical insight into the interaction of naringenin with human haemoglobin: A combined spectroscopic and computational modeling approaches

NASA Astrophysics Data System (ADS)

Maity, Subhajit; Chakraborty, Sandipan; Chakraborti, Abhay Sankar

2017-02-01

The present study demonstrates critical insight into the binding of a bioactive flavanone naringenin with normal human haemoglobin (NHb). Both spectrophotometric and spectrofluorimetric studies reveal that naringenin interacts with NHb. The binding affinity constant and number of binding sites appear to be approximately (1.5 ± 0.2) × 104 M-1 and 1, respectively. Static quenching seems to be an important factor in binding process, as evident from steady-state and time-resolved fluorescence spectroscopic studies. Far UV circular dichroism spectroscopy depicts that binding of naringenin to NHb causes no change in the secondary structure of the protein, which is also evident from Fourier transform infrared spectroscopic study. Free energy change (ΔG0) for naringenin-NHb interaction, determined by spectroscopic and isothermal calorimetric method, appears to be -5.67 kcal/mol and -6.90 kcal/mol, respectively, and is close to the docking energy -6.84 kcal/mol. Molecular docking suggests that naringenin binds near the cavity of the tetrameric heme protein, forming hydrogen bonds with surrounding amino acid residues. The binding site is away from the heme moieties, implicating naringenin binding does not affect the oxygen binding capacity of NHb, which makes the protein a suitable carrier of the flavonoid.

Individual differences in working memory capacity and workload capacity.

PubMed

Yu, Ju-Chi; Chang, Ting-Yun; Yang, Cheng-Ta

2014-01-01

We investigated the relationship between working memory capacity (WMC) and workload capacity (WLC). Each participant performed an operation span (OSPAN) task to measure his/her WMC and three redundant-target detection tasks to measure his/her WLC. WLC was computed non-parametrically (Experiments 1 and 2) and parametrically (Experiment 2). Both levels of analyses showed that participants high in WMC had larger WLC than those low in WMC only when redundant information came from visual and auditory modalities, suggesting that high-WMC participants had superior processing capacity in dealing with redundant visual and auditory information. This difference was eliminated when multiple processes required processing for only a single working memory subsystem in a color-shape detection task and a double-dot detection task. These results highlighted the role of executive control in integrating and binding information from the two working memory subsystems for perceptual decision making.

A Single Rainbow Trout Cobalamin-binding Protein Stands in for Three Human Binders

PubMed Central

Greibe, Eva; Fedosov, Sergey; Sorensen, Boe S.; Højrup, Peter; Poulsen, Steen S.; Nexo, Ebba

2012-01-01

Cobalamin uptake and transport in mammals are mediated by three cobalamin-binding proteins: haptocorrin, intrinsic factor, and transcobalamin. The nature of cobalamin-binding proteins in lower vertebrates remains to be elucidated. The aim of this study was to characterize the cobalamin-binding proteins of the rainbow trout (Oncorhynchus mykiss) and to compare their properties with those of the three human cobalamin-binding proteins. High cobalamin-binding capacity was found in trout stomach (210 pmol/g), roe (400 pmol/g), roe fluid (390 nmol/liter), and plasma (2500 nmol/liter). In all cases, it appeared to be the same protein based on analysis of partial sequences and immunological responses. The trout cobalamin-binding protein was purified from roe fluid, sequenced, and further characterized. Like haptocorrin, the trout cobalamin-binding protein was stable at low pH and had a high binding affinity for the cobalamin analog cobinamide. Like haptocorrin and transcobalamin, the trout cobalamin-binding protein was present in plasma and recognized ligands with altered nucleotide moiety. Like intrinsic factors, the trout cobalamin-binding protein was present in the stomach and resisted degradation by trypsin and chymotrypsin. It also resembled intrinsic factor in the composition of conserved residues in the primary cobalamin-binding site in the C terminus. The trout cobalamin-binding protein was glycosylated and displayed spectral properties comparable with those of haptocorrin and intrinsic factor. In conclusion, only one soluble cobalamin-binding protein was identified in the rainbow trout, a protein that structurally behaves like an intermediate between the three human cobalamin-binding proteins. PMID:22872637

Quantification of oil binding capacity of structuring fats: A novel method and its application.

PubMed

Omonov, Tolibjon S; Bouzidi, Laziz; Narine, Suresh S

2010-09-01

A robust, well-defined and reproducible method to accurately measure the oil binding capacity (OBC) of structuring fats was developed. The method was validated using two oil/fat model systems, i.e., fully hydrogenated canola oil (FHCO) in canola oil (CO) (FHCO/CO) and fully hydrogenated soybean oil (FHSO) in CO (FHSO/CO). The mixtures were crystallized from the melt down to three different temperatures (15, 25 and 35 degrees C) at constant rates of cooling and the OBC was measured after different periods of storage time. The critical concentration of hard fat at which the solid fat network is stable and effectively binds oil has been also measured for mixtures crystallized at temperatures close to room temperature, i.e., 25 degrees C. Crystal structure, melting behavior, microstructure, and solid fat content of these binary systems have been investigated in relation to the OBC of the solid fat network using X-ray diffraction (XRD), differential scanning calorimetry (DSC), polarized light microscopy (PLM), and wide-line pulsed nuclear magnetic resonance (pNMR) techniques. The two model systems exhibited similar trends in OBC over time, a behavior attributed to their similar TAG composition and polymorphism. However, relatively smaller OBC values were achieved by the CO/FHSO compared to CO/FHCO samples, largely due to differences in their solid network structure. Four successive decreasing linear segments, identifying successive mechanisms of oil migration/binding, were observed in the experimental OBC versus fat weight fraction curves. The critical concentration of hard fat, at which the solid fat network is effective in binding oil, was also determined and found to be approximately 6wt% for both systems. Copyright 2010 Elsevier Ireland Ltd. All rights reserved.

Temperature and pH Dual-Responsive Core-Brush Nanocomposite for Enrichment of Glycoproteins.

PubMed

Jiang, Lingdong; Messing, Maria E; Ye, Lei

2017-03-15

In this report, we present a novel modular approach to the immobilization of a high density of boronic acid ligands on thermoresponsive block copolymer brushes for effective enrichment of glycoproteins via their synergistic multiple covalent binding with the immobilized boronic acids. Specifically, a two-step, consecutive surface-initiated atom transfer radical polymerization (SI-ATRP) was employed to graft a flexible block copolymer brush, pNIPAm-b-pGMA, from an initiator-functionalized nanosilica surface, followed by postpolymerization modification of the pGMA moiety with sodium azide. Subsequently, an alkyne-tagged boronic acid (PCAPBA) was conjugated to the polymer brush via a Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC) click reaction, leading to a silica-supported polymeric hybrid material, Si@pNIPAm-b-pBA, with a potent glycol binding affinity. The obtained core-brush nanocomposite was systematically characterized with regard to particle size, morphology, organic content, brush density, and number of immobilized boronic acids. We also studied the characteristics of glycoprotein binding of the nanocomposite under different conditions. The nanocomposite showed high binding capacities for ovalbumin (OVA) (98.0 mg g -1 ) and horseradish peroxidase (HRP) (26.8 mg g -1 ) in a basic buffer (pH 9.0) at 20 °C. More importantly, by adjusting the pH and temperature, the binding capacities of the nanocomposite can be tuned, which is meaningful for the separation of biological molecules. In general, the synthetic approach developed for the fabrication of block copolymer brushes in the nanocomposite opened new opportunities for the design of more functional hybrid materials that will be useful in bioseparation and biomedical applications.

Optimization of exopolysaccharide production from Pseudomonas stutzeri AS22 and examination of its metal-binding abilities.

PubMed

Maalej, H; Hmidet, N; Boisset, C; Buon, L; Heyraud, A; Nasri, M

2015-02-01

To investigate the effect of culture conditions and medium components on exopolysaccharide (EPS) production by Pseudomonas stutzeri AS22 and to access the EPS performance as a metal-binding exopolysaccharide. The EPS production conditions of Ps. stutzeri AS22 in submerged culture were optimized using two approaches for EPS quantification, and its metal-binding capacity was evaluated using both single and mixed metal ions systems. Maximum EPS level was achieved after 24 h of incubation at 30°C with an initial pH of 8.0, 250 rev min(-1) stirring level and 10% inoculum size. 50 g l(-1) starch, 5 g l(-1) yeast extract, 0.5 g l(-1) NaCl, 1.4 g l(-1) K2 HPO4, 0.4 g l(-1) MgSO4, 0.4 g l(-1) CaCl2 and 1 g l(-1) mannose were found to be the most suitable carbon, nitrogen, mineral and additional carbohydrate sources, respectively. From metal-binding experiments, the crude EPS showed interesting metal adsorption capacity adopting the order Pb > Co > Fe > Cu > Cd. Lead was preferentially biosorbed with a maximal uptake of 460 mg g(-1) crude EPS. Under the optimal culture requirements, EPS level reached 10.2 g l(-1) after 24 h of fermentation, seven times more than the production under initial conditions. According to the metal-binding assay, the crude EPS has potential to be used as a novel biosorbent in the treatment of heavy metals-contaminated water. Our results are interesting in terms of yield as well as efficiency for the potential use of the Ps. stutzeri exopolysaccharide as a metal-absorbent polymer in the bioremediation field. © 2014 The Society for Applied Microbiology.

The amino terminal end determines the stability and assembling capacity of eukaryotic ribosomal stalk proteins P1 and P2.

PubMed

Camargo, Hendricka; Nusspaumer, Gretel; Abia, David; Briceño, Verónica; Remacha, Miguel; Ballesta, Juan P G

2011-05-01

The eukaryotic ribosomal proteins P1 and P2 bind to protein P0 through their N-terminal domain to form the essential ribosomal stalk. A mutational analysis points to amino acids at positions 2 and 3 as determinants for the drastic difference of Saccharomyces cerevisiae P1 and P2 half-life, and suggest different degradation mechanisms for each protein type. Moreover, the capacity to form P1/P2 heterodimers is drastically affected by mutations in the P2β four initial amino acids, while these mutations have no effect on P1β. Binding of P2β and, to a lesser extent, P1β to the ribosome is also seriously affected showing the high relevance of the amino acids in the first turn of the NTD α-helix 1 for the stalk assembly. The negative effect of some mutations on ribosome binding can be reversed by the presence of the second P1/P2 couple in the ribosome, indicating a stabilizing structural influence between the two heterodimers. Unexpectedly, some mutations totally abolish heterodimer formation but allow significant ribosome binding and, therefore, a previous P1 and P2 association seems not to be an absolute requirement for stalk assembly. Homology modeling of the protein complexes suggests that the mutated residues can affect the overall protein conformation. © The Author(s) 2011. Published by Oxford University Press.

Endosomal escape and siRNA delivery with cationic shell crosslinked knedel-like nanoparticles with tunable buffering capacities

PubMed Central

Shrestha, Ritu; Elsabahy, Mahmoud; Florez-Malaver, Stephanie; Samarajeewa, Sandani; Wooley, Karen L.

2012-01-01

Cationic shell crosslinked knedel-like nanoparticles (cSCKs) have emerged as a highly efficient transfection agent for nucleic acids delivery. In this study, a new class of cSCKs with tunable buffering capacities has been developed by altering the amounts of histamines and primary amines incorporated into their crosslinked shell regions. The effect of histamine content of these nanoparticles with a hydrodynamic diameter of ca. 20 nm, on the siRNA-binding affinity, cytotoxicity, immunogenicity, and transfection efficiency was investigated. The modification of cSCKs with histamine was found to reduce the siRNA-binding affinity and cellular binding. On the other hand, it significantly reduced the toxicity and immunogenicity of the nanoparticles with subsequent increase in the transfection efficiency. In addition, escape from endosomes was facilitated by having two species of low and high pKas (i.e. histamine and primary amine groups, respectively), as demonstrated by the potentiometric titration experiments and the effect of bafilomycin A1, an inhibitor of the endosomal acidification, on the transfection efficiency of cSCKs. Histamine modification of 15 mol% was a threshold, above which cSCKs with higher histamine content completely lost the ability to bind siRNA and to transfect cells. This study highlights the potential of histamine incorporation to augment the gene silencing activity of cationic nanoparticles, reduce their toxicity, and increase their biocompatibility, which is of particular importance in the design of nucleic acids delivery vectors. PMID:22901966

Nanostructured Block Polymer Membranes as High Capacity Adsorbers for the Capture of Metal Ions from Water

NASA Astrophysics Data System (ADS)

Boudouris, Bryan; Weidman, Jacob; Mulvenna, Ryan; Phillip, William

The efficient removal of metal ions from aqueous streams is of significant import in applications ranging from industrial waste treatment to the purification of drinking water. An emerging paradigm associated with this separation is one that utilizes membrane adsorbers as a means by which to bind metal salt contaminants. Here, we demonstrate that the casting of an A-B-C triblock polymer using the self-assembly and non-solvent induced phase separation (SNIPS) methodology results in a nanoporous membrane geometry. The nature of the triblock polymer affords an extremely high density of binding sites within the membrane. As such, we demonstrate that the membranes with binding capacities equal to that of state-of-the-art packed bed columns. Moreover, because the affinity of the C moiety can be tuned, highly selective binding events can occur based solely on the chemistry of the block polymer and the metal ions in solution (i.e., in a manner that is independent of the size of the metal ions). Due to these combined facts, these membranes efficiently remove heavy metal (e.g., lead- and cadmium-based) salts from contaminated water streams with greater than 95% efficiency. Finally, we show that the membranes can be regenerated through a simple treatment in order to provide long-lasting adsorber systems as well. Thus, it is anticipated that these nanostructured triblock polymer membranes are a platform by which to obtain next-generation water purification processes.

Boronic acid-modified magnetic materials for antibody purification

PubMed Central

Dhadge, Vijaykumar L.; Hussain, Abid; Azevedo, Ana M.; Aires-Barros, Raquel; Roque, Ana C. A.

2014-01-01

Aminophenyl boronic acids can form reversible covalent ester interactions with cis-diol-containing molecules, serving as a selective tool for binding glycoproteins as antibody molecules that possess oligosaccharides in both the Fv and Fc regions. In this study, amino phenyl boronic acid (APBA) magnetic particles (MPs) were applied for the magnetic separation of antibody molecules. Iron oxide MPs were firstly coated with dextran to avoid non-specific binding and then with 3-glycidyloxypropyl trimethoxysilane to allow further covalent coupling of APBA (APBA_MP). When contacted with pure protein solutions of human IgG (hIgG) and bovine serum albumin (BSA), APBA_MP bound 170 ± 10 mg hIgG g−1 MP and eluted 160 ± 5 mg hIgG g−1 MP, while binding only 15 ± 5 mg BSA g−1 MP. The affinity constant for the interaction between hIgG and APBA_MP was estimated as 4.9 × 105 M−1 (Ka) with a theoretical maximum capacity of 492 mg hIgG adsorbed g−1 MP (Qmax), whereas control particles bound a negligible amount of hIgG and presented an estimated theoretical maximum capacity of 3.1 mg hIgG adsorbed g−1 MP (Qmax). APBA_MPs were also tested for antibody purification directly from CHO cell supernatants. The particles were able to bind 98% of IgG loaded and to recover 95% of pure IgG (purity greater than 98%) at extremely mild conditions. PMID:24258155

Differential processing of the two subunits of human choriogonadotropin (hCG) by granulosa cells. I. Preparation and characterization of selectively labeled hCG

DOE Office of Scientific and Technical Information (OSTI.GOV)

Landefeld, T.D.; Byrne, M.D.; Campbell, K.L.

1981-12-01

The alpha- and beta-subunits of hCG were radioiodinated and recombined with unlabeled complementary subunits. The resultant recombined hormones, selectively labeled in either the alpha- or beta-subunit, were separated from unrecombined subunit by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, extracted with Triton X-100, and characterized by binding analysis. The estimates of maximum binding (active fraction) of the two resultant selectively labeled, recombined hCG preparations, determined with excess receptor were 0.41 and 0.59. These values are similar to those obtained when hCG is labeled as an intact molecule. The specific activities of the recombined preparations were estimated by four different methods, and themore » resulting values were used in combination with the active fraction estimates to determine the concentrations of active free and bound hormone. Binding analyses were run using varying concentrations of both labeled and unlabeled hormone. Estimates of the equilibrium dissociation binding constant (Kd) and receptor capacity were calculated in three different ways. The mean estimates of capacity (52.6 and 52.7 fmol/mg tissue) and Kd (66.6 and 65.7 pM) for the two preparations were indistinguishable. Additionally, these values were similar to values reported previously for hCG radioiodinated as an intact molecule. The availability of well characterized, selectively labeled hCG preparations provides new tools for studying the mechanism of action and the target cell processing of the subunits of this hormone.« less

Transformation of glucocorticoid receptors bound to the antagonist RU 486: Effects of alkaline phosphatase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gruol, D.J.; Wolfe, K.A.

1990-08-28

RU 486 is a synthetic steroid that binds avidly to glucocorticoid receptors without promoting their transformation into activated transcription factors. A significant part of this behavior has been shown to be due to a failure of the RU 486 bound receptor to be efficiently released from a larger (sedimenting at 8-9 S) multimeric complex containing the 90-kDa heat shock protein. The studies have found that in vitro at 15{degree}C the RU 486-receptor was slowly released from the 8-9S complex and converted into a DNA binding protein by a process that could be blocked by sodium fluoride. Moreover, this transition wasmore » significantly accelerated by treatment with alkaline phosphatase. High-resolution anion-exchange chromatography showed that the profile of receptor subspecies released from the 8-9S complex was different for the RU 486 bound receptor when compared to the receptor occupied by the agonist triamcinolone acetonide. Production of the earliest eluting receptor form (peak A) was inhibited with RU 486. Treatment of the Ru 486-receptor with alkaline phosphatase increased the formation of the peak A subspecies as well as the capacity of receptor to bind DNA-cellulose. Taken together, the results indicate that phosphorylation of the receptor or a tightly bound factor contributes to defining the capacity with which individual steroids can promote dissociation of the 8-9S complex and conversion of the glucocorticoid receptor into a DNA-binding protein.« less

Incommensurate Graphene Foam as a High Capacity Lithium Intercalation Anode

PubMed Central

Paronyan, Tereza M.; Thapa, Arjun Kumar; Sherehiy, Andriy; Jasinski, Jacek B.; Jangam, John Samuel Dilip

2017-01-01

Graphite’s capacity of intercalating lithium in rechargeable batteries is limited (theoretically, 372 mAh g−1) due to low diffusion within commensurately-stacked graphene layers. Graphene foam with highly enriched incommensurately-stacked layers was grown and applied as an active electrode in rechargeable batteries. A 93% incommensurate graphene foam demonstrated a reversible specific capacity of 1,540 mAh g−1 with a 75% coulombic efficiency, and an 86% incommensurate sample achieves above 99% coulombic efficiency exhibiting 930 mAh g−1 specific capacity. The structural and binding analysis of graphene show that lithium atoms highly intercalate within weakly interacting incommensurately-stacked graphene network, followed by a further flexible rearrangement of layers for a long-term stable cycling. We consider lithium intercalation model for multilayer graphene where capacity varies with N number of layers resulting LiN+1C2N stoichiometry. The effective capacity of commonly used carbon-based rechargeable batteries can be significantly improved using incommensurate graphene as an anode material. PMID:28059110

Global workspace dynamics: cortical "binding and propagation" enables conscious contents.

PubMed

Baars, Bernard J; Franklin, Stan; Ramsoy, Thomas Zoega

2013-01-01

A global workspace (GW) is a functional hub of binding and propagation in a population of loosely coupled signaling elements. In computational applications, GW architectures recruit many distributed, specialized agents to cooperate in resolving focal ambiguities. In the brain, conscious experiences may reflect a GW function. For animals, the natural world is full of unpredictable dangers and opportunities, suggesting a general adaptive pressure for brains to resolve focal ambiguities quickly and accurately. GW theory aims to understand the differences between conscious and unconscious brain events. In humans and related species the cortico-thalamic (C-T) core is believed to underlie conscious aspects of perception, thinking, learning, feelings of knowing (FOK), felt emotions, visual imagery, working memory, and executive control. Alternative theoretical perspectives are also discussed. The C-T core has many anatomical hubs, but conscious percepts are unitary and internally consistent at any given moment. Over time, conscious contents constitute a very large, open set. This suggests that a brain-based GW capacity cannot be localized in a single anatomical hub. Rather, it should be sought in a functional hub - a dynamic capacity for binding and propagation of neural signals over multiple task-related networks, a kind of neuronal cloud computing. In this view, conscious contents can arise in any region of the C-T core when multiple input streams settle on a winner-take-all equilibrium. The resulting conscious gestalt may ignite an any-to-many broadcast, lasting ∼100-200 ms, and trigger widespread adaptation in previously established networks. To account for the great range of conscious contents over time, the theory suggests an open repertoire of binding coalitions that can broadcast via theta/gamma or alpha/gamma phase coupling, like radio channels competing for a narrow frequency band. Conscious moments are thought to hold only 1-4 unrelated items; this small focal capacity may be the biological price to pay for global access. Visuotopic maps in cortex specialize in features like color, retinal size, motion, object identity, and egocentric/allocentric framing, so that a binding coalition for the sight of a rolling billiard ball in nearby space may resonate among activity maps of LGN, V1-V4, MT, IT, as well as the dorsal stream. Spatiotopic activity maps can bind into coherent gestalts using adaptive resonance (reentry). Single neurons can join a dominant coalition by phase tuning to regional oscillations in the 4-12 Hz range. Sensory percepts may bind and broadcast from posterior cortex, while non-sensory FOKs may involve prefrontal and frontotemporal areas. The anatomy and physiology of the hippocampal complex suggest a GW architecture as well. In the intact brain the hippocampal complex may support conscious event organization as well as episodic memory storage.

Global Workspace Dynamics: Cortical “Binding and Propagation” Enables Conscious Contents

PubMed Central

Baars, Bernard J.; Franklin, Stan; Ramsoy, Thomas Zoega

2013-01-01

A global workspace (GW) is a functional hub of binding and propagation in a population of loosely coupled signaling elements. In computational applications, GW architectures recruit many distributed, specialized agents to cooperate in resolving focal ambiguities. In the brain, conscious experiences may reflect a GW function. For animals, the natural world is full of unpredictable dangers and opportunities, suggesting a general adaptive pressure for brains to resolve focal ambiguities quickly and accurately. GW theory aims to understand the differences between conscious and unconscious brain events. In humans and related species the cortico-thalamic (C-T) core is believed to underlie conscious aspects of perception, thinking, learning, feelings of knowing (FOK), felt emotions, visual imagery, working memory, and executive control. Alternative theoretical perspectives are also discussed. The C-T core has many anatomical hubs, but conscious percepts are unitary and internally consistent at any given moment. Over time, conscious contents constitute a very large, open set. This suggests that a brain-based GW capacity cannot be localized in a single anatomical hub. Rather, it should be sought in a functional hub – a dynamic capacity for binding and propagation of neural signals over multiple task-related networks, a kind of neuronal cloud computing. In this view, conscious contents can arise in any region of the C-T core when multiple input streams settle on a winner-take-all equilibrium. The resulting conscious gestalt may ignite an any-to-many broadcast, lasting ∼100–200 ms, and trigger widespread adaptation in previously established networks. To account for the great range of conscious contents over time, the theory suggests an open repertoire of binding1 coalitions that can broadcast via theta/gamma or alpha/gamma phase coupling, like radio channels competing for a narrow frequency band. Conscious moments are thought to hold only 1–4 unrelated items; this small focal capacity may be the biological price to pay for global access. Visuotopic maps in cortex specialize in features like color, retinal size, motion, object identity, and egocentric/allocentric framing, so that a binding coalition for the sight of a rolling billiard ball in nearby space may resonate among activity maps of LGN, V1-V4, MT, IT, as well as the dorsal stream. Spatiotopic activity maps can bind into coherent gestalts using adaptive resonance (reentry). Single neurons can join a dominant coalition by phase tuning to regional oscillations in the 4–12 Hz range. Sensory percepts may bind and broadcast from posterior cortex, while non-sensory FOKs may involve prefrontal and frontotemporal areas. The anatomy and physiology of the hippocampal complex suggest a GW architecture as well. In the intact brain the hippocampal complex may support conscious event organization as well as episodic memory storage. PMID:23974723

Human lactoferrin binding to Porphyromonas gingivalis, Prevotella intermedia and Prevotella melaninogenica.

PubMed

Kalfas, S; Andersson, M; Edwardsson, S; Forsgren, A; Naidu, A S

1991-12-01

Human isolates of Porphyromonas gingivalis (n = 16), Prevotella intermedia n = 82) and Prevotella melaninogenica (n = 18) from diseased periodontal pockets were examined for interaction with human lactoferrin (HLf) in a standardized 125I-labeled protein binding assay. The highest HLf binding was found in P. intermedia strains, followed by P. gingivalis and P. melaninogenica. Further characterization of the interaction was performed with 1 representative strain from each species. HLf binding to P. gingivalis reached a saturation instantly and was optimal at pH 5.0-6.5. The corresponding values for P. melaninogenica were 90 min and pH 3.0-5.5. The HLf binding to the 2 strains seem to be nonspecific. In contrast, P. intermedia demonstrated specific binding, and a time-saturability within 60 min with an optimal uptake at pH 6.0-7.5. Scatchard analysis implied 45,000 receptors per cell with an affinity constant of 5.5 x 10(-7) M on P. intermedia strain 4H. The binding capacity in all 3 strains was affected by the culture medium. HLf binding components in these strains were susceptible to heat or proteases. Binding was eliminated in P. gingivalis and was enhanced in P. intermedia and P. melaninogenica by periodate treatment. Unlabeled HLf or bovine lactoferrin effectively displaced labeled HLf binding. Various proteins and carbohydrates did not inhibit HLf binding. Our data suggest that HLf binds to these periodontitis-associated species and that this mechanism is distinct from the previously known ligand interactions in oral bacteria.

Thermodynamics of the binding of L-arabinose and of D-galactose to the L-arabinose-binding protein of Escherichia coli.

PubMed

Fukada, H; Sturtevant, J M; Quiocho, F A

1983-11-10

The thermodynamics of the binding of L-arabinose and of D-galactose to the L-arabinose-binding protein of Escherichia coli have been studied by isothermal and scanning calorimetry. The binding reaction with arabinose is characterized by an enthalpy change of -15.3 +/- 0.5 kcal mol-1 at 25 degrees C, and a large decrease in apparent heat capacity, amounting to -0.44 +/- 0.05 kcal K-1 mol-1, which is constant over the temperature range 8 to 30 degrees C. Very similar results were obtained with D-galactose. These calorimetric results have been combined with binding constants determined by equilibrium dialysis (Clark, A. F., Gerken, T. A., and Hogg, R. W. (1982) Biochemistry 21, 2227-2233) to obtain free energy and entropy changes over the range 5 to 30 degrees C, and by extrapolation to 60 degrees C. The protein undergoes reversible unfolding on being heated with an increase in enthalpy at 53.5 degrees C of 151.8 +/- 1.1 kcal mol-1 (169.2 +/- 1.2 kcal mol-1 at 59.0 degrees C) and in apparent heat capacity of 3.16 +/- 0.07 kcal K-1 mol-1. In the presence of arabinose, the unfolding enthalpy is increased to 200.7 +/- 1.8 kcal mol-1 at 59.0 degrees C, the increase being due to the enthalpy of dissociation of the ligand which amounts to 31 kcal mol-1 at the unfolding temperature. The unfolding temperature is increased by the presence of excess arabinose or galactose, an effect which is due solely to displacement by the added ligand of the unfolding-dissociation equilibrium. The thermodynamic data are discussed in connection with the detailed structural information available for this system from x-ray crystallography (Newcomer, M. E., Gilliland, G. L. and Quiocho, F. A. (1981) J. Biol. Chem. 256, 13213-13217, and references cited therein).

Long-Lasting Impairment of mGluR5-Activated Intracellular Pathways in the Striatum After Withdrawal of Cocaine Self-Administration

PubMed Central

Hoffmann, Hanne Mette; Crouzin, Nadine; Moreno, Estefanía; Raivio, Noora; Fuentes, Silvia; McCormick, Peter J.; Vignes, Michel

2017-01-01

Abstract Background: Cocaine addiction continues to be a major heath concern, and despite public health intervention there is a lack of efficient pharmacological treatment options. A newly identified potential target are the group I metabotropic glutamate receptors, with allosteric modulators showing particular promise. Methods: We evaluated the capacity of group I metabotropic glutamate receptors to induce functional responses in ex vivo striatal slices from rats with (1) acute cocaine self-administration, (2) chronic cocaine self-administration, and (3) 60 days cocaine self-administration withdrawal by Western blot and extracellular recordings of synaptic transmission. Results: We found that striatal group I metabotropic glutamate receptors are the principal mediator of the mGluR1/5 agonist (RS)-3,5-dihydroxyphenylglycine-induced cAMP responsive-element binding protein phosphorylation. Both acute and chronic cocaine self-administration blunted group I metabotropic glutamate receptor effects on cAMP responsive-element binding protein phosphorylation in the striatum, which correlated with the capacity to induce long-term depression, an effect that was maintained 60 days after chronic cocaine self-administration withdrawal. In the nucleus accumbens, the principal brain region mediating the rewarding effects of drugs, chronic cocaine self-administration blunted group I metabotropic glutamate receptor stimulation of extracellular signal-regulated protein kinases 1/2 and cAMP responsive-element binding protein. Interestingly, the group I metabotropic glutamate receptor antagonist/inverse-agonist, 2-methyl-6-(phenylethynyl)pyridine hydrochloride, led to a specific increase in cAMP responsive-element binding protein phosphorylation after chronic cocaine self-administration, specifically in the nucleus accumbens, but not in the striatum. Conclusions: Prolonged cocaine self-administration, through withdrawal, leads to a blunting of group I metabotropic glutamate receptor responses in the striatum. In addition, specifically in the accumbens, group I metabotropic glutamate receptor signaling to cAMP responsive-element binding protein shifts from an agonist-induced to an antagonist-induced cAMP responsive-element binding protein phosphorylation. PMID:27744406

Insights into cellulase-lignin non-specific binding revealed by computational redesign of the surface of green fluorescent protein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Haarmeyer, Carolyn N.; Smith, Matthew D.; Chundawat, Shishir P. S.

Biological-mediated conversion of pretreated lignocellulosic biomass to biofuels and biochemicals is a promising avenue towards energy sustainability. However, a critical impediment to the commercialization of cellulosic biofuel production is the high cost of cellulase enzymes needed to deconstruct biomass into fermentable sugars. One major factor driving cost is cellulase adsorption and inactivation in the presence of lignin, yet we currently have a poor understanding of the protein structure-function relationships driving this adsorption. In this work, we have systematically investigated the role of protein surface potential on lignin adsorption using a model monomeric fluorescent protein. We have designed and experimentally characterizedmore » 16 model protein variants spanning the physiological range of net charge (-24 to +16 total charges) and total charge density (0.28 to 0.40 charges per sequence length) typical for natural proteins. Protein designs were expressed, purified, and subjected to in silico and in vitro biophysical measurements to evaluate the relationship between protein surface potential and lignin adsorption properties. The designs were comparable to model fluorescent protein in terms of thermostability and heterologous expression yield, although the majority of the designs unexpectedly formed homodimers. Protein adsorption to lignin was studied at two different temperatures using Quartz Crystal Microbalance with Dissipation Monitoring and a subtractive mass balance assay. We found a weak correlation between protein net charge and protein-binding capacity to lignin. No other single characteristic, including apparent melting temperature and 2nd virial coefficient, showed correlation with lignin binding. Analysis of an unrelated cellulase dataset with mutations localized to a family I carbohydrate-binding module showed a similar correlation between net charge and lignin binding capacity. Altogether, our study provides strategies to identify highly active, low lignin-binding cellulases by either rational design or by computational screening genomic databases.« less

Selective Capture of Cesium and Thallium from Natural Waters and Simulated Wastes with Copper Ferrocyanide Functionalized Mesoporous Silica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sangvanich, Thanapon; Sukwarotwat, Vichaya; Wiacek, Robert J.

2010-10-15

Copper(II) ferrocyanide immobilized inside mesoporous silica MCM-41 supports (Cu-FC-EDA-SAMMSTM) has been evaluated against iron(III) hexacyanoferrate(II) (insoluble Prussian blue) for the sorption of cesium (Cs+) and thallium (Tl+) from natural waters and simulated wastes. The affinities (in term of distribution coefficients, Kd) of both sorbents for Cs and Tl were measured as a function of solution pH, competing cations, and matrices. For the entire pH studied (pH 0.1 to 7.3), Cu-FC-EDA-SAMMS had higher affinities for Cs and Tl (one to two orders of magnitude higher Kd) than Prussian blue and was less negatively impacted by the solution pH, competing cations, andmore » matrices. The adsorption isotherms and kinetics of the two sorbents for Cs and/or Tl were also determined in seawater and simulated acid and alkaline wastes. SAMMS outperformed Prussian blue in terms of maximum adsorption capacity (e.g., 21.7 versus 2.6 mg Cs/g in acid waste stimulant, pH 1.1), and rate (e.g., over 95 wt% of Cs was removed after 2 minutes with SAMMS, while only 75 wt% was removed with Prussian blue). The lower affinity, capacity, and rate of Cs and Tl sorption on Prussian blue than those on Cu-FC-EDA-SAMMS were attributed to the molecular pore sizes, which restrict mass transport, and the insoluble Cs abducts of the Prussian blue, which restrict the ability of neighboring binding sites to further bind Cs ions. On the other hand, the large pores of SAMMS not only enable faster diffusion and faster binding chemistry, but they also allow isolation of binding sites so that one Cs binding event does not impact further Cs binding. In addition, iron (Fe) dissolved from insoluble Prussian blue over 10-fold of that from Cu-FC-EDA-SAMMS after 24 hours of contact time, indicating poorer material stability of Prussian blue.« less

Insights into cellulase-lignin non-specific binding revealed by computational redesign of the surface of green fluorescent protein

DOE PAGES

Haarmeyer, Carolyn N.; Smith, Matthew D.; Chundawat, Shishir P. S.; ...

2016-10-17

Biological-mediated conversion of pretreated lignocellulosic biomass to biofuels and biochemicals is a promising avenue towards energy sustainability. However, a critical impediment to the commercialization of cellulosic biofuel production is the high cost of cellulase enzymes needed to deconstruct biomass into fermentable sugars. One major factor driving cost is cellulase adsorption and inactivation in the presence of lignin, yet we currently have a poor understanding of the protein structure-function relationships driving this adsorption. In this work, we have systematically investigated the role of protein surface potential on lignin adsorption using a model monomeric fluorescent protein. We have designed and experimentally characterizedmore » 16 model protein variants spanning the physiological range of net charge (-24 to +16 total charges) and total charge density (0.28 to 0.40 charges per sequence length) typical for natural proteins. Protein designs were expressed, purified, and subjected to in silico and in vitro biophysical measurements to evaluate the relationship between protein surface potential and lignin adsorption properties. The designs were comparable to model fluorescent protein in terms of thermostability and heterologous expression yield, although the majority of the designs unexpectedly formed homodimers. Protein adsorption to lignin was studied at two different temperatures using Quartz Crystal Microbalance with Dissipation Monitoring and a subtractive mass balance assay. We found a weak correlation between protein net charge and protein-binding capacity to lignin. No other single characteristic, including apparent melting temperature and 2nd virial coefficient, showed correlation with lignin binding. Analysis of an unrelated cellulase dataset with mutations localized to a family I carbohydrate-binding module showed a similar correlation between net charge and lignin binding capacity. Altogether, our study provides strategies to identify highly active, low lignin-binding cellulases by either rational design or by computational screening genomic databases.« less

Insights into cellulase-lignin non-specific binding revealed by computational redesign of the surface of green fluorescent protein.

PubMed

Haarmeyer, Carolyn N; Smith, Matthew D; Chundawat, Shishir P S; Sammond, Deanne; Whitehead, Timothy A

2017-04-01

Biological-mediated conversion of pretreated lignocellulosic biomass to biofuels and biochemicals is a promising avenue toward energy sustainability. However, a critical impediment to the commercialization of cellulosic biofuel production is the high cost of cellulase enzymes needed to deconstruct biomass into fermentable sugars. One major factor driving cost is cellulase adsorption and inactivation in the presence of lignin, yet we currently have a poor understanding of the protein structure-function relationships driving this adsorption. In this work, we have systematically investigated the role of protein surface potential on lignin adsorption using a model monomeric fluorescent protein. We have designed and experimentally characterized 16 model protein variants spanning the physiological range of net charge (-24 to +16 total charges) and total charge density (0.28-0.40 charges per sequence length) typical for natural proteins. Protein designs were expressed, purified, and subjected to in silico and in vitro biophysical measurements to evaluate the relationship between protein surface potential and lignin adsorption properties. The designs were comparable to model fluorescent protein in terms of thermostability and heterologous expression yield, although the majority of the designs unexpectedly formed homodimers. Protein adsorption to lignin was studied at two different temperatures using Quartz Crystal Microbalance with Dissipation Monitoring and a subtractive mass balance assay. We found a weak correlation between protein net charge and protein-binding capacity to lignin. No other single characteristic, including apparent melting temperature and 2nd virial coefficient, showed correlation with lignin binding. Analysis of an unrelated cellulase dataset with mutations localized to a family I carbohydrate-binding module showed a similar correlation between net charge and lignin binding capacity. Overall, our study provides strategies to identify highly active, low lignin-binding cellulases by either rational design or by computational screening genomic databases. Biotechnol. Bioeng. 2017;114: 740-750. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Fluorescence Immunofiltration Assay of Brucella Melitensis.

DTIC Science & Technology

1995-01-01

second urease -labelled antibody directed against fluorescein. The assay system is useful for measuring protein, virus and bacteria in aqueous...binding site for the signal-generating urease -labelled antibody, it is a highly fluorescent molecule and has signal-generating capacity of its own

Characterization and localization of arginine vasotocin receptors in the brain and kidney of an amphibian

DOE Office of Scientific and Technical Information (OSTI.GOV)

Boyd, S.K.

1987-01-01

Because arginine vasotocin (AVT) activates male sexual behaviors in the rough-skinned newt (Taricha granulosa), quantitative autoradiography with radiolabeled arginine vasopressin (/sup 3/H-AVP) was used to localize and characterize putative AVT receptors in the brain of this amphibian. Binding of /sup 3/H-AVP to sites within the medial pallium was saturable, specific, reversible, of high affinity and low capacity. These binding sites appear to represent authentic central nervous system receptors for AVT. Furthermore, ligand specificity for the binding sites in this amphibian differs from that reported for AVP binding sites in rat brains. Dense concentrations of specific binding sites were located inmore » the olfactory nerve as it entered the olfactory bulb within the medial pallium, dorsal pallium, and amygdala pars lateralis of the telencephalon, and in the tegmental region of the medulla. Concentrations of binding sites differed significantly among various brain regions. A comparison of male and female newts collected during the breeding season revealed no sexual dimorphism. These areas may represent site(s) of action where AVT elicits sexual behaviors in male T. granulosa.« less

Characterization of the increased binding of acetaldehyde to red blood cells in alcoholics.

PubMed

Hernández-Muñoz, R; Baraona, E; Blacksberg, I; Lieber, C S

1989-10-01

Using equilibrium dialysis, we found that acetaldehyde, at the levels commonly occurring after ethanol ingestion, did not bind detectably to plasma proteins, but there was significant binding to red blood cells, more in alcoholics than in nonalcoholics. The binding to red blood cells was inhibited by pyridoxal phosphate and N-ethylmaleimide, suggesting adduction to amino and thiol groups. Binding kinetics were consistent with at least two sites. The one with the highest affinity for acetaldehyde corresponded to hemoglobin. Its affinity and Bmax were not changed in alcoholics, but these binding sites accounted for only 44% of the sites available in the red blood cells of alcoholics and 80% of those in controls. Moreover, this binding was not inhibited by N-ethylmaleimide. There was no detectable binding to red cell ghosts. Nonprotein binding was then assessed by changes in NADH produced by the addition of protein-free fractions of the cells to an alcohol dehydrogenase system in equilibrium; this revealed a second binder of lower affinity, larger capacity and with sensitivity to both inhibitors. This binding (possibly due to thiazolidine formation with cysteine) was enhanced in alcoholics, whose red blood cell cysteine content was doubled. Levels of red blood cell cysteine and acetaldehyde remained high for 2 weeks after withdrawal. Because of the prolonged persistence after withdrawal, these changes may provide new markers of alcoholism.

Binding of cholesterol and bile acid to hemicelluloses from rice bran.

PubMed

Hu, Guohua; Yu, Wenjian

2013-06-01

The objective of this study was to investigate the possibility of using hemicellulose from rice bran to scavenge cholesterol and bile acid in vitro study. This paper demonstrates that rice bran hemicellulose A (RBHA), rice bran hemicellulose B (RBHB) and rice bran hemicellulose C (RBHC) have the potential for binding cholesterol and bile acid. The quantity of cholesterol and bile acid bound varies from one rice bran fibre to another. As it can be inferred from the results of the study, RBHB was characterized by the highest capacity for cholesterol binding, followed by RBHC and RBHA. Binding of cholesterol and bile acid to rice bran insoluble dietary fibre (RBDF) and cellulose from rice bran was found to be poor. Lignin from rice bran was the least active fraction for binding cholesterol and bile acid. This confirms that the RBHB preparation from defatted rice bran has great potential in food applications, especially in the development of functional foods.

Sulfated glycopeptide nanostructures for multipotent protein activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Sungsoo S.; Fyrner, Timmy; Chen, Feng

Biological systems have evolved to utilize numerous proteins with capacity to bind polysaccharides for the purpose of optimizing their function. A well-known subset of these proteins with binding domains for the highly diverse sulfated polysaccharides are important growth factors involved in biological development and tissue repair. We report here on supramolecular sulfated glycopeptide nanostructures, which display a trisulfated monosaccharide on their surfaces and bind five critical proteins with different polysaccharide-binding domains. Binding does not disrupt the filamentous shape of the nanostructures or their internal β-sheet backbone, but must involve accessible adaptive configurations to interact with such different proteins. The glycopeptidemore » nanostructures amplified signalling of bone morphogenetic protein 2 significantly more than the natural sulfated polysaccharide heparin, and promoted regeneration of bone in the spine with a protein dose that is 100-fold lower than that required in the animal model. These highly bioactive nanostructures may enable many therapies in the future involving proteins.« less

Mechanisms of Zn(II) binded to collagen and its effect on the capacity of eco-friendly Zn-Cr combination tanning system.

PubMed

Cao, Shan; Liu, Bing; Cheng, Baozhen; Lu, Fuping; Wang, Yanping; Li, Yu

2017-01-05

The eco-friendly combination tanning process has been developed to reduce chromium in existing researches, which is based on zinc tanning agents. This can be considered as a less-chrome substitute for current tanning process. To gain deeper understanding of the binding mechanisms of zinc-collagen interaction, which are affected by tanning pH, experiments have been carried out. Analysis in this paper reveals how chemical bonds from the collagen's main function groups combine with zinc. XPS and NIR data was analyzed for further understanding of where the zinc binding sites lie on collagen fibers at different pH. The results indicate that high pH is helpful to amino-binding sites while low pH promotes carboxyl-binding sites on collagen fibers. Furthermore, from the effect of Zinc-chrome combination tanning, we can see that the new method reduces the chromium dosage in tanning process compared to the conventional chrome tanning method. Copyright © 2016 Elsevier B.V. All rights reserved.

Phosphatidylglycerol directs binding and inhibitory action of EIIAGlc protein on the maltose transporter.

PubMed

Bao, Huan; Duong, Franck

2013-08-16

The signal-transducing protein EIIA(Glc) belongs to the phosphoenolpyruvate carbohydrate phosphotransferase system. In its dephosphorylated state, EIIA(Glc) is a negative regulator for several permeases, including the maltose transporter MalFGK2. How EIIA(Glc) is targeted to the membrane, how it interacts with the transporter, and how it inhibits sugar uptake remain obscure. We show here that acidic phospholipids together with the N-terminal tail of EIIA(Glc) are essential for the high affinity binding of the protein to the transporter. Using protein docking prediction and chemical cross-linking, we demonstrate that EIIA(Glc) binds to the MalK dimer, interacting with both the nucleotide-binding and the C-terminal regulatory domains. Dissection of the ATPase cycle reveals that EIIA(Glc) does not affect the binding of ATP but rather inhibits the capacity of MalK to cleave ATP. We propose a mechanism of maltose transport inhibition by this central amphitropic regulatory protein.

Binding the Mammalian High Mobility Group Protein AT-hook 2 to AT-Rich Deoxyoligonucleotides: Enthalpy-Entropy Compensation

PubMed Central

Joynt, Suzanne; Morillo, Victor; Leng, Fenfei

2009-01-01

HMGA2 is a DNA minor-groove binding protein. We previously demonstrated that HMGA2 binds to AT-rich DNA with very high binding affinity where the binding of HMGA2 to poly(dA-dT)2 is enthalpy-driven and to poly(dA)poly(dT) is entropy-driven. This is a typical example of enthalpy-entropy compensation. To further study enthalpy-entropy compensation of HMGA2, we used isothermal-titration-calorimetry to examine the interactions of HMGA2 with two AT-rich DNA hairpins: 5′-CCAAAAAAAAAAAAAAAGCCCCCGCTTTTTTTTTTTTTTTGG-3′ (FL-AT-1) and 5′-CCATATATATATATATAGCCCCCGCTATATATATATATATGG-3′ (FL-AT-2). Surprisingly, we observed an atypical isothermal-titration-calorimetry-binding curve at low-salt aqueous solutions whereby the apparent binding-enthalpy decreased dramatically as the titration approached the end. This unusual behavior can be attributed to the DNA-annealing coupled to the ligand DNA-binding and is eliminated by increasing the salt concentration to ∼200 mM. At this condition, HMGA2 binding to FL-AT-1 is entropy-driven and to FL-AT-2 is enthalpy-driven. Interestingly, the DNA-binding free energies for HMGA2 binding to both hairpins are almost temperature independent; however, the enthalpy-entropy changes are dependent on temperature, which is another aspect of enthalpy-entropy compensation. The heat capacity change for HMGA2 binding to FL-AT-1 and FL-AT-2 are almost identical, indicating that the solvent displacement and charge-charge interaction in the coupled folding/binding processes for both binding reactions are similar. PMID:19450485

Binding biological motion and visual features in working memory.

PubMed

Ding, Xiaowei; Zhao, Yangfan; Wu, Fan; Lu, Xiqian; Gao, Zaifeng; Shen, Mowei

2015-06-01

Working memory mechanisms for binding have been examined extensively in the last decade, yet few studies have explored bindings relating to human biological motion (BM). Human BM is the most salient and biologically significant kinetic information encountered in everyday life and is stored independently from other visual features (e.g., colors). The current study explored 3 critical issues of BM-related binding in working memory: (a) how many BM binding units can be retained in working memory, (b) whether involuntarily object-based binding occurs during BM binding, and (c) whether the maintenance of BM bindings in working memory requires attention above and beyond that needed to maintain the constituent dimensions. We isolated motion signals of human BM from non-BM sources by using point-light displays as to-be-memorized BM and presented the participants colored BM in a change detection task. We found that working memory capacity for BM-color bindings is rather low; only 1 or 2 BM-color bindings could be retained in working memory regardless of the presentation manners (Experiments 1-3). Furthermore, no object-based encoding took place for colored BM stimuli regardless of the processed dimensions (Experiments 4 and 5). Central executive attention contributes to the maintenance of BM-color bindings, yet maintaining BM bindings in working memory did not require more central attention than did maintaining the constituent dimensions in working memory (Experiment 6). Overall, these results suggest that keeping BM bindings in working memory is a fairly resource-demanding process, yet central executive attention does not play a special role in this cross-module binding. (c) 2015 APA, all rights reserved).

The interaction of flavonoid-lysozyme and the relationship between molecular structure of flavonoids and their binding activity to lysozyme.

PubMed

Yang, Ran; Yu, Lanlan; Zeng, Huajin; Liang, Ruiling; Chen, Xiaolan; Qu, Lingbo

2012-11-01

In this work, the interactions of twelve structurally different flavonoids with Lysozyme (Lys) were studied by fluorescence quenching method. The interaction mechanism and binding properties were investigated. It was found that the binding capacities of flavonoids to Lys were highly depend on the number and position of hydrogen, the kind and position of glycosyl. To explore the selectivity of the bindings of flavonoids with Lys, the structure descriptors of the flavonoids were calculated under QSAR software package of Cerius2, the quantitative relationship between the structures of flavonoids and their binding activities to Lys (QSAR) was performed through genetic function approximation (GFA) regression analysis. The QSAR regression equation was K(A) = 37850.460 + 1630.01Dipole +3038.330HD-171.795MR. (r = 0.858, r(CV)(2) = 0.444, F((11,3)) = 7.48), where K(A) is binding constants, Dipole, HD and MR was dipole moment, number of hydrogen-bond donor and molecular refractivity, respectively. The obtained results make us understand better how the molecular structures influencing their binding to protein which may open up new avenues for the design of the most suitable flavonoids derivatives with structure variants.

Binders and Hosts for High-Capacity Lithium-ion Battery Anodes

NASA Astrophysics Data System (ADS)

Dufficy, Martin Kyle

Lithium-ion batteries (LIBs) are universal electrochemical energy storage devices that have revolutionized our mobile society. Nonetheless, societal and technological advances drive consumer demand for LIBs with enhanced electrochemical performance, such as higher charge capacity and longer life, compared to conventional LIBs. One method to enhance LIB performance is to replace graphite, the industry standard anode since commercialization of LIBs in 1991, with high-charge capacity materials. Implementing high-capacity anode materials such as tin, silicon, and manganese vanadates, to LIBs presents challenges; Li-insertion is destructive to anode framework, and increasing capacity increases structural strains that pulverize anode materials and results in a short-cycle life. This thesis reports on various methods to extended the cycle life of high-capacity materials. Most of the work is conducted on nano-sized anode materials to reduce Li and electron transport pathway length (facilitating charge-transfer) and reduce strains from volume expansions (preserving anode structure). The first method involves encapsulating tin particles into a graphene-containing carbon nanofiber (CNF) matrix. The composite-CNF matrix houses tin particles to assume strains from tin-volume expansions and produces favorable surface-electrolyte chemistries for stable charge-discharge cycling. Before tin addition, graphene-containing CNFs are produced and assessed as anode materials for LIBs. Graphene addition to CNFs improves electronic and mechanical properties of CNFs. Furthermore, the 2-D nature of graphene provides Li-binding sites to enhance composite-CNF both first-cycle and high-rate capacities > 150% when compared to CNFs in the absence of graphene. With addition of Sn, we vary loadings and thermal production temperature to elucidate structure-composition relationships of tin and graphene-containing CNF electrodes that lead to increased capacity retention. Of note, electrodes containing ≤ 20 wt% tin result in small tin (metallic and tin oxide) particles (≤ 15 nm) within the composite-CNF matrix, which yield long cycle-lives; large reversible capacities of ˜ 600 mAh g-1 are observed at 0.2-C rates, while capacities of ˜ 400 mAh g-1 (double the capacity of CNFs) are observed after hundreds of cycles at 2-C rates. The second method comprises an approach to enhance the cycle life of silicon anodes. Many researchers believe that Si is the future anode material of LIBs, and Si is capable of providing a much needed boost in overall cell performance. Silicon has the highest known charge capacity at ˜ 3579 mAh g-1, nearly an order of magnitude larger than graphite (372 mAh g-1). In attempt to realize the entire capacity of Si anodes, we use binding agents to prolong cycle life. Binding agents enhance capacity retention via favorable interactions with cell components such as active materials and electrolytes. In this study, we introduce galactomannans (specifically, guar) as viable, inexpensive, biopolymer binders for Si electrodes. In attempt to elucidate the role of the binder in Si electrodes, we study guar-electrode and -electrolyte interactions that lead to electrochemical performance enhancements. We recognize that there are deficiencies in guar-silicon systems, which we address in our following approach. Notably, we develop a guar-derived binder to increase the strength and conductivity of Si-based electrodes by crosslinking guar and carbon black dispersions. The crosslinked binders, in effect, enhance electrode adhesion and hinder electrode cracking by self-healing. This study monitors gelation via rheological methods and assesses effects of crosslinking density on physical and electrochemical properties. Lastly, we consider a vacancy-induced manganese vanadate as high-capacity, high-power anodes for LIBs. Rather than assessing nanoparticles, we tailored molecular structure to enhance electrochemical performances. X-ray diffraction studies enable us to suggest a Li-insertion mechanism, where Li travels through large channels created by defects in the crystal structure. The ensuing manganese vanadate structure produces a stable framework that results in stable cycling of hundreds of cycles.

The uranium from seawater program at PNNL: Overview of marine testing, adsorbent characterization, adsorbent durability, adsorbent toxicity, and deployment studies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gill, Gary A.; Kuo, Li -Jung; Janke, Christopher James

The Pacific Northwest National Laboratory's (PNNL) Marine Science Laboratory (MSL) located along the coast of Washington State is evaluating the performance of uranium adsorption materials being developed for seawater extraction under realistic marine conditions with natural seawater. Two types of exposure systems were employed in this program: flow-through columns for testing of fixed beds of individual fibers and pellets and a recirculating water flume for testing of braided adsorbent material. Testing consists of measurements of the adsorption of uranium and other elements from seawater as a function of time, typically 42 to 56 day exposures, to determine the adsorbent capacitymore » and adsorption rate (kinetics). Analysis of uranium and other trace elements collected by the adsorbents was conducted following strong acid digestion of the adsorbent with 50% aqua regia using either Inductively Coupled Plasma Optical Emission Spectrometry (ICP-OES) or Inductively Coupled Plasma Mass Spectrometer (ICP-MS). The ORNL 38H adsorbent had a 56 day adsorption capacity of 3.30 ± 0.68 g U/ kg adsorbent (normalized to a salinity of 35 psu), a saturation adsorption capacity of 4.89 ± 0.83 g U/kg of adsorbent material (normalized to a salinity of 35 psu) and a half-saturation time of 28 10 days. The AF1 adsorbent material had a 56 day adsorption capacity of 3.9 ± 0.2 g U/kg adsorbent material (normalized to a salinity of 35 psu), a saturation capacity of 5.4 ± 0.2 g U/kg adsorbent material (normalized to a salinity of 35 psu) and a half saturation time of 23 2 days. The ORNL amidoxime-based adsorbent materials are not specific for uranium, but also adsorb other elements from seawater. The major doubly charged cations in seawater (Ca and Mg) account for a majority of the cations adsorbed (61% by mass and 74% by molar percent). For the ORNL AF1 adsorbent material, U is the 4th most abundant element adsorbed by mass and 7th most abundant by molar percentage. Marine testing at Woods Hole Oceanographic Institution with the ORNL AF1 adsorbent produced 15% and 55% higher adsorption capacities than observed at PNNL for column and flume testing, respectively. Variations in competing ions may be the explanation for the regional differences. In addition to marine testing, a number of other efforts are underway to characterize adsorbents and impacts of deployment on the marine environment. Highlights include: Hydrodynamic modelling predicts that a farm of adsorbent materials will likely have minimal effect on ocean currents and removal of uranium and other elements from seawater when densities are < 1800 braids/km 2. A decrease in U adsorption capacity of up to 30% was observed after 42 days of exposure due to biofouling when the ORNL braided adsorbent AI8 was exposed to raw seawater in a flume in the presence of light. An identical raw seawater exposure with no light exposure showed little or no impact to adsorption capacity from biofouling. No toxicity was observed with column effluents of any absorbent materials tested to date. Toxicity could be induced with some non amidoxime-based absorbents only when the ratio of solid absorbent to test media was increased to highly unrealistic levels. Thermodynamic modeling of the seawater-amidoxime adsorbent was performed using the geochemical modeling program PHREEQC. Modeling of the binding of Ca, Mg, Fe, Ni, Cu, U, and V from batch interactions with seawater across a variety of concentrations of the amidoxime binding group reveal that when binding sites are limited (1 x 10 -8 binding sites/kg seawater), vanadium heavily out-competes other ions for the amidoxime sites. In contrast, when binding sites are abundant magnesium and calcium dominate the total percentage of metals bound to the sorbent.« less

The uranium from seawater program at PNNL: Overview of marine testing, adsorbent characterization, adsorbent durability, adsorbent toxicity, and deployment studies

DOE PAGES

Gill, Gary A.; Kuo, Li -Jung; Janke, Christopher James; ...

2016-02-07

The Pacific Northwest National Laboratory's (PNNL) Marine Science Laboratory (MSL) located along the coast of Washington State is evaluating the performance of uranium adsorption materials being developed for seawater extraction under realistic marine conditions with natural seawater. Two types of exposure systems were employed in this program: flow-through columns for testing of fixed beds of individual fibers and pellets and a recirculating water flume for testing of braided adsorbent material. Testing consists of measurements of the adsorption of uranium and other elements from seawater as a function of time, typically 42 to 56 day exposures, to determine the adsorbent capacitymore » and adsorption rate (kinetics). Analysis of uranium and other trace elements collected by the adsorbents was conducted following strong acid digestion of the adsorbent with 50% aqua regia using either Inductively Coupled Plasma Optical Emission Spectrometry (ICP-OES) or Inductively Coupled Plasma Mass Spectrometer (ICP-MS). The ORNL 38H adsorbent had a 56 day adsorption capacity of 3.30 ± 0.68 g U/ kg adsorbent (normalized to a salinity of 35 psu), a saturation adsorption capacity of 4.89 ± 0.83 g U/kg of adsorbent material (normalized to a salinity of 35 psu) and a half-saturation time of 28 10 days. The AF1 adsorbent material had a 56 day adsorption capacity of 3.9 ± 0.2 g U/kg adsorbent material (normalized to a salinity of 35 psu), a saturation capacity of 5.4 ± 0.2 g U/kg adsorbent material (normalized to a salinity of 35 psu) and a half saturation time of 23 2 days. The ORNL amidoxime-based adsorbent materials are not specific for uranium, but also adsorb other elements from seawater. The major doubly charged cations in seawater (Ca and Mg) account for a majority of the cations adsorbed (61% by mass and 74% by molar percent). For the ORNL AF1 adsorbent material, U is the 4th most abundant element adsorbed by mass and 7th most abundant by molar percentage. Marine testing at Woods Hole Oceanographic Institution with the ORNL AF1 adsorbent produced 15% and 55% higher adsorption capacities than observed at PNNL for column and flume testing, respectively. Variations in competing ions may be the explanation for the regional differences. In addition to marine testing, a number of other efforts are underway to characterize adsorbents and impacts of deployment on the marine environment. Highlights include: Hydrodynamic modelling predicts that a farm of adsorbent materials will likely have minimal effect on ocean currents and removal of uranium and other elements from seawater when densities are < 1800 braids/km 2. A decrease in U adsorption capacity of up to 30% was observed after 42 days of exposure due to biofouling when the ORNL braided adsorbent AI8 was exposed to raw seawater in a flume in the presence of light. An identical raw seawater exposure with no light exposure showed little or no impact to adsorption capacity from biofouling. No toxicity was observed with column effluents of any absorbent materials tested to date. Toxicity could be induced with some non amidoxime-based absorbents only when the ratio of solid absorbent to test media was increased to highly unrealistic levels. Thermodynamic modeling of the seawater-amidoxime adsorbent was performed using the geochemical modeling program PHREEQC. Modeling of the binding of Ca, Mg, Fe, Ni, Cu, U, and V from batch interactions with seawater across a variety of concentrations of the amidoxime binding group reveal that when binding sites are limited (1 x 10 -8 binding sites/kg seawater), vanadium heavily out-competes other ions for the amidoxime sites. In contrast, when binding sites are abundant magnesium and calcium dominate the total percentage of metals bound to the sorbent.« less

Nature differences of humic acids fractions induced by extracted sequence as explanatory factors for binding characteristics of heavy metals.

PubMed

Shi, Wenjing; Lü, Changwei; He, Jiang; En, He; Gao, Manshu; Zhao, Boyi; Zhou, Bin; Zhou, Haijun; Liu, Hualin; Zhang, Yu

2018-06-15

The composition and structure of Humic acid (HA) is so heterogeneous that it brings significant barriers to investigate the interaction between HA and heavy metal ions. The isolation of HA with relatively homogeneity is a key to reveal the binding mechanisms between HA and heavy metals. In this work, ten HA fractions (HAs) were obtained by sequential alkali extraction procedure and nature differences of the extracted HAs were considered as explanatory factors for binding characteristics of Cu 2+ , Pb 2+ and Cd 2+ . The results indicate that more large molecular weight (MW) HA subunits, less carboxyl and phenolic group contents, weaker aromaticity and polarity were measured with increasing extractions, inducing weaker binding capacity of HAs. Ligand binding and bi-Langmuir models indicated that the sorption capacity and binding affinity of earlier extracted HAs were higher than the latter ones. The peak area changes at 3427, 1599, and 619 cm -1 pre- and post-adsorption in FTIR spectra suggested carboxyl, phenolic and nitrogen-containing groups were involved in the adsorption process. At the same time, the peak area difference between HAs and HAs-metal (ΔS) of phenolic groups were 8.22-20.50, 6.81-21.11 and 10.66-19.80% for Cu 2+ , Pb 2+ and Cd 2+ , respectively, ΔS of carboxyl groups 6.64-17.03, 8.96-16.82 and 9.45-17.85% for Cu 2+ , Pb 2+ and Cd 2+ , respectively, ΔS of nitrogen-containing groups 0.33-0.48, 0.20-1.38 and 0.31-0.59% for Cu 2+ , Pb 2+ and Cd 2+ , respectively. ΔS of phenolic and carboxyl groups were larger than those of nitrogen-containing groups, implying that these two groups were the predominant binding sites suppliers for metal ions, which were also supported by the results of correlation analysis. This work is helpful to insight the environmental impacts of natural organic matter and the fate of heavy metals in natural environment. Copyright © 2018 Elsevier Inc. All rights reserved.

Bovine Leukemia Virus SU Protein Interacts with Zinc, and Mutations within Two Interacting Regions Differently Affect Viral Fusion and Infectivity In Vivo

PubMed Central

Gatot, Jean-Stéphane; Callebaut, Isabelle; Van Lint, Carine; Demonté, Dominique; Kerkhofs, Pierre; Portetelle, Daniel; Burny, Arsène; Willems, Luc; Kettmann, Richard

2002-01-01

Bovine leukemia virus (BLV) and human T-cell lymphotropic virus type 1 (HTLV-1) belong to the genus of deltaretroviruses. Their entry into the host cell is supposed to be mediated by interactions of the extracellular (SU) envelope glycoproteins with cellular receptors. To gain insight into the mechanisms governing this process, we investigated the ability of SU proteins to interact with specific ligands. In particular, by affinity chromatography, we have shown that BLV SU protein specifically interacted with zinc ions. To identify the protein domains involved in binding, 16 peptides distributed along the sequence were tested. Two of them appeared to be able to interact with zinc. To unravel the role of these SU regions in the biology of the virus, mutations were introduced into the env gene of a BLV molecular clone in order to modify residues potentially interacting with zinc. The fusogenic capacity of envelope mutated within the first zinc-binding region (104 to 123) was completely abolished. Furthermore, the integrity of this domain was also required for in vivo infectivity. In contrast, mutations within the second zinc-binding region (218 to 237) did not hamper the fusogenic capacity; indeed, the syncytia were even larger. In sheep, mutations in region 218 to 237 did not alter infectivity or viral spread. Finally, we demonstrated that the envelope of the related HTLV-1 was also able to bind zinc. Interestingly, zinc ions were found to be associated with the receptor-binding domain (RBD) of Friend murine leukemia virus (Fr-MLV) SU glycoprotein, further supporting their relevance in SU structure. Based on the sequence similarities shared with the Fr-MLV RBD, whose three-dimensional structure has been experimentally determined, we located the BLV zinc-binding peptide 104-123 on the opposite side of the potential receptor-binding surface. This observation supports the hypothesis that zinc ions could mediate interactions of the SU RBD either with the C-terminal part of SU, thereby contributing to the SU structural integrity, or with a partner(s) different from the receptor. PMID:12134000

The effects of methylglyoxal-bis(guanylhydrazone) on spermine binding and transport in liver mitochondria.

PubMed

Toninello, A; Via, L D; Di Noto, V; Mancon, M

1999-12-15

This study evaluated the effect of the anticancer drug methylglyoxal-bis(guanylhydrazone) (MGBG) on the binding of the polyamine spermine to the mitochondrial membrane and its transport into the inner compartment of this organelle. Spermine binding was studied by applying a new thermodynamic treatment of ligand-receptor interactions (Di Noto et al., Macromol Theory Simul 5: 165-181, 1996). Results showed that MGBG inhibited the binding of spermine to the site competent for the first step in polyamine transport; the interaction of spermine with this site, termed S1, also mediates the inhibitory effect of the polyamine on the mitochondrial permeability transition (Dalla Via et al., Biochim Biophys Acta 1284: 247-252, 1996). In the presence of 1 mM MGBG, the binding capacity and affinity of this site were reduced by about 2.6-fold; on the contrary, the binding capacity of the S2 site, which is most likely responsible for the internalization of cytoplasmic proteins (see Dalla Via et al., reference cited above), increased by about 1.3-fold, and its binding affinity remained unaffected. MGBG also inhibited the initial rate of spermine transport in a dose-dependent manner by establishing apparently sigmoidal kinetics. Consequently, the total extent of spermine accumulation inside mitochondria was inhibited. This inhibition in transport seems to reflect a conformational change at the level of the channel protein constituting the polyamine transport system, rather than competitive inhibition at the inner active site of the channel, thereby excluding the possibility that the polyamine and drug use the same transport pathway. Furthermore, it is suggested that, in the presence of MGBG, the S2 site is able to participate in residual spermine transport. MGBG also strongly inhibits deltapH-dependent spermine efflux, resulting in a complete block in the bidirectional flux of the polyamine and its sequestration inside the matrix space. The effects of MGBG on spermine accumulation are consistent with in vivo disruption of the regulator of energy metabolism and replication of the mitochondrial genome.

The Binding Site of Human Adenosine Deaminase for Cd26/Dipeptidyl Peptidase IV

PubMed Central

Richard, Eva; Arredondo-Vega, Francisco X.; Santisteban, Ines; Kelly, Susan J.; Patel, Dhavalkumar D.; Hershfield, Michael S.

2000-01-01

Human, but not murine, adenosine deaminase (ADA) forms a complex with the cell membrane protein CD26/dipeptidyl peptidase IV. CD26-bound ADA has been postulated to regulate extracellular adenosine levels and to modulate the costimulatory function of CD26 on T lymphocytes. Absence of ADA–CD26 binding has been implicated in causing severe combined immunodeficiency due to ADA deficiency. Using human–mouse ADA hybrids and ADA point mutants, we have localized the amino acids critical for CD26 binding to the helical segment 126–143. Arg142 in human ADA and Gln142 in mouse ADA largely determine the capacity to bind CD26. Recombinant human ADA bearing the R142Q mutation had normal catalytic activity per molecule, but markedly impaired binding to a CD26+ ADA-deficient human T cell line. Reduced CD26 binding was also found with ADA from red cells and T cells of a healthy individual whose only expressed ADA has the R142Q mutation. Conversely, ADA with the E217K active site mutation, the only ADA expressed by a severely immunodeficient patient, showed normal CD26 binding. These findings argue that ADA binding to CD26 is not essential for immune function in humans. PMID:11067872

Binding of /sup 3/H-acetylcholine to cholinergic receptors in bovine cerebral arteries

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shimohama, S.; Tsukahara, T.; Taniguchi, T.

Cholinergic receptor sites in bovine cerebral arteries were analyzed using radioligand binding techniques with the cholinergic agonist, /sup 3/H-acetylcholine (ACh), as the ligand. Specific binding of /sup 3/H-ACh to membrane preparations of bovine cerebral arteries was saturable, of two binding sites, with dissociation constant (K/sub D/) values of 0.32 and 23.7 nM, and maximum binding capacity (Bmax) values of 67 and 252 fmol/mg protein, respectively. Specific binding of /sup 3/H-ACh was displaced effectively by muscarinic cholinergic agents and less effectively by nicotinic cholinergic agents. IC/sub 50/ values of cholinergic drugs for /sup 3/H-ACh binding were as follows: atropine, 38.5 nM;more » ACh, 59.8 nM; oxotremorine, 293 nM; scopolamine 474 nM; carbamylcholine, 990 nM. IC/sub 50/ values of nicotinic cholinergic agents such as nicotine, cytisine and ..cap alpha..-bungarotoxin exceeded 50 ..mu..M. Choline acetyltransferase activity was 1.09 nmol/mg protein/hour in the cerebral arteries. These findings suggest that the cholinergic nerves innervate the bovine cerebral arteries and that there are at least two classes of ACh binding sites of different affinities on muscarinic reporters in these arteries. 18 references, 2 figures, 2 tables.« less

[(3)H]8-Ethyl-4-methyl-2-phenyl-(8R)-4,5,7,8-tetrahydro-1H-imidazo[2,1-i]-purin-5-one ([(3)H]PSB-11), a novel high-affinity antagonist radioligand for human A(3) adenosine receptors.

PubMed

Müller, Christa E; Diekmann, Martina; Thorand, Mark; Ozola, Vita

2002-02-11

This study describes the preparation and binding properties of [(3)H]PSB-11, a novel, potent, and selective antagonist radioligand for human A(3) adenosine receptors (ARs). [(3)H]PSB-11 binding to membranes of Chinese hamster ovary (CHO) cells expressing the human A(3) AR was saturable and reversible. Saturation experiments showed that [(3)H]PSB-11 labeled a single class of binding sites with high affinity (K(D)=4.9 nM) and limited capacity (B(max)=3500 fmol/mg of protein). PSB-11 is highly selective versus the other adenosine receptor subtypes. The new radioligand shows an extraordinarily low degree of non-specific binding rendering it a very useful tool for studying the (patho)physiological roles of A(3 )ARs.

Nanoparticles engineered to bind cellular motors for efficient delivery.

PubMed

Dalmau-Mena, Inmaculada; Del Pino, Pablo; Pelaz, Beatriz; Cuesta-Geijo, Miguel Ángel; Galindo, Inmaculada; Moros, María; de la Fuente, Jesús M; Alonso, Covadonga

2018-03-30

Dynein is a cytoskeletal molecular motor protein that transports cellular cargoes along microtubules. Biomimetic synthetic peptides designed to bind dynein have been shown to acquire dynamic properties such as cell accumulation and active intra- and inter-cellular motion through cell-to-cell contacts and projections to distant cells. On the basis of these properties dynein-binding peptides could be used to functionalize nanoparticles for drug delivery applications. Here, we show that gold nanoparticles modified with dynein-binding delivery sequences become mobile, powered by molecular motor proteins. Modified nanoparticles showed dynamic properties, such as travelling the cytosol, crossing intracellular barriers and shuttling the nuclear membrane. Furthermore, nanoparticles were transported from one cell to another through cell-to-cell contacts and quickly spread to distant cells through cell projections. The capacity of these motor-bound nanoparticles to spread to many cells and increasing cellular retention, thus avoiding losses and allowing lower dosage, could make them candidate carriers for drug delivery.

Interaction of berberine with human platelet. alpha. sub 2 adrenoceptors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hui, Ka Kit; Yu, Jun Liang; Chan, Wai Fong A.

1991-01-01

Berberine was found to inhibit competitively the specific binding of ({sup 3}H)-yohimbine. The displacement curve was parallel to those of clonidine, epinephrine, norepinephrine, with the rank order of potency (IC{sub 50}) being clonidine {gt} epinephrine {gt} norepinephrine (14.5 {mu}M) = berberine. Increasing concentrations of berberine from 0.1 {mu}M to 10 {mu}M inhibited ({sup 3}H)-yohimbine binding, shifting the saturation binding curve to the right without decreasing the maximum binding capacity. In platelet cyclic AMP accumulation experiments, berberine at concentrations of 0.1 {mu}M to 0.1 mM inhibited the cAMP accumulation induced by 10 {mu}M prostaglandin E{sub 1} in a dose dependent manner,more » acting as an {alpha}{sub 2} adrenoceptor agonist. In the presence of L-epinephrine, berberine blocked the inhibitory effect of L-epinephrine behaving as an {alpha}{sub 2} adrenoceptor antagonist.« less

Proceedings of the 2010 AFMS Medical Research Symposium. Volume 5. Nursing Track: Abstracts and Presentations

DTIC Science & Technology

2011-03-15

management, toxicology/health risks (e.g., particulates nanomaterials, radiation, etc.), monitoring disease trends , other areas of preventive medicine...will include hematocrit, hemoglobin, mean corpuscle volume, iron, total iron binding capacity, Ferritin , and soluble transferring receptor. The

A Simplified Representation of the Chemical Nature and Reactions of Soil Humus.

ERIC Educational Resources Information Center

Stevenson, F. J.; Olsen, R. A.

1989-01-01

Presented is a comprehensible structural representation of humic substances. A number of important roles of soil organic matter, including contribution to the cation-exchange capacity, binding of pesticides, and formation of complexes with micronutrient cations, are illustrated. (Author/CW)

Biochar Improves Performance of Plants for Mine Soil Revegetation

EPA Science Inventory

Biochar (the solid by-product of pyrolysis of biomass), has the potential to improve plant performance for revegetation of mine soils by improving soil chemistry, fertility, moisture holding capacity and by binding heavy metals. We investigated the effect of gasified conifer sof...

Watermelon consumption improves inflammation and antioxidant capacity in rats fed an atherogenic diet.

PubMed

Hong, Mee Young; Hartig, Nicole; Kaufman, Katy; Hooshmand, Shirin; Figueroa, Arturo; Kern, Mark

2015-03-01

Cardiovascular disease (CVD) is the leading cause of death in the United States. Watermelon, rich in antioxidants and other bioactive components, may be a viable method to improve CVD risk factors through reduced oxidative stress. The purpose of the study was to determine the effects of watermelon powder consumption on lipid profiles, antioxidant capacity, and inflammation in dextran sodium sulfate (DSS)-treated rats fed an atherogenic diet. We hypothesized that watermelon would increase antioxidant capacity and reduce blood lipids and inflammation through modulation of related gene expression. Forty male-weanling (21 days old) Sprague-Dawley rats were divided into 4 groups (10 per group, total N = 40) in a 2 diets (control or 0.33% watermelon) × 2 treatments (with or without DSS) factorial design using an atherogenic diet. Watermelon-fed groups exhibited significantly lower serum triglycerides, total cholesterol, and low-density lipoprotein cholesterol (P< .05). C-reactive protein levels were significantly lower in watermelon-fed rats than the control (P= .001). In addition, oxidative stress as measured by thiobarbituric acid reactive substances was significantly lower in watermelon groups (P= .001). Total antioxidant capacity, superoxide dismutase, and catalase activities were greater in watermelon groups (P< .05). Aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and lactate dehydrogenase were significantly lower in DSS-treated rats when watermelon was consumed (P< .05). Fatty acid synthase, 3-hydroxy-3methyl-glutaryl-CoA reductase, sterol regulatory element-binding protein 1, sterol regulatory element-binding protein 2, and cyclooxygenase-2 gene expression was significantly downregulated in the watermelon group without DSS (P< .05). These findings indicate that watermelon improves risk factors for CVD in rats through better lipid profiles, lower inflammation, and greater antioxidant capacity by altering gene expression for lipid metabolism. Copyright © 2015 Elsevier Inc. All rights reserved.

Anchoring antibodies to membranes using a diphtheria toxin T domain-ZZ fusion protein as a pH sensitive membrane anchor.

PubMed

Nizard, P; Liger, D; Gaillard, C; Gillet, D

1998-08-14

We have constructed a fusion protein, T-ZZ, in which the IgG-Fc binding protein ZZ was fused to the C-terminus of the diphtheria toxin transmembrane domain (T domain). While soluble at neutral pH, T-ZZ retained the capacity of the T domain to bind to phospholipid membranes at acidic pH. Once anchored to the membrane, the ZZ part of the protein was capable of binding mouse monoclonal or rabbit polyclonal IgG. Our results show that the T-ZZ protein can function as a pH sensitive membrane anchor for the linkage of IgG to the membrane of lipid vesicles, adherent and non-adherent cells.

Prevention of chinosol absorption by rubber stoppers used to seal glass vials containing tuberculin PPD Mantoux solutions.

PubMed

Landi, S; Held, H R

1965-01-01

Chinosol (8-quinolinol sulfate), which is employed as an antimicrobial agent in tuberculin PPD solutions used for the Mantoux test, is known to disappear from these solutions after storage. It has been established that the loss of this preservative from tuberculin solutions dispensed in glass vials is caused by the rubber stoppers used to seal the vials. All the rubber stoppers tested absorbed Chinosol very readily.The nature of the binding of Chinosol by a rubber stopper is both chemical (irreversible) and physical (reversible). The capacity to bind Chinosol chemically was determined for 12 types of rubber stopper, and was found to vary from 0 to more than 25% by weight of the stopper. This phenomenon can be attributed mainly to metal ingredients in the stopper. The capacity of stoppers to bind Chinosol physically is expressed quantitatively by a partition coefficient. This was determined for 7 types of rubber stopper and found to be high when compared with the partition coefficients of other preservatives.We have shown that all stoppers presaturated in Chinosol can be equilibrated against a buffered solution containing 0.01% Chinosol. Equilibrated silicone, white, and red oxiglazed stoppers can be used satisfactorily for sealing multi-dose vials of tuberculin PPD containing 0.01% Chinosol.

Prevention of Chinosol absorption by rubber stoppers used to seal glass vials containing tuberculin PPD Mantoux solutions

PubMed Central

Landi, S.; Held, H. R.

1965-01-01

Chinosol (8-quinolinol sulfate), which is employed as an antimicrobial agent in tuberculin PPD solutions used for the Mantoux test, is known to disappear from these solutions after storage. It has been established that the loss of this preservative from tuberculin solutions dispensed in glass vials is caused by the rubber stoppers used to seal the vials. All the rubber stoppers tested absorbed Chinosol very readily. The nature of the binding of Chinosol by a rubber stopper is both chemical (irreversible) and physical (reversible). The capacity to bind Chinosol chemically was determined for 12 types of rubber stopper, and was found to vary from 0 to more than 25% by weight of the stopper. This phenomenon can be attributed mainly to metal ingredients in the stopper. The capacity of stoppers to bind Chinosol physically is expressed quantitatively by a partition coefficient. This was determined for 7 types of rubber stopper and found to be high when compared with the partition coefficients of other preservatives. We have shown that all stoppers presaturated in Chinosol can be equilibrated against a buffered solution containing 0.01% Chinosol. Equilibrated silicone, white, and red oxiglazed stoppers can be used satisfactorily for sealing multi-dose vials of tuberculin PPD containing 0.01% Chinosol. ImagesFIG. 2 PMID:5294923

Highly selective isolation and purification of heme proteins in biological samples using multifunctional magnetic nanospheres.

PubMed

Liu, Yating; Li, Yan; Wei, Yun

2014-12-01

Magnetic particles with suitable surface modification are capable of binding proteins selectively, and magnetic separations have advantages of rapidity, convenience, and high selectivity. In this paper, new magnetic nanoparticles modified with imidazolium ionic liquid (Fe3O4 @SiO2 @ILs) were successfully fabricated. N-Methylimidazolium was immobilized onto silica-coated magnetic nanoparticles via γ-chloropropyl modification as a magnetic nanoadsorbent for heme protein separation. The particle size was about 90 nm without significant aggregation during the preparation process. Hemoglobin as one of heme proteins used in this experiment was compared with other nonheme proteins. It has been found that the magnetic nanoparticles can be used for more rapid, efficient, and specific adsorption of hemoglobin with a binding capacity as high as 5.78 mg/mg. In comparison with other adsorption materials of proteins in the previous reports, Fe3 O4 @SiO2 @ILs magnetic nanoparticles exhibit the excellent performance in isolation of heme proteins with higher binding capacity and selectivity. In addition, a short separation time makes the functionalized nanoparticles suitable for purifying unstable proteins, as well as having other potential applications in a variety of biomedical fields. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Comparison between methods using copper, lanthanum, and colorimetry for the determination of the cation exchange capacity of plant cell walls.

PubMed

Wehr, J Bernhard; Blamey, F Pax C; Menzies, Neal W

2010-04-28

The determination of the cation exchange capacity (CEC) of plant cell walls is important for many physiological studies. We describe the determination of cell wall CEC by cation binding, using either copper (Cu) or lanthanum (La) ions, and by colorimetry. Both cations are strongly bound by cell walls, permitting fast and reproducible determinations of the CEC of small samples. However, the dye binding methods using two cationic dyes, Methylene Blue and Toluidine Blue, overestimated the CEC several-fold. Column and centrifugation methods are proposed for CEC determination by Cu or La binding; both provide similar results. The column method involves packing plant material (2-10 mg dry mass) in a chromatography column (10 mL) and percolating with 20 bed volumes of 1 mM La or Cu solution, followed by washing with deionized water. The centrifugation method uses a suspension of plant material (1-2 mL) that is centrifuged, and the pellet is mixed three times with 10 pellet volumes of 1 mM La or Cu solution followed by centrifugation and final washing with deionized water. In both methods the amount of La or Cu bound to the material was determined by spectroscopic methods.

Active-State Model of a Dopamine D2 Receptor - Gαi Complex Stabilized by Aripiprazole-Type Partial Agonists

PubMed Central

Kling, Ralf C.; Tschammer, Nuska; Lanig, Harald; Clark, Timothy; Gmeiner, Peter

2014-01-01

Partial agonists exhibit a submaximal capacity to enhance the coupling of one receptor to an intracellular binding partner. Although a multitude of studies have reported different ligand-specific conformations for a given receptor, little is known about the mechanism by which different receptor conformations are connected to the capacity to activate the coupling to G-proteins. We have now performed molecular-dynamics simulations employing our recently described active-state homology model of the dopamine D2 receptor-Gαi protein-complex coupled to the partial agonists aripiprazole and FAUC350, in order to understand the structural determinants of partial agonism better. We have compared our findings with our model of the D2R-Gαi-complex in the presence of the full agonist dopamine. The two partial agonists are capable of inducing different conformations of important structural motifs, including the extracellular loop regions, the binding pocket and, in particular, intracellular G-protein-binding domains. As G-protein-coupling to certain intracellular epitopes of the receptor is considered the key step of allosterically triggered nucleotide-exchange, it is tempting to assume that impaired coupling between the receptor and the G-protein caused by distinct ligand-specific conformations is a major determinant of partial agonist efficacy. PMID:24932547

Efficient synthesis of molecularly imprinted polymers with bio-recognition sites for the selective separation of bovine hemoglobin.

PubMed

Zhang, Zulei; Li, Lei

2018-06-01

We developed a facile approach to the construction of bio-recognition sites in silica nanoparticles for efficient separation of bovine hemoglobin based on amino-functionalized silica nanoparticles grafting by 3-aminopropyltriethoxylsilane providing hydrogen bonds with bovine hemoglobin through surface molecularly imprinting technology. The resulting amino-functionalized silica surface molecularly imprinted polymers were characterized using scanning electron microscope, transmission electronic microscopy, Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, and thermogravimetric analysis. Results showed that the as-synthesized imprinted polymers exhibited spherical morphology and favorable thermal stability. The binding adsorption experiments showed that the imprinted polymers can reach equilibrium within 1 h. The Langmuir isotherm and pseudo-second-order kinetic model fitted the adsorption data well. Meanwhile, the imprinted polymers possessed a maximum binding capacity up to 90.3 mg/g and highly selectivity for the recognition of bovine hemoglobin. Moreover, such high binding capacity and selectivity retained after eight cycles, indicating the good stability and reusability of the imprinted polymers. Finally, successful application in the selective recognition of bovine hemoglobin from a real bovine blood sample indicated that the imprinted polymers displayed great potentials in efficient purification and separation of target proteins. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The Periplasmic Cyclodextrin Binding Protein CymE from Klebsiella oxytoca and Its Role in Maltodextrin and Cyclodextrin Transport

PubMed Central

Pajatsch, Markus; Gerhart, Maria; Peist, Ralf; Horlacher, Reinhold; Boos, Winfried; Böck, August

1998-01-01

Klebsiella oxytoca M5a1 has the capacity to transport and to metabolize α-, β- and γ-cyclodextrins. Cyclodextrin transport is mediated by the products of the cymE, cymF, cymG, cymD, and cymA genes, which are functionally homologous to the malE, malF, malG, malK, and lamB gene products of Escherichia coli. CymE, which is the periplasmic binding protein, has been overproduced and purified. By substrate-induced fluorescence quenching, the binding of ligands was analyzed. CymE bound α-cyclodextrin, β-cyclodextrin, and γ-cyclodextrin, with dissociation constants (Kd) of 0.02, 0.14 and 0.30 μM, respectively, and linear maltoheptaose, with a Kd of 70 μM. In transport experiments, α-cyclodextrin was taken up by the cym system of K. oxytoca three to five times less efficiently than maltohexaose by the E. coli maltose system. Besides α-cyclodextrin, maltohexaose was also taken up by the K. oxytoca cym system, but because of the inability of maltodextrins to induce the cym system, growth of E. coli mal mutants on linear maltodextrin was not observed when the cells harbored only the cym uptake system. Strains which gained this capacity by mutation could easily be selected, however. PMID:9573146

Analyte induced water adsorbability in gas phase biosensors: the influence of ethinylestradiol on the water binding protein capacity.

PubMed

Snopok, Borys; Kruglenko, Ivanna

2015-05-07

An ultra-sensitive gas phase biosensor/tracer/bio-sniffer is an emerging technology platform designed to provide real-time information on air-borne analytes, or those in liquids, through classical headspace analysis. The desired bio-sniffer measures gaseous 17α- ethinylestradiol (ETED) as frequency changes on a quartz crystal microbalance (QCM), which is a result of the interactions of liquid sample components in the headspace (ETED and water) with a biorecognition layer. The latter was constructed by immobilization of polyclonal antiserum against a phenolic A-ring of estrogenic receptors through protein A. The QCM response exhibited stretched exponential kinetics of negative frequency shifts with reversible and "irreversible" components of mass uptake onto the sensor surface in static headspace conditions when exposed to water solutions of ETED over the sensor working range, from 10(-10) to 10(-17) g L(-1). It was shown that the variations in the QCM response characteristics are due to the change of the water-binding capacity of the sensing layer induced by protein transformations initiated by the binding of ETED molecules. This result is well correlated with the natural physiological function of estrogens in controlling the homeostasis of body fluids in living beings.

Kafirin adsorption on ion-exchange resins: isotherm and kinetic studies.

PubMed

Kumar, Prashant; Lau, Pei Wen; Kale, Sandeep; Johnson, Stuart; Pareek, Vishnu; Utikar, Ranjeet; Lali, Arvind

2014-08-22

Kafirin is a natural, hydrophobic and celiac safe prolamin protein obtained from sorghum seeds. Today kafirin is found to be useful in designing delayed delivery systems and coatings of pharmaceuticals and nutraceuticals where its purity is important and this can be obtained by adsorptive chromatography. This study is the first scientific insight into the isotherm and kinetic studies of kafirin adsorption on anion- and cation-exchange resins for practical applications in preparative scale chromatography. Adsorption isotherms of kafirin were determined for five anion- and two cation-exchange resins in batch systems. Isotherm parameters such as maximum binding capacity and dissociation constant were determined from Langmuir isotherm, and adsorptive capacity and affinity constant from Freundlich isotherm. Langmuir isotherm was found to fit the adsorption equilibrium data well. Batch uptake kinetics for kafirin adsorption on these resins was also carried out and critical parameters including the diffusion coefficient, film mass transfer coefficient, and Biot number for film-pore diffusion model were calculated. Both the isotherm and the kinetic parameters were considered for selection of appropriate resin for kafirin purification. UNOsphere Q (78.26 mg/ml) and Toyopearl SP-650M (57.4 mg/ml) were found to offer better kafirin binding capacities and interaction strength with excellent uptake kinetics under moderate operating conditions. With these adsorbents, film diffusion resistance was found to be major governing factor for adsorption (Bi<10 and δ<1). Based on designer objective function, UNOsphere Q was found be best adsorbent for binding of kafirin. The data presented is valuable for designing large scale preparative adsorptive chromatographic kafirin purification systems. Copyright © 2014 Elsevier B.V. All rights reserved.

Enhancing Adsorption Capacity while Maintaining Specific Recognition Performance of Mesoporous Silica: A Novel Imprinting Strategy with Amphiphilic Ionic Liquid as Surfactant.

PubMed

Ding, Shichao; Li, Zhiling; Cheng, Yuan; Du, Chunbao; Gao, Junfeng; Zhang, Yong-Wei; Zhang, Nan; Li, Zhaotong; Chang, Ninghui; Hu, Xiaoling

2018-06-21

In order to facilitate the broad applications of molecular recognition materials in biomedical areas, it is critical to enhance their adsorption capacity while maintaining their excellent recognition performance. In this work, we designed and synthesized well-defined peptide-imprinted mesoporous silica (PIMS) for specific recognition of an immunostimulating hexapeptide from human casein (IHHC) by using amphiphilic ionic liquid as the surfactant to anchor IHHC via a combination of one step sol-gel method and docking oriented imprinting approach. Thereinto, theoretical calculation was employed to reveal the multiple binding interactions and dual-template configuration between amphiphilic ionic liquid and IHHC. The fabricated PIMS was characterized and an in-depth analysis of specific recognition mechanism was conducted. Results revealed that both adsorption and recognition capabilities of PIMS far exceeded that of the NIMS's. More significantly, the PIMS exhibited a superior binding capacity (60.5 mg g-1), which could increase 18.9% than the previous work. The corresponding imprinting factor and selectivity coefficient could reach up to 4.51 and 3.30, respectively. The PIMS also possessed lickety-split kinetic binding for IHHC, which the equilibrium time was only 10 min. All of these merits were due to the high surface area and the synergistic effect of multiple interactions (including hydrogen bonding, π-π stacking, ion-ion electrostatic interactions and van der Waals interactions, etc.) between PIMS and IHHC in imprinted sites. The present work suggests the potential application of PIMS for large-scale and high-effective separation of IHHC, which may lead to their broad applications in drug/gene deliver, biosensors, catalyst and so on. © 2018 IOP Publishing Ltd.

Modulation of thyroid hormone receptors by non-thyroidal stimuli

DOE Office of Scientific and Technical Information (OSTI.GOV)

ErkenBrack, D.E.; Clemons, G.K.

1988-01-01

The ability of non-thyroidal stimuli to affect the binding affinity and capacity of solubilized nuclear receptors for thyroid hormones was studied in a normal homeostatic system (erythropoiesis) and a pathobiologic one (lung-ozone interaction). No significant effects on affinity were found, as Kd control values for receptors derived from rat bone marrow averaged 57 (+/- 28) pM while experimental (hypoxic) values averaged 89 (+/- 55) pM. Kd control values in rat lung were found to average 142 (+/- 22) pM while average values derived from experimental protocols with ozone and methimazole were 267 (+/- 44) pM and 161 (+/- 35) pMmore » respectively. Finally, Kd control values for receptors derived from cultured MEL cells averaged 19 (+/- 2.6) pM while experimental values during exposure to DMSO or IGF1 were 23 (+/- 3.6) pM and 26 (+/- 11) pM respectively. In contrast, binding capacity (expressed as fmoles of hormone bound per unit protein of solubilized receptor) was markedly perturbed in several tissues by various agents: ozone effects on lung were shown by an average control value of 3.3 (+/- 0.4) as opposed to an experimental average of 28 (+/- 1.9); and hypoxia effects on erythroid tissue were displayed by an average control value of 0.7 (+/- 0.07) as opposed to the experimental figure of 1.8 (+/- 0.03). In cultured MEL cells, binding capacity was seen to be increased from control values of 388 (+/- 15) sites/cell to 1243 (+/- 142) sites/cell after DMSO exposure and 2002 (+/- 10) sites/cell after IGF1 exposure. Parallel experiments done with receptors derived from rat liver yielded values similar to those reported by other investigators and were unaffected by the experimental agents.« less

Antigenic Determinants of the Bilobal Cockroach Allergen Bla g 2.

PubMed

Woodfolk, Judith A; Glesner, Jill; Wright, Paul W; Kepley, Christopher L; Li, Mi; Himly, Martin; Muehling, Lyndsey M; Gustchina, Alla; Wlodawer, Alexander; Chapman, Martin D; Pomés, Anna

2016-01-29

Bla g 2 is a major indoor cockroach allergen associated with the development of asthma. Antigenic determinants on Bla g 2 were analyzed by mutagenesis based on the structure of the allergen alone and in complex with monoclonal antibodies that interfere with IgE antibody binding. The structural analysis revealed mechanisms of allergen-antibody recognition through cation-π interactions. Single and multiple Bla g 2 mutants were expressed in Pichia pastoris and purified. The triple mutant K132A/K251A/F162Y showed an ∼100-fold reduced capacity to bind IgE, while preserving the native molecular fold, as proven by x-ray crystallography. This mutant was still able to induce mast cell release. T-cell responses were assessed by analyzing Th1/Th2 cytokine production and the CD4(+) T-cell phenotype in peripheral blood mononuclear cell cultures. Although T-cell activating capacity was similar for the KKF mutant and Bla g 2 based on CD25 expression, the KKF mutant was a weaker inducer of the Th2 cytokine IL-13. Furthermore, this mutant induced IL-10 from a non-T-cell source at higher levels that those induced by Bla g 2. Our findings demonstrate that a rational design of site-directed mutagenesis was effective in producing a mutant with only 3 amino acid substitutions that maintained the same fold as wild type Bla g 2. These residues, which were involved in IgE antibody binding, endowed Bla g 2 with a T-cell modulatory capacity. The antigenic analysis of Bla g 2 will be useful for the subsequent development of recombinant allergen vaccines. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Synthesis and characterization of low viscosity carbon dioxide binding organic liquids for flue gas clean up

DOE Office of Scientific and Technical Information (OSTI.GOV)

Koech, Phillip K.; Malhotra, Deepika; Heldebrant, David J.

2015-01-01

Climate change is partly attributed to global anthropogenic carbon dioxide (CO2) emission to the atmosphere. These environmental effects can be mitigated by CO2 capture, utilization and storage. Alkanolamine solvents, such as monoethanolamine (MEA), which bind CO2 as carbamates or bicarbonate salts are used for CO2 capture in niche applications. These solvents consist of approximately 30 wt% of MEA in water, exhibiting a low, CO2-rich viscosity, fast kinetics and favorable thermodynamics. However, these solvents have low CO2 capacity and high heat capacity of water, resulting in prohibitively high costs of thermal solvent regeneration. Effective capture of the enormous amounts of CO2more » produced by coal-fired plants requires a material with high CO2 capacity and low regeneration energy requirements. To this end, several water-lean transformational solvents systems have been developed in order to reduce these energy penalties. These technologies include nano-material organic hybrids (NOHMs), task-specific, protic and conventional ionic liquids, phase change solvents. As part of an ongoing program in our group, we have developed new water lean transformational solvents known as CO2 binding organic liquids (CO2BOLs) which have the potential to be energy efficient CO2 capture solvents. These solvents, also known as switchable ionic liquids meaning, are organic solvents that can reversibly transform from non- ionic to ionic form and back. The zwitterionic state in these liquids is formed when low polarity non-ionic alkanolguanidines or alkanolamidines react with CO2 or SO2 to form ionic liquids with high polarity. These polar ionic liquids can be thermally converted to the less polar non-ionic solvent by releasing CO2.« less

Development, characterization, and in vitro evaluation of phosphatidylcholine-sodium cholate-based nanoparticles for siRNA delivery to MCF-7 human breast cancer cells

NASA Astrophysics Data System (ADS)

Pérez, Sebastián Ezequiel; Gándola, Yamila; Carlucci, Adriana Mónica; González, Lorena

2015-03-01

Phosphatidylcholine-sodium cholate (SC)-based nanoparticles were designed, characterized, and evaluated as plausible oligonucleotides delivery systems. For this purpose, formulation of the systems was optimized to obtain low cytotoxic vehicles with high siRNA-loading capacity and acceptable transfection ability. Mixtures of soybean phosphatidylcholine (SPC) and SC were prepared at different molar ratios with 2 % w/v total concentration; distilled water and two different buffers were used as dispersion medium. Nanoparticles below 150 nm were observed showing spherical shape which turned smaller in diameter as the SC molar proportion increased, accounting for small unilamellar vesicles when low proportions of SC were present in the formulation, but clear mixed micellar solutions at higher SC percentages. Macroscopic characteristics along with physico-chemical parameters values supported the presence of these types of structures. SYBR green displacement assays demonstrated an important oligonucleotide binding that increased as bile salt relative content got higher. Within the same molar ratio, nanoparticles showed the following binding efficiency order: pH 7.4 > pH 5.0 > distilled water. siRNA-loading capacity assays confirmed the higher siRNA binding by the mixed micelles containing higher SC proportion; moreover, the complexes formed were smaller as the SC:SPC ratio increased. Considering cytotoxicity and siRNA-loading capacity, 1:2 and 1:4 SPC:SC formulations were selected for further biological assays. Nanoparticles prepared in any of the three media were able to induce dsRNA uptake and efficiently transfect RNA for gene silencing, for the compositions prepared in buffer pH 5.0 being the most versatile.

New insights into the structure-function relationships and therapeutic applications of cholera-like enterotoxins.

PubMed

Hirst, Timothy R; Fraser, Sylvia; Soriani, Marco; Aman, A Tholib; de, Haan Lolke; Hearn, Arron; Merritt, Ethan

2002-02-01

Cholera toxin and E. coli heat-labile enterotoxin are structurally homologous proteins comprised of an enzymatically active A-subunit and five B-subunits that bind with high affinity to GM1-ganglioside receptors found on the surface of mammalian cells. The B-subunits have long been thought of simply as trafficking vehicles that trigger entry and subsequent delivery of the 'toxic' A-subunit into cells. Indeed, such is the capacity of the B-subunits to enter cells, that they have been developed as generic carriers for attachment and delivery of a variety of peptides into mammalian cells. However, the B-subunits also appear to possess discrete 'signalling functions', that induce both transcription factor and cell activation. These are thought to be directly responsible for the potent immunomodulatory properties of the B-subunits, and have resulted in their use as adjuvants and as agents to suppress inflammatory immune disorders. The relationship between the signalling properties of the B-subunits and their capacity to act as trafficking vehicles has remained unclear. In an effort to understand the structural requirements for these two functions, a set of mutant B-subunits, with amino acid substitutions at position His-57, have been generated and studied. Importantly, such mutant B-subunits retain an ability to bind with high affinity to GM1 and to traffic into cells, but have entirely lost their capacity to activate immune cell populations. Thus, while binding via GM1 appears to be sufficient to trigger cellular uptake it is not sufficient to activate signal transduction. The His-57 region is therefore speculated to be actively engaged in triggering signalling events, possibly via cognate interaction with other cell surface molecules.

Antigenic Determinants of the Bilobal Cockroach Allergen Bla g 2*

PubMed Central

Woodfolk, Judith A.; Glesner, Jill; Wright, Paul W.; Kepley, Christopher L.; Li, Mi; Himly, Martin; Muehling, Lyndsey M.; Gustchina, Alla; Wlodawer, Alexander; Chapman, Martin D.; Pomés, Anna

2016-01-01

Bla g 2 is a major indoor cockroach allergen associated with the development of asthma. Antigenic determinants on Bla g 2 were analyzed by mutagenesis based on the structure of the allergen alone and in complex with monoclonal antibodies that interfere with IgE antibody binding. The structural analysis revealed mechanisms of allergen-antibody recognition through cation-π interactions. Single and multiple Bla g 2 mutants were expressed in Pichia pastoris and purified. The triple mutant K132A/K251A/F162Y showed an ∼100-fold reduced capacity to bind IgE, while preserving the native molecular fold, as proven by x-ray crystallography. This mutant was still able to induce mast cell release. T-cell responses were assessed by analyzing Th1/Th2 cytokine production and the CD4+ T-cell phenotype in peripheral blood mononuclear cell cultures. Although T-cell activating capacity was similar for the KKF mutant and Bla g 2 based on CD25 expression, the KKF mutant was a weaker inducer of the Th2 cytokine IL-13. Furthermore, this mutant induced IL-10 from a non-T-cell source at higher levels that those induced by Bla g 2. Our findings demonstrate that a rational design of site-directed mutagenesis was effective in producing a mutant with only 3 amino acid substitutions that maintained the same fold as wild type Bla g 2. These residues, which were involved in IgE antibody binding, endowed Bla g 2 with a T-cell modulatory capacity. The antigenic analysis of Bla g 2 will be useful for the subsequent development of recombinant allergen vaccines. PMID:26644466

Comprehensive approach to study complement C4 in systemic lupus erythematosus: Gene polymorphisms, protein levels and functional activity.

PubMed

Tsang-A-Sjoe, M W P; Bultink, I E M; Korswagen, L A; van der Horst, A; Rensink, I; de Boer, M; Hamann, D; Voskuyl, A E; Wouters, D

2017-12-01

Genetic variation of the genes encoding complement component C4 is strongly associated with systemic lupus erythematosus (SLE), a chronic multi-organ auto-immune disease. This study examined C4 and its isotypes on a genetic, protein, and functional level in 140 SLE patients and 104 healthy controls. Gene copy number (GCN) variation, silencing CT-insertion, and the retroviral HERV-K(C4) insertion) were analyzed with multiplex ligation-dependent probe amplification. Increased susceptibility to SLE was found for low GCN (≪2) of C4A. Serositis was the only clinical manifestation associated with low C4A GCN. One additional novel silencing mutation in the C4A gene was found by Sanger sequencing. This mutation causes a premature stop codon in exon 11. Protein concentrations of C4 isoforms C4A and C4B were determined with ELISA and were significantly lower in SLE patients compared to healthy controls. To study C4 isotypes on a functional level, a new C4 assay was developed, which distinguishes C4A from C4B by its binding capacity to amino or hydroxyl groups, respectively. This assay showed high correlation with ELISA and detected crossing over of Rodgers and Chido antigens in 3.2% (8/244) of individuals. The binding capacity of available C4 to its substrates was unaffected in SLE. Our study provides, for the first time, a complete overview of C4 in SLE from genetic variation to binding capacity using a novel test. As this test detects crossing over of Rodgers and Chido antigens, it will allow for more accurate measurement of C4 in future studies. Copyright © 2017 Elsevier Ltd. All rights reserved.

Frequency of Hereditary Hemochromatosis (HFE) Gene Mutations in Egyptian Beta Thalassemia Patients and its Relation to Iron Overload.

PubMed

Enein, Azza Aboul; El Dessouky, Nermine A; Mohamed, Khalda S; Botros, Shahira K A; Abd El Gawad, Mona F; Hamdy, Mona; Dyaa, Nehal

2016-06-15

This study aimed to detect the most common HFE gene mutations (C282Y, H63D, and S56C) in Egyptian beta thalassemia major patients and its relation to their iron status. The study included 50 beta thalassemia major patients and 30 age and sex matched healthy persons as a control group. Serum ferritin, serum iron and TIBC level were measured. Detection of the three HFE gene mutations (C282Y, H63D and S65C) was done by PCR-RFLP analysis. Confirmation of positive cases for the mutations was done by sequencing. Neither homozygote nor carrier status for the C282Y or S65C alleles was found. The H63D heterozygous state was detected in 5/50 (10%) thalassemic patients and in 1/30 (3.3%) controls with no statistically significant difference between patients and control groups (p = 0.22). Significantly higher levels of the serum ferritin and serum iron in patients with this mutation (p = 001). Our results suggest that there is an association between H63D mutation and the severity of iron overload in thalassemic patients.

Tribo-mechanical and electrical properties of boron-containing coatings

NASA Astrophysics Data System (ADS)

Qian, Jincheng

The development of new hard protective coatings with advanced performance is very important for progress in a variety of scientific and industrial fields. Application of hard protective coatings can significantly improve the performance of parts and components, extend their service life, and save energy in many industrial applications including aerospace, automotive, manufacturing, and other industries. In addition, the multifunctionality of protective coatings is also required in many other application fields such as optics, microelectronics, biomedical, magnetic storage media, etc. Therefore, protective coatings with enhanced tribo-mechanical and corrosion properties as well as other functions are in demand. The coating characteristics can be adjusted by controlling the microstructure at different scales. For example, films with nanostructures, such as superlattice, nanocolumn, and nanocomposite systems, exhibit distinctive characteristics compared to single-phase materials. They show superior tribo-mechanical properties due to the presence of strong interfaces, and different functions can be achieved due to the multi-phase characteristics. Boron-containing materials with their excellent mechanical properties and interesting electronic characteristics are good candidates for functional hard protective coatings. For instance, cubic boron nitride (c-BN), boron carbide (B1-xCx), and titanium diboride (TiB 2) are well known for their high hardness, high thermal stability, and high chemical inertness. An interesting example is the boron carbon nitride (BCN) compound that possesses many attractive properties because its structure is similar to that of carbon (graphite and diamond) and of boron nitride (BN in hexagonal and cubic phases). The main goal of this work is to further develop the family of Boron-containing films including B1-xCx, Ti-B-C, and BCN films fabricated by magnetron sputtering, and to enhance their performance by controlling their microstructure on the nanoscale. Their tribo-mechanical, corrosion, and electrical properties are studied in relation to the composition and microstructure, aiming at enhancing their performance for multi-functional protective coating applications via microstructural design. First, B1-xCx (0 < x < 1) films with tailored tribo-mechanical properties were deposited by magnetron sputtering using one graphite and two boron targets. The hardness of the B1-xC x films was found to reach 25 GPa both for boron-rich and carbon-rich films, and the friction coefficient and wear rate can be adjusted from 0.66 to 0.13 and from 6.4x10-5 mm3/Nm to 1.3x10 -7 mm3/Nm, respectively, by changing the carbon content from 19 to 76 at.%. The hardness variation is closely related to the microstructure, and the low friction and wear rate of the B0.24C0.76 film are due to the high portion of an amorphous carbon phase. Moreover, application of the B0.81C0.19 film improves the corrosion resistance of the M2 steel substrate significantly, indicated by the decrease of the corrosion current by almost four orders of magnitude. Based on the optimization of the B1-xCx films, nanostructured Ti-B-C films with different compositions were deposited by adding titanium by simultaneously sputtering a titanium diboride target. We found that the film microstructure features TiB2 nanocrystallites embedded in an amorphous boron carbide matrix. The film hardness varies from 33 to 42 GPa with different titanium contents, which is related to the changes in microstructure, namely, the size and concentration of the TiB2 nanocrystallites. The friction coefficient and wear rate are in the ranges of 0.37-0.73 and of 3.3x10-6-5.7x10-5 mm3/Nm, respectively, which are affected by the mechanical properties and the surface chemical states of the films. By applying the Ti-B-C films, the corrosion resistance of the M2 steel substrate is significantly enhanced as documented by a reduction of the corrosion current density by two orders of magnitude. BCN films were synthesized by magnetron sputtering using a single B 4C target in an N2: Ar gas mixture. The BCN films exhibit an amorphous structure and contain a mixture of B-C, B-N, and C-N bonds. The films show p-type conductivity with an optical band gap of 1.0 eV. Subsequently, ZnO nanorods were grown on the BCN films using hydrothermal synthesis to form BCN/ZnO nanorods p-n heterojunctions. The performance of the junctions is evaluated by the I-V characterization, which shows a rectification behavior with a rectification ratio of 1500 at the bias voltages of +/-5 V.

Naturally occurring mutations in the human 5-lipoxygenase gene promoter that modify transcription factor binding and reporter gene transcription.

PubMed Central

In, K H; Asano, K; Beier, D; Grobholz, J; Finn, P W; Silverman, E K; Silverman, E S; Collins, T; Fischer, A R; Keith, T P; Serino, K; Kim, S W; De Sanctis, G T; Yandava, C; Pillari, A; Rubin, P; Kemp, J; Israel, E; Busse, W; Ledford, D; Murray, J J; Segal, A; Tinkleman, D; Drazen, J M

1997-01-01

Five lipoxygenase (5-LO) is the first committed enzyme in the metabolic pathway leading to the synthesis of the leukotrienes. We examined genomic DNA isolated from 25 normal subjects and 31 patients with asthma (6 of whom had aspirin-sensitive asthma) for mutations in the known transcription factor binding regions and the protein encoding region of the 5-LO gene. A family of mutations in the G + C-rich transcription factor binding region was identified consisting of the deletion of one, deletion of two, or addition of one zinc finger (Sp1/Egr-1) binding sites in the region 176 to 147 bp upstream from the ATG translation start site where there are normally 5 Sp1 binding motifs in tandem. Reporter gene activity directed by any of the mutant forms of the transcription factor binding region was significantly (P < 0.05) less effective than the activity driven by the wild type transcription factor binding region. Electrophoretic mobility shift assays (EMSAs) demonstrated the capacity of wild type and mutant transcription factor binding regions to bind nuclear extracts from human umbilical vein endothelial cells (HUVECs). These data are consistent with a family of mutations in the 5-LO gene that can modify reporter gene transcription possibly through differences in Sp1 and Egr-1 transactivation. PMID:9062372

Naturally occurring mutations in the human 5-lipoxygenase gene promoter that modify transcription factor binding and reporter gene transcription.

PubMed

In, K H; Asano, K; Beier, D; Grobholz, J; Finn, P W; Silverman, E K; Silverman, E S; Collins, T; Fischer, A R; Keith, T P; Serino, K; Kim, S W; De Sanctis, G T; Yandava, C; Pillari, A; Rubin, P; Kemp, J; Israel, E; Busse, W; Ledford, D; Murray, J J; Segal, A; Tinkleman, D; Drazen, J M

1997-03-01

Five lipoxygenase (5-LO) is the first committed enzyme in the metabolic pathway leading to the synthesis of the leukotrienes. We examined genomic DNA isolated from 25 normal subjects and 31 patients with asthma (6 of whom had aspirin-sensitive asthma) for mutations in the known transcription factor binding regions and the protein encoding region of the 5-LO gene. A family of mutations in the G + C-rich transcription factor binding region was identified consisting of the deletion of one, deletion of two, or addition of one zinc finger (Sp1/Egr-1) binding sites in the region 176 to 147 bp upstream from the ATG translation start site where there are normally 5 Sp1 binding motifs in tandem. Reporter gene activity directed by any of the mutant forms of the transcription factor binding region was significantly (P < 0.05) less effective than the activity driven by the wild type transcription factor binding region. Electrophoretic mobility shift assays (EMSAs) demonstrated the capacity of wild type and mutant transcription factor binding regions to bind nuclear extracts from human umbilical vein endothelial cells (HUVECs). These data are consistent with a family of mutations in the 5-LO gene that can modify reporter gene transcription possibly through differences in Sp1 and Egr-1 transactivation.

The intervening domain from MeCP2 enhances the DNA affinity of the methyl binding domain and provides an independent DNA interaction site.

PubMed

Claveria-Gimeno, Rafael; Lanuza, Pilar M; Morales-Chueca, Ignacio; Jorge-Torres, Olga C; Vega, Sonia; Abian, Olga; Esteller, Manel; Velazquez-Campoy, Adrian

2017-01-31

Methyl-CpG binding protein 2 (MeCP2) preferentially interacts with methylated DNA and it is involved in epigenetic regulation and chromatin remodelling. Mutations in MeCP2 are linked to Rett syndrome, the leading cause of intellectual retardation in girls and causing mental, motor and growth impairment. Unstructured regions in MeCP2 provide the plasticity for establishing interactions with multiple binding partners. We present a biophysical characterization of the methyl binding domain (MBD) from MeCP2 reporting the contribution of flanking domains to its structural stability and dsDNA interaction. The flanking disordered intervening domain (ID) increased the structural stability of MBD, modified its dsDNA binding profile from an entropically-driven moderate-affinity binding to an overwhelmingly enthalpically-driven high-affinity binding. Additionally, ID provided an additional site for simultaneously and autonomously binding an independent dsDNA molecule, which is a key feature linked to the chromatin remodelling and looping activity of MeCP2, as well as its ability to interact with nucleosomes replacing histone H1. The dsDNA interaction is characterized by an unusually large heat capacity linked to a cluster of water molecules trapped within the binding interface. The dynamics of disordered regions together with extrinsic factors are key determinants of MeCP2 global structural properties and functional capabilities.

The intervening domain from MeCP2 enhances the DNA affinity of the methyl binding domain and provides an independent DNA interaction site

PubMed Central

Claveria-Gimeno, Rafael; Lanuza, Pilar M.; Morales-Chueca, Ignacio; Jorge-Torres, Olga C.; Vega, Sonia; Abian, Olga; Esteller, Manel; Velazquez-Campoy, Adrian

2017-01-01

Methyl-CpG binding protein 2 (MeCP2) preferentially interacts with methylated DNA and it is involved in epigenetic regulation and chromatin remodelling. Mutations in MeCP2 are linked to Rett syndrome, the leading cause of intellectual retardation in girls and causing mental, motor and growth impairment. Unstructured regions in MeCP2 provide the plasticity for establishing interactions with multiple binding partners. We present a biophysical characterization of the methyl binding domain (MBD) from MeCP2 reporting the contribution of flanking domains to its structural stability and dsDNA interaction. The flanking disordered intervening domain (ID) increased the structural stability of MBD, modified its dsDNA binding profile from an entropically-driven moderate-affinity binding to an overwhelmingly enthalpically-driven high-affinity binding. Additionally, ID provided an additional site for simultaneously and autonomously binding an independent dsDNA molecule, which is a key feature linked to the chromatin remodelling and looping activity of MeCP2, as well as its ability to interact with nucleosomes replacing histone H1. The dsDNA interaction is characterized by an unusually large heat capacity linked to a cluster of water molecules trapped within the binding interface. The dynamics of disordered regions together with extrinsic factors are key determinants of MeCP2 global structural properties and functional capabilities. PMID:28139759

Aminoglycosylation Can Enhance the G-Quadruplex Binding Activity of Epigallocatechin

PubMed Central

Bai, Li-Ping; Ho, Hing-Man; Ma, Dik-Lung; Yang, Hui; Fu, Wai-Chung; Jiang, Zhi-Hong

2013-01-01

With the aim of enhancing G-quadruplex binding activity, two new glucosaminosides (16, 18) of penta-methylated epigallocatechin were synthesized by chemical glycosylation. Subsequent ESI-TOF-MS analysis demonstrated that these two glucosaminoside derivatives exhibit much stronger binding activity to human telomeric DNA and RNA G-quadruplexes than their parent structure (i.e., methylated EGC) (14) as well as natural epigallocatechin (EGC, 6). The DNA G-quadruplex binding activity of 16 and 18 is even more potent than strong G-quadruplex binder quercetin, which has a more planar structure. These two synthetic compounds also showed a higher binding strength to human telomeric RNA G-quadruplex than its DNA counterpart. Analysis of the structure-activity relationship revealed that the more basic compound, 16, has a higher binding capacity with DNA and RNA G-quadruplexes than its N-acetyl derivative, 18, suggesting the importance of the basicity of the aminoglycoside for G-quadruplex binding activity. Molecular docking simulation predicted that the aromatic ring of 16 π-stacks with the aromatic ring of guanine nucleotides, with the glucosamine moiety residing in the groove of G-quadruplex. This research indicates that glycosylation of natural products with aminosugar can significantly enhance their G-quadruplex binding activities, thus is an effective way to generate small molecules targeting G-quadruplexes in nucleic acids. In addition, this is the first report that green tea catechin can bind to nucleic acid G-quadruplex structures. PMID:23335983

Fluctuating capacity and advance decision-making in Bipolar Affective Disorder - Self-binding directives and self-determination.

PubMed

Gergel, Tania; Owen, Gareth S

2015-01-01

For people with Bipolar Affective Disorder, a self-binding (advance) directive (SBD), by which they commit themselves to treatment during future episodes of mania, even if unwilling, can seem the most rational way to deal with an imperfect predicament. Knowing that mania will almost certainly cause enormous damage to themselves, their preferred solution may well be to allow trusted others to enforce treatment and constraint, traumatic though this may be. No adequate provision exists for drafting a truly effective SBD and efforts to establish such provision are hampered by very valid, but also paralysing ethical, clinical and legal concerns. Effectively, the autonomy and rights of people with bipolar are being 'protected' through being denied an opportunity to protect themselves. From a standpoint firmly rooted in the clinical context and experience of mania, this article argues that an SBD, based on a patient-centred evaluation of capacity to make treatment decisions (DMC-T) and grounded within the clinician-patient relationship, could represent a legitimate and ethically coherent form of self-determination. After setting out background information on fluctuating capacity, mania and advance directives, this article proposes a framework for constructing such an SBD, and considers common objections, possible solutions and suggestions for future research. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Fluctuating capacity and advance decision-making in Bipolar Affective Disorder — Self-binding directives and self-determination

PubMed Central

Gergel, Tania; Owen, Gareth S.

2015-01-01

For people with Bipolar Affective Disorder, a self-binding (advance) directive (SBD), by which they commit themselves to treatment during future episodes of mania, even if unwilling, can seem the most rational way to deal with an imperfect predicament. Knowing that mania will almost certainly cause enormous damage to themselves, their preferred solution may well be to allow trusted others to enforce treatment and constraint, traumatic though this may be. No adequate provision exists for drafting a truly effective SBD and efforts to establish such provision are hampered by very valid, but also paralysing ethical, clinical and legal concerns. Effectively, the autonomy and rights of people with bipolar are being ‘protected’ through being denied an opportunity to protect themselves. From a standpoint firmly rooted in the clinical context and experience of mania, this article argues that an SBD, based on a patient-centred evaluation of capacity to make treatment decisions (DMC-T) and grounded within the clinician–patient relationship, could represent a legitimate and ethically coherent form of self-determination. After setting out background information on fluctuating capacity, mania and advance directives, this article proposes a framework for constructing such an SBD, and considers common objections, possible solutions and suggestions for future research. PMID:25939286

Estimation of methacrylate monolith binding capacity from pressure drop data.

PubMed

Podgornik, Aleš; Smrekar, Vida; Krajnc, Peter; Strancar, Aleš

2013-01-11

Convective chromatographic media comprising of membranes and monoliths represent an important group of chromatographic supports due to their flow-unaffected chromatographic properties and consequently fast separation and purification even of large biological macromolecules. Consisting of a single piece of material, common characterization procedures based on analysis of a small sample assuming to be representative for the entire batch, cannot be applied. Because of that, non-invasive characterization methods are preferred. In this work pressure drop was investigated for an estimation of dynamic binding capacity (DBC) of proteins and plasmid DNA for monoliths with different pore sizes. It was demonstrated that methacrylate monolith surface area is reciprocally proportional to pore diameter and that pressure drop on monolith is reciprocally proportional to square pore size demonstrating that methacrylate monolith microstructure is preserved by changing pore size. Based on these facts mathematical formalism has been derived predicting that DBC is in linear correlation with the square root of pressure drop. This was experimentally confirmed for ion-exchange and hydrophobic interactions for proteins and plasmid DNA. Furthermore, pressure drop was also applied for an estimation of DBC in grafted layers of different thicknesses as estimated from the pressure drop data. It was demonstrated that the capacity is proportional to the estimated grafted layer thickness. Copyright © 2012 Elsevier B.V. All rights reserved.

One amino acid in mouse activated factor VII defines its endothelial protein C receptor (EPCR) binding and modulates its EPCR-dependent hemostatic activity in vivo.

PubMed

Pavani, G; Zintner, S M; Ivanciu, L; Small, J C; Stafford, K A; Szeto, J H; Margaritis, P

2017-03-01

Essentials The lack of factor (F) VIIa-endothelial protein C receptor (EPCR) binding in mice is unresolved. A single substitution of Leu4 to Phe in mouse FVIIa (mFVIIa) enables its interaction with EPCR. mFVIIa with a Phe4 shows EPCR binding-dependent enhanced hemostatic function in vivo vs. mFVIIa. Defining the FVIIa-EPCR interaction in mice allows for further investigating its biology in vivo. Background Human activated factor VII (hFVIIa), which is used in hemophilia treatment, binds to the endothelial protein C (PC) receptor (EPCR) with unclear hemostatic consequences. Interestingly, mice lack the activated FVII (FVIIa)-EPCR interaction. Therefore, to investigate the hemostatic consequences of this interaction in hemophilia, we previously engineered a mouse FVIIa (mFVIIa) molecule that bound mouse EPCR (mEPCR) by using three substitutions from mouse PC (mPC), i.e. Leu4→Phe, Leu8→Met, and Trp9→Arg. The resulting molecule, mFVIIa-FMR, modeled the EPCR-binding properties of hFVIIa and showed enhanced hemostatic capacity in hemophilic mice versus mFVIIa. These data implied a role of EPCR in the action of hFVIIa in hemophilia treatment. However, the substitutions in mFVIIa-FMR only broadly defined the sequence determinants for its mEPCR interaction and enhanced function in vivo. Objectives To determine the individual contributions of mPC Phe4, Met8 and Arg9 to the in vitro/in vivo properties of mFVIIa-FMR. Methods The mEPCR-binding properties of single amino acid variants of mFVIIa or mPC at position 4, 8 or 9 were investigated. Results and conclusions Phe4 in mFVIIa or mPC was solely critical for interaction with mEPCR. In hemophilic mice, administration of mFVIIa harboring a Phe4 resulted in a 1.9-2.5-fold increased hemostatic capacity versus mFVIIa that was EPCR binding-dependent. This recapitulated previous observations made with triple-mutant mFVIIa-FMR. As Leu8 is crucial for hFVIIa-EPCR binding, we describe the sequence divergence of this interaction in mice, now allowing its further characterization in vivo. We also illustrate that modulation of the EPCR-FVIIa interaction may lead to improved FVIIa therapeutics. © 2016 International Society on Thrombosis and Haemostasis.

Computational Docking of the Isomers of Nonylphenol to the Ligand Binding Domain of the Estrogen Receptor

EPA Science Inventory

Nonylphenols are environmentally persistent endocrine disrupting chemicals. They exist in the environment as complex mixtures containing many nonylphenol isomers. Environmental mixtures of nonylphenols, along with a few single isomers have been tested for their capacity to inte...

RAPID SCREENING OF ENVIRONMENTAL CHEMICALS FOR ESTROGEN RECEPTOR BINDING CAPACITY

EPA Science Inventory

Over the last few years, an increased awareness of endocrine disrupting chemicals (EDCs) and their potential to affect wildlife and humans has produced a demand for practical screening methods to identify endocrine activity in a wide range of environmental and industrial chemical...

The influence of different techniques in characterizing human antibodies to cow's milk proteins

PubMed Central

McCaffery, T. D.; Kraft, S. C.; Rothberg, R. M.

1972-01-01

Sera from 760 subjects with and without inflammatory bowel disease (IBD) were studied selectively using both primary and secondary antibody assay techniques and different cow's milk antigens. Techniques which demonstrate antibody–antigen binding revealed that the incidence, amount and immunoglobulin class of detectable antibody to bovine serum albumin (BSA) were not significantly different among IBD and control subjects. Only 13 of the 138 sera with the most anti-BSA by primary binding techniques had the capacity to precipitate spontaneously either BSA or antigens in raw (RSM) and pasteurized (PSM) skimmed milk. In passive haemagglutination studies, 41% of these 138 sera had the capacity to agglutinate BSA-coated erythrocytes, while the respective figures for RSM and PSM were 56% and 77%. Only in studies employing the passive haemagglutination of RSM-coated erythrocytes were high titres found more frequently in sera from patients with IBD than in sera from control subjects. Taken as a whole, this study fails to provide evidence for the pathogenetic significance of milk antibodies in IBD. PMID:4625158

One-dimensional surface-imprinted polymeric nanotubes for specific biorecognition by initiated chemical vapor deposition (iCVD).

PubMed

Ince, Gozde Ozaydin; Armagan, Efe; Erdogan, Hakan; Buyukserin, Fatih; Uzun, Lokman; Demirel, Gokhan

2013-07-24

Molecular imprinting is a powerful, generic, and cost-effective technique; however, challenges still remain related to the fabrication and development of these systems involving nonhomogeneous binding sites, insufficient template removing, incompatibility with aqueous media, low rebinding capacity, and slow mass transfer. The vapor-phase deposition of polymers is a unique technique because of the conformal nature of coating and offers new possibilities in a number of applications including sensors, microfluidics, coating, and bioaffinity platforms. Herein, we demonstrated a simple but versatile concept to generate one-dimensional surface-imprinted polymeric nanotubes within anodic aluminum oxide (AAO) membranes based on initiated chemical vapor deposition (iCVD) technique for biorecognition of immunoglobulin G (IgG). It is reported that the fabricated surface-imprinted nanotubes showed high binding capacity and significant specific recognition ability toward target molecules compared with the nonimprinted forms. Given its simplicity and universality, the iCVD method can offer new possibilities in the field of molecular imprinting.

Understanding the interacted mechanism between carbonation and chloride aerosol attack in ordinary Portland cement concrete

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Jun; Qiu, Qiwen; Chen, Xiaochi

An experimental study is carried out with the aim to understand the interacted mechanism between carbonation and chloride aerosol attack in ordinary Portland cement (OPC) concrete. Effects of carbonation on the chloride profile, the chloride binding capacity and the chloride diffusion coefficient are evaluated. Besides, effect of chloride aerosol attack on the carbonation rate is investigated. Concrete specimens with three water-to-cement ratios (0.38, 0.47 and 0.53) are fabricated in this work. Tested results demonstrate that carbonation remarkably affects the chloride profile, reduces the chloride binding capacity, and also accelerates the rate of chloride ion diffusion of concrete. Besides, the presencemore » of chloride aerosol can lead to lower the carbonation depth and increase the pH value of carbonated concrete. Microscopic properties such as morphology, porosity, and pore size distribution for the contaminated concretes are explored by scanning electron microscope and mercury intrusion porosimetry, which provide strong evidence to these research findings.« less

Chemical modeling of acid-base properties of soluble biopolymers derived from municipal waste treatment materials.

PubMed

Tabasso, Silvia; Berto, Silvia; Rosato, Roberta; Marinos, Janeth Alicia Tafur; Ginepro, Marco; Zelano, Vincenzo; Daniele, Pier Giuseppe; Montoneri, Enzo

2015-02-04

This work reports a study of the proton-binding capacity of biopolymers obtained from different materials supplied by a municipal biowaste treatment plant located in Northern Italy. One material was the anaerobic fermentation digestate of the urban wastes organic humid fraction. The others were the compost of home and public gardening residues and the compost of the mix of the above residues, digestate and sewage sludge. These materials were hydrolyzed under alkaline conditions to yield the biopolymers by saponification. The biopolymers were characterized by 13C NMR spectroscopy, elemental analysis and potentiometric titration. The titration data were elaborated to attain chemical models for interpretation of the proton-binding capacity of the biopolymers obtaining the acidic sites concentrations and their protonation constants. The results obtained with the models and by NMR spectroscopy were elaborated together in order to better characterize the nature of the macromolecules. The chemical nature of the biopolymers was found dependent upon the nature of the sourcing materials.

Chemical Modeling of Acid-Base Properties of Soluble Biopolymers Derived from Municipal Waste Treatment Materials

PubMed Central

Tabasso, Silvia; Berto, Silvia; Rosato, Roberta; Tafur Marinos, Janeth Alicia; Ginepro, Marco; Zelano, Vincenzo; Daniele, Pier Giuseppe; Montoneri, Enzo

2015-01-01

This work reports a study of the proton-binding capacity of biopolymers obtained from different materials supplied by a municipal biowaste treatment plant located in Northern Italy. One material was the anaerobic fermentation digestate of the urban wastes organic humid fraction. The others were the compost of home and public gardening residues and the compost of the mix of the above residues, digestate and sewage sludge. These materials were hydrolyzed under alkaline conditions to yield the biopolymers by saponification. The biopolymers were characterized by 13C NMR spectroscopy, elemental analysis and potentiometric titration. The titration data were elaborated to attain chemical models for interpretation of the proton-binding capacity of the biopolymers obtaining the acidic sites concentrations and their protonation constants. The results obtained with the models and by NMR spectroscopy were elaborated together in order to better characterize the nature of the macromolecules. The chemical nature of the biopolymers was found dependent upon the nature of the sourcing materials. PMID:25658795

Effect of the Programmed Nutrition Beef Program on moisture retention of cooked ground beef patties and enhanced strip loins.

PubMed

2015-02-01

This study evaluated the influence of the Programmed Nutrition Beef Program and exogenous growth promotants (ExGP) on water holding capacity characteristics of enhanced beef strip loins. Sixty, frozen strip loins, arranged in a 2 × 2 factorial treatment arrangement with dietary program serving as the first factor and use of ExGP as the second factor, were thawed, injected with an enhancement solution, and stored for 7 days. Loins from ExGP cattle possessed the ability to bind more (P < 0.05) water before pumping and bind less (P < 0.05) water after pumping and storage. Loin pH across treatments was similar (P > 0.10) before injection, but increased post-injection and after storage (P < 0.01). Treatments did not affect loin purge loss, steak cook loss, and expressible moisture (P > 0.10). The Programmed Nutrition Beef Program and use of ExGPs minimally impacted water holding capacity of enhanced frozen/thawed beef strip loins.

Systematic research on the pretreatment of peptides for quantitative proteomics using a C₁₈ microcolumn.

PubMed

Zhai, Linhui; Chang, Cheng; Li, Ning; Duong, Duc M; Chen, Hao; Deng, Zixin; Yang, Jian; Hong, Xuechuan; Zhu, Yunping; Xu, Ping

2013-08-01

Reversed phase microcolumns have been widely used for peptide pretreatment to desalt and remove interferences before tandem LC-MS in proteomics studies. However, few studies have characterized the effects of experimental parameters as well as column characteristics on the composition of identified peptides. In this study, several parameters including the concentration of ACN in washing buffer, the microcolumn's purification effect, the peptide recovery rate, and the dynamic-binding capacity were characterized in detail, based upon stable isotope labeling by amino acids in a cell culture quantitative approach. The results showed that peptide losses can be reduced with low ACN concentration in washing buffers resulting in a recovery rate of approximately 82%. Furthermore, the effects of ACN concentration and loading amount on the properties of identified peptides were also evaluated. We found that the dynamic-binding capacity of the column was approximately 26 μg. With increased loading amounts, more hydrophilic peptides were replaced by hydrophobic peptides. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Mutations in the substrate binding site of human heat-shock protein 70 indicate specific interaction with HLA-DR outside the peptide binding groove

PubMed Central

Rohrer, Karin M; Haug, Markus; Schwörer, Daniela; Kalbacher, Hubert; Holzer, Ursula

2014-01-01

Heat-shock protein 70 (Hsp70)–peptide complexes are involved in MHC class I-and II-restricted antigen presentation, enabling enhanced activation of T cells. As shown previously, mammalian cytosolic Hsp70 (Hsc70) molecules interact specifically with HLA-DR molecules. This interaction might be of significance as Hsp70 molecules could transfer bound antigenic peptides in a ternary complex into the binding groove of HLA-DR molecules. The present study provides new insights into the distinct interaction of Hsp70 with HLA-DR molecules. Using a quantitative binding assay, it could be demonstrated that a point mutation of amino acids alanine 406 and valine 438 in the substrate binding pocket led to reduced peptide binding compared with the wild-type Hsp70 whereas HLA-DR binding remains unaffected. The removal of the C-terminal lid neither altered the substrate binding capacity nor the Hsp70 binding characteristics to HLA-DR. A truncated variant lacking the nucleotide binding domain showed no binding interactions with HLA-DR. Furthermore, the truncated ATPase subunit of constitutively expressed Hsc70 revealed similar binding affinities to HLA-DR compared with the complete Hsc70. Hence, it can be assumed that the Hsp70–HLA-DR interaction takes place outside the peptide binding groove and is attributed to the ATPase domain of HSP70 molecules. The Hsp70-chaperoned peptides might thereby be directly transferred into the binding groove of HLA-DR, so enabling enhanced presentation of the peptide on antigen-presenting cells and leading to an improved proliferation of responding T cells as shown previously. PMID:24428437

Respiratory Syncytial Virus (RSV): Neutralizing Antibody, a Correlate of Immune Protection.

PubMed

Piedra, Pedro A; Hause, Anne M; Aideyan, Letisha

2016-01-01

Assays that measure RSV-specific neutralizing antibody activity are very useful for evaluating vaccine candidates, performing seroprevalence studies, and detecting infection. Neutralizing antibody activity is normally measured by a plaque reduction neutralization assay or by a microneutralization assay with or without complement. These assays measure the functional capacity of serum (or other fluids) to neutralize virus infectivity in cells as compared to ELISA assays that only measure the binding capacity against an antigen. This chapter discusses important elements in standardization of the RSV-specific microneutralization assay for use in the laboratory.

[Hemoglobin oxygen transport capacity in surgical endotoxicosis ].

PubMed

Poryadin, G V; Vlasov, A P; Trofimov, V A; Vlasova, T I; Kamkina, O V; Grigoryev, A G; Vlasov, P A

2016-01-01

In surgical endointoxication hemoglobin oxygen transport capacity of red blood cells (hemoglobin affinity ligands: the ability to bind and release ligands) is reduced and is associated with the severity of endogenous intoxication. Violation of oxygen transport function of hemoglobin at endogenous intoxication is associated with conformational changes of a biomolecule, and its possible influence on reactive oxygen species, which confirmed in experiments in vitro: under the influence of oxygen-iron ascorbate ability of hemoglobin deteriorates. Largely similar structural and functional changes in hemoglobin occur in patients with surgical endotoxicosis.

Modification of Herbicide Binding to Photosystem II in Two Biotypes of Senecio vulgaris L

PubMed Central

Pfister, Klaus; Radosevich, Steven R.; Arntzen, Charles J.

1979-01-01

The present study compares the binding and inhibitory activity of two photosystem II inhibitors: 3-(3,4-dichlorophenyl)-1,1-dimethylurea (diuron [DCMU]) and 2-chloro-4-(ethylamine)-6-(isopropyl amine)-S-triazene (atrazine). Chloroplasts isolated from naturally occurring triazine-susceptible and triazine-resistant biotypes of common groundsel (Senecio vulgaris L.) showed the following characteristics. (a) Diuron strongly inhibited photosynthetic electron transport from H2O to 2,6-dichlorophenolindophenol in both biotypes. Strong inhibition by atrazine was observed only with the susceptible chloroplasts. (b) Hill plots of electron transport inhibition data indicate a noncooperative binding of one inhibitor molecule at the site of action for both diuron and atrazine. (c) Susceptible chloroplasts show a strong diuron and atrazine binding (14C-radiolabel assays) with binding constants (K) of 1.4 × 10−8 molar and 4 × 10−8 molar, respectively. In the resistant chloroplasts the diuron binding was slightly decreased (K = 5 × 10−8 molar), whereas no specific atrazine binding was detected. (d) In susceptible chloroplasts, competitive binding between radioactively labeled diuron and non-labeled atrazine was observed. This competition was absent in the resistant chloroplasts. We conclude that triazine resistance of both intact plants and isolated chloroplasts of Senecio vulgaris L. is based upon a minor modification of the protein in the photosystem II complex which is responsible for herbicide binding. This change results in a specific loss of atrazine (triazine)-binding capacity. PMID:16661120

Sex pheromone recognition and characterization of three pheromone-binding proteins in the legume pod borer, Maruca vitrata Fabricius (Lepidoptera: Crambidae)

PubMed Central

Mao, Aping; Zhou, Jing; Bin Mao; Zheng, Ya; Wang, Yufeng; Li, Daiqin; Wang, Pan; Liu, Kaiyu; Wang, Xiaoping; Ai, Hui

2016-01-01

Pheromone-binding proteins (PBPs) are essential for the filtering, binding and transporting of sex pheromones across sensillum lymph to membrane-associated pheromone receptors of moths. In this study, three novel PBP genes were expressed in Escherichia coli to examine their involvement in the sex pheromone perception of Maruca vitrata. Fluorescence binding experiments indicated that MvitPBP1-3 had strong binding affinities with four sex pheromones. Moreover, molecular docking results demonstrated that six amino acid residues of three MvitPBPs were involved in the binding of the sex pheromones. These results suggested that MvitPBP1-3 might play critical roles in the perception of female sex pheromones. Additionally, the binding capacity of MvitPBP3 with the host-plant floral volatiles was high and was similar to that of MvitGOBP2. Furthermore, sequence alignment and docking analysis showed that both MvitGOBP2 and MvitPBP3 possessed an identical key binding site (arginine, R130/R140) and a similar protein pocket structure around the binding cavity. Therefore, we hypothesized that MvitPBP3 and MvitGOBP2 might have synergistic roles in binding different volatile ligands. In combination, the use of synthetic sex pheromones and floral volatiles from host-plant may be used in the exploration for more efficient monitoring and integrated management strategies for the legume pod borer in the field. PMID:27698435

Interaction between rose bengal and different protein components.

PubMed

Tseng, S C; Zhang, S H

1995-07-01

Bindings of rose bengal to several proteins were determined by Sephadex G-75 chromatography. Their respective blocking effect against dye uptake was demonstrated in an assay using a rabbit corneal epithelial cell layer. The total binding capacity of nonmucin proteins was measured using fluorometry and Scatchard analysis. The results showed that albumin, lactoferrin, transferrin, and lysozyme could--but serum prealbumin, IgA, carboxymethyl cellulose (CMC), and Sepharose 4B-purified porcine stomach mucin (PSM) could not--bind rose bengal. Lysozyme formed precipitates with rose bengal. Sufficient concentrations of albumin, lactoferrin, transferrin, or lysozyme premixed with rose bengal could block dye uptake by cells, but IgA and serum prealbumin could not. Premixed PSM was not as effective as precoated PSM in blocking dye uptake. The dissociation constant (Kd) was 1.2 x 10(-7) M, 3.6 x 10(-7) M, 3.9 x 10(-7) M, and 1.6 x 10(-6) M for albumin, transferrin, lactoferrin, and lysozyme, respectively. Based on these values, the total maximal binding capacity of nonmucin proteins in normal 7-microliters tears was extrapolated to be 0.249 micrograms rose bengal, which is too small to explain the negative staining of rose bengal on the normal ocular surface. Rose bengal, but not fluorescein, could interact with carbohydrate-containing Sephadex, CMC, and PSM to slow down its elution via Sephadex column chromatography. Therefore, the normal negative staining to rose bengal might be caused by the blocking effect of preocular mucus tear layer, which serves as a diffusion barrier. Rose bengal remains a unique dye for detecting the protective function of the preocular mucus tear.

DockoMatic: automated peptide analog creation for high throughput virtual screening.

PubMed

Jacob, Reed B; Bullock, Casey W; Andersen, Tim; McDougal, Owen M

2011-10-01

The purpose of this manuscript is threefold: (1) to describe an update to DockoMatic that allows the user to generate cyclic peptide analog structure files based on protein database (pdb) files, (2) to test the accuracy of the peptide analog structure generation utility, and (3) to evaluate the high throughput capacity of DockoMatic. The DockoMatic graphical user interface interfaces with the software program Treepack to create user defined peptide analogs. To validate this approach, DockoMatic produced cyclic peptide analogs were tested for three-dimensional structure consistency and binding affinity against four experimentally determined peptide structure files available in the Research Collaboratory for Structural Bioinformatics database. The peptides used to evaluate this new functionality were alpha-conotoxins ImI, PnIA, and their published analogs. Peptide analogs were generated by DockoMatic and tested for their ability to bind to X-ray crystal structure models of the acetylcholine binding protein originating from Aplysia californica. The results, consisting of more than 300 simulations, demonstrate that DockoMatic predicts the binding energy of peptide structures to within 3.5 kcal mol(-1), and the orientation of bound ligand compares to within 1.8 Å root mean square deviation for ligand structures as compared to experimental data. Evaluation of high throughput virtual screening capacity demonstrated that Dockomatic can collect, evaluate, and summarize the output of 10,000 AutoDock jobs in less than 2 hours of computational time, while 100,000 jobs requires approximately 15 hours and 1,000,000 jobs is estimated to take up to a week. Copyright © 2011 Wiley Periodicals, Inc.

Determination of drug and fatty acid binding capacity to pluronic f127 in microemulsions.

PubMed

James-Smith, Monica A; Shekhawat, Dushyant; Moudgil, Brij M; Shah, Dinesh O

2007-02-13

We propose that one can deduce very insightful information regarding the drug and fatty acid binding capacity of microemulsions through simple turbidity experiments. Pluronic F127-based oil-in-water microemulsions of various compositions were synthesized and titrated to turbidity with concentrated amitriptyline, an antidepressant drug. We observed that, above certain Pluronic F127 concentrations, turbidity was never observed, irrespective of how much amitriptyline was added to the microemulsion. We also observed that whenever sodium caprylate fatty acid was not included in the microemulsion formulation, turbidity never occurred. On the basis of these findings, we were able to determine the point at which all sodium caprylate present in the microemulsion formulation was bound to the F127 in the microemulsion (i.e., no fatty acid was free in the bulk in monomer form). By the same logic we were also able to determine how much amitriptyline was binding to the microemulsions. We also measured the dynamic surface tension, foamability, and fabric wetting time of the microemulsion formulations to further prove the hypothesis that all fatty acid is bound to the F127 in the microemulsion above a critical Pluronic F127 concentration. On the basis of this research, we have concluded that there are approximately 11 molecules of sodium caprylate fatty acid bound per molecule of Pluronic F127 and approximately 12 molecules of amitriptyline bound per molecule of Pluronic F127 in the optimal microemulsion formulation. These findings give us valuable information about the charge density at the oil/water interface and about the mechanism of binding of the drug to the microemulsion.

Effect of micronization on the physicochemical properties of insoluble dietary fiber from citrus (Citrus junos Sieb. ex Tanaka) pomace.

PubMed

Ye, Fayin; Tao, Bingbing; Liu, Jia; Zou, Yan; Zhao, Guohua

2016-04-01

The aim of this work was to study the effect of micronization (mechanical and jet grindings) on the physicochemical properties of the insoluble dietary fiber from citrus pomace in comparison with ordinary grinding. The results showed that micronization treatment effectively pulverized the IDF-CP powders to micron scale and significantly increased the soluble dietary fiber content (p < 0.05). Compared with mechanical grinding, jet grinding was more effective in size reduction and resulted in IDF-CP powders with narrower particle size distributions. Micronized IDF-CP powders had smaller particle size, smoother surface, higher fluidity, cation-exchange capacity, and metal cation binding capacity values, but lower water holding capacity, oil holding capacity, and swelling capacity values. These functional properties were significantly dependent on surface area and particle size (D0.5). The present study suggested that micronization treatments could modify functional properties of IDF-CP powders, which promotes their use in food applications. © The Author(s) 2015.

Analysis of the heat capacity of nanoclusters of FCC metals on the example of Al, Ni, Cu, Pd, and Au

NASA Astrophysics Data System (ADS)

Gafner, Yu. Ya.; Gafner, S. L.; Zamulin, I. S.; Redel, L. V.; Baidyshev, V. S.

2015-06-01

The heat capacity of ideal nickel, copper, gold, aluminum, and palladium fcc clusters with diameter of up to 6 nm has been studied in the temperature range of 150-800 K in terms of the molecular-dynamics theory using a tight-binding potential. The heat capacity of individual metallic nanoclusters has been found to exceed that characteristic of the bulk state, but by no more than 16-20%, even in the case of very small clusters. To explain the discrepancy between the simulated data and the experimental results on the compacted metals, aluminum and palladium samples with 80% theoretical density have also been investigated. Based on the simulation results and analysis of the experimental data, it has been established that the increased heat capacity of the compacted nanomaterials does not depend on the enhanced heat capacity of the individual clusters but rather, can be due to either the disordered state of the nanomaterial or a significant content of impurities (mainly, hydrogen).

Adaptive capacity of fishing communities at marine protected areas: a case study from the Colombian Pacific.

PubMed

Moreno-Sánchez, Rocío del Pilar; Maldonado, Jorge Higinio

2013-12-01

Departing from a theoretical methodology, we estimate empirically an index of adaptive capacity (IAC) of a fishing community to the establishment of marine protected areas (MPAs). We carried out household surveys, designed to obtain information for indicators and sub-indicators, and calculated the IAC. Moreover, we performed a sensitivity analysis to check for robustness of the results. Our findings show that, despite being located between two MPAs, the fishing community of Bazán in the Colombian Pacific is highly vulnerable and that the socioeconomic dimension of the IAC constitutes the most binding dimension for building adaptive capacity. Bazán is characterized by extreme poverty, high dependence on resources, and lack of basic public infrastructure. Notwithstanding, social capital and local awareness about ecological conditions may act as enhancers of adaptive capacity. The establishment of MPAs should consider the development of strategies to confer adaptive capacity to local communities highly dependent on resource extraction.

Working memory for visual features and conjunctions in schizophrenia.

PubMed

Gold, James M; Wilk, Christopher M; McMahon, Robert P; Buchanan, Robert W; Luck, Steven J

2003-02-01

The visual working memory (WM) storage capacity of patients with schizophrenia was investigated using a change detection paradigm. Participants were presented with 2, 3, 4, or 6 colored bars with testing of both single feature (color, orientation) and feature conjunction conditions. Patients performed significantly worse than controls at all set sizes but demonstrated normal feature binding. Unlike controls, patient WM capacity declined at set size 6 relative to set size 4. Impairments with subcapacity arrays suggest a deficit in task set maintenance: Greater impairment for supercapacity set sizes suggests a deficit in the ability to selectively encode information for WM storage. Thus, the WM impairment in schizophrenia appears to be a consequence of attentional deficits rather than a reduction in storage capacity.

Heat and pressure treatments effects on peanut allergenicity

USDA-ARS?s Scientific Manuscript database

Peanut allergy is recognized as one of the most severe food allergies. The aim of this study was to investigate the changes in IgE binding capacity of peanut proteins produced by thermal-processing methods, including autoclaving. Immunoreactivity to raw and thermally processed peanut extracts was ev...

Binding Thermodynamics of Ferredoxin:NADP+ Reductase: Two Different Protein Substrates and One Energetics

PubMed Central

Martínez-Júlvez, Marta; Medina, Milagros; Velázquez-Campoy, Adrián

2009-01-01

Abstract The thermodynamics of the formation of binary and ternary complexes between Anabaena PCC 7119 FNR and its substrates, NADP+ and Fd, or Fld, has been studied by ITC. Despite structural dissimilarities, the main difference between Fd and Fld binding to FNR relates to hydrophobicity, reflected in different binding heat capacity and number of water molecules released from the interface. At pH 8, the formation of the binary complexes is both enthalpically and entropically driven, accompanied by the protonation of at least one ionizable group. His299 FNR has been identified as the main responsible for the proton exchange observed. However, at pH 10, where no protonation occurs and intrinsic binding parameters can be obtained, the formation of the binary complexes is entropically driven, with negligible enthalpic contribution. Absence of the FMN cofactor in Fld does not alter significantly the strength of the interaction, but considerably modifies the enthalpic and entropic contributions, suggesting a different binding mode. Ternary complexes show negative cooperativity (6-fold and 11-fold reduction in binding affinity, respectively), and an increase in the enthalpic contribution (more favorable) and a decrease in the entropic contribution (less favorable), with regard to the binary complexes energetics. PMID:19527656

Steroid production and estrogen binding in flowers of Gladiolus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Adler, J.H.; Wolfe, G.R.; Janik, J.R.

1987-04-01

The bioconversion of /sup 3/H-cholesterol to steroids was examined in excised tissue from the pistils and bracts of Gladiolus. Ovary-ovule and stigma-style tissues produce a compound with chromatographic properties on reverse phase HPLC similar to 17..beta..-estradiol (E/sub 2/). The stigma-style fraction also produced a compound that chromatographed similarly to progesterone. Bracts and the oxidation controls produced no radiolabeled compounds which were chromatographically similar to E/sub 2/. An endogenous E/sub 2/ binding protein was partially characterized from the ovules. The protein binds E/sub 2/, estriol, and diethylstilbesterol whereas testosterone and progesterone do not bind. The total specific binding capacities in themore » cytosolic and nuclear fractions are 1.6 and 2.2 femtomoles of estradiol per mg of tissue. The dissociation constant is 1.1 x 10/sup -9/ M/sup -1/ for both subcellular fractions. The protein-estradiol complex has a sedimentation coefficient of 4.7 +/- 0.1S. The tissue specific biosynthesis of estrogens and the presence of a steroid binding protein similar to a Type 1 estrogen receptor found in mammals is suggestive of a role for steroids in pistil ontogeny.« less

Pulsed ultraviolet light reduces immunoglobulin E binding to Atlantic white shrimp (Litopenaeus setiferus) extract.

PubMed

Shriver, Sandra; Yang, Wade; Chung, Si-Yin; Percival, Susan

2011-07-01

Pulsed ultraviolet light (PUV), a novel food processing and preservation technology, has been shown to reduce allergen levels in peanut and soybean samples. In this study, the efficacy of using PUV to reduce the reactivity of the major shrimp allergen, tropomyosin (36-kDa), and to attenuate immunoglobulin E (IgE) binding to shrimp extract was examined. Atlantic white shrimp (Litopenaeus setiferus) extract was treated with PUV (3 pulses/s, 10 cm from light source) for 4 min. Tropomyosin was compared in the untreated, boiled, PUV-treated and [boiled+PUV]-treated samples, and changes in the tropomyosin levels were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). IgE binding of the treated extract was analyzed via immunoblot and enzyme-linked immunosorbent assay (ELISA) using pooled human plasma containing IgE antibodies against shrimp allergens. Results showed that levels of tropomyosin and IgE binding were reduced following PUV treatment. However, boiling increased IgE binding, while PUV treatment could offset the increased allergen reactivity caused by boiling. In conclusion, PUV treatment reduced the reactivity of the major shrimp allergen, tropomyosin, and decreased the IgE binding capacity of the shrimp extract.

Interaction of the dietary pigment curcumin with hemoglobin: energetics of the complexation.

PubMed

Basu, Anirban; Kumar, Gopinatha Suresh

2014-08-01

Thermodynamics of the interaction of the chemotherapeutic and chemopreventive dietary pigment, curcumin, with hemoglobin was studied by isothermal titration calorimetry. The binding was characterized to be exothermic. At 293.15 K, the equilibrium constant for curcumin-Hb complexation was found to be (4.88 ± 0.06) × 10(5) M(-1). The binding stoichiometry was calculated to be 1.08 ± 0.05, confirming a 1:1 complexation. The binding was driven by a large negative standard molar enthalpy change (ΔH(0) = -118.45 ± 0.05 kJ mol(-1)) and an unfavorable standard molar entropy change (TΔS(0) = -86.53 ± 0.01 kJ mol(-1)) at 293.15 K. Increasing the temperature favoured the binding, and the magnitude of the negative standard molar heat capacity change suggested the involvement of significant hydrophobic forces in the binding process. With increasing salt concentration, the magnitude of the equilibrium constant decreased slightly; and the complexation mostly involved non-polyelectrolytic forces contributing about 92-94% of the standard molar Gibbs energy change. DSC studies revealed that curcumin binding caused a partial unfolding of the protein.

Spectroscopic and microcalorimetric studies on the molecular binding of food colorant acid red 27 with deoxyribonucleic acid.

PubMed

Basu, Anirban; Kumar, Gopinatha Suresh

2016-08-01

Interaction of the food colorant acid red 27 with double stranded DNA was investigated using spectroscopic and calorimetric methods. Absorbance and fluorescence studies suggested an intimate binding interaction between the dye and DNA. The quantum efficiency value testified an effective energy transfer from the DNA base pairs to the dye molecules. Minor groove displacement assay with Hoechst 33258 revealed that the binding occurs in the minor groove of DNA. Circular dichroism studies revealed that acid red 27 induces moderate conformational perturbations in DNA. Results of calorimetric studies suggested that the complexation process was driven largely by positive entropic contribution with a smaller favorable enthalpy contribution. The equilibrium constant of the binding was calculated to be (3.04 ± 0.09) × 10(4)  M(-1) at 298.15 K. Negative heat capacity value along with the enthalpy-entropy compensation phenomenon established the involvement of dominant hydrophobic forces in the binding process. Differential scanning calorimetry studies presented evidence for an increased thermal stability of DNA on binding of acid red 27. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Influences of urea, pH and metal ions on the interaction between cepharanthine and lysozyme by steady state fluorescence spectroscopy

NASA Astrophysics Data System (ADS)

Yang, Yumin; Li, Daojin; Xu, Chen

2015-03-01

The study on the binding mode of drug with protein is important to understand the pharmacokinetics and toxicity of the drug as well as the relationship of structure and function of the protein. In the study, the interaction between cepharanthine and lysozyme (Lys) in aqueous solution was first investigated by fluorescence spectroscopic techniques at pH 7.4. The obtained quenching rate constant and binding constant indicated the static quenching mechanism and medium binding force. The effect of cepharanthine on the conformation of Lys was analyzed using synchronous fluorescence and three-dimensional (3D) fluorescence. In addition, the effect of urea on the interaction of cepharanthine with Lys was studied and the binding capacity of cepharanthine to the denatured Lys deceases dramatically, as compared with that of cepharanthine to native Lys. Moreover, influence of pH on the interaction of cepharanthine with Lys was investigated. As compared with that at pH 7.4, the binding abilities of the drug to Lys under other pH conditions (pH 9.0, 5.5, 3.5, and 1.9) deceased. Furthermore, the effect of metal ions on the binding constant of cepharanthine with Lys was investigated.

Modulation of [3H]DAGO binding by substance P (SP) and SP fragments in the mouse brain and spinal cord via MU1 interactions.

PubMed

Krumins, S A; Kim, D C; Seybold, V S; Larson, A A

1989-01-01

Binding of [3H]DAGO to fresh, frozen or beta-funaltrexamine (beta-FNA) pretreated membranes of mouse brain and spinal cord was extensively studied using substance P (SP) or SP fragments as potential competitors and/or modulators. The objective was to determine whether SP exerts its analgesic effect by interacting with mu opioid receptors. The affinity of DAGO was reduced and binding capacity was increased in the presence of SP or the N-terminal SP fragments SP(1-9) and SP(1-4) but not the C-terminal SP fragment SP(5-11). Because sub-nanomolar concentrations of SP or N-terminal SP fragments displaced [3H] DAGO binding to a minor but detectable degree, it is suggested that SP interacts with mu 1 sites through its N-terminus portion. The effect of SP on DAGO binding was less in the spinal cord compared to the rest of the brain. Modulation of DAGO binding by SP was enhanced in the brain after pretreatment of membranes with the narcotic antagonist beta-FNA. These results suggest a novel mechanism for the analgesic action of SP.

Multispectroscopic and calorimetric studies on the binding of the food colorant tartrazine with human hemoglobin.

PubMed

Basu, Anirban; Suresh Kumar, Gopinatha

2016-11-15

Interaction of the food colorant tartrazine with human hemoglobin was studied using multispectroscopic and microcalorimetric techniques to gain insights into the binding mechanism and thereby the toxicity aspects. Hemoglobin spectrum showed hypochromic changes in the presence of tartrazine. Quenching of the fluorescence of hemoglobin occurred and the quenching mechanism was through a static mode as revealed from temperature dependent and time-resolved fluorescence studies. According to the FRET theory the distance between β-Trp37 of hemoglobin and bound tartrazine was evaluated to be 3.44nm. Synchronous fluorescence studies showed that tartrazine binding led to alteration of the microenvironment around the tryptophans more in comparison to tyrosines. 3D fluorescence and FTIR data provided evidence for conformational changes in the protein on binding. Circular dichroism studies revealed that the binding led to significant loss in the helicity of hemoglobin. The esterase activity assay further complemented the circular dichroism data. Microcalorimetric study using isothermal titration calorimetry revealed the binding to be exothermic and driven largely by positive entropic contribution. Dissection of the Gibbs energy change proposed the protein-dye complexation to be dominated by non-polyelectrolytic forces. Negative heat capacity change also corroborated the involvement of hydrophobic forces in the binding process. Copyright © 2016 Elsevier B.V. All rights reserved.

Stereoselective binding of doxazosin enantiomers to plasma proteins from rats, dogs and humans in vitro

PubMed Central

Sun, Jia-an; Kong, De-zhi; Zhen, Ya-qin; Li, Qing; Zhang, Wei; Zhang, Jiang-hua; Yin, Zhi-wei; Ren, Lei-ming

2013-01-01

Aim: (±)Doxazosin is a long-lasting inhibitor of α1-adrenoceptors that is widely used to treat benign prostatic hyperplasia and lower urinary tract symptoms. In this study we investigated the stereoselective binding of doxazosin enantiomers to the plasma proteins of rats, dogs and humans in vitro. Methods: Human, dog and rat plasma were prepared. Equilibrium dialysis was used to determine the plasma protein binding of each enantiomer in vitro. Chiral HPLC with fluorescence detection was used to measure the drug concentrations on each side of the dialysis membrane bag. Results: Both the enantiomers were highly bound to the plasma proteins of rats, dogs and humans [(−)doxazosin: 89.4%–94.3%; (+)doxazosin: 90.9%–95.4%]. (+)Doxazosin exhibited significantly higher protein binding capacities than (−)doxazosin in all the three species, and the difference in the bound concentration (Cb) between the two enantiomers was enhanced as their concentrations were increased. Although the percentage of the plasma protein binding in the dog plasma was significantly lower than that in the human plasma at 400 and 800 ng/mL, the corrected percentage of plasma protein binding was dog>human>rat. Conclusion: (−)Doxazosin and (+)doxazosin show stereoselective plasma protein binding with a significant species difference among rats, dogs and humans. PMID:24241343

Altered binding of /sup 125/I-labeled calmodulin to a 46. 5-kilodalton protein in skin fibroblasts cultured from patients with cystic fibrosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tallant, E.A.; Wallace, R.W.

1987-02-01

The levels of calmodulin and calmodulin-binding proteins have been determined in cultured skin fibroblasts from patients with cystic fibrosis (CF) and age- and sex-matched controls. Calmodulin ranged from 0.20 to 0.76 microgram/mg protein; there was no difference between calmodulin concentration in fibroblasts from CF patients and controls. Calmodulin-binding proteins of 230, 212, 204, 164, 139, 70, 59, 46.5, and 41 kD were identified. A protein with a mobility identical to the 59-kD calmodulin-binding protein was labeled by antiserum against calmodulin-dependent phosphatase. Although Ca/sup 2 +//calmodulin-dependent phosphatase activity was detected, there was no different in activity between control and CF fibroblastsmore » or in the level of phosphatase protein as determined by radioimmunoassay. Lower amounts of /sup 125/I-calmodulin were bound to the 46.5-kD calmodulin-binding protein in CF fibroblasts as compared with controls. The 46.5-kD calmodulin-binding protein may be reduced in CF fibroblasts or its structure may be altered resulting in a reduced binding capacity and/or affinity for calmodulin and perhaps reflecting, either directly or indirectly, the genetic defect responsible for cystic fibrosis.« less

Molecular mechanisms of immunosuppression by cyclosporins.

PubMed

Zenke, G; Baumann, G; Wenger, R; Hiestand, P; Quesniaux, V; Andersen, E; Schreier, M H

1993-06-23

Despite the successful clinical application of the immunosuppressive drug cyclosporin A (CsA, Sandimmun), its precise mechanism of action in the process of T cell activation remains elusive. CsA binds to the high-affinity cytosolic receptor cyclophilin whose peptidyl-prolyl cis-trans isomerase activity is inhibited upon binding. The linkage of this effect with the inhibition of the T cell receptor-mediated signal transduction pathway, which leads to a suppression of lymphokine gene transcription, is still unclear. We analyzed the relationship between cyclophilin-binding and immunosuppressive activity (e.g., effect on IL-2 transcription) of cyclosporin derivatives in vitro. The results show that binding to cyclophilin is required, but not sufficient for immunosuppression. Cyclosporin analogues which completely lack immunosuppressive activity but fully retained their cyclophilin-binding capacity antagonize the immunosuppressive activity of CsA. These derivatives inhibit the isomerase activity of cyclophilin, which clearly demonstrates that inhibition of the cyclophilin isomerase activity does not lead to immunosuppression. In analogy to the other immunosuppressants of microbial origin, FK-506 and rapamycin, a specific structure of the "effector" domain of CsA, which is unrelated to the cyclophilin-binding domain, determines the biological activity. In the nucleus, CsA interferes with the DNA-binding of inducible transcription factors to their respective DNA motifs within lymphokine promoters by affecting intracellular translocation of transcription factor subunits.

Different types of working memory binding in epilepsy patients with unilateral anterior temporal lobectomy.

PubMed

van Geldorp, Bonnie; Bouman, Zita; Hendriks, Marc P H; Kessels, Roy P C

2014-03-01

The medial temporal lobe is an important structure for long-term memory formation, but its role in working memory is less clear. Recent studies have shown hippocampal involvement during working memory tasks requiring binding of information. It is yet unclear whether this is limited to tasks containing spatial features. The present study contrasted three binding conditions and one single-item condition in patients with unilateral anterior temporal lobectomy. A group of 43 patients with temporal lobectomy (23 left; 20 right) and 20 matched controls were examined with a working memory task assessing spatial relational binding (object-location), non-spatial relational binding (object-object), conjunctive binding (object-colour) and working memory for single items. We varied the delay period (3 or 6s), as there is evidence showing that delay length may modulate working memory performance. The results indicate that performance was worse for patients than for controls in both relational binding conditions, whereas patients were unimpaired in conjunctive binding. Single-item memory was found to be marginally impaired, due to a deficit on long-delay trials only. In conclusion, working memory binding deficits are found in patients with unilateral anterior temporal lobectomy. The role of the medial temporal lobe in working memory is not limited to tasks containing spatial features. Rather, it seems to be involved in relational binding, but not in conjunctive binding. The medial temporal lobe gets involved when working memory capacity does not suffice, for example when relations have to be maintained or when the delay period is long. Copyright © 2014 Elsevier Inc. All rights reserved.

Enhancing lithium-sulphur battery performance by strongly binding the discharge products on amino-functionalized reduced graphene oxide.

PubMed

Wang, Zhiyu; Dong, Yanfeng; Li, Hongjiang; Zhao, Zongbin; Wu, Hao Bin; Hao, Ce; Liu, Shaohong; Qiu, Jieshan; Lou, Xiong Wen David

2014-09-25

Lithium-sulphur batteries are one very appealing power source with high energy density. But their practical use is still hindered by several issues including short lifespan, low efficiency and safety concern from the lithium anode. Polysulphide dissolution and insulating nature of sulphur are generally considered responsible for the capacity degradation. However, the detachment of discharge products, that is, highly polar lithium sulphides, from nonpolar carbon matrix (for example, graphene) has been rarely studied as one critical factor. Here we report the strongly covalent stabilization of sulphur and its discharge products on amino-functionalized reduced graphene oxide that enables stable capacity retention of 80% for 350 cycles with high capacities and excellent high-rate response up to 4 C. The present study demonstrates a feasible and effective strategy to solve the long-term cycling difficulty for lithium-sulphur batteries and also helps to understand the capacity decay mechanism involved.

Biophysical constraints on the computational capacity of biochemical signaling networks

NASA Astrophysics Data System (ADS)

Wang, Ching-Hao; Mehta, Pankaj

Biophysics fundamentally constrains the computations that cells can carry out. Here, we derive fundamental bounds on the computational capacity of biochemical signaling networks that utilize post-translational modifications (e.g. phosphorylation). To do so, we combine ideas from the statistical physics of disordered systems and the observation by Tony Pawson and others that the biochemistry underlying protein-protein interaction networks is combinatorial and modular. Our results indicate that the computational capacity of signaling networks is severely limited by the energetics of binding and the need to achieve specificity. We relate our results to one of the theoretical pillars of statistical learning theory, Cover's theorem, which places bounds on the computational capacity of perceptrons. PM and CHW were supported by a Simons Investigator in the Mathematical Modeling of Living Systems Grant, and NIH Grant No. 1R35GM119461 (both to PM).

Nanoscale characteristics of triacylglycerol oils: phase separation and binding energies of two-component oils to crystalline nanoplatelets.

PubMed

MacDougall, Colin J; Razul, M Shajahan; Papp-Szabo, Erzsebet; Peyronel, Fernanda; Hanna, Charles B; Marangoni, Alejandro G; Pink, David A

2012-01-01

Fats are elastoplastic materials with a defined yield stress and flow behavior and the plasticity of a fat is central to its functionality. This plasticity is given by a complex tribological interplay between a crystalline phase structured as crystalline nanoplatelets (CNPs) and nanoplatelet aggregates and the liquid oil phase. Oil can be trapped within microscopic pores within the fat crystal network by capillary action, but it is believed that a significant amount of oil can be trapped by adsorption onto crystalline surfaces. This, however, remains to be proven. Further, the structural basis for the solid-liquid interaction remains a mystery. In this work, we demonstrate that the triglyceride liquid structure plays a key role in oil binding and that this binding could potentially be modulated by judicious engineering of liquid triglyceride structure. The enhancement of oil binding is central to many current developments in this area since an improvement in the health characteristics of fat and fat-structured food products entails a reduction in the amount of crystalline triacylglycerols (TAGs) and a relative increase in the amount of liquid TAGs. Excessive amounts of unbound, free oil, will lead to losses in functionality of this important food component. Engineering fats for enhanced oil binding capacity is thus central to the design of more healthy food products. To begin to address this, we modelled the interaction of triacylglycerol oils, triolein (OOO), 1,2-olein elaidin (OOE) and 1,2-elaidin olein (EEO) with a model crystalline nanoplatelet composed of tristearin in an undefined polymorphic form. The surface of the CNP in contact with the oil was assumed to be planar. We considered pure OOO and mixtures of OOO + OOE and OOO + EEO with 80% OOO. The last two cases were taken as approximations to high oleic sunflower oil (HOSO). The intent was to investigate whether phase separation on a nanoscale took place. We defined an "oil binding capacity" parameter, B(Q,Q'), relating a state Q to a reference state Q'. We used atomic scale molecular dynamics in the NVT ensemble and computed averages over 1-5 ns. We found that the probability of the OOE phase separating into a layer on the surface of the CNP compared to being retained randomly in an OOO + OOE mix were approximately equal. However, we found that it was probable that the EEO component of an OOO + EEO mix would phase separate and coat the surface of the CNP. These results suggest a mechanism whereby many-component oils undergo phase separation on a nanoscale so as to create a transition oil region between the surface of the CNP and the bulk major oil component (OOO in the case considered here) so as to create the appropriate oil binding capacity for the use to which it is put.

A substitutional mutation in the DNA binding domain of the androgen receptor causes complete androgen insensitivity syndrome.

PubMed

Komori, S; Sakata, K; Kasumi, H; Tsuji, Y; Hamada, K; Koyama, K

1999-10-01

DNA analysis of the androgen receptor gene in a patient with complete androgen insensitivity syndrome identified a substitutional mutation (tyrosine converted to cysteine at position 571) in the DNA binding domain. In vitro transfection experiments with the patients' androgen receptor gene, indicated normal expression of the androgen receptor in transfected COS-7 cells compared to the wild type gene. There was also no evidence of impaired thermal stability of the 5 alpha-dihydrotestosterone-androgen receptor complex. However, the capacity of the androgen receptor to activate target gene transcription was found to be completely disrupted in a luciferase assay. These results confirmed that only one substitutional mutation in the DNA binding domain was related to the pathogenesis of the complete androgen insensitivity syndrome.

Intrinsic thermodynamics of ethoxzolamide inhibitor binding to human carbonic anhydrase XIII

PubMed Central

2012-01-01

Background Human carbonic anhydrases (CAs) play crucial role in various physiological processes including carbon dioxide and hydrocarbon transport, acid homeostasis, biosynthetic reactions, and various pathological processes, especially tumor progression. Therefore, CAs are interesting targets for pharmaceutical research. The structure-activity relationships (SAR) of designed inhibitors require detailed thermodynamic and structural characterization of the binding reaction. Unfortunately, most publications list only the observed thermodynamic parameters that are significantly different from the intrinsic parameters. However, only intrinsic parameters could be used in the rational design and SAR of the novel compounds. Results Intrinsic binding parameters for several inhibitors, including ethoxzolamide, trifluoromethanesulfonamide, and acetazolamide, binding to recombinant human CA XIII isozyme were determined. The parameters were the intrinsic Gibbs free energy, enthalpy, entropy, and the heat capacity. They were determined by titration calorimetry and thermal shift assay in a wide pH and temperature range to dissect all linked protonation reaction contributions. Conclusions Precise determination of the inhibitor binding thermodynamics enabled correct intrinsic affinity and enthalpy ranking of the compounds and provided the means for SAR analysis of other rationally designed CA inhibitors. PMID:22676044

Weak partitioning chromatography for anion exchange purification of monoclonal antibodies.

PubMed

Kelley, Brian D; Tobler, Scott A; Brown, Paul; Coffman, Jonathan L; Godavarti, Ranga; Iskra, Timothy; Switzer, Mary; Vunnum, Suresh

2008-10-15

Weak partitioning chromatography (WPC) is an isocratic chromatographic protein separation method performed under mobile phase conditions where a significant amount of the product protein binds to the resin, well in excess of typical flowthrough operations. The more stringent load and wash conditions lead to improved removal of more tightly binding impurities, although at the cost of a reduction in step yield. The step yield can be restored by extending the column load and incorporating a short wash at the end of the load stage. The use of WPC with anion exchange resins enables a two-column cGMP purification platform to be used for many different mAbs. The operating window for WPC can be easily established using high throughput batch-binding screens. Under conditions that favor very strong product binding, competitive effects from product binding can give rise to a reduction in column loading capacity. Robust performance of WPC anion exchange chromatography has been demonstrated in multiple cGMP mAb purification processes. Excellent clearance of host cell proteins, leached Protein A, DNA, high molecular weight species, and model virus has been achieved. (c) 2008 Wiley Periodicals, Inc.

Thermodynamic stability of carbonic anhydrase: measurements of binding affinity and stoichiometry using ThermoFluor.

PubMed

Matulis, Daumantas; Kranz, James K; Salemme, F Raymond; Todd, Matthew J

2005-04-05

ThermoFluor (a miniaturized high-throughput protein stability assay) was used to analyze the linkage between protein thermal stability and ligand binding. Equilibrium binding ligands increase protein thermal stability by an amount proportional to the concentration and affinity of the ligand. Binding constants (K(b)) were measured by examining the systematic effect of ligand concentration on protein stability. The precise ligand effects depend on the thermodynamics of protein stability: in particular, the unfolding enthalpy. An extension of current theoretical treatments was developed for tight binding inhibitors, where ligand effect on T(m) can also reveal binding stoichiometry. A thermodynamic analysis of carbonic anhydrase by differential scanning calorimetry (DSC) enabled a dissection of the Gibbs free energy of stability into enthalpic and entropic components. Under certain conditions, thermal stability increased by over 30 degrees C; the heat capacity of protein unfolding was estimated from the dependence of calorimetric enthalpy on T(m). The binding affinity of six sulfonamide inhibitors to two isozymes (human type 1 and bovine type 2) was analyzed by both ThermoFluor and isothermal titration calorimetry (ITC), resulting in a good correlation in the rank ordering of ligand affinity. This combined investigation by ThermoFluor, ITC, and DSC provides a detailed picture of the linkage between ligand binding and protein stability. The systematic effect of ligands on stability is shown to be a general tool to measure affinity.

Increased thyrotropin binding in hyperfunctioning thyroid nodules.

PubMed

Müller-Gärtner, H W; Schneider, C; Bay, V; Tadt, A; Rehpenning, W; de Heer, K; Jessel, M

1987-08-01

The object of this study was to investigate TSH receptors in hyperfunctioning thyroid nodules (HFN). In HFN, obtained from seven patients, 125-I-TSH binding as determined by equilibrium binding analysis on particulate membrane preparations, was found to be significantly increased as compared with normal thyroid tissues (five patients; P less than 0.001). Scatchard analysis of TSH-binding revealed two kinds of binding sites for both normal thyroid tissue and HFN, and displayed significantly increased association constants of high- and low-affinity binding sites in HFN (Ka = 11.75 +/- 6.8 10(9) M-1, P less than 0.001 and Ka = 2.1 +/- 1.0 10(7) M-1, P less than 0.025; x +/- SEM) as compared with normal thyroid tissue (Ka = 0.25 +/- 0.06 10(9) M-1, Ka = 0.14 +/- 0.03 10(7) M-1; x +/- SEM). The capacity of the high-affinity binding sites in HFN was found to be decreased (1.8 +/- 1.1 pmol/mg protein, x +/- SEM) in comparison with normal thyroid tissue (4.26 +/- 1.27 pmol/mg protein; x +/- SEM). TSH-receptor autoradiography applied to cryostatic tissue sections confirmed increased TSH binding of the follicular epithelium in HFN. These data suggest that an increased affinity of TSH-receptor sites in HFN in iodine deficient areas may be an important event in thyroid autonomy.

Characterization of Rose Bengal binding to sinusoidal and bile canalicular plasma membrane from rat liver.

PubMed

Yachi, K; Sugiyama, Y; Sawada, Y; Iga, T; Ikeda, Y; Toda, G; Hanano, M

1989-01-16

The binding of Rose bengal, a model organic anion, to sinusoidal and bile canalicular membrane fractions isolated from rat liver was compared. The fluorescence change of Rose bengal after being bound to liver plasma membranes was utilized for measuring the binding. The dissociation constants (Kd = 0.1-0.12 microM) and the binding capacities (n = 11-15 nmol/mg protein) for Rose bengal are comparable between the two membrane fractions, although the n value for sinusoidal membrane is somewhat larger than that for bile canalicular membrane. The Rose bengal binding to both membrane fractions was inhibited by various organic anions at relatively low concentrations, i.e., the half-inhibition concentrations (IC50) for Indocyanine green, sulfobromophthalein, Bromophenol blue and 1-anilino-8-naphthalene sulfonate were 0.1, 100, 1.5-2.5 and 100 microM, respectively, while taurocholate did not inhibit the Rose bengal binding to either membrane fraction at these low concentration ranges. The type of inhibition of sulfobromophthalein and Indocyanine green for Rose bengal binding is different between the two membrane domains. That is, in sinusoidal and bile canalicular membrane fractions, these organic anions exhibit mixed-type and competitive-type inhibition, respectively. It was suggested that the fluorescence method using Rose bengal may provide a simple method for detecting the specific organic anion binding protein(s) in the liver plasma membrane.

Sedimentation properties in density gradients correspond with levels of sperm DNA fragmentation, chromatin compaction and binding affinity to hyaluronic acid.

PubMed

Torabi, Forough; Binduraihem, Adel; Miller, David

2017-03-01

Mature spermatozoa bind hyaluronic acid in the extracellular matrix via hyaladherins. Immature spermatozoa may be unable to interact because they do not express the appropriate hyaladherins on their surface. Fresh human semen samples were fractionated using differential density gradient centrifugation (DDGC) and the ability of these fractions to bind hyaluronic acid was evaluated. The presence of sperm hyaladherins was also assessed. CD44 was located mainly on the acrosome and equatorial segment and became more restricted to the equatorial segment in capacitated spermatozoa. Hyaluronic acid-TRITC (hyaluronic acid conjugated with tetramethylrhodamine isothiocyanante), a generic hyaluronic-acid-binding reagent, labelled the membrane and the neck region, particularly after capacitation. Sperm populations obtained after DDGC or after interaction with hyaluronic acid were assessed for DNA fragmentation and chromatin maturity. Strong relationships between both measures and sperm sedimentation and hyaluronic-acid-binding profiles were revealed. Capacitation enhanced hyaluronic acid binding of both DDGC-pelleted sperm and sperm washed free of seminal fluid. In conclusion, hyaladherins were detected on human sperm and a higher capacity for sperm hyaluronic-acid-binding was shown to correspond with their DDGC sedimentation profiles and with lower levels of DNA fragmentation and better chromatin maturity. Capacitation induced changes in the distribution and presence of hyaladherins may enhance hyaluronic-acid-binding. Copyright © 2016 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

Hypophysectomy eliminates and growth hormone (GH) maintains the midpregnancy elevation in GH receptor and serum binding protein in the mouse

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sanchez-Jimenez, F.; Fielder, P.J.; Martinez, R.R.

1990-02-01

({sup 125}I)Iodomouse GH (({sup 125}I)iodo-mGH) binding to samples of serum and hepatic microsomal membranes was measured in hypophysectomized pregnant, sham-operated pregnant, intact pregnant, and intact adult virgin mice. Surgeries were carried out on day 11 of pregnancy, and the animals were killed on day 14. The binding of mGH to both serum and hepatic microsomal membranes of intact virgin mice was much lower than to those of intact pregnant mice. In hypophysectomized mice, the mGH-binding capacity of both serum and hepatic microsomes decreased to values similar to those of nonpregnant mice. No significant differences were observed between intact and sham-operatedmore » pregnant animals in the maternal serum mGH concentration, the serum GH-binding protein concentration, or the hepatic GH receptor concentration. GH receptor and binding protein-encoding mRNAs were also higher in intact and sham-operated pregnant mice than in virgin and hypophysectomized mice. Hypophysectomized mice were treated with 200 micrograms/day bovine GH, administered by osmotic minipump; after 3 days of treatment, a significant elevation of hepatic GH receptor and serum GH-binding protein levels was observed. These results demonstrate an up-regulation of hepatic GH receptors and serum GH-binding protein by GH during pregnancy in the mouse.« less

Cooperativity and oligomeric status of cardiac muscarinic cholinergic receptors.

PubMed

Park, Paul S-H; Sum, Chi Shing; Pawagi, Asha B; Wells, James W

2002-04-30

Muscarinic cholinergic receptors can appear to be more numerous when labeled by [(3)H]quinuclidinylbenzilate (QNB) than by N-[(3)H]methylscopolamine (NMS). The nature of the implied heterogeneity has been studied with M(2) receptors in detergent-solubilized extracts of porcine atria. The relative capacity for [(3)H]NMS and [(3)H]QNB was about 1 in digitonin-cholate, 0.56 in cholate-NaCl, and 0.44 in Lubrol-PX. Adding digitonin to extracts in cholate-NaCl increased the absolute capacity for both radioligands, and the relative capacity increased to near 1. The latency cannot be attributed to a chemically impure radioligand, instability of the receptor, an irreversible effect of NMS, or a failure to reach equilibrium. Binding at near-saturating concentrations of [(3)H]QNB in cholate-NaCl or Lubrol-PX was blocked fully by unlabeled NMS, which therefore appeared to inhibit noncompetitively at sites inaccessible to radiolabeled NMS. Such an effect is inconsistent with the notion of functionally distinct, noninterconverting, and mutually independent sites. Both the noncompetitive effect of NMS on [(3)H]QNB and the shortfall in capacity for [(3)H]NMS can be described quantitatively in terms of cooperative interactions within a receptor that is at least tetravalent; no comparable agreement is possible with a receptor that is only di- or trivalent. The M(2) muscarinic receptor therefore appears to comprise at least four interacting sites, presumably within a tetramer or larger array, and ligands appear to bind in a cooperative manner under at least some conditions.

Development and characterization of methacrylate-based hydrazide monoliths for oriented immobilization of antibodies.

PubMed

Brne, P; Lim, Y-P; Podgornik, A; Barut, M; Pihlar, B; Strancar, A

2009-03-27

Convective interaction media (CIM; BIA Separations) monoliths are attractive stationary phases for use in affinity chromatography because they enable fast affinity binding, which is a consequence of convectively enhanced mass transport. This work focuses on the development of novel CIM hydrazide (HZ) monoliths for the oriented immobilization of antibodies. Adipic acid dihydrazide (AADH) was covalently bound to CIM epoxy monoliths to gain hydrazide groups on the monolith surface. Two different antibodies were afterwards immobilized to hydrazide functionalized monolithic columns and prepared columns were tested for their selectivity. One column was further tested for the dynamic binding capacity.

12 CFR Appendix C to Part 202 - Sample Notification Forms

Code of Federal Regulations, 2013 CFR

2013-01-01

... Taken and Statement of Reasons Statement of Credit Denial, Termination or Change Date: Applicant's Name... report. Your credit score can change, depending on how the information in your consumer report changes..., national origin, sex, marital status, age (provided the applicant has the capacity to enter into a binding...

Recognition and binding of β-lactam antibiotics to bovine serum albumin by frontal affinity chromatography in combination with spectroscopy and molecular docking.

PubMed

Li, Qian; Zhang, Tianlong; Bian, Liujiao

2016-03-01

Serum albumins are the most abundant carrier proteins in blood plasma and participate in the binding and transportation of various exogenous and endogenous compounds in the body. This work was designed to investigate the recognition and binding of three typical β-lactam antibiotics including penicillin G (Pen G), penicillin V (Pen V) and cefalexin (Cef) with bovine serum albumin (BSA) by frontal affinity chromatography in combination with UV-vis absorption spectra, fluorescence emission spectra, binding site marker competitive experiment and molecular docking under simulated physiological conditions. The results showed that a BSA only bound with one antibiotic molecule in the binding process, and the binding constants for Pen G-BSA, Pen V-BSA and Cef-BSA complexes were 4.22×10(1), 4.86×10(2) and 3.32×10(3) (L/mol), respectively. All the three β-lactam antibiotics were mainly inserted into the subdomain IIA (binding site 1) of BSA by hydrogen bonds and Van der Waals forces. The binding capacity between the antibiotics and BSA was closely related to the functional groups and flexibility of side chains in antibiotics. This study provided an important insight into the molecular recognition and binding interaction of BSA with β-lactam antibiotics, which may be a useful guideline for the innovative clinical medications and new antibiotic designs with effective pharmacological properties. Copyright © 2016 Elsevier B.V. All rights reserved.

Definition of IgG- and albumin-binding regions of streptococcal protein G.

PubMed

Akerström, B; Nielsen, E; Björck, L

1987-10-05

Protein G, the immunoglobin G-binding surface protein of group C and G streptococci, also binds serum albumin. The albumin-binding site on protein G is distinct from the immunoglobulin G-binding site. By mild acid hydrolysis of the papain-liberated protein G fragment (35 kDa), a 28-kDa fragment was produced which retained full immunoglobulin G-binding activity (determined by Scatchard plotting) but had lost all albumin-binding capacity. A protein G (65 kDa), isolated after cloning and expression of the protein G gene in Escherichia coli, had comparable affinity to immunoglobulin G (5-10 X 10(10)M-1), but much higher affinity to albumin than the 35- and 28-kDa protein G fragments (31, 2.6, and 0 X 10(9)M-1, respectively). The amino-terminal amino acid sequences of the 65-, 35-, and 28-kDa fragments allowed us to exactly locate the three fragments in an overall sequence map of protein G, based on the partial gene sequences published by Guss et al. (Guss, B., Eliasson, M., Olsson, A., Uhlen, M., Frej, A.-K., Jörnvall, H., Flock, J.-I., and Lindberg, M. (1986) EMBO J. 5, 1567-1575) and Fahnestock et al. (Fahnestock, S. R., Alexander, P., Nagle, J., and Filpula, D. (1986) J. Bacteriol. 167, 870-880). In this map could then be deduced the location of three homologous albumin-binding regions and three homologous immunoglobulin G-binding regions.

The pathogen-related yeast protein Pry1, a member of the CAP protein superfamily, is a fatty acid-binding protein

PubMed Central

Darwiche, Rabih; Mène-Saffrané, Laurent; Gfeller, David; Asojo, Oluwatoyin A.; Schneiter, Roger

2017-01-01

Members of the CAP superfamily (cysteine-rich secretory proteins, antigen 5, and pathogenesis-related 1 proteins), also known as SCP superfamily (sperm-coating proteins), have been implicated in many physiological processes, including immune defenses, venom toxicity, and sperm maturation. Their mode of action, however, remains poorly understood. Three proteins of the CAP superfamily, Pry1, -2, and -3 (pathogen related in yeast), are encoded in the Saccharomyces cerevisiae genome. We have shown previously that Pry1 binds cholesterol in vitro and that Pry function is required for sterol secretion in yeast cells, indicating that members of this superfamily may generally bind sterols or related small hydrophobic compounds. On the other hand, tablysin-15, a CAP protein from the horsefly Tabanus yao, has been shown to bind leukotrienes and free fatty acids in vitro. Therefore, here we assessed whether the yeast Pry1 protein binds fatty acids. Computational modeling and site-directed mutagenesis indicated that the mode of fatty acid binding is conserved between tablysin-15 and Pry1. Pry1 bound fatty acids with micromolar affinity in vitro, and its function was essential for fatty acid export in cells lacking the acyl-CoA synthetases Faa1 and Faa4. Fatty acid binding of Pry1 is independent of its capacity to bind sterols, and the two sterol- and fatty acid-binding sites are nonoverlapping. These results indicate that some CAP family members, such as Pry1, can bind different lipids, particularly sterols and fatty acids, at distinct binding sites, suggesting that the CAP domain may serve as a stable, secreted protein domain that can accommodate multiple ligand-binding sites. PMID:28365570

The GM2 Glycan Serves as a Functional Coreceptor for Serotype 1 Reovirus

PubMed Central

Liu, Yan; Blaum, Bärbel S.; Reiter, Dirk M.; Feizi, Ten; Dermody, Terence S.; Stehle, Thilo

2012-01-01

Viral attachment to target cells is the first step in infection and also serves as a determinant of tropism. Like many viruses, mammalian reoviruses bind with low affinity to cell-surface carbohydrate receptors to initiate the infectious process. Reoviruses disseminate with serotype-specific tropism in the host, which may be explained by differential glycan utilization. Although α2,3-linked sialylated oligosaccharides serve as carbohydrate receptors for type 3 reoviruses, neither a specific glycan bound by any reovirus serotype nor the function of glycan binding in type 1 reovirus infection was known. We have identified the oligosaccharide portion of ganglioside GM2 (the GM2 glycan) as a receptor for the attachment protein σ1 of reovirus strain type 1 Lang (T1L) using glycan array screening. The interaction of T1L σ1 with GM2 in solution was confirmed using NMR spectroscopy. We established that GM2 glycan engagement is required for optimal infection of mouse embryonic fibroblasts (MEFs) by T1L. Preincubation with GM2 specifically inhibited type 1 but not type 3 reovirus infection of MEFs. To provide a structural basis for these observations, we defined the mode of receptor recognition by determining the crystal structure of T1L σ1 in complex with the GM2 glycan. GM2 binds in a shallow groove in the globular head domain of T1L σ1. Both terminal sugar moieties of the GM2 glycan, N-acetylneuraminic acid and N-acetylgalactosamine, form contacts with the protein, providing an explanation for the observed specificity for GM2. Viruses with mutations in the glycan-binding domain display diminished hemagglutination capacity, a property dependent on glycan binding, and reduced capacity to infect MEFs. Our results define a novel mode of virus-glycan engagement and provide a mechanistic explanation for the serotype-dependent differences in glycan utilization by reovirus. PMID:23236285

The GM2 glycan serves as a functional coreceptor for serotype 1 reovirus.

PubMed

Reiss, Kerstin; Stencel, Jennifer E; Liu, Yan; Blaum, Bärbel S; Reiter, Dirk M; Feizi, Ten; Dermody, Terence S; Stehle, Thilo

2012-01-01

Viral attachment to target cells is the first step in infection and also serves as a determinant of tropism. Like many viruses, mammalian reoviruses bind with low affinity to cell-surface carbohydrate receptors to initiate the infectious process. Reoviruses disseminate with serotype-specific tropism in the host, which may be explained by differential glycan utilization. Although α2,3-linked sialylated oligosaccharides serve as carbohydrate receptors for type 3 reoviruses, neither a specific glycan bound by any reovirus serotype nor the function of glycan binding in type 1 reovirus infection was known. We have identified the oligosaccharide portion of ganglioside GM2 (the GM2 glycan) as a receptor for the attachment protein σ1 of reovirus strain type 1 Lang (T1L) using glycan array screening. The interaction of T1L σ1 with GM2 in solution was confirmed using NMR spectroscopy. We established that GM2 glycan engagement is required for optimal infection of mouse embryonic fibroblasts (MEFs) by T1L. Preincubation with GM2 specifically inhibited type 1 but not type 3 reovirus infection of MEFs. To provide a structural basis for these observations, we defined the mode of receptor recognition by determining the crystal structure of T1L σ1 in complex with the GM2 glycan. GM2 binds in a shallow groove in the globular head domain of T1L σ1. Both terminal sugar moieties of the GM2 glycan, N-acetylneuraminic acid and N-acetylgalactosamine, form contacts with the protein, providing an explanation for the observed specificity for GM2. Viruses with mutations in the glycan-binding domain display diminished hemagglutination capacity, a property dependent on glycan binding, and reduced capacity to infect MEFs. Our results define a novel mode of virus-glycan engagement and provide a mechanistic explanation for the serotype-dependent differences in glycan utilization by reovirus.

Binding strength-associated toxicity reduction by birnessite and hydroxyapatite in Pb and Cd contaminated sediments.

PubMed

Lee, Seungbae; An, Jinsung; Kim, Young-Jin; Nam, Kyoungphile

2011-02-28

In situ stabilization of sediment-bound heavy metals has been proposed as an alternative to ex situ treatment due to the concerns on ecosystem disturbance and remediation cost. The present study was conducted to test the performance of birnessite, hydroxyapatite, and zeolite as stabilizing agents for Pb and Cd in sediment. The heavy metal binding capacity and strength of the stabilizing agents were determined by analyzing Langmuir model parameters. The three agents showed the similar binding capacity (i.e., maximum monolayer sorption constant, K(a)) ranging from 1.13 to 3.62×10(5) mg/kg for Pb and 1.07 to 1.33×10(5) mg/kg for Cd. In contrast, binding strength (i.e., binding energy constant, b) of birnessite and hydroxyapatite was about one order higher than that of zeolite. This is further supported by five-step sequential extraction data: more than 99 and 70% of freshly spiked Pb and Cd were present as not-readily extractable fractions in birnessite and hydroxyapatite, respectively while the fractions were 17.9 and 14.1% in zeolite. Toxicity Characteristic Leaching Procedure (TCLP) test was also conducted to verify the effectiveness of the heavy metal-stabilizing ability of birnessite and hydroxyapatite. Birnessite successfully retained both Pb and Cd against the leaching solution, satisfying the TCLP extract concentration limits (i.e., 5 and 1 mg/L, respectively). However, hydroxyapatite released about 223.7 mg/L of Cd into the solution, which greatly exceeded the limit. The toxicity test with Hyalella azteca showed that their survival rate increased by 92.5-100% when birnessite or hydroxyapatite was added to Pb- or Cd-spiked sediment as a stabilizing agent. Our data demonstrate the potential use of birnessite and hydroxyapatite as an effective in situ remediation means for heavy metal-contaminated sediment with minimal risk to the aquatic ecosystem. Copyright © 2011 Elsevier B.V. All rights reserved.

Kinetic intermediates of unfolding of dimeric prostatic phosphatase.

PubMed

Kuciel, Radosława; Mazurkiewicz, Aleksandra; Dudzik, Paulina

2007-01-01

Kinetics of guanidine hydrochloride (GdnHCl)-induced unfolding of human prostatic acid phosphatase (hPAP), a homodimer of 50 kDa subunit molecular mass was investigated with enzyme activity measurements, capacity for binding an external hydrophobic probe, 1-anilinonaphtalene-8-sulfonate (ANS), accessibility of thiols to reaction with 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) and 2-(4'-maleimidylanilino)naphthalene-6-sulfonate (MIANS) and ability to bind Congo red dye. Kinetic analysis was performed to describe a possible mechanism of hPAP unfolding and dissociation that leads to generation of an inactive monomeric intermediate that resembles, in solution of 1.25 M GdnHCl pH 7.5, at 20 degrees C, in equilibrium, a molten globule state. The reaction of hPAP inactivation in 1.25 M GdnHCl followed first order kinetics with the reaction rate constant 0.0715 +/- 0.0024 min(-1) . The rate constants of similar range were found for the pseudo-first-order reactions of ANS and Congo red binding: 0.0366 +/- 0.0018 min(-1) and 0.0409 +/- 0.0052 min(-1), respectively. Free thiol groups, inaccessible in the native protein, were gradually becoming, with the progress of unfolding, exposed for the reactions with DTNB and MIANS, with the pseudo-first-order reaction rate constants 0.327 +/- 0.014 min(-1) and 0.216 +/- 0.010 min(-1), respectively. The data indicated that in the course of hPAP denaturation exposure of thiol groups to reagents took place faster than the enzyme inactivation and exposure of the protein hydrophobic surface. This suggested the existence of a catalytically active, partially unfolded, but probably dimeric kinetic intermediate in the process of hPAP unfolding. On the other hand, the protein inactivation was accompanied by exposure of a hydrophobic, ANS-binding surface, and with an increased capacity to bind Congo red. Together with previous studies these results suggest that the stability of the catalytically active conformation of the enzyme depends mainly on the dimeric structure of the native hPAP.

S-Glutathionylation Regulates Inflammatory Activities of S100A9*

PubMed Central

Lim, Su Yin; Raftery, Mark J.; Goyette, Jesse; Geczy, Carolyn L.

2010-01-01

Reactive oxygen species generated by activated neutrophils can cause oxidative stress and tissue damage. S100A8 (A8) and S100A9 (A9), abundant in neutrophil cytoplasm, are exquisitely sensitive to oxidation, which may alter their functions. Murine A8 is a neutrophil chemoattractant, but it suppresses leukocyte transmigration in the microcirculation when S-nitrosylated. Glutathione (GSH) modulates intracellular redox, and S-glutathionylation can protect susceptible proteins from oxidative damage and regulate function. We characterized S-glutathionylation of A9; GSSG and GSNO generated S-glutathionylated A8 (A8-SSG) and A9 (A9-SSG) in vitro, whereas only A9-SSG was detected in cytosol of neutrophils activated with phorbol myristate acetate (PMA) but not with fMLP or opsonized zymosan. S-Glutathionylation exposed more hydrophobic regions in Zn2+-bound A9 but did not alter Zn2+ binding affinity. A9-SSG had reduced capacity to form heterocomplexes with A8, but the arachidonic acid binding capacities of A8/A9 and A8/A9-SSG were similar. A9 and A8/A9 bind endothelial cells; S-glutathionylation reduced binding. We found little effect of A9 or A9-SSG on neutrophil CD11b/CD18 expression or neutrophil adhesion to endothelial cells. However, A9, A9-SSG and A8/A9 promoted neutrophil adhesion to fibronectin but, in the presence of A8, A9-mediated adhesion was abrogated by glutathionylation. S-Glutathionylation of A9 may protect its oxidation to higher oligomers and reduce neutrophil binding to the extracellular matrix. This may regulate the magnitude of neutrophil migration in the extravasculature, and together with the functional changes we reported for S-nitrosylated A8, particular oxidative modifications of these proteins may limit tissue damage in acute inflammation. PMID:20223829

Characterization of epitope specificities of reference antibodies used for the quantification of the birch pollen allergen Bet v 1.

PubMed

Brier, S; Le Mignon, M; Jain, K; Lebrun, C; Peurois, F; Kellenberger, C; Bordas-Le Floch, V; Mascarell, L; Nony, E; Moingeon, P

2018-05-01

Accurate allergen quantification is needed to document the consistency of allergen extracts used for immunotherapy. Herein, we characterize the epitope specificities of two monoclonal antibodies used in an ELISA for the quantification of the major birch pollen allergen Bet v 1, established as a reference by the BSP090 European project. The ability of mAbs 5B4 and 6H4 to recognize Bet v 1 isoforms was addressed by immunochromatography. The capacity of each mAb to compete with patients' IgE for binding to Bet v 1 was measured by ELISA inhibition. Epitope mapping was performed by pepscan analysis, site-directed mutagenesis, and hydrogen/deuterium exchange-mass spectrometry. The 5B4 epitope corresponds to a peptide sequence (I56-K68) overlapping with the binding sites of patients' serum IgEs. Mutation of residues P59, E60, and K65 abolishes 5B4 binding to Bet v 1 and reduces the level of IgE recognition. In contrast, 6H4 recognizes a conformational epitope lying opposite to the 5B4 binding site, involving residues located in segments I44-K55 and R70-F79. Substitution of E45 reduces the binding capacity of 6H4, confirming that it is critical for the interaction. Both mAbs interact with >90% of Bet v 1 content present in the birch pollen extract, while displaying a weak cross-reactivity with other allergens of the PR-10 family. MAbs 5B4 and 6H4 recognize structurally distinct epitopes present in the vast majority of Bet v 1 isoforms. These results support the relevance as a reference method of the Bet v 1-specific quantitative ELISA adopted by the European Pharmacopoeia. © 2017 EAACI and John Wiley and Sons A/S. Published by John Wiley and Sons Ltd.

Polymeric brushes as functional templates for immobilizing ribonuclease A: study of binding kinetics and activity.

PubMed

Cullen, Sean P; Liu, Xiaosong; Mandel, Ian C; Himpsel, Franz J; Gopalan, Padma

2008-02-05

The ability to immobilize proteins with high binding capacities on surfaces while maintaining their activity is critical for protein microarrays and other biotechnological applications. We employed poly(acrylic acid) (PAA) brushes as templates to immobilize ribonuclease A (RNase A), which is commonly used to remove RNA from plasmid DNA preparations. The brushes are grown by surface-anchored atom-transfer radical polymerization (ATRP) initiators. RNase A was immobilized by both covalent esterification and a high binding capacity metal-ion complexation method to PAA brushes. The polymer brushes immobilized 30 times more enzyme compared to self-assembled monolayers. As the thickness of the brush increases, the surface density of the RNase A increases monotonically. The immobilization was investigated by ellipsometry, X-ray photoelectron spectroscopy (XPS), thermogravimetric analysis (TGA), and near-edge X-ray absorption fine structure spectroscopy (NEXAFS). The activity of the immobilized RNase A was determined using UV absorbance. As much as 11.0 microg/cm(2) of RNase A was bound to PAA brushes by metal-ion complexation compared to 5.8 microg/cm(2) by covalent immobilization which is 30 and 16 times the estimated mass bound in a monolayer. The calculated diffusion coefficient D was 0.63 x 10(-14) cm(2)/s for metal-ion complexation and 0.71 x 10(-14) cm(2)/s for covalent immobilization. Similar values of D indicate that the binding kinetics is similar, but the thermodynamic equilibrium coverage varies with the binding chemistry. Immobilization kinetics and thermodynamics were characterized by ellipsometry for both methods. A maximum relative activity of 0.70-0.80 was reached between five and nine monolayers of the immobilized enzyme. However, the relative activity for covalent immobilization was greater than that of metal-ion complexation. Covalent esterification resulted in similar temperature dependence as free enzyme, whereas metal-ion complexation showed no temperature dependence indicating a significant change in conformation.

Depigmented allergoids reveal new epitopes with capacity to induce IgG blocking antibodies.

PubMed

López-Matas, M Angeles; Gallego, Mayte; Iraola, Víctor; Robinson, Douglas; Carnés, Jerónimo

2013-01-01

The synthesis of allergen-specific blocking IgGs that interact with IgE after allergen immunotherapy (SIT) has been related to clinical efficacy. The objectives were to investigate the epitope specificity of IgG-antibodies induced by depigmented-polymerized (Dpg-Pol) allergoids and unmodified allergen extracts, and examine IgE-blocking activity of induced IgG-antibodies. Rabbits were immunized with native and Dpg-Pol extracts of birch pollen, and serum samples were obtained. Recognition of linear IgG-epitopes of Bet v 1 and Bet v 2 and the capacity of these IgG-antibodies to block binding of human-IgE was determined. Serum from rabbits immunized with native extracts recognised 11 linear epitopes from Bet v 1, while that from Dpg-Pol-immunized animals recognised 8. For Bet v 2, 8 epitopes were recognized by IgG from native immunized animals, and 9 from Dpg-Pol immunized one. Dpg-Pol and native immunized serum did not always recognise the same epitopes, but specific-IgG from both could block human-IgE binding sites for native extract. Depigmented-polymerized birch extract stimulates the synthesis of specific IgG-antibodies which recognize common but also novel epitopes compared with native extracts. IgG-antibodies induced by Dpg-Pol effectively inhibit human-IgE binding to allergens which may be part of the mechanism of action of SIT.

Roles of Trypanosoma cruzi calreticulin in parasite-host interactions and in tumor growth.

PubMed

Ramírez, Galia; Valck, Carolina; Aguilar, Lorena; Kemmerling, Ulrike; López-Muñoz, Rodrigo; Cabrera, Gonzalo; Morello, Antonio; Ferreira, Jorge; Maya, Juan Diego; Galanti, Norbel; Ferreira, Arturo

2012-10-01

In Latin America, there are about 10-12 million people infected with Trypanosoma cruzi, the agent of Chagas' disease, one of the most important neglected tropical parasitism. Identification of molecular targets, specific for the aggressor or host cells or both, may be useful in the development of pharmacological and/or immunological therapeutic tools. Classic efforts in Chagas' disease explore those strategies. Although the immune system frequently controls parasite aggressions, sterile immunity is seldom achieved and chronic interactions are thus established. However, laboratory-modified immunologic probes aimed at selected parasite targets, may be more effective than their unmodified counterparts. Calreticulin (CRT) from vertebrates is a calcium binding protein, present mainly in the endoplasmic reticulum (ER), where it directs the conformation of proteins and controls calcium levels. We have isolated, gene-cloned, expressed and characterized T. cruzi calreticulin (TcCRT). Upon infection, the parasite can translocate this molecule from the ER to the surface, where it inhibits both the classical and lectin complement pathways. Moreover, by virtue of its capacity to bind and inactivate first complement component C1, it promotes parasite infectivity. These two related properties reside in the central domain of this molecule. A different domain, amino terminal, binds to endothelial cells, thus inhibiting their angiogenic capacity. Since tumor growth depends, to a large extent on angiogenesis, their growth is also inhibited. Copyright © 2012 Elsevier Ltd. All rights reserved.

Comparison of deferasirox and deferoxamine effects on iron overload and immunological changes in patients with blood transfusion-dependent β-thalassemia.

PubMed

Al-Kuraishy, Hayder M; Al-Gareeb, Ali I

2017-01-01

Beta-thalassemias are a cluster of inherited (autosomal recessive) hematological disorders prevalent in the Mediterranean area due to defects in synthesis of β chains of hemoglobin. The aim of present study was to compare the effects of deferasirox and deferoxamine on iron overload and immunological changes in patients with blood transfusion-dependent β-thalassemia major and intermedia. This study involved 64 patients with known cases of β-thalassemia major or intermedia that has been treated with blood transfusion and iron chelators. Serum ferritin, serum iron, serum total iron binding, unsaturated iron-binding capacity (UIBC), and immunological parameters were assessed in deferoxamine and deferasirox-treated patients. In deferoxamine-treated patients, serum ferritin levels were high (8160.33 ± 233.75 ng/dL) compared to deferasirox-treated patients (3000.62 ± 188.23 ng/dL; P < 0.0001), also there were significant differences in serum iron, total iron-binding capacity and UIBC ( P < 0.0001) in deferasirox-treated patients compared to deferoxamine-treated patients. Immunological changes between two treated groups showed insignificant differences in levels of complements (C3 and C4) and immunoglobulin levels (IgM, IgG, and IgA) P > 0.05. This study indicated that deferasirox is more effective than deferoxamine regarding the iron overload but not in the immunological profile in patients with blood transfusion-dependent β-thalassemia.

Laminin-411 Is a Vascular Ligand for MCAM and Facilitates TH17 Cell Entry into the CNS

PubMed Central

Flanagan, Ken; Fitzgerald, Kent; Baker, Jeanne; Regnstrom, Karin; Gardai, Shyra; Bard, Frederique; Mocci, Simonetta; Seto, Pui; You, Monica; Larochelle, Catherine; Prat, Alexandre; Chow, Samuel; Li, Lauri; Vandevert, Chris; Zago, Wagner; Lorenzana, Carlos; Nishioka, Christopher; Hoffman, Jennifer; Botelho, Raquel; Willits, Christopher; Tanaka, Kevin; Johnston, Jennifer; Yednock, Ted

2012-01-01

TH17 cells enter tissues to facilitate pathogenic autoimmune responses, including multiple sclerosis (MS). However, the adhesion molecules involved in the unique migratory capacity of TH17 cells, into both inflamed and uninflamed tissues remain unclear. Herein, we characterize MCAM (CD146) as an adhesion molecule that defines human TH17 cells in the circulation; following in vitro restimulation of human memory T cells, nearly all of the capacity to secrete IL-17 is contained within the population of cells expressing MCAM. Furthermore, we identify the MCAM ligand as laminin 411, an isoform of laminin expressed within the vascular endothelial basement membranes under inflammatory as well as homeotstatic conditions. Purified MCAM-Fc binds to laminin 411 with an affinity of 27 nM, and recognizes vascular basement membranes in mouse and human tissue. MCAM-Fc binding was undetectable in tissue from mice with targeted deletion of laminin 411, indicating that laminin 411 is a major tissue ligand for MCAM. An anti-MCAM monoclonal antibody, selected for inhibition of laminin binding, as well as soluble MCAM-Fc, inhibited T cell adhesion to laminin 411 in vitro. When administered in vivo, the antibody reduced TH17 cell infiltration into the CNS and ameliorated disease in an animal model of MS. Our data suggest that MCAM and laminin 411 interact to facilitate TH17 cell entry into tissues and promote inflammation. PMID:22792325

Fermented inulin hydrolysate by Bifidobacterium breve as cholesterol binder in functional food application

NASA Astrophysics Data System (ADS)

Melanie, Hakiki; Susilowati, Agustine; Maryati, Yati

2017-01-01

Inulin hydrolysate is a result of inulin hydrolysis by inulinase enzyme of Scopulariopsis sp.-CBS1 fungi isolated from dahlia tuber skin in the formation of fructooligosaccharides (FOS) as dietary fiber. Inulin hydrolysate fermented by Bifidobacterium breve has a potential as cholesterol binder in digestive system due to dietary fiber content in inulin. This study was conducted to evaluate the best cholesterol binding capacity by the variation of lactic acid bacteria (LAB) culture concentration of 10%, 20% and 30% (v/v), respectively. Fermentation process were conducted with inulin hydrolysate concentration of 25% (w/v), skim milk 7,5% (w/v) and various LAB culture concentration at 40 °C for 0, 12, 24, 36 and 48 hours. The results showed that the variation of LAB culture concentrations affect the cholesterol binding ability in fermented inulin hydrolysate. The fermentation process with 10% LAB culture concentration at 40°C for 48 hours resulted in the highest cholesterol binding capacity (CBC) of 13,69 mg/g at pH 7and 14,44 mg/g at pH 2 with composition of total acids of 0,787%, soluble dietary fiber of 0,396%, insoluble dietary fiber of 5,47%, total solids of 14,476%, total sugars of 472,484 mg/mL, reducing sugar of 92 mg/mL and total plate count (TPC) of 7,278 log CFU/mL, respectively.

Variability in tannin content, chemistry and activity in a diverse group of tannin containing sorghum cultivars.

PubMed

Kaufman, Rhett C; Herald, Thomas J; Bean, Scott R; Wilson, Jeff D; Tuinstra, Mitch R

2013-03-30

Tannins are large polyphenolic polymers and are known to bind proteins, limiting their digestibility, but are also excellent antioxidants. Numerous studies investigating the functional properties of sorghum tannin have been conducted by comparing grain samples from different sorghum lines without considering the other intrinsic characteristics of the grain. The purpose of this study was to remove the confounding intrinsic factors present in the endosperm so the effect of the tannins could be evaluated utilizing a unique decortication/reconstitution procedure. The tannin content of the 14 cultivars tested ranged from 2.3 to 67.2 catechin equivalents. The bran fractions were studied for their impact on protein binding and antioxidant capacity. Protein digestibility by pepsin ranged from 8% to 58% at the highest tannin level addition. Protein binding ranged from 3.11 to 16.33 g blue bovine serum albumin kg⁻¹ bran. Antioxidant capacity ranged from 81.33 to 1122.54 µmol Trolox equivalents g⁻¹ bran. High-performance size-exclusion chromatography detailed molecular size distributions of the tannin polymers and relationship to tannin functionality. The tannin content and composition play a significant role in determining tannin functionality. These differences will allow for selections of high-tannin sorghums with consideration of the biological activities of the tannins. Published 2012. This article is a U.S. Government work and is in the public domain in the USA.

Energetic basis on interactions between ferredoxin and ferredoxin NADP{sup +} reductase at varying physiological conditions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kinoshita, Misaki; Kim, Ju Yaen; Kume, Satoshi

In spite of a number of studies to characterize ferredoxin (Fd):ferredoxin NADP{sup +} reductase (FNR) interactions at limited conditions, detailed energetic investigation on how these proteins interact under near physiological conditions and its linkage to FNR activity are still lacking. We herein performed systematic Fd:FNR binding thermodynamics using isothermal titration calorimetry (ITC) at distinct pH (6.0 and 8.0), NaCl concentrations (0–200 mM), and temperatures (19–28 °C) for mimicking physiological conditions in chloroplasts. Energetically unfavorable endothermic enthalpy changes were accompanied by Fd:FNR complexation at all conditions. This energetic cost was compensated by favorable entropy changes, balanced by conformational and hydrational entropy. Increases inmore » the NaCl concentration and pH weakened interprotein affinity due to the less contribution of favorable entropy change regardless of energetic gains from enthalpy changes, suggesting that entropy drove complexation and modulated affinity. Effects of temperature on binding thermodynamics were much smaller than those of pH and NaCl. NaCl concentration and pH-dependent enthalpy and heat capacity changes provided clues for distinct binding modes. Moreover, decreases in the enthalpy level in the Hammond's postulate-based energy landscape implicated kinetic advantages for FNR activity. All these energetic interplays were comprehensively demonstrated by the driving force plot with the enthalpy-entropy compensation which may serve as an energetic buffer against outer stresses. We propose that high affinity at pH 6.0 may be beneficial for protection from proteolysis of Fd and FNR in rest states, and moderate affinity at pH 8.0 and proper NaCl concentrations with smaller endothermic enthalpy changes may contribute to increase FNR activity. - Highlights: • Energetics of Fd:FNR binding were examined by considering physiological conditions. • NaCl and pH affect energetically Fd:FNR binding with minimal effects of temperature. • Enthalpy and heat capacity may modulate binding kinetics and modes for FNR activity. • Entropy drives complexation by overcoming unfavorable enthalpy and tunes affinity. • Driving force plot reveals condition-dependent energetic interplays for complexation.« less

Mapping of monoclonal antibody- and receptor-binding domains on human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) using a surface plasmon resonance-based biosensor.

PubMed

Laricchia-Robbio, L; Liedberg, B; Platou-Vikinge, T; Rovero, P; Beffy, P; Revoltella, R P

1996-10-01

An automated surface plasmon resonance (SPR)-based biosensor system has been used for mapping antibody and receptor-binding regions on the recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) molecule. A rabbit antimouse IgG1-Fc antibody (RAM.Fc) was coupled to an extended carboxymethylated-hydrogel matrix attached to a gold surface in order to capture an anti-rhGM-CSF monoclonal antibody (MAb) injected over the sensing layer. rhGM-CSF was subsequently injected and allowed to bind to this antibody. Multisite binding assays were then performed, by flowing sequentially other antibodies and peptides over the surface, and the capacity of the latter to interact with the entrapped rhGM-CSF in a multimolecular complex was monitored in real time with SPR. Eleven MAb (all IgG1K), were analyzed: respectively, four antipeptide MAb raised against three distinct epitopes of the cytokine (two clones against residues 14-24, that includes part of the first alpha-helix toward the N-terminal region; one clone against peptide 30-41, an intrahelical loop; and one clone against residues 79-91, including part of the third alpha-helix) and seven antiprotein MAbs raised against the entire rhGM-CSF, whose target native epitopes are still undetermined. In addition, the binding capacity to rhGM-CSF of a synthetic peptide, corresponding to residues 238-254 of the extracellular human GM-CSF receptor alpha-chain, endowed with rhGM-CSF binding activity, was tested. The results from experiments performed with the biosensor were compared with those obtained by a sandwich enzyme-linked immunosorbent assay (ELISA), using the same reagents. The features of the biosensor technology (fully automated, measure in real time, sharpened yes/no response, less background disturbances, no need for washing step or labeling of the reagent) offered several advantages in these studies of MAb immunoreactivity and epitope mapping, giving a much better resolution and enabling more distinct epitopes to be identified over ELISA.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Steiner, B.; Cousot, D.; Trzeciak, A.

The platelet glycoprotein IIb-IIIa complex (GP IIb-IIIa) is a member of the integrin receptor family that recognizes adhesive proteins containing the Arg-Gly-Asp (RGD) sequence. In the present study the binding characteristics of the synthetic hexapeptide Tyr-Asn-Arg-Gly-Asp-Ser (YNRGDS, a sequence present in the fibrinogen alpha-chain at position 570-575) to purified GP IIb-IIIa were determined by equilibrium dialysis. The binding of 125I-YNRGDS to GP IIb-IIIa was specific, saturable, and reversible. The apparent dissociation constant was 1.0 +/- 0.2 microM, and the maximal binding capacity was 0.92 +/- 0.02 mol of 125I-YNRGDS/mol of GP IIb-IIIa, indicating that GP IIb-IIIa contains a single bindingmore » site for RGD peptides. The binding of 125I-YNRGDS to purified GP IIb-IIIa showed many of the characteristics of fibrinogen binding to activated platelets: the binding was inhibited by fibrinogen, by the monoclonal antibody A2A9, and by the dodecapeptide from the C terminus of the fibrinogen gamma-chain. In addition, the binding of 125I-YNRGDS to GP IIb-IIIa was divalent cation-dependent. Our data suggest that two divalent cation binding sites must be occupied for YNRGDS to bind: one site is specific for calcium and is saturated at 1 microM free Ca2+, whereas the other site is less specific and reaches saturation at millimolar concentrations of either Ca2+ or Mg2+. The results of the present study support the hypothesis that the RGD domains within the adhesive proteins are responsible for their binding to GP IIb-IIIa.« less

Effects of DDT and Triclosan on Tumor-cell Binding Capacity and Cell-Surface Protein Expression of Human Natural Killer Cells

PubMed Central

Hurd-Brown, Tasia; Udoji, Felicia; Martin, Tamara; Whalen, Margaret M.

2012-01-01

1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane (DDT) and triclosan (TCS) are organochlorine (OC) compounds that contaminate the environment, are found in human blood, and have been shown to decrease the tumor-cell killing (lytic) function of human natural killer (NK) cells. NK cells defend against tumor cells and virally infected cells. They bind to these targets, utilizing a variety of cell surface proteins. This study examined concentrations of DDT and TCS that decrease lytic function for alteration of NK binding to tumor targets. Levels of either compound that caused loss of binding function were then examined for effects on expression of cell-surface proteins needed for binding. NK cells exposed to 2.5 μM DDT for 24 h (which caused a greater than 55% loss of lytic function) showed a decrease in NK binding function of about 22%, and a decrease in CD16 cell-surface protein of 20%. NK cells exposed to 5 μM TCS for 24 h showed a decrease in ability to bind tumor cells of 37% and a decrease in expression of CD56 of about 34%. This same treatment caused a decrease in lytic function of greater than 87%. These results indicated that only a portion of the loss of NK lytic function seen with exposures to these compounds could be accounted for by loss of binding function. They also showed that loss of binding function is accompanied by a loss cell-surface proteins important in binding function. PMID:22729613

The complex nature of calcium cation interactions with phospholipid bilayers

PubMed Central

Melcrová, Adéla; Pokorna, Sarka; Pullanchery, Saranya; Kohagen, Miriam; Jurkiewicz, Piotr; Hof, Martin; Jungwirth, Pavel; Cremer, Paul S.; Cwiklik, Lukasz

2016-01-01

Understanding interactions of calcium with lipid membranes at the molecular level is of great importance in light of their involvement in calcium signaling, association of proteins with cellular membranes, and membrane fusion. We quantify these interactions in detail by employing a combination of spectroscopic methods with atomistic molecular dynamics simulations. Namely, time-resolved fluorescent spectroscopy of lipid vesicles and vibrational sum frequency spectroscopy of lipid monolayers are used to characterize local binding sites of calcium in zwitterionic and anionic model lipid assemblies, while dynamic light scattering and zeta potential measurements are employed for macroscopic characterization of lipid vesicles in calcium-containing environments. To gain additional atomic-level information, the experiments are complemented by molecular simulations that utilize an accurate force field for calcium ions with scaled charges effectively accounting for electronic polarization effects. We demonstrate that lipid membranes have substantial calcium-binding capacity, with several types of binding sites present. Significantly, the binding mode depends on calcium concentration with important implications for calcium buffering, synaptic plasticity, and protein-membrane association. PMID:27905555

DNA/RNA binding and anticancer/antimicrobial activities of polymer-copper(II) complexes

NASA Astrophysics Data System (ADS)

Lakshmipraba, Jagadeesan; Arunachalam, Sankaralingam; Riyasdeen, Anvarbatcha; Dhivya, Rajakumar; Vignesh, Sivanandham; Akbarsha, Mohammad Abdulkader; James, Rathinam Arthur

2013-05-01

Water soluble polymer-copper(II) complexes with various degrees of coordination in the polymer chain were synthesized and characterized by elemental analysis, IR, UV-visible and EPR spectra. The DNA/RNA binding behavior of these polymer-copper(II) complexes was examined by UV-visible absorption, emission and circular dichroism spectroscopic methods, and cyclic voltammetry techniques. The binding of the polymer-copper(II) complexes with DNA/RNA was mainly through intercalation but some amount of electrostatic interaction was also observed. This binding capacity increased with the degree of coordination of the complexes. The polymer-copper(II) complex having the highest degree of coordination was subjected to analysis of cytotoxic and antimicrobial properties. The cytotoxicity study indicated that the polymer-copper(II) complexes affected the viability of MCF-7 mammary carcinoma cells, and the cells responded to the treatment with mostly through apoptosis although a few cells succumbed to necrosis. The antimicrobial screening showed activity against some human pathogens.

An unfolded protein-induced conformational switch activates mammalian IRE1

PubMed Central

Acosta-Alvear, Diego; Nguyen, Hieu T; Lee, Crystal P; Chu, Feixia

2017-01-01

The unfolded protein response (UPR) adjusts the cell’s protein folding capacity in the endoplasmic reticulum (ER) according to need. IRE1 is the most conserved UPR sensor in eukaryotic cells. It has remained controversial, however, whether mammalian and yeast IRE1 use a common mechanism for ER stress sensing. Here, we show that similar to yeast, human IRE1α’s ER-lumenal domain (hIRE1α LD) binds peptides with a characteristic amino acid bias. Peptides and unfolded proteins bind to hIRE1α LD’s MHC-like groove and induce allosteric changes that lead to its oligomerization. Mutation of a hydrophobic patch at the oligomerization interface decoupled peptide binding to hIRE1α LD from its oligomerization, yet retained peptide-induced allosteric coupling within the domain. Importantly, impairing oligomerization of hIRE1α LD abolished IRE1’s activity in living cells. Our results provide evidence for a unifying mechanism of IRE1 activation that relies on unfolded protein binding-induced oligomerization. PMID:28971800

A glucose-responsive insulin therapy protects animals against hypoglycemia

PubMed Central

Yang, Ruojing; Wu, Margaret; Lin, Songnian; Nargund, Ravi P.; Li, Xinghai; Kelly, Theresa; Yan, Lin; Dai, Ge; Qian, Ying; Dallas-yang, Qing; Fischer, Paul A.; Cui, Yan; Shen, Xiaolan; Huo, Pei; Feng, Danqing Dennis; Erion, Mark D.; Kelley, David E.

2018-01-01

Hypoglycemia is commonly associated with insulin therapy, limiting both its safety and efficacy. The concept of modifying insulin to render its glucose-responsive release from an injection depot (of an insulin complexed exogenously with a recombinant lectin) was proposed approximately 4 decades ago but has been challenging to achieve. Data presented here demonstrate that mannosylated insulin analogs can undergo an additional route of clearance as result of their interaction with endogenous mannose receptor (MR), and this can occur in a glucose-dependent fashion, with increased binding to MR at low glucose. Yet, these analogs retain capacity for binding to the insulin receptor (IR). When the blood glucose level is elevated, as in individuals with diabetes mellitus, MR binding diminishes due to glucose competition, leading to reduced MR-mediated clearance and increased partitioning for IR binding and consequent glucose lowering. These studies demonstrate that a glucose-dependent locus of insulin clearance and, hence, insulin action can be achieved by targeting MR and IR concurrently. PMID:29321379

Structural and immunologic characterization of bovine, horse, and rabbit serum albumins

PubMed Central

Majorek, Karolina A.; Porebski, Przemyslaw J.; Dayal, Arjun; Zimmerman, Matthew D.; Jablonska, Kamila; Stewart, Alan J.; Chruszcz, Maksymilian; Minor, Wladek

2012-01-01

Serum albumin (SA) is the most abundant plasma protein in mammals. SA is a multifunctional protein with extraordinary ligand binding capacity, making it a transporter molecule for a diverse range of metabolites, drugs, nutrients, metals and other molecules. Due to its ligand binding properties, albumins have wide clinical, pharmaceutical, and biochemical applications. Albumins are also allergenic, and exhibit a high degree of cross-reactivity due to significant sequence and structure similarity of SAs from different organisms. Here we present crystal structures of albumins from cattle (BSA), horse (ESA) and rabbit (RSA) serums. The structural data are correlated with the results of immunological studies of SAs. We also analyze the conservation or divergence of structures and sequences of SAs in the context of their potential allergenicity and cross-reactivity. In addition, we identified a previously uncharacterized ligand binding site in the structure of RSA, and calcium binding sites in the structure of BSA, which is the first serum albumin structure to contain metal ions. PMID:22677715

RIPiT-Seq: A high-throughput approach for footprinting RNA:protein complexes

PubMed Central

Singh, Guramrit; Ricci, Emiliano P.; Moore, Melissa J.

2013-01-01

Development of high-throughput approaches to map the RNA interaction sites of individual RNA binding proteins (RBPs) transcriptome-wide is rapidly transforming our understanding of post-transcriptional gene regulatory mechanisms. Here we describe a ribonucleoprotein (RNP) footprinting approach we recently developed for identifying occupancy sites of both individual RBPs and multi-subunit RNP complexes. RNA:protein immunoprecipitation in tandem (RIPiT) yields highly specific RNA footprints of cellular RNPs isolated via two sequential purifications; the resulting RNA footprints can then be identified by high-throughput sequencing (Seq). RIPiT-Seq is broadly applicable to all RBPs regardless of their RNA binding mode and thus provides a means to map the RNA binding sites of RBPs with poor inherent ultraviolet (UV) crosslinkability. Further, among current high-throughput approaches, RIPiT has the unique capacity to differentiate binding sites of RNPs with overlapping protein composition. It is therefore particularly suited for studying dynamic RNP assemblages whose composition evolves as gene expression proceeds. PMID:24096052

The complex nature of calcium cation interactions with phospholipid bilayers

NASA Astrophysics Data System (ADS)

Melcrová, Adéla; Pokorna, Sarka; Pullanchery, Saranya; Kohagen, Miriam; Jurkiewicz, Piotr; Hof, Martin; Jungwirth, Pavel; Cremer, Paul S.; Cwiklik, Lukasz

2016-12-01

Understanding interactions of calcium with lipid membranes at the molecular level is of great importance in light of their involvement in calcium signaling, association of proteins with cellular membranes, and membrane fusion. We quantify these interactions in detail by employing a combination of spectroscopic methods with atomistic molecular dynamics simulations. Namely, time-resolved fluorescent spectroscopy of lipid vesicles and vibrational sum frequency spectroscopy of lipid monolayers are used to characterize local binding sites of calcium in zwitterionic and anionic model lipid assemblies, while dynamic light scattering and zeta potential measurements are employed for macroscopic characterization of lipid vesicles in calcium-containing environments. To gain additional atomic-level information, the experiments are complemented by molecular simulations that utilize an accurate force field for calcium ions with scaled charges effectively accounting for electronic polarization effects. We demonstrate that lipid membranes have substantial calcium-binding capacity, with several types of binding sites present. Significantly, the binding mode depends on calcium concentration with important implications for calcium buffering, synaptic plasticity, and protein-membrane association.

Combined effects of a thymic peptide, thymopoietin and myasthenic patient sera in rat myotube culture.

PubMed

Eymard, B; Aimé, C; Cottin, C; Morel, E; Goldstein, G; Bach, J F; Berrih-Aknin, S

1992-10-01

We investigated in a rat myotube assay the combined effect of 26 myasthenic (MG) patient sera and a thymic peptide, thymopoietin (Tpo) which had previously been shown to bind Torpedo and human AChR and to compete with alpha-bungarotoxin (alpha-Bgt) binding. Cultures were first exposed to Tpo alone for 3 h (0.3, 7.5, 15 nM), then MG sera (5% final dilution) were added for an additional 18 h. Reduction in the amount of 125I-alpha-Bgt binding sites in the presence of various concentrations of Tpo were similar with control sera and in all the patients with low or undetectable anti-AChR Ab (11 cases). In cultures exposed to Tpo and sera with high anti-AChR Ab titre (15 cases), Tpo and anti-AChR Ab have an additive capacity to reduce the number of alpha-Bgt binding sites. The results are compatible with the hypothesis that anti-AChR Ab and Tpo could impair neuromuscular transmission by complementary mechanisms.