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Sample records for alanine aminotransferase creatine

  1. Alanine aminotransferase controls seed dormancy in barley

    PubMed Central

    Sato, Kazuhiro; Yamane, Miki; Yamaji, Nami; Kanamori, Hiroyuki; Tagiri, Akemi; Schwerdt, Julian G.; Fincher, Geoffrey B.; Matsumoto, Takashi; Takeda, Kazuyoshi; Komatsuda, Takao

    2016-01-01

    Dormancy allows wild barley grains to survive dry summers in the Near East. After domestication, barley was selected for shorter dormancy periods. Here we isolate the major seed dormancy gene qsd1 from wild barley, which encodes an alanine aminotransferase (AlaAT). The seed dormancy gene is expressed specifically in the embryo. The AlaAT isoenzymes encoded by the long and short dormancy alleles differ in a single amino acid residue. The reduced dormancy allele Qsd1 evolved from barleys that were first domesticated in the southern Levant and had the long dormancy qsd1 allele that can be traced back to wild barleys. The reduced dormancy mutation likely contributed to the enhanced performance of barley in industrial applications such as beer and whisky production, which involve controlled germination. In contrast, the long dormancy allele might be used to control pre-harvest sprouting in higher rainfall areas to enhance global adaptation of barley. PMID:27188711

  2. Intramitochondrial localization of alanine aminotransferase in rat-liver mitochondria: comparison with glutaminase and aspartate aminotransferase.

    PubMed

    Masola, B; Devlin, T M

    1995-12-01

    The removal of the outer mitochondrial membrane and hence of constituents of the intermembrane space in rat-liver mitochondria using digitonin showed that phosphate-dependent glutaminase, alanine and aspartate aminotransferase were localized in the mitoplasts. Further fractionation of mitoplasts following their sonication resulted in 90% of glutaminase, 98% of alanine aminotransferase and 48% of aspartate aminotransferase being recovered in the soluble fraction while the remainder of each enzyme was recovered in the sonicated vesicles fraction. These results indicated that glutaminase and alanine aminotransferase were soluble matrix enzymes, the little of each enzyme recovered in the sonicated vesicles fraction being probably due to entrapment in the vesicles. Aspartate aminotransferase had dual localization, in the inner membrane and matrix with the high specific activity in sonicated vesicles confirming its association with the membrane. Activation experiments suggested that the membrane-bound enzyme was localized on the inner side of the inner mitochondrial membrane.

  3. Crystal Structures of Aedes Aegypt Alanine Glyoxylate Aminotransferase

    SciTech Connect

    Han,Q.; Robinson, H.; Gao, Y.; Vogelaar, N.; Wilson, S.; Rizzi, M.; Li, J.

    2006-01-01

    Mosquitoes are unique in having evolved two alanine glyoxylate aminotransferases (AGTs). One is 3-hydroxykynurenine transaminase (HKT), which is primarily responsible for catalyzing the transamination of 3-hydroxykynurenine (3-HK) to xanthurenic acid (XA). Interestingly, XA is used by malaria parasites as a chemical trigger for their development within the mosquito. This 3-HK to XA conversion is considered the major mechanism mosquitoes use to detoxify the chemically reactive and potentially toxic 3-HK. The other AGT is a typical dipteran insect AGT and is specific for converting glyoxylic acid to glycine. Here we report the 1.75{angstrom} high-resolution three-dimensional crystal structure of AGT from the mosquito Aedes aegypti (AeAGT) and structures of its complexes with reactants glyoxylic acid and alanine at 1.75 and 2.1{angstrom} resolution, respectively. This is the first time that the three-dimensional crystal structures of an AGT with its amino acceptor, glyoxylic acid, and amino donor, alanine, have been determined. The protein is dimeric and adopts the type I-fold of pyridoxal 5-phosphate (PLP)-dependent aminotransferases. The PLP co-factor is covalently bound to the active site in the crystal structure, and its binding site is similar to those of other AGTs. The comparison of the AeAGT-glyoxylic acid structure with other AGT structures revealed that these glyoxylic acid binding residues are conserved in most AGTs. Comparison of the AeAGT-alanine structure with that of the Anopheles HKT-inhibitor complex suggests that a Ser-Asn-Phe motif in the latter may be responsible for the substrate specificity of HKT enzymes for 3-HK.

  4. Crystal structures of Aedes aegypti alanine glyoxylate aminotransferase.

    PubMed

    Han, Qian; Robinson, Howard; Gao, Yi Gui; Vogelaar, Nancy; Wilson, Scott R; Rizzi, Menico; Li, Jianyong

    2006-12-01

    Mosquitoes are unique in having evolved two alanine glyoxylate aminotransferases (AGTs). One is 3-hydroxykynurenine transaminase (HKT), which is primarily responsible for catalyzing the transamination of 3-hydroxykynurenine (3-HK) to xanthurenic acid (XA). Interestingly, XA is used by malaria parasites as a chemical trigger for their development within the mosquito. This 3-HK to XA conversion is considered the major mechanism mosquitoes use to detoxify the chemically reactive and potentially toxic 3-HK. The other AGT is a typical dipteran insect AGT and is specific for converting glyoxylic acid to glycine. Here we report the 1.75A high-resolution three-dimensional crystal structure of AGT from the mosquito Aedes aegypti (AeAGT) and structures of its complexes with reactants glyoxylic acid and alanine at 1.75 and 2.1A resolution, respectively. This is the first time that the three-dimensional crystal structures of an AGT with its amino acceptor, glyoxylic acid, and amino donor, alanine, have been determined. The protein is dimeric and adopts the type I-fold of pyridoxal 5-phosphate (PLP)-dependent aminotransferases. The PLP co-factor is covalently bound to the active site in the crystal structure, and its binding site is similar to those of other AGTs. The comparison of the AeAGT-glyoxylic acid structure with other AGT structures revealed that these glyoxylic acid binding residues are conserved in most AGTs. Comparison of the AeAGT-alanine structure with that of the Anopheles HKT-inhibitor complex suggests that a Ser-Asn-Phe motif in the latter may be responsible for the substrate specificity of HKT enzymes for 3-HK.

  5. Alanine Aminotransferase Variants Conferring Diverse NUE Phenotypes in Arabidopsis thaliana

    PubMed Central

    McAllister, Chandra H.; Good, Allen G.

    2015-01-01

    Alanine aminotransferase (AlaAT, E.C. 2.6.1.2), is a pyridoxal-5’-phosphate-dependent (PLP) enzyme that catalyzes the reversible transfer of an amino group from alanine to 2-oxoglutarate to produce glutamate and pyruvate, or vice versa. It has been well documented in both greenhouse and field studies that tissue-specific over-expression of AlaAT from barley (Hordeum vulgare, HvAlaAT) results in a significant increase in plant NUE in both canola and rice. While the physical phenotypes associated with over-expression of HvAlaAT have been well characterized, the role this enzyme plays in vivo to create a more N efficient plant remains unknown. Furthermore, the importance of HvAlaAT, in contrast to other AlaAT enzyme homologues in creating this phenotype has not yet been explored. To address the role of AlaAT in NUE, AlaAT variants from diverse sources and different subcellular locations, were expressed in the wild-type Arabidopsis thaliana Col-0 background and alaat1;2 (alaat1-1;alaat2-1) knockout background in various N environments. The analysis and comparison of both the physical and physiological properties of AlaAT over-expressing transgenic plants demonstrated significant differences between plants expressing the different AlaAT enzymes under different external conditions. This analysis indicates that the over-expression of AlaAT variants other than HvAlaAT in crop plants could further increase the NUE phenotype(s) previously observed. PMID:25830496

  6. Alanine aminotransferase variants conferring diverse NUE phenotypes in Arabidopsis thaliana.

    PubMed

    McAllister, Chandra H; Good, Allen G

    2015-01-01

    Alanine aminotransferase (AlaAT, E.C. 2.6.1.2), is a pyridoxal-5'-phosphate-dependent (PLP) enzyme that catalyzes the reversible transfer of an amino group from alanine to 2-oxoglutarate to produce glutamate and pyruvate, or vice versa. It has been well documented in both greenhouse and field studies that tissue-specific over-expression of AlaAT from barley (Hordeum vulgare, HvAlaAT) results in a significant increase in plant NUE in both canola and rice. While the physical phenotypes associated with over-expression of HvAlaAT have been well characterized, the role this enzyme plays in vivo to create a more N efficient plant remains unknown. Furthermore, the importance of HvAlaAT, in contrast to other AlaAT enzyme homologues in creating this phenotype has not yet been explored. To address the role of AlaAT in NUE, AlaAT variants from diverse sources and different subcellular locations, were expressed in the wild-type Arabidopsis thaliana Col-0 background and alaat1;2 (alaat1-1;alaat2-1) knockout background in various N environments. The analysis and comparison of both the physical and physiological properties of AlaAT over-expressing transgenic plants demonstrated significant differences between plants expressing the different AlaAT enzymes under different external conditions. This analysis indicates that the over-expression of AlaAT variants other than HvAlaAT in crop plants could further increase the NUE phenotype(s) previously observed.

  7. Serum Alanine Aminotransferase Levels, Hematocrit Rate and Body Weight Correlations Before and After Hemodialysis Session

    PubMed Central

    Lopes, Edmundo Pessoa; Sette, Luis Henrique B. C.; Sette, Jorge Bezerra C.; Luna, Carlos F.; Andrade, Amaro M.; Moraes, Maviael; Sette, Paulo C. A.; Menezes, Roberto; Cavalcanti, Rui L.; Conceição, Sergio C.

    2009-01-01

    PURPOSE To evaluate alanine aminotransferase levels before and after a hemodialysis session and to correlate these values with the hematocrit rate and weight loss during hemodialysis. PATIENTS AND METHODS The serum alanine aminotransferase levels, hematocrit rate and body weight were measured and correlated before and after a single hemodialysis session for 146 patients with chronic renal failure. An receiver operating characteristic (ROC) curve for the serum alanine aminotransferase levels collected before and after hemodialysis was plotted to identify hepatitis C virus-infected patients. RESULTS The mean weight loss of the 146 patients during hemodialysis was 5.3% (p < 0.001). The mean alanine aminotransferase levels before and after hemodialysis were 18.8 and 23.9 IU/, respectively, denoting a significant 28.1% increase. An equally significant increase of 16.4% in the hematocrit rate also occurred after hemodialysis. The weight loss was inversely correlated with the rise in both the alanine aminotransferase level (r = 0.3; p < 0.001) and hematocrit rate (r = 0.5; p < 0.001). A direct correlation was found between the rise in alanine aminotransferase levels and the hematocrit during the hemodialysis session (r = 0.4; p < 0.001). Based on the ROC curve, the upper limit of the normal alanine aminotransferase level should be reduced by 40% relative to the upper limit of normal if the blood samples are collected before the hemodialysis session or by 60% if blood samples are collected after the session. CONCLUSION In the present study, significant elevations in the serum alanine aminotransferase levels and hematocrit rates occurred in parallel to a reduction in body weight after the hemodialysis session. These findings suggest that one of the factors for low alanine aminotransferase levels prior to hemodialysis could be hemodilution in patients with chronic renal failure. PMID:19841699

  8. Observations of alanine aminotransferase and aspartate aminotransferase in THRIVE studies treated orally with ximelagatran.

    PubMed

    Harenberg, Job; Jörg, Ingrid; Weiss, Christel

    2006-01-01

    Treatment of acute venous thromboembolism (VTE) and prophylaxis of recurrent events has been investigated in the THRIVE (THRombin Inhibitor in Venous Thrombe Embolism) Treatment and the THRIVE III trial using the oral direct thrombin inhibitor ximelagatran. Alanine aminotransferase (ALAT) increased in 9.6% and 6.4% of patients in the THRIVE Treatment and THRIVE III trials, respectively. The authors analysed the time course of the ALAT and in additionally of aspartate aminotransferase (ASAT) in blood from 52 and 23 patients participating in the THRIVE Treatment and the THRIVE III trials in Germany. Analysis of variance for repeated measures and t test were performed. In the THRIVE Treatment trial, ALAT was significantly higher at week 2 for enoxaparin/warfarin (p => .0039, t test) and at months 3 and 6 for ximelagatran (p = .0453, p = .0014, respectively). ASAT and ASAT/ALAT ratio values did not increase and not differ for both groups. In the THRIVE III trial, ALAT and ASAT did not increase and did not differ compared to the comparator placebo. 2 x 36 mg Ximelagatran, induced higher ALAT values at months 3 and 6 compared to 2 x 24 mg ximelagatran (p = .0105, p = .0063, respectively). ASAT did not differ between the two doses of ximelagatran. The ASAT/ALAT ratios were lower at week 2 for enoxaparin/warfarin (t-test, p = .0032) and at month 3 and 6 for 2 x 36 mg versus warfarin or 2 x 24 mg Ximelagatran (p between .0187 and .0002). The authors conclude that ALAT increases dose dependently during therapy with ximelagatran. The less frequent and lower increase of ASAT values compared to ALAT values indicates a nontoxic effect of ximelagatran on liver cells.

  9. Correlation of serum alanine aminotransferase and aspartate aminotransferase with coronary heart disease

    PubMed Central

    Shen, Jianying; Zhang, Jingying; Wen, Jing; Ming, Qiang; Zhang, Ji; Xu, Yawei

    2015-01-01

    Objective: This study aimed to explore the relationship between different risk factors (especially serum alanine aminotransferase [ALT] and aspartate aminotransferase [AST]) and coronary heart disease (CHD). Methods: A total of 610 inpatients were recruited. Initial coronary angiography (CAG) was performed to evaluate the severity of coronary lesions. On the basis of findings from CAG, patients were divided into control group (n=260) and CHD group (n=350). Logistic regression analysis was employed for the evaluation of clinical characteristics and biochemical parameters, aiming to explore the relationship between risk factors (including AST and ALT) and CHD. Results: Results showed type 2 diabetes, hypertension, dyslipidemia, smoking and family history of CHD were clinical risk factors of CHD. Laboratory examinations showed the serum levels of triglycerides, low-density lipoprotein, AST and ALT in CHD group were significantly higher than those in control group (P<0.05). Of these parameters, the AST was 50.98±8.12 U/L in CHD group and 20.14±3.94 U/L in control group (P<0.01); the ALT was 42.31±8.34 U/L in CHD group and 18.25±6.38 U/L in control group (P<0.01). Conclusion: The serum levels of AST and ALT in CHD patients are higher than those in controls. High serum AST and ALT are biochemical markers which can be used to predict the severity of CHD and are also independent risk factors of CHD. PMID:26064360

  10. Beta-alanine/alpha-ketoglutarate aminotransferase for 3-hydroxypropionic acid production

    DOEpatents

    Jessen, Holly Jean [Chanhassen, MN; Liao, Hans H [Eden Prairie, MN; Gort, Steven John [Apple Valley, MN; Selifonova, Olga V [Plymouth, MN

    2011-10-04

    The present disclosure provides novel beta-alanine/alpha ketoglutarate aminotransferase nucleic acid and protein sequences having increased biological activity. Also provided are cells containing such enzymes, as well as methods of their use, for example to produce malonyl semialdehyde and downstream products thereof, such as 3-hydroxypropionic acid and derivatives thereof.

  11. Beta-alanine/alpha-ketoglutarate aminotransferase for 3-hydroxypropionic acid production

    SciTech Connect

    Jessen, Holly Jean; Liao, Hans H; Gort, Steven John; Selifonova, Olga V

    2014-11-18

    The present disclosure provides novel beta-alanine/alpha ketoglutarate aminotransferase nucleic acid and protein sequences having increased biological activity. Also provided are cells containing such enzymes, as well as methods of their use, for example to produce malonyl semialdehyde and downstream products thereof, such as 3-hydroxypropionic acid and derivatives thereof.

  12. Repeated Supramaximal Exercise-Induced Oxidative Stress: Effect of β-Alanine Plus Creatine Supplementation

    PubMed Central

    Belviranli, Muaz; Okudan, Nilsel; Revan, Serkan; Balci, Serdar; Gokbel, Hakki

    2016-01-01

    Background: Carnosine is a dipeptide formed from the β-alanine and histidine amino acids and found in mainly in the brain and muscle, especially fast twitch muscle. Carnosine and creatine has an antioxidant effect and carnosine accounts for about 10% of the muscle's ability to buffer the H+ ions produced by exercise. Objectives: The aim of the study was to investigate the effects of beta alanine and/or creatine supplementation on oxidant and antioxidant status during repeated Wingate tests (WTs). Patients and Methods: Forty four sedentary males participated in the study. Participants performed three 30s WTs with 2 minutes rest between exercise bouts. After the first exercise session, the subjects were assigned to one of four groups: Placebo, Creatine, Beta-alanine and Beta-alanine plus creatine. Participants ingested twice per day for 22 consecutive days, then four times per day for the following 6 days. After the supplementation period the second exercise session was applied. Blood samples were taken before and immediately after the each exercise session for the analysis of oxidative stress and antioxidant markers. Results: Malondialdehyde levels and superoxide dismutase activities were affected by neither supplementation nor exercise. During the pre-supplementation session, protein carbonyl reduced and oxidized glutathione (GSH and GSSG) levels increased immediately after the exercise. However, during the post-supplementation session GSH and GSSG levels increased in beta-alanine and beta-alanine plus creatine groups immediately after the exercise compared to pre-exercise. In addition, during the post-supplementation session total antioxidant capacity increased in beta-alanine group immediately after the exercise. Conclusions: Beta-alanine supplementation has limited antioxidant effect during the repeated WTs. PMID:27217925

  13. Protein Homeostasis Defects of Alanine-Glyoxylate Aminotransferase: New Therapeutic Strategies in Primary Hyperoxaluria Type I

    PubMed Central

    Pey, Angel L.; Albert, Armando; Salido, Eduardo

    2013-01-01

    Alanine-glyoxylate aminotransferase catalyzes the transamination between L-alanine and glyoxylate to produce pyruvate and glycine using pyridoxal 5′-phosphate (PLP) as cofactor. Human alanine-glyoxylate aminotransferase is a peroxisomal enzyme expressed in the hepatocytes, the main site of glyoxylate detoxification. Its deficit causes primary hyperoxaluria type I, a rare but severe inborn error of metabolism. Single amino acid changes are the main type of mutation causing this disease, and considerable effort has been dedicated to the understanding of the molecular consequences of such missense mutations. In this review, we summarize the role of protein homeostasis in the basic mechanisms of primary hyperoxaluria. Intrinsic physicochemical properties of polypeptide chains such as thermodynamic stability, folding, unfolding, and misfolding rates as well as the interaction of different folding states with protein homeostasis networks are essential to understand this disease. The view presented has important implications for the development of new therapeutic strategies based on targeting specific elements of alanine-glyoxylate aminotransferase homeostasis. PMID:23956997

  14. Identification and expression analyses of the alanine aminotransferase (AlaAT) gene family in poplar seedlings.

    PubMed

    Xu, Zhiru; Ma, Jing; Qu, Chunpu; Hu, Yanbo; Hao, Bingqing; Sun, Yan; Liu, Zhongye; Yang, Han; Yang, Chengjun; Wang, Hongwei; Li, Ying; Liu, Guanjun

    2017-04-05

    Alanine aminotransferase (AlaAT, E.C.2.6.1.2) catalyzes the reversible conversion of pyruvate and glutamate to alanine and α-oxoglutarate. The AlaAT gene family has been well studied in some herbaceous plants, but has not been well characterized in woody plants. In this study, we identified four alanine aminotransferase homologues in Populus trichocarpa, which could be classified into two subgroups, A and B. AlaAT3 and AlaAT4 in subgroup A encode AlaAT, while AlaAT1 and AlaAT2 in subgroup B encode glutamate:glyoxylate aminotransferase (GGAT), which catalyzes the reaction of glutamate and glyoxylate to α-oxoglutarate and glycine. Four AlaAT genes were cloned from P. simonii × P. nigra. PnAlaAT1 and PnAlaAT2 were expressed predominantly in leaves and induced by exogenous nitrogen and exhibited a diurnal fluctuation in leaves, but was inhibited in roots. PnAlaAT3 and PnAlaAT4 were mainly expressed in roots, stems and leaves, and was induced by exogenous nitrogen. The expression of PnAlaAT3 gene could be regulated by glutamine or its related metabolites in roots. Our results suggest that PnAlaAT3 gene may play an important role in nitrogen metabolism and is regulated by glutamine or its related metabolites in the roots of P. simonii × P. nigra.

  15. Identification and expression analyses of the alanine aminotransferase (AlaAT) gene family in poplar seedlings

    PubMed Central

    Xu, Zhiru; Ma, Jing; Qu, Chunpu; Hu, Yanbo; Hao, Bingqing; Sun, Yan; Liu, Zhongye; Yang, Han; Yang, Chengjun; Wang, Hongwei; Li, Ying; Liu, Guanjun

    2017-01-01

    Alanine aminotransferase (AlaAT, E.C.2.6.1.2) catalyzes the reversible conversion of pyruvate and glutamate to alanine and α-oxoglutarate. The AlaAT gene family has been well studied in some herbaceous plants, but has not been well characterized in woody plants. In this study, we identified four alanine aminotransferase homologues in Populus trichocarpa, which could be classified into two subgroups, A and B. AlaAT3 and AlaAT4 in subgroup A encode AlaAT, while AlaAT1 and AlaAT2 in subgroup B encode glutamate:glyoxylate aminotransferase (GGAT), which catalyzes the reaction of glutamate and glyoxylate to α-oxoglutarate and glycine. Four AlaAT genes were cloned from P. simonii × P. nigra. PnAlaAT1 and PnAlaAT2 were expressed predominantly in leaves and induced by exogenous nitrogen and exhibited a diurnal fluctuation in leaves, but was inhibited in roots. PnAlaAT3 and PnAlaAT4 were mainly expressed in roots, stems and leaves, and was induced by exogenous nitrogen. The expression of PnAlaAT3 gene could be regulated by glutamine or its related metabolites in roots. Our results suggest that PnAlaAT3 gene may play an important role in nitrogen metabolism and is regulated by glutamine or its related metabolites in the roots of P. simonii × P. nigra. PMID:28378825

  16. Serum γ-Glutamyltransferase, Alanine Aminotransferase and Aspartate Aminotransferase Activity in Healthy Blood Donor of Different Ethnic Groups in Gorgan

    PubMed Central

    Mehrpouya, Masoumeh; Pourhashem, Zeinab

    2016-01-01

    Introduction Measure of liver enzymes may help to increase safety of blood donation for both blood donor and recipient. Determination of liver enzymes may prepare valuable clinical information. Aim To assess serum γ-Glutamyltransferase (GGT), Alanine Aminotransferase (ALT), and Aspartate Aminotransferase (AST) activities in healthy blood donors in different ethnic groups in Gorgan. Materials and Methods This study was performed in 450 healthy male blood donors, in three ethnic groups (Fars, Sistanee and Turkman) who attended Gorgan blood transfusion center. Liver enzymes (GGT, ALT and AST) were determined. Results Serum AST and ALT in three ethnic groups were significant except for serum GGT levels. There was significant correlation between family histories of liver disease and systolic blood pressure and AST in Fars, and GGT in Sistanee ethnic groups. Conclusion Several factors, such as age, family history of diabetes mellitus, family history of liver disease and smoking habit had no effect on some liver enzymes in different ethnic groups in this area. Variation of AST, ALT, and GGT enzyme activities in healthy subjects was associated with some subjects in our study groups. According to our study, it suggests that screening of AST and GGT enzymes in subjects with family history of liver disease is necessary in different ethnic groups. PMID:27630834

  17. PPAR{alpha} regulates the hepatotoxic biomarker alanine aminotransferase (ALT1) gene expression in human hepatocytes

    SciTech Connect

    Thulin, Petra; Rafter, Ingalill; Stockling, Kenneth; Tomkiewicz, Celine; Norjavaara, Ensio; Aggerbeck, Martine; Hellmold, Heike; Ehrenborg, Ewa; Andersson, Ulf; Cotgreave, Ian; Glinghammar, Bjoern

    2008-08-15

    In this work, we investigated a potential mechanism behind the observation of increased aminotransferase levels in a phase I clinical trial using a lipid-lowering drug, the peroxisome proliferator-activated receptor (PPAR) {alpha} agonist, AZD4619. In healthy volunteers treated with AZD4619, serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities were elevated without an increase in other markers for liver injury. These increases in serum aminotransferases have previously been reported in some patients receiving another PPAR{alpha} agonist, fenofibrate. In subsequent in vitro studies, we observed increased expression of ALT1 protein and mRNA in human hepatocytes after treatment with fenofibric acid. The PPAR effect on ALT1 expression was shown to act through a direct transcriptional mechanism involving at least one PPAR response element (PPRE) in the proximal ALT1 promoter, while no effect of fenofibrate and AZD4619 was observed on the ALT2 promoter. Binding of PPARs to the PPRE located at - 574 bp from the transcriptional start site was confirmed on both synthetic oligonucleotides and DNA in hepatocytes. These data show that intracellular ALT expression is regulated by PPAR agonists and that this mechanism might contribute to increased ALT activity in serum.

  18. Inhibition study of alanine aminotransferase enzyme using sequential online capillary electrophoresis analysis.

    PubMed

    Liu, Lina; Chen, Yuanfang; Yang, Li

    2014-12-15

    We report the study of several inhibitors on alanine aminotransferase (ALT) enzyme using sequential online capillary electrophoresis (CE) assay. Using metal ions (Na(+) and Mg(2+)) as example inhibitors, we show that evolution of the ALT inhibition reaction can be achieved by automatically and simultaneously monitoring the substrate consumption and product formation as a function of reaction time. The inhibition mechanism and kinetic constants of ALT inhibition with succinic acid and two traditional Chinese medicines were derived from the sequential online CE assay. Our study could provide valuable information about the inhibition reactions of ALT enzyme.

  19. Alanine Aminotransferase Elevation in Obese Infants and Children: A Marker of Early Onset Non Alcoholic Fatty Liver Disease

    PubMed Central

    Engelmann, Guido; Hoffmann, Georg Friedrich; Grulich-Henn, Juergen; Teufel, Ulrike

    2014-01-01

    Background: Elevated aminotransferases serve as surrogate markers of non-alcoholic fatty liver disease, a feature commonly associated with the metabolic syndrome. Studies on the prevalence of fatty liver disease in obese children comprise small patient samples or focus on those patients with liver enzyme elevation. Objectives: We have prospectively analyzed liver enzymes in all overweight and obese children coming to our tertiary care centre. Patients and Methods: In a prospective study 224 healthy, overweight or obese children aged 1 - 12 years were examined. Body Mass Index-Standard Deviation Score, alanine aminotransferase, aspartate aminotransferase and gamma-glutamyl-transpeptidase were measured. Results: Elevated alanine aminotransferase was observed in 29% of children. 26 % of obese and 30 % of overweight children had liver enzyme elevations. Obese children had significantly higher alanine aminotransferase levels than overweight children (0.9 vs. 0.7 times the Upper Limit of Normal; P = 0.04). Conclusions: Elevation of liver enzymes appears in 29 % obese children in a tertiary care centre. Absolute alanine aminotransferase levels are significantly higher in obese than in overweight children. Even obese children with normal liver enzymes show signs of fatty liver disease as demonstrated by liver enzymes at the upper limit of normal. PMID:24748893

  20. Effect of creatine and beta-alanine supplementation on performance and endocrine responses in strength/power athletes.

    PubMed

    Hoffman, Jay; Ratamess, Nicholas; Kang, Jie; Mangine, Gerald; Faigenbaum, Avery; Stout, Jeffrey

    2006-08-01

    The effects of creatine and creatine plus beta-alanine on strength, power, body composition, and endocrine changes were examined during a 10-wk resistance training program in collegiate football players. Thirty-three male subjects were randomly assigned to either a placebo (P), creatine (C), or creatine plus beta-alanine (CA) group. During each testing session subjects were assessed for strength (maximum bench press and squat), power (Wingate anaerobic power test, 20-jump test), and body composition. Resting blood samples were analyzed for total testosterone, cortisol, growth hormone, IGF-1, and sex hormone binding globulin. Changes in lean body mass and percent body fat were greater (P < 0.05) in CA compared to C or P. Significantly greater strength improvements were seen in CA and C compared to P. Resting testosterone concentrations were elevated in C, however, no other significant endocrine changes were noted. Results of this study demonstrate the efficacy of creatine and creatine plus beta-alanine on strength performance. Creatine plus beta-alanine supplementation appeared to have the greatest effect on lean tissue accruement and body fat composition.

  1. Effective disposal of nitrogen waste in blood-fed Aedes aegypti mosquitoes requires alanine aminotransferase

    PubMed Central

    Mazzalupo, Stacy; Isoe, Jun; Belloni, Virginia; Scaraffia, Patricia Y.

    2016-01-01

    To better understand the mechanisms responsible for the success of female mosquitoes in their disposal of excess nitrogen, we investigated the role of alanine aminotransferase (ALAT) in blood-fed Aedes aegypti. Transcript and protein levels from the 2 ALAT genes were analyzed in sucrose- and blood-fed A. aegypti tissues. ALAT1 and ALAT2 exhibit distinct expression patterns in tissues during the first gonotrophic cycle. Injection of female mosquitoes with either double-stranded RNA (dsRNA)-ALAT1 or dsRNA ALAT2 significantly decreased mRNA and protein levels of ALAT1 or ALAT2 in fat body, thorax, and Malpighian tubules compared with dsRNA firefly luciferase-injected control mosquitoes. The silencing of either A. aegypti ALAT1 or ALAT2 caused unexpected phenotypes such as a delay in blood digestion, a massive accumulation of uric acid in the midgut posterior region, and a significant decrease of nitrogen waste excretion during the first 48 h after blood feeding. Concurrently, the expression of genes encoding xanthine dehydrogenase and ammonia transporter (Rhesus 50 glycoprotein) were significantly increased in tissues of both ALAT1- and ALAT2-deficient females. Moreover, perturbation of ALAT1 and ALAT2 in the female mosquitoes delayed oviposition and reduced egg production. These novel findings underscore the efficient mechanisms that blood-fed mosquitoes use to avoid ammonia toxicity and free radical damage.—Mazzalupo, S., Isoe, J., Belloni, V., Scaraffia, P. Y. Effective disposal of nitrogen waste in blood-fed Aedes aegypti mosquitoes requires alanine aminotransferase. PMID:26310269

  2. Abdominal obesity validates the association between elevated alanine aminotransferase and newly diagnosed diabetes mellitus.

    PubMed

    Yueh, Chen-Yu; Yang, Yao-Hsu; Sung, Yi-Ting; Lee, Li-Wen

    2014-01-01

    To examine how elevated alanine aminotransferase (ALT) could be associated with newly diagnosed diabetes mellitus. We conducted a cross-sectional analysis on a mass health examination. The odds ratios (ORs) for diabetes mellitus and newly diagnosed diabetes mellitus were compared between people with and without abdominal obesity, together with and without elevated ALT levels. 5499 people were included in this study. Two hundred fifty two (4.6%) fulfilled the diagnosis of diabetes mellitus with 178 (3.2%) undiagnosed before. Metabolic syndrome was vigorously associated with diabetes mellitus and newly diagnosed diabetes mellitus (12.4% vs. 1.4% and 9.0% vs. 0.9%), but elevated ALT alone was not. However, coexisting with obesity, elevated ALTs were robustly associated with diabetes mellitus and newly diagnosed diabetes mellitus. For the incidence of newly diagnosed diabetes mellitus, in comparison to non-obese people with normal ALT (1.7%, OR = 1), obese people especially with elevated ALT levels had significantly higher ORs (obese with ALT ≤ 40 U/L: 4.7%, OR 1.73, 95% CI 1.08-2.77, P 0.023; ALT 41-80 U/L: 6.8%, OR 2.06, 95% CI 1.20-3.55, P 0.009; ALT 81-120 U/L: 8.8%, OR 3.07, 95% CI 1.38-6.84, P 0.006; ALT > 120 U/L: 18.2%, OR 7.44, 95% CI 3.04-18.18, P < 0.001). Abdominal obesity validates the association between elevated alanine aminotransferase and diabetes mellitus and newly diagnosed diabetes mellitus. People with abdominal obesity, especially with coexisting elevated ALT levels should be screened for undiagnosed diabetes mellitus.

  3. Irritable Bowel Syndrome May Be Associated with Elevated Alanine Aminotransferase and Metabolic Syndrome

    PubMed Central

    Lee, Seung-Hwa; Kim, Kwang-Min; Joo, Nam-Seok

    2016-01-01

    Purpose Recent studies have revealed close relationships between hepatic injury, metabolic pathways, and gut microbiota. The microorganisms in the intestine also cause irritable bowel syndrome (IBS). The aim of this study was to examine whether IBS was associated with elevated hepatic enzyme [alanine aminotransferase (ALT) and aspartate aminotransferase (AST)], gamma-glutamyl transferase (γ-GT) levels, and metabolic syndrome (MS). Materials and Methods This was a retrospective, cross-sectional, case-control study. The case and control groups comprised subjects who visited our health promotion center for general check-ups from June 2010 to December 2010. Of the 1127 initially screened subjects, 83 had IBS according to the Rome III criteria. The control group consisted of 260 age- and sex-matched subjects without IBS who visited our health promotion center during the same period. Results Compared to control subjects, patients with IBS showed significantly higher values of anthropometric parameters (body mass index, waist circumference), liver enzymes, γ-GT, and lipid levels. The prevalences of elevated ALT (16.9% vs. 7.7%; p=0.015) and γ-GT (24.1% vs. 11.5%; p=0.037) levels were significantly higher in patients with IBS than in control subjects. A statistically significant difference was observed in the prevalence of MS between controls and IBS patients (12.7% vs. 32.5%; p<0.001). The relationships between elevated ALT levels, MS, and IBS remained statistically significant after controlling for potential confounding factors. Conclusion On the basis of our study results, IBS may be an important condition in certain patients with elevated ALT levels and MS. PMID:26632395

  4. Structure of GroEL in Complex with an Early Folding Intermediate of Alanine Glyoxylate Aminotransferase*

    PubMed Central

    Albert, Armando; Yunta, Cristina; Arranz, Rocío; Peña, Álvaro; Salido, Eduardo; Valpuesta, José María; Martín-Benito, Jaime

    2010-01-01

    Primary hyperoxaluria type 1 is a rare autosomal recessive disease caused by mutations in the alanine glyoxylate aminotransferase gene (AGXT). We have previously shown that P11L and I340M polymorphisms together with I244T mutation (AGXT-LTM) represent a conformational disease that could be amenable to pharmacological intervention. Thus, the study of the folding mechanism of AGXT is crucial to understand the molecular basis of the disease. Here, we provide biochemical and structural data showing that AGXT-LTM is able to form non-native folding intermediates. The three-dimensional structure of a complex between the bacterial chaperonin GroEL and a folding intermediate of AGXT-LTM mutant has been solved by cryoelectron microscopy. The electron density map shows the protein substrate in a non-native extended conformation that crosses the GroEL central cavity. Addition of ATP to the complex induces conformational changes on the chaperonin and the internalization of the protein substrate into the folding cavity. The structure provides a three-dimensional picture of an in vivo early ATP-dependent step of the folding reaction cycle of the chaperonin and supports a GroEL functional model in which the chaperonin promotes folding of the AGXT-LTM mutant protein through forced unfolding mechanism. PMID:20056599

  5. A Micro-Platinum Wire Biosensor for Fast and Selective Detection of Alanine Aminotransferase

    PubMed Central

    Thuy, Tran Nguyen Thanh; Tseng, Tina T.-C.

    2016-01-01

    In this study, a miniaturized biosensor based on permselective polymer layers (overoxidized polypyrrole (Ppy) and Nafion®) modified and enzyme (glutamate oxidase (GlutOx)) immobilized micro-platinum wire electrode for the detection of alanine aminotransferase (ALT) was fabricated. The proposed ALT biosensor was measured electrochemically by constant potential amperometry at +0.7 V vs. Ag/AgCl. The ALT biosensor provides fast response time (~5 s) and superior selectivity towards ALT against both negatively and positively charged species (e.g., ascorbic acid (AA) and dopamine (DA), respectively). The detection range of the ALT biosensor is found to be 10–900 U/L which covers the range of normal ALT levels presented in the serum and the detection limit and sensitivity are found to be 8.48 U/L and 0.059 nA/(U/L·mm2) (N = 10), respectively. We also found that one-day storage of the ALT biosensor at −20 °C right after the sensor being fabricated can enhance the sensor sensitivity (1.74 times higher than that of the sensor stored at 4 °C). The ALT biosensor is stable after eight weeks of storage at −20 °C. The sensor was tested in spiked ALT samples (ALT activities: 20, 200, 400, and 900 U/L) and reasonable recoveries (70%~107%) were obtained. PMID:27240366

  6. Histological and Clinical Characteristics of Patients with Chronic Hepatitis C and Persistently Normal Alanine Aminotransferase Levels

    PubMed Central

    Guzman, Grace

    2014-01-01

    Patients with chronic hepatitis C virus (HCV) infection and persistently normal alanine aminotransferase (PNALT) are generally described to have mild liver disease. The aim of this study was to compare clinical and histological features in HCV-infected patients with PNALT and elevated ALT. Patients presenting to the University of Illinois Medical Center, Chicago, who had biopsy proven HCV, an ALT measurement at the time of liver biopsy, at least one additional ALT measurement over the next 12 months, and liver biopsy slides available for review were identified. PNALT was defined as ALT ≤ 30 on at least 2 different occasions over 12 months. Of 1200 patients with HCV, 243 met the study criteria. 13% (32/243) of patients had PNALT while 87% (211/243) had elevated ALT. Significantly more patients with PNALT had advanced fibrosis (F3 and F4) compared to those with elevated ALT (P = 0.007). There was no significant difference in the histology activity index score as well as mean inflammatory score between the two groups. In conclusion, in a well-characterized cohort of patients at a tertiary medical center, PNALT did not distinguish patients with mild liver disease. PMID:24891947

  7. Elevated Aspartate and Alanine Aminotransferase Levels and Natural Death among Patients with Methamphetamine Dependence

    PubMed Central

    Kuo, Chian-Jue; Tsai, Shang-Ying; Liao, Ya-Tang; Conwell, Yeates; Lee, Wen-Chung; Huang, Ming-Chyi; Lin, Shih-Ku; Chen, Chiao-Chicy; Chen, Wei J.

    2012-01-01

    Background Methamphetamine is one of the fastest growing illicit drugs worldwide, causing multiple organ damage and excessive natural deaths. The authors aimed to identify potential laboratory indices and clinical characteristics associated with natural death through a two-phase study. Methods Methamphetamine-dependent patients (n = 1,254) admitted to a psychiatric center in Taiwan between 1990 and 2007 were linked with a national mortality database for causes of death. Forty-eight subjects died of natural causes, and were defined as the case subjects. A time-efficient sex- and age-matched nested case-control study derived from the cohort was conducted first to explore the potential factors associated with natural death through a time-consuming standardized review of medical records. Then the identified potential factors were evaluated in the whole cohort to validate the findings. Results In phase I, several potential factors associated with natural death were identified, including aspartate aminotransferase (AST), alanine aminotransferase (ALT), comorbid alcohol use disorder, and the prescription of antipsychotic drugs. In phase II, these factors were confirmed in the whole cohort using survival analysis. For the characteristics at the latest hospital admission, Cox proportional hazards models showed that the adjusted hazard ratios for natural death were 6.75 (p<0.001) in the group with markedly elevated AST (>80 U/L) and 2.66 (p<0.05) in the group with mildly elevated AST (40–80 U/L), with reference to the control group (<40 U/L). As for ALT, the adjusted hazard ratios were 5.41 (p<0.001), and 1.44 (p>0.05). Comorbid alcohol use disorder was associated with an increased risk of natural death, whereas administration of antipsychotic drugs was not associated with lowered risk. Conclusions This study highlights the necessity of intensive follow-up for those with elevated AST and ALT levels and comorbid alcohol use disorder for preventing excessive natural

  8. Association between Serum Uric Acid and Elevated Alanine Aminotransferase in the General Population

    PubMed Central

    Chen, Shuang; Guo, Xiaofan; Yu, Shasha; Sun, Guozhe; Yang, Hongmei; Li, Zhao; Sun, Yingxian

    2016-01-01

    Background: Both the serum uric acid (SUA) level and elevated alanine aminotransferase (ALT) are related to metabolic syndrome. However, the association between SUA and elevated ALT has not been elucidated in the general population. The objective of this study was to investigate the association between SUA and elevated ALT in the general population of China; Methods: A total of 11,572 adults (≥35 years of age) participated in this survey. Elevated ALT was defined as >40 U/L. SUA ≥ 7.0 mg/dL in males or ≥6.0 mg/dL in females was defined as hyperuricemia. SUA within the reference range was divided into quartiles, and its associations with elevated ALT were evaluated by logistic regressions; Results: A total of 7.4% participants had elevated ALT. The prevalence of hyperuricemia was 14.9% in males and 7.3% in females. There was a significantly positive dose-response association between SUA levels and the prevalence of elevated ALT. After adjusting for potential confounders, a positive relationship for elevated ALT was observed in subjects with hyperuricemia (odds ratio [OR]: 2.032, 95% confidence interval [CI]: 1.443–2.861 for men; OR: 2.045, 95% CI: 1.221–3.425 for women, both p < 0.05). Within the reference range, the association between SUA and elevated ALT persisted in the fourth quartile (OR: 1.467, 95% CI: 1.063–2.025 for men; OR: 1.721, 95% CI: 1.146–2.585 for women, both p < 0.05); Conclusions: Our results indicated that an increased SUA level, even within the reference range, was independently associated with elevated ALT in Chinese adults. PMID:27563918

  9. Association between Elevated Alanine Aminotransferase and Urosepsis in Children with Acute Pyelonephritis

    PubMed Central

    Kim, Dongwan; Lee, Sung Hyun; Ryoo, Eell; Cho, Hye Kyung; Kim, Yun Mi

    2016-01-01

    Purpose The aim of this study is to investigate the association between elevated alanine aminotransferase (ALT) and urosepsis in children with acute pyelonephritis (APN). Methods We retrospectively identified all children who were managed in our hospital with APN during a decade period. In our study a diagnosis of APN was defined as having a positive urine culture and a positive (99m)Tc-dimercaptosuccinic acid scintigraphy. We compared those with elevated ALT and those with normal ALT according to the following variables: age, gender, duration of fever prior to admission, presence of hypotension, C-reactive protein (CRP), creatinine, presence of anemia, white blood cells count, platelet count, blood culture result, and grades of vesicoureteral reflux. In addition, the correlation between elevated ALT and positive blood culture was analyzed in detail. Results A total of 996 children were diagnosed with APN, of which 883 were included in the study. ALT was elevated in 81 children (9.2%). In the analysis of demographic characteristics, the number of children with elevated ALT was higher in children between 0 to 3 months, boys, and in those with positive blood culture (p=0.002, 0.036, and 0.010, respectively). In multivariate analysis of variables associated with positive blood culture, age younger than 3 months, elevated ALT, elevated CRP, and elevated creatinine showed statistical significance (p=0.004, 0.030, 0.043, and 0.044, respectively). Conclusion Our study demonstrates the association between elevated ALT and increased prevalence of urosepsis in addition to elevated CRP, elevated creatinine, and age younger than 3 months in children with APN. PMID:27066449

  10. Reconfiguration of N Metabolism upon Hypoxia Stress and Recovery: Roles of Alanine Aminotransferase (AlaAT) and Glutamate Dehydrogenase (GDH)

    PubMed Central

    Diab, Houssein; Limami, Anis M.

    2016-01-01

    In the context of climatic change, more heavy precipitation and more frequent flooding and waterlogging events threaten the productivity of arable farmland. Furthermore, crops were not selected to cope with flooding- and waterlogging-induced oxygen limitation. In general, low oxygen stress, unlike other abiotic stresses (e.g., cold, high temperature, drought and saline stress), received little interest from the scientific community and less financial support from stakeholders. Accordingly, breeding programs should be developed and agronomical practices should be adapted in order to save plants’ growth and yield—even under conditions of low oxygen availability (e.g., submergence and waterlogging). The prerequisite to the success of such breeding programs and changes in agronomical practices is a good knowledge of how plants adapt to low oxygen stress at the cellular and the whole plant level. In the present paper, we summarized the recent knowledge on metabolic adjustment in general under low oxygen stress and highlighted thereafter the major changes pertaining to the reconfiguration of amino acids syntheses. We propose a model showing (i) how pyruvate derived from active glycolysis upon hypoxia is competitively used by the alanine aminotransferase/glutamate synthase cycle, leading to alanine accumulation and NAD+ regeneration. Carbon is then saved in a nitrogen store instead of being lost through ethanol fermentative pathway. (ii) During the post-hypoxia recovery period, the alanine aminotransferase/glutamate dehydrogenase cycle mobilizes this carbon from alanine store. Pyruvate produced by the reverse reaction of alanine aminotransferase is funneled to the TCA cycle, while deaminating glutamate dehydrogenase regenerates, reducing equivalent (NADH) and 2-oxoglutarate to maintain the cycle function. PMID:27258319

  11. Creatine

    MedlinePlus

    ... loss. Inherited nerve damage (hereditary motor and sensory neuropathy). Early research in people with inherited nerve damage ... concern that combining creatine with caffeine and the herb ephedra (also called Ma Huang) might increase the ...

  12. The aspartate aminotransferase-to-alanine aminotransferase ratio predicts all-cause and cardiovascular mortality in patients with type 2 diabetes

    PubMed Central

    Zoppini, Giacomo; Cacciatori, Vittorio; Negri, Carlo; Stoico, Vincenzo; Lippi, Giuseppe; Targher, Giovanni; Bonora, Enzo

    2016-01-01

    Abstract An increased aspartate aminotransferase-to-alanine aminotransferase ratio (AAR) has been widely used as a marker of advanced hepatic fibrosis. Increased AAR was also shown to be significantly associated with the risk of developing cardiovascular (CV) disease. The aim of this study was to assess the relationship between the AAR and mortality risk in a well-characterized cohort of patients with type 2 diabetes. A cohort of 2529 type 2 diabetic outpatients was followed-up for 6 years to collect cause-specific mortality. Cox regression analyses were modeled to estimate the independent association between AAR and the risk of all-cause and CV mortality. Over the 6-year follow-up period, 12.1% of patients died, 47.5% of whom from CV causes. An increased AAR, but not its individual components, was significantly associated with an increased risk of all-cause (adjusted-hazard risk 1.83, confidence interval [CI] 95% 1.14–2.93, P = 0.012) and CV (adjusted-hazard risk 2.60, CI 95% 1.38–4.90, P < 0.003) mortality after adjustment for multiple clinical risk factors and potential confounding variables. The AAR was independently associated with an increased risk of both all-cause and CV mortality in patients with type 2 diabetes. These findings suggest that an increased AAR may reflect more systemic derangements that are not simply limited to liver damage. Further studies are needed to elucidate the pathophysiological implications of an increased AAR. PMID:27787357

  13. Liver peroxisomal alanine:glyoxylate aminotransferase and the effects of mutations associated with Primary Hyperoxaluria Type I: An overview.

    PubMed

    Oppici, Elisa; Montioli, Riccardo; Cellini, Barbara

    2015-09-01

    Liver peroxisomal alanine:glyoxylate aminotransferase (AGT) (EC 2.6.1.44) catalyses the conversion of l-alanine and glyoxylate to pyruvate and glycine, a reaction that allows glyoxylate detoxification. Inherited mutations on the AGXT gene encoding AGT lead to Primary Hyperoxaluria Type I (PH1), a rare disorder characterized by the deposition of calcium oxalate crystals primarily in the urinary tract. Here we describe the results obtained on the biochemical features of AGT as well as on the molecular and cellular effects of polymorphic and pathogenic mutations. A complex scenario on the molecular pathogenesis of PH1 emerges in which the co-inheritance of polymorphic changes and the condition of homozygosis or compound heterozygosis are two important factors that determine the enzymatic phenotype of PH1 patients. All the reported data represent relevant steps toward the understanding of genotype/phenotype correlations, the prediction of the response of the patients to the available therapies, and the development of new therapeutic approaches. This article is part of a Special Issue entitled: Cofactor-dependent proteins: evolution, chemical diversity and bio-applications.

  14. BarR, an Lrp-type transcription factor in Sulfolobus acidocaldarius, regulates an aminotransferase gene in a β-alanine responsive manner.

    PubMed

    Liu, Han; Orell, Alvaro; Maes, Dominique; van Wolferen, Marleen; Lindås, Ann-Christin; Bernander, Rolf; Albers, Sonja-Verena; Charlier, Daniel; Peeters, Eveline

    2014-05-01

    In archaea, nothing is known about the β-alanine degradation pathway or its regulation. In this work, we identify and characterize BarR, a novel Lrp-like transcription factor and the first one that has a non-proteinogenic amino acid ligand. BarR is conserved in Sulfolobus acidocaldarius and Sulfolobus tokodaii and is located in a divergent operon with a gene predicted to encode β-alanine aminotransferase. Deletion of barR resulted in a reduced exponential growth rate in the presence of β-alanine. Furthermore, qRT-PCR and promoter activity assays demonstrated that BarR activates the expression of the adjacent aminotransferase gene, but only upon β-alanine supplementation. In contrast, auto-activation proved to be β-alanine independent. Heterologously produced BarR is an octamer in solution and forms a single complex by interacting with multiple sites in the 170 bp long intergenic region separating the divergently transcribed genes. In vitro, DNA binding is specifically responsive to β-alanine and site-mutant analyses indicated that β-alanine directly interacts with the ligand-binding pocket. Altogether, this work contributes to the growing body of evidence that in archaea, Lrp-like transcription factors have physiological roles that go beyond the regulation of α-amino acid metabolism.

  15. Ceruloplasmin, a reliable marker of fibrosis in chronic hepatitis B virus patients with normal or minimally raised alanine aminotransferase

    PubMed Central

    Zeng, Da-Wu; Dong, Jing; Jiang, Jia-Ji; Zhu, Yue-Yong; Liu, Yu-Rui

    2016-01-01

    AIM To develop a non-invasive model to evaluate significant fibrosis and cirrhosis by investigating the association between serum ceruloplasmin (CP) levels and liver fibrosis in chronic hepatitis B (CHB) patients with normal or minimally raised alanine aminotransferase (ALT). METHODS Serum samples and liver biopsy were obtained from 193 CHB patients with minimally raised or normal ALT who were randomly divided into a training group (n = 97) and a validation group (n = 96). Liver histology was evaluated by the METAVIR scoring system. Receiver operator characteristic curves were applied to the diagnostic value of CP for measuring liver fibrosis in CHB patients. Spearman rank correlation analyzed the relationship between CP and liver fibrosis. A non-invasive model was set up through multivariate logistic regression analysis. RESULTS Serum CP levels individualized various fibrosis stages via area under the curve (AUC) values. Multivariate analysis revealed that CP levels were significantly related to liver cirrhosis. Combining CP with serum GGT levels, a CG model was set up to predict significant fibrosis and liver cirrhosis in CHB patients with normal or minimally raised ALT. The AUC, sensitivity, specificity, positive predictive value, and negative predictive value were 0.84, 83.1%, 78.6%, 39.6%, and 96.5% to predict liver cirrhosis, and 0.789, 80.26%, 68.38%, 62.25%, and 84.21% to predict significant fibrosis. This model expressed a higher AUC than FIB-4 (age, ALT, aspartate aminotransferase, platelets) and GP (globulin, platelets) models to predict significant fibrosis (P = 0.019 and 0.022 respectively) and revealed a dramatically greater AUC than FIB-4 (P = 0.033) to predict liver cirrhosis. CONCLUSION The present study showed that CP was independently and negatively associated with liver fibrosis. Furthermore, we developed a novel promising model (CG), based on routine serum markers, for predicting liver fibrosis in CHB patients with normal or minimally raised

  16. Genetic engineering of improved nitrogen use efficiency in rice by the tissue-specific expression of alanine aminotransferase.

    PubMed

    Shrawat, Ashok K; Carroll, Rebecka T; DePauw, Mary; Taylor, Gregory J; Good, Allen G

    2008-09-01

    Summary Nitrogen is quantitatively the most essential nutrient for plants and a major factor limiting crop productivity. One of the critical steps limiting the efficient use of nitrogen is the ability of plants to acquire it from applied fertilizer. Therefore, the development of crop plants that absorb and use nitrogen more efficiently has been a long-term goal of agricultural research. In an attempt to develop nitrogen-efficient plants, rice (Oryza sativa L.) was genetically engineered by introducing a barley AlaAT (alanine aminotransferase) cDNA driven by a rice tissue-specific promoter (OsAnt1). This modification increased the biomass and grain yield significantly in comparison with control plants when plants were well supplied with nitrogen. Compared with controls, transgenic rice plants also demonstrated significant changes in key metabolites and total nitrogen content, indicating increased nitrogen uptake efficiency. The development of crop plants that take up and assimilate nitrogen more efficiently would not only improve the use of nitrogen fertilizers, resulting in lower production costs, but would also have significant environmental benefits. These results are discussed in terms of their relevance to the development of strategies to engineer enhanced nitrogen use efficiency in crop plants.

  17. A Novel Pathway for Metabolism of the Cardiovascular Risk Factor Homoarginine by alanine:glyoxylate aminotransferase 2

    PubMed Central

    Rodionov, Roman N.; Oppici, Elisa; Martens-Lobenhoffer, Jens; Jarzebska, Natalia; Brilloff, Silke; Burdin, Dmitrii; Demyanov, Anton; Kolouschek, Anne; Leiper, James; Maas, Renke; Cellini, Barbara; Weiss, Norbert; Bode-Böger, Stefanie M.

    2016-01-01

    Low plasma concentrations of L-homoarginine are associated with an increased risk of cardiovascular events, while homoarginine supplementation is protective in animal models of metabolic syndrome and stroke. Catabolism of homoarginine is still poorly understood. Based on the recent findings from a Genome Wide Association Study we hypothesized that homoarginine can be metabolized by alanine:glyoxylate aminotransferase 2 (AGXT2). We purified human AGXT2 from tissues of AGXT2 transgenic mice and demonstrated its ability to metabolize homoarginine to 6-guanidino-2-oxocaproic acid (GOCA). After incubation of HepG2 cells overexpressing AGXT2 with isotope-labeled homoarginine-d4 we were able to detect labeled GOCA in the medium. We injected wild type mice with labeled homoarginine and detected labeled GOCA in the plasma. We found that AGXT2 knockout (KO) mice have higher homoarginine and lower GOCA plasma levels as compared to wild type mice, while the reverse was true for AGXT2 transgenic (Tg) mice. In summary, we experimentally proved the presence of a new pathway of homoarginine catabolism – its transamination by AGXT2 with formation of GOCA and demonstrated that endogenous AGXT2 is required for maintenance of homoarginine levels in mice. Our findings may lead to development of novel therapeutic approaches for cardiovascular pathologies associated with homoarginine deficiency. PMID:27752063

  18. Dose-Response Relationship between Alanine Aminotransferase Levels within the Reference Interval and Metabolic Syndrome in Chinese Adults

    PubMed Central

    Wu, Peipei; Chen, Qicai; Chen, Lili; Zhang, Pengpeng; Xiao, Juan; Chen, Xiaoxiao; Liu, Meng

    2017-01-01

    Purpose Elevation in serum alanine aminotransferase (ALT) levels is a biomarker for metabolic syndrome (MS); however, the relationship has not been fully investigated within the reference interval of ALT levels. Our objective was to explore the relationship between serum ALT levels within the reference interval and MS in Chinese adults. Materials and Methods This cross-sectional study included 16028 adults, who attended routine health check-ups at Shengli Oilfield Central Hospital from January 2006 to March 2012. The reference interval of serum ALT level was defined as less than 40 U/L. Logistic regression models and restricted cubic spline were used to evaluate the association of ALT with MS. Results The prevalence of MS in the total population was 13.7% (6.4% for females and 18.4% for males). Multiple logistic regression showed that ALT levels were positively associated with MS after adjustment for potential confounding factors. The odds ratio of MS in the top quartile was 4.830 [95% confidence interval (CI): 2.980–7.829] in females and 3.168 (95% CI: 2.649–3.790) in males, compared with the ALT levels in the bottom quartile. The restricted cubic spline models revealed a positive non-linear dose-response relationship between ALT levels and the risk of MS in women (p for nonlinearity was 0.0327), but a positive linear dose-response relationship in men (p for nonlinearity was 0.0659). Conclusion Serum ALT levels within the reference interval are positively associated with MS in a dose-response manner. Elevated ALT levels, even within the reference interval, may reflect early dysmetabolic changes. PMID:27873509

  19. Prevalence of elevated alanine-aminotransferase (ALT) among US adolescents and associated factors: NHANES 1999-2004

    PubMed Central

    Fraser, Abigail; Longnecker, Matthew P.; Lawlor, Debbie A

    2007-01-01

    Background & aims Non-alcoholic fatty liver disease (NAFLD) is a common cause of liver disease in children and adolescents. The majority of studies of NAFLD in children have been in select populations of the clinically obese. Study aims were to estimate the prevalence of elevated alanine-aminotransferase (ALT, as a marker of NAFLD) in a general contemporary adolescent population and to identify leading risk factors for ALT elevation (> 30 U/L). Methods We analysed data of adolescent participants (age 12-19, N=5586) in NHANES 1999-2004, a representative sample of the civilian non-institutionalized U.S population. Results The prevalence of elevated ALT (>30 U/L) was 7∙4% among white adolescents, 11∙5%, among Mexican Americans, and 6∙0%, among black adolescents. It was prevalent in 12∙4% of males compared to 3∙5% of females. Multivariable associations with elevated ALT were found for sex (OR male versus female = 7∙7, 95%CI: 3∙9, 15∙1), ethnicity (OR black versus white=0∙6, 95%CI: 0∙3, 1∙3; OR Mexican American versus white=1∙6, 95%CI: 1∙0, 2∙6), waist circumference (OR per 1 SD=1∙4, 95%CI: 1∙0, 2∙0), and fasting insulin (OR per 1 SD=1∙ 6, 95%CI: 1∙ 2, 2∙ 1). Age, C-reactive protein and triglycerides were also positively, and socio-economic position inversely associated with elevated ALT. The magnitude of associations with ALT was similar across ethnic groups. Conclusions ALT is associated with waist circumference and insulin resistance even in a young population. These characteristics could be utilized to identify adolescents who may benefit from screening for NAFLD, offering an opportunity to prevent disease progression at an early age. PMID:18054554

  20. Diet and the frequency of the alanine:glyoxylate aminotransferase Pro11Leu polymorphism in different human populations.

    PubMed

    Caldwell, Elizabeth F; Mayor, Lianne R; Thomas, Mark G; Danpure, Christopher J

    2004-11-01

    The intermediary metabolic enzyme alanine:glyoxylate aminotransferase (AGT) contains a Pro11Leu polymorphism that decreases its catalytic activity by a factor of three and causes a small proportion to be mistargeted from its normal intracellular location in the peroxisomes to the mitochondria. These changes are predicted to have significant effects on the synthesis and excretion of the metabolic end-product oxalate and the deposition of insoluble calcium oxalate in the kidney and urinary tract. Based on the evolution of AGT targeting in mammals, we have previously hypothesised that this polymorphism would be advantageous for individuals who have a meat-rich diet, but disadvantageous for those who do not. If true, the frequency distribution of Pro11Leu in different extant human populations should have been shaped by their dietary history so that it should be more common in populations with predominantly meat-eating ancestral diets than it is in populations in which the ancestral diets were predominantly vegetarian. In the present study, we have determined frequency of Pro11Leu in 11 different human populations with divergent ancestral dietary lifestyles. We show that the Pro11Leu allelic frequency varies widely from 27.9% in the Saami, a population with a very meat-rich ancestral diet, to 2.3% in Chinese, who are likely to have had a more mixed ancestral diet. FST analysis shows that the differences in Pro11Leu frequency between some populations (particularly Saami vs Chinese) was very high when compared with neutral loci, suggesting that its frequency might have been shaped by dietary selection pressure.

  1. Genetic variations in the alanine-glyoxylate aminotransferase 2 (AGXT2) gene and dimethylarginines levels in rheumatoid arthritis.

    PubMed

    Dimitroulas, Theodoros; Hodson, James; Panoulas, Vasileios F; Sandoo, Aamer; Smith, Jacqueline; Kitas, George

    2017-03-29

    Rheumatoid arthritis (RA) is associated with high rates of cardiovascular events mainly due to coronary and cerebrovascular atherosclerotic disease. Asymmetric (ADMA) and symmetric (SDMA) dimethylarginines are endogenous inhibitors of nitric oxide synthase and have been repeatedly linked with adverse cardiovascular outcomes in the general population and various disease settings. Alanine-glyoxylate aminotransferase 2 (AGTX2) is considered an alternative metabolic pathway contributing to the clearance of dimethylarginines in humans. The aim of the current study was to investigate the effect of specific AGXT-2 gene polymorphisms on circulating levels of ADMA or SDMA in patients with RA. Serum ADMA and SDMA levels were measured in 201 individuals with RA [median age: 67 years (IQR: 59-73), 155 females]. Two single nucleotide polymorphisms (SNPs) in the AGXT-2 gene-rs37369 and rs28305-were genotyped. Distributions of SDMA and ADMA were skewed, hence comparisons across the gene polymorphisms were performed using Kruskal-Wallis tests, and summarized using medians and interquartile ranges. Univariable analysis did not demonstrate a significant difference in the levels of SDMA or ADMA amongst the different genotypic groups of either rs37369AGXT2 (p = 0.800, 0.977) or rs28305AGXT2 (p = 0.463, 0.634). In multivariable analyses, ADMA levels were found to be significantly associated with erythrocyte sedimentation rate and estimated glomerular filtration rate, whilst SDMA levels were significantly associated with estimated glomerular filtration rate and quantitative insulin sensitivity check index. After adjustments for these factors, the relationship between the AGXT2 gene variants and both ADMA and SDMA remained non-significant. Our study in a well-characterized RA population did not show an association between serum concentrations of dimethylarginines and genetic variants of the AGXT2 gene.

  2. Identification of mutations associated with peroxisome-to-mitochondrion mistargeting of alanine/glyoxylate aminotransferase in primary hyperoxaluria type 1

    PubMed Central

    1990-01-01

    We have previously shown that in some patients with primary hyperoxaluria type 1 (PH1), disease is associated with mistargeting of the normally peroxisomal enzyme alanine/glyoxylate aminotransferase (AGT) to mitochondria (Danpure, C.J., P.J. Cooper, P.J. Wise, and P.R. Jennings. J. Cell Biol. 108:1345-1352). We have synthesized, amplified, cloned, and sequenced AGT cDNA from a PH1 patient with mitochondrial AGT (mAGT). This identified three point mutations that cause amino acid substitutions in the predicted AGT protein sequence. Using PCR and allele-specific oligonucleotide hybridization, a range of PH1 patients and controls were screened for these mutations. This revealed that all eight PH1 patients with mAGT carried at least one allele with the same three mutations. Two were homozygous for this allele and six were heterozygous. In at least three of the heterozygotes, it appeared that only the mutant allele was expressed. All three mutations were absent from PH1 patients lacking mAGT. One mutation encoding a Gly----Arg substitution at residue 170 was not found in any of the control individuals. However, the other two mutations, encoding Pro----Leu and Ile----Met substitutions at residues 11 and 340, respectively, cosegregated in the normal population at an allelic frequency of 5-10%. In an individual homozygous for this allele (substitutions at residues 11 and 340) only a small proportion of AGT appeared to be rerouted to mitochondria. It is suggested that the substitution at residue 11 generates an amphiphilic alpha-helix with characteristics similar to recognized mitochondrial targeting sequences, the full functional expression of which is dependent upon coexpression of the substitution at residue 170, which may induce defective peroxisomal import. PMID:1703535

  3. ALT (Alanine Aminotransferase) Test

    MedlinePlus

    ... to help recognize heart or muscle injury. ALT values are often compared to the results of other tests such as alkaline phosphatase (ALP) , total protein , and bilirubin to help determine which form of liver disease is present. ALT is often used to monitor the treatment ...

  4. Trihalomethane exposure and biomonitoring for the liver injury indicator, alanine aminotransferase, in the United States population (NHANES 1999–2006)

    PubMed Central

    Burch, James B.; Everson, Todd M.; Seth, Ratanesh K.; Wirth, Michael D.; Chatterjee, Saurabh

    2015-01-01

    Exposure to trihalomethanes (or THMs: chloroform, bromoform, bromodichloromethane, and dibromochloromethane [DBCM]) formed via drinking water disinfection has been associated with adverse reproductive outcomes and cancers of the digestive or genitourinary organs. However, few studies have examined potential associations between THMs and liver injury in humans, even though experimental studies suggest that these agents exert hepatotoxic effects, particularly among obese individuals. This study examined participants in the National Health and Nutrition Examination Survey (1999–2006, N = 2781) to test the hypothesis that THMs are associated with liver injury as assessed by alanine aminotransferase (ALT) activity in circulation. Effect modification by body mass index (BMI) or alcohol consumption also was examined. Associations between blood THM concentrations and ALT activity were assessed using unconditional multiple logistic regression to calculate prevalence odds ratios (ORs) with 95% confidence intervals (CIs) for exposure among cases with elevated ALT activity (men: >40 IU/L, women: >30 IU/L) relative to those with normal ALT, after adjustment for variables that may confound the relationship between ALT and THMs. Compared to controls, cases were 1.35 times more likely (95% CI: 1.02, 1.79) to have circulating DBCM concentrations exceeding median values in the population. There was little evidence for effect modification by BMI, although the association varied by alcohol consumption. Among non-drinkers, cases were more likely than controls to be exposed to DBCM (OR: 3.30, 95% CI: 1.37–7.90), bromoform (OR: 2.88, 95% CI: 1.21–6.81), or brominated THMs (OR: 4.00, 95% CI: 1.31–12.1), but no association was observed among participants with low, or moderate to heavy alcohol consumption. Total THM levels exceeding benchmark exposure limits continue to be reported both in the United States and globally. Results from this study suggest a need for further

  5. SAFETY study: Alanine aminotransferase cutoff values are set too high for reliable detection of pediatric chronic liver disease

    PubMed Central

    Schwimmer, Jeffrey B.; Dunn, Winston; Norman, Gregory J.; Pardee, Perrie E.; Middleton, Michael S.; Kerkar, Nanda; Sirlin, Claude B.

    2010-01-01

    Background & Aims The appropriate alanine aminotransferase (ALT) threshold value to use for diagnosis of chronic liver disease in children is unknown. We sought to develop sex-specific, biology-based, pediatric ALT thresholds. Methods The screening ALT for elevation in today’s youth (SAFETY) study collected observational data from acute care children’s hospitals, the national health and nutrition examination survey (NHANES, 1999–2006), overweight children with and without non-alcoholic fatty liver disease (NAFLD), and children with chronic Hepatitis B virus (HBV) or Hepatitis C virus (HCV) infections. The study compared the sensitivity and specificity of ALT thresholds currently used by children’s hospitals versus study-derived, sex-specific, biology-based, ALT thresholds for detecting children with NAFLD, HCV, or HBV. Results The median upper limit of ALT at children’s hospitals was 53 U/L (range, 30–90). The 95th percentile levels for ALT in healthy weight, metabolically normal, liver disease-free, NHANES pediatric participants were 25.8 U/L (boys) and 22.1 U/L (girls). The concordance statistics of these NHANES-derived thresholds for liver disease detection were 0.85 (95% confidence interval [CI] 0.74–0.96) in boys and 0.91 (95% CI 0.83–0.99) in girls for NAFLD, 0.80 (95% CI 0.70–0.91) in boys and 0.79 (95% CI 0.69–0.89) in girls for HBV, and 0.86 (95% CI 0.77–0.95) in boys and 0.84 (95% CI 0.75–0.93) in girls for HCV. Using current children’s hospitals ALT thresholds, the median sensitivity for detection of NAFLD, HBV, and HCV ranged from 32% to 48%; median specificity was 92% (boys) and 96% (girls). Using NHANES-derived thresholds, the sensitivities were 72% (boys) and 82% (girls); specificities were 79% (boys) and 85% (girls). Conclusions The upper limit of ALT used in children’s hospitals varies widely and is set too high to reliably detect chronic liver disease. Biology-based thresholds provide higher sensitivity and only

  6. Effects of 28 days of beta-alanine and creatine monohydrate supplementation on aerobic power, ventilatory and lactate thresholds, and time to exhaustion.

    PubMed

    Zoeller, R F; Stout, J R; O'kroy, J A; Torok, D J; Mielke, M

    2007-09-01

    The effect of beta-alanine (beta-Ala) alone or in combination with creatine monohydrate (Cr) on aerobic exercise performance is unknown. The purpose of this study was to examine the effects of 4 weeks of beta-Ala and Cr supplementation on indices of endurance performance. Fifty-five men (24.5 +/- 5.3 yrs) participated in a double-blind, placebo-controlled study and randomly assigned to one of 4 groups; placebo (PL, n = 13), creatine (Cr, n = 12), beta-alanine (beta-Ala, n = 14), or beta-alanine plus creatine (CrBA, n = 16). Prior to and following supplementation, participants performed a graded exercise test on a cycle ergometer to determine VO(2peak), time to exhaustion (TTE), and power output, VO(2), and percent VO(2peak) associated with VT and LT. No significant group effects were found. However, within groups, a significant time effect was observed for CrBa on 5 of the 8 parameters measured. These data suggest that CrBA may potentially enhance endurance performance.

  7. Analysis of alanine aminotransferase in various organs of soybean (Glycine max) and in dependence of different nitrogen fertilisers during hypoxic stress.

    PubMed

    Rocha, Marcio; Sodek, Ladaslav; Licausi, Francesco; Hameed, Muhammad Waqar; Dornelas, Marcelo Carnier; van Dongen, Joost T

    2010-10-01

    Alanine aminotransferase (AlaAT) catalyses the reversible conversion of pyruvate and glutamate into alanine and oxoglutarate. In soybean, two subclasses were identified, each represented by two highly similar members. To investigate the role of AlaAT during hypoxic stress in soybean, changes in transcript level of both subclasses were analysed together with the enzyme activity and alanine content of the tissue. Moreover, the dependency of AlaAT activity and gene expression was investigated in relation to the source of nitrogen supplied to the plants. Using semi-quantitative PCR, GmAlaAT genes were determined to be highest expressed in roots and nodules. Under normal growth conditions, enzyme activity of AlaAT was detected in all organs tested, with lowest activity in the roots. Upon waterlogging-induced hypoxia, AlaAT activity increased strongly. Concomitantly, alanine accumulated. During re-oxygenation, AlaAT activity remained high, but the transcript level and the alanine content decreased. Our results show a role for AlaAT in the catabolism of alanine during the initial period of re-oxygenation following hypoxia. GmAlaAT also responded to nitrogen availability in the solution during waterlogging. Ammonium as nitrogen source induced both gene expression and enzyme activity of AlaAT more than when nitrate was supplied in the nutrient solution. The work presented here indicates that AlaAT might not only be important during hypoxia, but also during the recovery phase after waterlogging, when oxygen is available to the tissue again.

  8. Relationship of creatine kinase, aspartate aminotransferase, lactate dehydrogenase, and proteinuria to cardiomyopathy in the owl monkey (Aotus vociferans)

    SciTech Connect

    Gozalo, Alfonso S.; Chavera, Alfonso; Montoya, Enrique J.; Takano, Juan; Weller, Richard E.

    2008-02-01

    The purpose of this study was to determine serum reference values for crea- tine kinase (CK), aspartate aminotransferase (AST), and lactate dehydroge- nase (LDH) in captive-born and wild-caught owl monkeys to assess their usefulness for diagnosing myocardial disease. Urine samples were also collected and semi-quantitative tests performed. There was no statistically significant difference between CK, AST, and LDH when comparing both groups. However, when comparing monkeys with proteinuria to those without proteinuria, a statistically significant difference in CK value was observed (P = 0.021). In addition, the CK/AST ratio revealed that 29% of the animals included in this study had values suggesting cardiac infarction. Grossly, cardiac concentric hypertrophy of the left ventricle and small, pitted kidneys were the most common findings. Microscopically, myocardial fibrosis, contraction band necrosis, hypertrophy and hyperplasia of coronary arteries, medium-sized renal arteries, and afferent glomerular arteriolae were the most significant lesions, along with increased mesangial matrix and hypercellularity of glomeruli, Bowman’s capsule, and peritubular space fibroplasia. These findings suggest that CK, AST, and LDH along with urinalysis provide a reliable method for diagnosing cardiomyopathies in the owl monkey. In addition, CK/AST ratio, proteinuria, and the observed histological and ultrastructural changes suggest that Aotus vociferans suffer from arterial hypertension and chronic myocardial infarction.

  9. A novel C-S lyase from the latex-producing plant Taraxacum brevicorniculatum displays alanine aminotransferase and l-cystine lyase activity.

    PubMed

    Munt, Oliver; Prüfer, Dirk; Schulze Gronover, Christian

    2013-01-01

    We isolated a novel pyridoxal-5-phosphate-dependent l-cystine lyase from the dandelion Taraxacum brevicorniculatum. Real time qPCR analysis showed that C-S lyase from Taraxacum brevicorniculatum (TbCSL) mRNA is expressed in all plant tissues, although at relatively low levels in the latex and pedicel. The 1251 bp TbCSL cDNA encodes a protein with a calculated molecular mass of 46,127 kDa. It is homologous to tyrosine and alanine aminotransferases (AlaATs) as well as to an Arabidopsis thaliana carbon-sulfur lyase (C-S lyase) (SUR1), which has a role in glucosinolate metabolism. TbCSL displayed in vitrol-cystine lyase and AlaAT activities of 4 and 19nkatmg(-1) protein, respectively. However, we detected no in vitro tyrosine aminotransferase (TyrAT) activity and RNAi knockdown of the enzyme had no effect on phenotype, showing that TbCSL substrates might be channeled into redundant pathways. TbCSL is in vivo localized in the cytosol and functions as a C-S lyase or an aminotransferase in planta, but the purified enzyme converts at least two substrates specifically, and can thus be utilized for further in vitro applications.

  10. The enzymology of alanine aminotransferase (AlaAT) isoforms from Hordeum vulgare and other organisms, and the HvAlaAT crystal structure.

    PubMed

    Duff, Stephen M G; Rydel, Timothy J; McClerren, Amanda L; Zhang, Wenlan; Li, Jimmy Y; Sturman, Eric J; Halls, Coralie; Chen, Songyang; Zeng, Jiamin; Peng, Jiexin; Kretzler, Crystal N; Evdokimov, Artem

    2012-12-01

    In this paper we describe the expression, purification, kinetics and biophysical characterization of alanine aminotransferase (AlaAT) from the barley plant (Hordeum vulgare). This dimeric PLP-dependent enzyme is a pivotal element of several key metabolic pathways from nitrogen assimilation to carbon metabolism, and its introduction into transgenic plants results in increased yield. The enzyme exhibits a bi-bi ping-pong reaction mechanism with a K(m) for alanine, 2-oxoglutarate, glutamate and pyruvate of 3.8, 0.3, 0.8 and 0.2 mM, respectively. Barley AlaAT catalyzes the forward (alanine-forming) reaction with a k(cat) of 25.6 s(-1), the reverse (glutamate-forming) reaction with k(cat) of 12.1 s(-1) and an equilibrium constant of ~0.5. The enzyme is also able to utilize aspartate and oxaloacetate with ~10% efficiency as compared to the native substrates, which makes it much more specific than related bacterial/archaeal enzymes (that also have lower K(m) values). We have crystallized barley AlaAT in complex with PLP and l-cycloserine and solved the structure of this complex at 2.7 Å resolution. This is the first example of a plant AlaAT structure, and it reveals a canonical aminotransferase fold similar to structures of the Thermotoga maritima, Pyrococcus furiosus, and human enzymes. This structure bridges our structural understanding of AlaAT mechanism between three kingdoms of life and allows us to shed some light on the specifics of the catalysis performed by these proteins.

  11. CaAlaAT1 catalyzes the alanine: 2-oxoglutarate aminotransferase reaction during the resistance response against Tobacco mosaic virus in hot pepper.

    PubMed

    Kim, Ki-Jeong; Park, Chang-Jin; An, Jong-Min; Ham, Byung-Kook; Lee, Boo-Ja; Paek, Kyung-Hee

    2005-08-01

    Hot pepper (Capsicum annuum L. cv. Bugang) plants exhibit a hypersensitive response (HR) upon infection by Tobacco mosaic virus (TMV) pathotype P0. To elucidate molecular mechanism that underlies this resistance, hot pepper cv. Bugang leaves were inoculated with TMV-P0 and genes specifically up-regulated during the HR were isolated by differential screening. One of the clones, CaAlaAT1 encoding a putative alanine aminotransferase (EC 2.6.1.2) exhibited organ-specific expression pattern and the transcript accumulated abundantly in red (ripe) fruit tissues. CaAlaAT1 transcript was also induced in older leaves during senescence. The expression of CaAlaAT1 gene was increased in the incompatible interaction with TMV-P0 but was not in the compatible interaction with TMV-P1.2. When a strain of Xanthomonas campestris pv. vesicatoria (Xcv) carrying an AvrBs2 gene was infiltrated into the leaves of a pepper cv. ECW 20R carrying Bs2 resistance gene, a marked induction and maintenance of CaAlaAT1 gene expression was observed. The expression of CaAlaAT1 gene was triggered by salicylic acid (SA) and ethylene but not by methyl jasmonate (MeJA). CaAlaAT1 seemed to be localized mostly at the cytosol from the polyethylene glycol (PEG)-mediated transformation experiment. CaAlaAT1 seemed to catalyze alanine: 2-oxoglutarate aminotransferase (AKT) reaction, which was a main activity among the four activities in vitro, during the resistance response against TMV in hot pepper. These results suggest that CaAlaAT1, a protein known to be involved in metabolic reactions, might be one of the components in the plant's defense signal pathway against pathogens.

  12. Determination of Alanine Aminotransferase with an Electrochemical Nano Ir-C Biosensor for the Screening of Liver Diseases

    PubMed Central

    Hsueh, Chang-Jung; Wang, Joanne H.; Dai, Liming; Liu, Chung-Chiun

    2011-01-01

    Alanine aminotransaminase (ALT), is an enzyme that normally resides in serum and body tissues, especially in the liver. It is released into the serum as a result of tissue injury; hence the concentration of ALT in the serum may be increased with acute damage to hepatic cells. A single use, disposable biosensor, comprising iridium nano-particle as catalyst dispersed on carbon paste, has been developed for the determination of ALT concentration. The biosensor is based on quantifying H2O2 concentration produced by a serial of ALT enzymatic reactions. It operates well at room temperature in different physiological fluids: phosphate buffer, calf serum and human serum for ALT concentration of 0–544 ng/mL. Experimental results in human serum are compared to those obtained by spectrophotometric assays with excellent agreement. Therefore, the Ir/C biosensor shows good relationship on the dilution of concentrated ALT clinical applications. PMID:25586923

  13. Crystal structure of the S187F variant of human liver alanine: glyoxylate [corrected] aminotransferase associated with primary hyperoxaluria type I and its functional implications.

    PubMed

    Oppici, Elisa; Fodor, Krisztian; Paiardini, Alessandro; Williams, Chris; Voltattorni, Carla Borri; Wilmanns, Matthias; Cellini, Barbara

    2013-08-01

    The substitution of Ser187, a residue located far from the active site of human liver peroxisomal alanine:glyoxylate aminotransferase (AGT), by Phe gives rise to a variant associated with primary hyperoxaluria type I. Unexpectedly, previous studies revealed that the recombinant form of S187F exhibits a remarkable loss of catalytic activity, an increased pyridoxal 5'-phosphate (PLP) binding affinity and a different coenzyme binding mode compared with normal AGT. To shed light on the structural elements responsible for these defects, we solved the crystal structure of the variant to a resolution of 2.9 Å. Although the overall conformation of the variant is similar to that of normal AGT, we noticed: (i) a displacement of the PLP-binding Lys209 and Val185, located on the re and si side of PLP, respectively, and (ii) slight conformational changes of other active site residues, in particular Trp108, the base stacking residue with the pyridine cofactor moiety. This active site perturbation results in a mispositioning of the AGT-pyridoxamine 5'-phosphate (PMP) complex and of the external aldimine, as predicted by molecular modeling studies. Taken together, both predicted and observed movements caused by the S187F mutation are consistent with the following functional properties of the variant: (i) a 300- to 500-fold decrease in both the rate constant of L-alanine half-transamination and the kcat of the overall transamination, (ii) a different PMP binding mode and affinity, and (iii) a different microenvironment of the external aldimine. Proposals for the treatment of patients bearing S187F mutation are discussed on the basis of these results.

  14. Risk factors associated with hepatitis B or C markers or elevated alanine aminotransferase level among blood donors on a tropical island: the Guadeloupe experience.

    PubMed

    Fest, T; Viel, J F; Agis, F; Coffe, C; Dupond, J L; Hervé, P

    1992-10-01

    Donated blood is currently screened for hepatitis B surface antigen (HBsAg), antibody to hepatitis B core antigen (anti-HBc), antibody to hepatitis C virus (anti-HCV), and alanine aminotransferase (ALT) levels to prevent posttransfusion hepatitis. A prospective study of 2368 blood donors was carried out in Guadeloupe (French West Indies) with a view to determining the risk factors associated with serologic abnormalities. Blood donors included in the study had to complete a questionnaire. Statistical analysis was performed on the data thus obtained: 571 donations (24%) were positive for at least one of the four analyzed markers. The results were that 3.2 percent were positive for HBsAg, 22 percent for anti-HBc, and 0.8 percent for anti-HCV, and 1.4 percent had ALT > or = 45 IU per L. A good correlation was found between anti-HCV and elevated ALT. Transfusion history and two socioeconomic categories (working class, military personnel) were found to be risk factors. Other risk factors were lifelong residence in Guadeloupe (with risk increasing with the number of years), birthplace and current residence in the southern part of the island, and the existence of gastrointestinal discomfort unrelated to viral hepatitis (odds ratio = 2.98). The results of this study illustrate the difficulty of implementing a preventive policy against posttransfusion hepatitis in a tropical area. The unique epidemiologic situation of Guadeloupe as regards hepatitis B virus has led to more restrictive criteria for the acceptance of blood donors.

  15. The consensus-based approach for gene/enzyme replacement therapies and crystallization strategies: the case of human alanine-glyoxylate aminotransferase.

    PubMed

    Mesa-Torres, Noel; Yunta, Cristina; Fabelo-Rosa, Israel; Gonzalez-Rubio, Juana María; Sánchez-Ruiz, José M; Salido, Eduardo; Albert, Armando; Pey, Angel L

    2014-09-15

    Protein stability is a fundamental issue in biomedical and biotechnological applications of proteins. Among these applications, gene- and enzyme-replacement strategies are promising approaches to treat inherited diseases that may benefit from protein engineering techniques, even though these beneficial effects have been largely unexplored. In the present study we apply a sequence-alignment statistics procedure (consensus-based approach) to improve the activity and stability of the human AGT (alanine-glyoxylate aminotransferase) protein, an enzyme which causes PH1 (primary hyperoxaluria type I) upon mutation. By combining only five consensus mutations, we obtain a variant (AGT-RHEAM) with largely enhanced in vitro thermal and kinetic stability, increased activity, and with no side effects on foldability and peroxisomal targeting in mammalian cells. The structure of AGT-RHEAM reveals changes at the dimer interface and improved electrostatic interactions responsible for increased kinetic stability. Consensus-based variants maintained the overall protein fold, crystallized more easily and improved the expression as soluble proteins in two different systems [AGT and CIPK24 (CBL-interacting serine/threonine-protein kinase) SOS2 (salt-overly-sensitive 2)]. Thus the consensus-based approach also emerges as a simple and generic strategy to increase the crystallization success for hard-to-get protein targets as well as to enhance protein stability and function for biomedical applications.

  16. Gly161 mutations associated with Primary Hyperoxaluria Type I induce the cytosolic aggregation and the intracellular degradation of the apo-form of alanine:glyoxylate aminotransferase.

    PubMed

    Oppici, Elisa; Roncador, Alessandro; Montioli, Riccardo; Bianconi, Silvia; Cellini, Barbara

    2013-12-01

    Primary Hyperoxaluria Type I (PH1) is a severe rare disorder of metabolism due to inherited mutations on liver peroxisomal alanine:glyoxylate aminotransferase (AGT), a pyridoxal 5'-phosphate (PLP)-dependent enzyme whose deficiency causes the deposition of calcium oxalate crystals in the kidneys and urinary tract. PH1 is an extremely heterogeneous disease and there are more than 150 disease-causing mutations currently known, most of which are missense mutations. Moreover, the molecular mechanisms by which missense mutations lead to AGT deficiency span from structural, functional to subcellular localization defects. Gly161 is a highly conserved residue whose mutation to Arg, Cys or Ser is associated with PH1. Here we investigated the molecular bases of the AGT deficit caused by Gly161 mutations with expression studies in a mammalian cellular system paired with biochemical analyses on the purified recombinant proteins. Our results show that the mutations of Gly161 (i) strongly reduce the expression levels and the intracellular half-life of AGT, and (ii) make the protein in the apo-form prone to an electrostatically-driven aggregation in the cell cytosol. The coenzyme PLP, by shifting the equilibrium from the apo- to the holo-form, is able to reduce the aggregation propensity of the variants, thus partly decreasing the effect of the mutations. Altogether, these results shed light on the mechanistic details underlying the pathogenicity of Gly161 variants, thus expanding our knowledge of the enzymatic phenotypes leading to AGT deficiency.

  17. Diabetes-linked transcription factor HNF4α regulates metabolism of endogenous methylarginines and β-aminoisobutyric acid by controlling expression of alanine-glyoxylate aminotransferase 2

    PubMed Central

    Burdin, Dmitry V.; Kolobov, Alexey A.; Brocker, Chad; Soshnev, Alexey A.; Samusik, Nikolay; Demyanov, Anton V.; Brilloff, Silke; Jarzebska, Natalia; Martens-Lobenhoffer, Jens; Mieth, Maren; Maas, Renke; Bornstein, Stefan R.; Bode-Böger, Stefanie M.; Gonzalez, Frank; Weiss, Norbert; Rodionov, Roman N.

    2016-01-01

    Elevated levels of circulating asymmetric and symmetric dimethylarginines (ADMA and SDMA) predict and potentially contribute to end organ damage in cardiovascular diseases. Alanine-glyoxylate aminotransferase 2 (AGXT2) regulates systemic levels of ADMA and SDMA, and also of beta-aminoisobutyric acid (BAIB)-a modulator of lipid metabolism. We identified a putative binding site for hepatic nuclear factor 4 α (HNF4α) in AGXT2 promoter sequence. In a luciferase reporter assay we found a 75% decrease in activity of Agxt2 core promoter after disruption of the HNF4α binding site. Direct binding of HNF4α to Agxt2 promoter was confirmed by chromatin immunoprecipitation assay. siRNA-mediated knockdown of Hnf4a led to an almost 50% reduction in Agxt2 mRNA levels in Hepa 1–6 cells. Liver-specific Hnf4a knockout mice exhibited a 90% decrease in liver Agxt2 expression and activity, and elevated plasma levels of ADMA, SDMA and BAIB, compared to wild-type littermates. Thus we identified HNF4α as a major regulator of Agxt2 expression. Considering a strong association between human HNF4A polymorphisms and increased risk of type 2 diabetes our current findings suggest that downregulation of AGXT2 and subsequent impairment in metabolism of dimethylarginines and BAIB caused by HNF4α deficiency might contribute to development of cardiovascular complications in diabetic patients. PMID:27752141

  18. Diabetes-linked transcription factor HNF4α regulates metabolism of endogenous methylarginines and β-aminoisobutyric acid by controlling expression of alanine-glyoxylate aminotransferase 2.

    PubMed

    Burdin, Dmitry V; Kolobov, Alexey A; Brocker, Chad; Soshnev, Alexey A; Samusik, Nikolay; Demyanov, Anton V; Brilloff, Silke; Jarzebska, Natalia; Martens-Lobenhoffer, Jens; Mieth, Maren; Maas, Renke; Bornstein, Stefan R; Bode-Böger, Stefanie M; Gonzalez, Frank; Weiss, Norbert; Rodionov, Roman N

    2016-10-18

    Elevated levels of circulating asymmetric and symmetric dimethylarginines (ADMA and SDMA) predict and potentially contribute to end organ damage in cardiovascular diseases. Alanine-glyoxylate aminotransferase 2 (AGXT2) regulates systemic levels of ADMA and SDMA, and also of beta-aminoisobutyric acid (BAIB)-a modulator of lipid metabolism. We identified a putative binding site for hepatic nuclear factor 4 α (HNF4α) in AGXT2 promoter sequence. In a luciferase reporter assay we found a 75% decrease in activity of Agxt2 core promoter after disruption of the HNF4α binding site. Direct binding of HNF4α to Agxt2 promoter was confirmed by chromatin immunoprecipitation assay. siRNA-mediated knockdown of Hnf4a led to an almost 50% reduction in Agxt2 mRNA levels in Hepa 1-6 cells. Liver-specific Hnf4a knockout mice exhibited a 90% decrease in liver Agxt2 expression and activity, and elevated plasma levels of ADMA, SDMA and BAIB, compared to wild-type littermates. Thus we identified HNF4α as a major regulator of Agxt2 expression. Considering a strong association between human HNF4A polymorphisms and increased risk of type 2 diabetes our current findings suggest that downregulation of AGXT2 and subsequent impairment in metabolism of dimethylarginines and BAIB caused by HNF4α deficiency might contribute to development of cardiovascular complications in diabetic patients.

  19. The ribavirin analog ICN 17261 demonstrates reduced toxicity and antiviral effects with retention of both immunomodulatory activity and reduction of hepatitis-induced serum alanine aminotransferase levels.

    PubMed

    Tam, R C; Ramasamy, K; Bard, J; Pai, B; Lim, C; Averett, D R

    2000-05-01

    The demonstrated utility of the nucleoside analog ribavirin in the treatment of certain viral diseases can be ascribed to its multiple distinct properties. These properties may vary in relative importance in differing viral disease conditions and include the direct inhibition of viral replication, the promotion of T-cell-mediated immune responses via an enhanced type 1 cytokine response, and a reduction of circulating alanine aminotransferase (ALT) levels associated with hepatic injury. Ribavirin also has certain known toxicities, including the induction of anemia upon chronic administration. To determine if all these properties are linked, we compared the D-nucleoside ribavirin to its L-enantiomer (ICN 17261) with regard to these properties. Strong similarities were seen for these two compounds with respect to induction of type 1 cytokine bias in vitro, enhancement of type 1 cytokine responses in vivo, and the reduction of serum ALT levels in a murine hepatitis model. In contrast, ICN 17261 had no in vitro antiviral activity against a panel of RNA and DNA viruses, while ribavirin exhibited its characteristic activity profile. Importantly, the preliminary in vivo toxicology profile of ICN 17261 is significantly more favorable than that of ribavirin. Administration of 180 mg of ICN 17261 per kg of body weight to rats by oral gavage for 4 weeks generated substantial serum levels of drug but no observable clinical pathology, whereas equivalent doses of ribavirin induced a significant anemia and leukopenia. Thus, structural modification of ribavirin can dissociate its immunomodulatory properties from its antiviral and toxicologic properties, resulting in a compound (ICN 17261) with interesting therapeutic potential.

  20. Misfolding caused by the pathogenic mutation G47R on the minor allele of alanine:glyoxylate aminotransferase and chaperoning activity of pyridoxine.

    PubMed

    Montioli, Riccardo; Oppici, Elisa; Dindo, Mirco; Roncador, Alessandro; Gotte, Giovanni; Cellini, Barbara; Borri Voltattorni, Carla

    2015-10-01

    Liver peroxisomal alanine:glyoxylate aminotransferase (AGT), a pyridoxal 5'-phosphate (PLP) enzyme, exists as two polymorphic forms, the major (AGT-Ma) and the minor (AGT-Mi) haplotype. Deficit of AGT causes Primary Hyperoxaluria Type 1 (PH1), an autosomal recessive rare disease. Although ~one-third of the 79 disease-causing missense mutations segregates on AGT-Mi, only few of them are well characterized. Here for the first time the molecular and cellular defects of G47R-Mi are reported. When expressed in Escherichia coli, the recombinant purified G47R-Mi variant exhibits only a 2.5-fold reduction of its kcat, and its apo form displays a remarkably decreased PLP binding affinity, increased dimer-monomer equilibrium dissociation constant value, susceptibility to thermal denaturation and to N-terminal region proteolytic cleavage, and aggregation propensity. When stably expressed in a mammalian cell line, we found ~95% of the intact form of the variant in the insoluble fraction, and proteolyzed (within the N-terminal region) and aggregated forms both in the soluble and insoluble fractions. Moreover, the intact and nicked forms have a peroxisomal and a mitochondrial localization, respectively. Unlike what already seen for G41R-Mi, exposure of G47R-Mi expressing cells to pyridoxine (PN) remarkably increases the expression level and the specific activity in a dose-dependent manner, reroutes all the protein to peroxisomes, and rescues its functionality. Although the mechanism of the different effect of PN on the variants G47R-Mi and G41R-Mi remains elusive, the chaperoning activity of PN may be of value in the therapy of patients bearing the G47R mutation.

  1. Enzymological and mutational analysis of a complex primary hyperoxaluria type I phenotype involving alanine: Glyoxylate aminotransferase peroxisome-to-mitochondrion mistargeting and intraperoxisomal aggregation

    SciTech Connect

    Danpure, C.J.; Purdue, P.E.; Allsop, J.; Lumb, M.J.; Jennings, P.R. ); Scheinman, J.I. ); Mauer, S.M. ); Davidson, N.O. )

    1993-08-01

    Primary hyperoxaluri type 1 (PH1) is a rare autosomal recessive disease caused by a deficiency of the liver-specific peroxisomal enzyme alanine:glyoxylate aminotransferase (AGT). Three unrelated PH1 patients, who possess a novel complex phenotype, are described. At the enzymological level, this phenotype is characterized by a complete, or nearly complete, absence of AGT catalytic activity and reduced AGT immunoreactivity. Unlike normal individuals in whom the AGT is confined to the peroxisomal matrix, the immunoreactive AGT in these three patients was distributed approximately equally between the peroxisomes and mitochondria. The peroxisomal AGT appeared to be aggregated into amorphous core-like structures in which no other peroxisomal enzymes could be identified. Mutational analysis of the AGT gene showed that two of the three patients were compound heterozygotes for two previously unrecognized point mutations which caused Gly41[yields]Arg and Phe152[yields]Iso amino acid substitutions. The third patient was shown to be a compound heterozygote for the Gly41[yields]Arg mutation and a previously recognized Gly170[yields]Arg mutation. All three patients were homozygous for the Pro11[yields]Leu polymorphism that had been found previously with a high allelic frequency in normal populations. It is suggested the the Phe152[yields]Iso and Gly170[yields]Arg substitutions, which are only eighteen residues apart and located in the same highly conserved internal region of 58 amino acids, might be involved in the inhibition of peroxisomal targeting and/or import of AGT and, in combination with the Pro11[yields]Leu polymorphism, be responsible for its aberrant mitochondrial compartmentalization. On the other hand, the Gly41[yields]Arg substitution, either in combination with the Pro11[yields]Leu polymorphism or by itself, is predicted to be responsible for the intraperoxisomal aggregation of the AGT protein. 50 refs., 8 figs., 4 tabs.

  2. L-alanine-glyoxylate aminotransferase II of rat kidney and liver mitochondria possesses cysteine S-conjugate beta-lyase activity: a contributing factor to the nephrotoxicity/hepatotoxicity of halogenated alkenes?

    PubMed Central

    Cooper, Arthur J L; Krasnikov, Boris F; Okuno, Etsuo; Jeitner, Thomas M

    2003-01-01

    Several halogenated alkenes are metabolized in part to cysteine S-conjugates, which are mitochondrial toxicants of kidney and, to a lesser extent, other organs. Toxicity is due to cysteine S-conjugate beta-lyases, which convert the cysteine S-conjugate into pyruvate, ammonia and a reactive sulphur-containing fragment. A section of the human population is exposed to halogenated alkenes. To understand the health effects of such exposure, it is important to identify cysteine S-conjugate beta-lyases that contribute to mitochondrial damage. Mitochondrial aspartate aminotransferase [Cooper, Bruschi, Iriarte and Martinez-Carrion (2002) Biochem. J. 368, 253-261] and mitochondrial branched-chain aminotransferase [Cooper, Bruschi, Conway and Hutson (2003) Biochem. Pharmacol. 65, 181-192] exhibit beta-lyase activity toward S -(1,2-dichlorovinyl)-L-cysteine (the cysteine S-conjugate of trichloroethylene) and S -(1,1,2,2-tetrafluoroethyl)-L-cysteine (the cysteine S-conjugate of tetrafluoroethylene). Turnover leads to eventual inactivation of these enzymes. Here we report that mitochondrial L-alanine-glyoxylate aminotransferase II, which, in the rat, is most active in kidney, catalyses cysteine S-conjugate beta-lyase reactions with S -(1,1,2,2-tetrafluoroethyl)-L-cysteine, S -(1,2-dichlorovinyl)-L-cysteine and S -(benzothiazolyl-L-cysteine); turnover leads to inactivation. Previous workers showed that the reactive-sulphur-containing fragment released from S -(1,1,2,2-tetrafluoroethyl)-L-cysteine and S -(1,2-dichlorovinyl)-L-cysteine is toxic by acting as a thioacylating agent - particularly of lysine residues in nearby proteins. Toxicity, however, may also involve 'self-inactivation' of key enzymes. The present findings suggest that alanine-glyoxylate aminotransferase II may be an important factor in the well-established targeting of rat kidney mitochondria by toxic halogenated cysteine S-conjugates. Previous reports suggest that alanine-glyoxylate aminotransferase II is absent

  3. Alanine-glyoxylate aminotransferase 2 (AGXT2) polymorphisms have considerable impact on methylarginine and β-aminoisobutyrate metabolism in healthy volunteers.

    PubMed

    Kittel, Anja; Müller, Fabian; König, Jörg; Mieth, Maren; Sticht, Heinrich; Zolk, Oliver; Kralj, Ana; Heinrich, Markus R; Fromm, Martin F; Maas, Renke

    2014-01-01

    Elevated plasma concentrations of asymmetric (ADMA) and symmetric (SDMA) dimethylarginine have repeatedly been linked to adverse clinical outcomes. Both methylarginines are substrates of alanine-glyoxylate aminotransferase 2 (AGXT2). It was the aim of the present study to simultaneously investigate the functional relevance and relative contributions of common AGXT2 single nucleotide polymorphisms (SNPs) to plasma and urinary concentrations of methylarginines as well as β-aminoisobutyrate (BAIB), a prototypic substrate of AGXT2. In a cohort of 400 healthy volunteers ADMA, SDMA and BAIB concentrations were determined in plasma and urine using HPLC-MS/MS and were related to the coding AGXT2 SNPs rs37369 (p.Val140Ile) and rs16899974 (p.Val498Leu). Volunteers heterozygous or homozygous for the AGXT2 SNP rs37369 had higher SDMA plasma concentrations by 5% and 20% (p = 0.002) as well as higher BAIB concentrations by 54% and 146%, respectively, in plasma and 237% and 1661%, respectively, in urine (both p<0.001). ADMA concentrations were not affected by both SNPs. A haplotype analysis revealed that the second investigated AGXT2 SNP rs16899974, which was not significantly linked to the other AGXT2 SNP, further aggravates the effect of rs37369 with respect to BAIB concentrations in plasma and urine. To investigate the impact of the amino acid exchange p.Val140Ile, we established human embryonic kidney cell lines stably overexpressing wild-type or mutant (p.Val140Ile) AGXT2 protein and assessed enzyme activity using BAIB and stable-isotope labeled [²H₆]-SDMA as substrate. In vitro, the amino acid exchange of the mutant protein resulted in a significantly lower enzyme activity compared to wild-type AGXT2 (p<0.05). In silico modeling of the SNPs indicated reduced enzyme stability and substrate binding. In conclusion, SNPs of AGXT2 affect plasma as well as urinary BAIB and SDMA concentrations linking methylarginine metabolism to the common genetic trait of hyper

  4. Alanine-glyoxylate aminotransferase 2 (AGXT2) Polymorphisms Have Considerable Impact on Methylarginine and β-aminoisobutyrate Metabolism in Healthy Volunteers

    PubMed Central

    König, Jörg; Mieth, Maren; Sticht, Heinrich; Zolk, Oliver; Kralj, Ana; Heinrich, Markus R.; Fromm, Martin F.; Maas, Renke

    2014-01-01

    Elevated plasma concentrations of asymmetric (ADMA) and symmetric (SDMA) dimethylarginine have repeatedly been linked to adverse clinical outcomes. Both methylarginines are substrates of alanine-glyoxylate aminotransferase 2 (AGXT2). It was the aim of the present study to simultaneously investigate the functional relevance and relative contributions of common AGXT2 single nucleotide polymorphisms (SNPs) to plasma and urinary concentrations of methylarginines as well as β-aminoisobutyrate (BAIB), a prototypic substrate of AGXT2. In a cohort of 400 healthy volunteers ADMA, SDMA and BAIB concentrations were determined in plasma and urine using HPLC-MS/MS and were related to the coding AGXT2 SNPs rs37369 (p.Val140Ile) and rs16899974 (p.Val498Leu). Volunteers heterozygous or homozygous for the AGXT2 SNP rs37369 had higher SDMA plasma concentrations by 5% and 20% (p = 0.002) as well as higher BAIB concentrations by 54% and 146%, respectively, in plasma and 237% and 1661%, respectively, in urine (both p<0.001). ADMA concentrations were not affected by both SNPs. A haplotype analysis revealed that the second investigated AGXT2 SNP rs16899974, which was not significantly linked to the other AGXT2 SNP, further aggravates the effect of rs37369 with respect to BAIB concentrations in plasma and urine. To investigate the impact of the amino acid exchange p.Val140Ile, we established human embryonic kidney cell lines stably overexpressing wild-type or mutant (p.Val140Ile) AGXT2 protein and assessed enzyme activity using BAIB and stable-isotope labeled [2H6]-SDMA as substrate. In vitro, the amino acid exchange of the mutant protein resulted in a significantly lower enzyme activity compared to wild-type AGXT2 (p<0.05). In silico modeling of the SNPs indicated reduced enzyme stability and substrate binding. In conclusion, SNPs of AGXT2 affect plasma as well as urinary BAIB and SDMA concentrations linking methylarginine metabolism to the common genetic trait of hyper

  5. Recurrent truncating mutations in alanine-glyoxylate aminotransferase gene in two South Indian families with primary hyperoxaluria type 1 causing later onset end-stage kidney disease

    PubMed Central

    Dutta, A. K.; Paulose, B. K.; Danda, S.; Alexander, S.; Tamilarasi, V.; Omprakash, S.

    2016-01-01

    Primary hyperoxaluria type 1 is an autosomal recessive inborn error of metabolism due to liver-specific peroxisomal enzyme alanine-glyoxylate transaminase deficiency. Here, we describe two unrelated patients who were diagnosed to have primary hyperoxaluria. Homozygous c.445_452delGTGCTGCT (p.L151Nfs*14) (Transcript ID: ENST00000307503; human genome assembly GRCh38.p2) (HGMD ID CD073567) mutation was detected in both the patients and the parents were found to be heterozygous carriers. Our patients developed end-stage renal disease at 23 years and 35 years of age. However, in the largest series published from OxalEurope cohort, the median age of end-stage renal disease for null mutations carriers was 9.9 years, which is much earlier than our cases. Our patients had slower progressions as compared to three unrelated patients from North India and Pakistan, who had homozygous c.302T>C (p.L101P) (HGMD ID CM093792) mutation in exon 2. Further, patients need to be studied to find out if c.445_452delGTGCTGCT mutation represents a founder mutation in Southern India. PMID:27512303

  6. Creatine supplementation.

    PubMed

    Hall, Matthew; Trojian, Thomas H

    2013-01-01

    Creatine monohydrate is a dietary supplement that increases muscle performance in short-duration, high-intensity resistance exercises, which rely on the phosphocreatine shuttle for adenosine triphosphate. The effective dosing for creatine supplementation includes loading with 0.3 g·kg·d for 5 to 7 days, followed by maintenance dosing at 0.03 g·kg·d most commonly for 4 to 6 wk. However loading doses are not necessary to increase the intramuscular stores of creatine. Creatine monohydrate is the most studied; other forms such as creatine ethyl ester have not shown added benefits. Creatine is a relatively safe supplement with few adverse effects reported. The most common adverse effect is transient water retention in the early stages of supplementation. When combined with other supplements or taken at higher than recommended doses for several months, there have been cases of liver and renal complications with creatine. Further studies are needed to evaluate the remote and potential future adverse effects from prolonged creatine supplementation.

  7. Evolution of alanine:glyoxylate aminotransferase 1 peroxisomal and mitochondrial targeting. A survey of its subcellular distribution in the livers of various representatives of the classes Mammalia, Aves and Amphibia.

    PubMed

    Danpure, C J; Fryer, P; Jennings, P R; Allsop, J; Griffiths, S; Cunningham, A

    1994-08-01

    As part of a wider study on the molecular evolution of alanine:glyoxylate aminotransferase 1 (AGT1) intracellular compartmentalization, we have determined the subcellular distribution of immunoreactive AGT1, using postembedding protein A-gold immunoelectron microscopy, in the livers of various members of the classes Mammalia, Aves, and Amphibia. As far as organellar distribution is concerned, three categories could be distinguished. In members of the first category (type I), all, or nearly all, of the immunoreactive AGT1 was concentrated within the peroxisomes. In the second category (type II), AGT1 was found more evenly distributed in both peroxisomes and mitochondria. In the third category (type III), AGT1 was localized mainly within the mitochondria with much lower, but widely variable, amounts in the peroxisomes. Type I animals include the human, two great apes (gorilla, orangutan), two Old World monkeys (anubis baboon, Japanese macaque), a New World monkey (white-faced Saki monkey), a lago, morph (European rabbit), a bat (Seba's short-tailed fruit bat), two caviomorph rodents (guinea pig, orange-rumped agouti), and two Australian marsupials (koala, Bennett's wallaby). Type II animals include two New World monkeys (common marmoset, cotton-top tamarin), three prosimians (brown lemur, fat-tailed dwarf lemur, pygmy slow loris), five rodents (a hybrid crested porcupine, Colombian ground squirrel, laboratory rat, laboratory mouse, golden hamster), an American marsupial (grey short-tailed opossum), and a bird (raven). Type III animals include the large tree shrew, three insectivores (common Eurasian mole, European hedgehog, house shrew), four carnivores (domestic cat, ocelot, domestic dog, polecat ferret), and an amphibian (common frog). In addition to these categories, some animals (e.g. guinea pig, common frog) possessed significant amounts of cytosolic AGT1. Whereas the subcellular distribution of AGT1 in some orders (e.g. Insectivora and Carnivora) did not appear

  8. AGXT2: a promiscuous aminotransferase

    PubMed Central

    Rodionov, Roman N.; Jarzebska, Natalia; Weiss, Norbert; Lentz, Steven R.

    2014-01-01

    Alanine-glyoxylate aminotransferase 2 (AGXT2) is a multifunctional mitochondrial aminotransferase that was first identified in 1978. The physiological importance of AGXT2 was largely overlooked for three decades because AGXT2 is less active in glyoxylate metabolism than AGXT1, the enzyme that is deficient in primary hyperoxaluria type I. Recently, several novel functions of AGXT2 have been “rediscovered” in the setting of modern genomic and metabolomic studies. It is now apparent that AGXT2 has multiple substrates and products and that altered AGXT2 activity may contribute to the pathogenesis of cardiovascular, renal, neurological and hematological diseases. This article reviews the biochemical properties and physiological functions of AGXT2, its unique role at the intersection of key mitochondrial pathways, and its potential as a drug target. PMID:25294000

  9. Serum creatine kinase B subunit activity in diagnosis of acute myocardial infarction.

    PubMed Central

    Ljungdahl, L; Gerhardt, W; Hofvendahl, S

    1980-01-01

    The value of serum creatine kinase B subunit activity (CK B) in the diagnosis of acute myocardial infarction was studied in 238 consecutive cases. All were admitted to a coronary care unit because of suspected acute myocardial infarction. Serum CK B activity was determined by an immunoinhibition procedure, using a CK M subunit inhibiting antibody (anti-M). For the evaluation of serum CK B, patients were classified into acute myocardial infarction and non-acute myocardial infarction groups. This classification was based on electrocardiographic findings, on quantitative determinations of serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and total serum creatine kinase (CK) activities, and on qualitative electrophoretic determinations of serum CK and serum lactate dehydrogenase (LD) isoenzymes. The prevalence of acute myocardial infarction in the patient material was 0.47. Serum CK B subunit activity was found to be a highly selective indicator of acute myocardial infarction with a predictive value of a positive test result of 0.97 and a predictive value of a negative test result of 0.99. The serum CK B activity increased above the acute myocardial infarction discrimination limit within 12 hours from onset of symptoms. Two non-acute myocardial infarction patients, who were resuscitated after cardiac arrest, had increased serum CK B values caused by the transient presence of CK isoenzyme BB in serum. PMID:7378210

  10. Plasma aminotransferase concentrations in preterm infants.

    PubMed

    Victor, S; Dickinson, H; Turner, M A

    2011-03-01

    The aim of this study was to generate reference ranges for aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in preterm infants by describing the observed plasma concentration of these enzymes in babies born between 22 and 36 weeks' gestation. A service evaluation was conducted in babies admitted to two large neonatal intensive care units in the UK. 7006 blood samples from 1860 infants admitted to the two units between 2004 and 2008 were included. Extremely premature infants had high plasma enzyme activities when compared to babies at a later corrected gestational age. This may be due to more severe illness immediately after birth.

  11. Antiretroviral Drugs and Risk of Chronic Alanine Aminotransferase Elevation in Human Immunodeficiency Virus (HIV)-Monoinfected Persons: The Data Collection on Adverse Events of Anti-HIV Drugs Study

    PubMed Central

    Kovari, Helen; Sabin, Caroline A.; Ledergerber, Bruno; Ryom, Lene; Reiss, Peter; Law, Matthew; Pradier, Christian; Dabis, Francois; d'Arminio Monforte, Antonella; Smith, Colette; de Wit, Stephane; Kirk, Ole; Lundgren, Jens D.; Weber, Rainer

    2016-01-01

    Background. Although human immunodeficiency virus (HIV)-positive persons on antiretroviral therapy (ART) frequently have chronic liver enzyme elevation (cLEE), the underlying cause is often unclear. Methods. Data Collection on Adverse Events of Anti-HIV Drugs (D:A:D) Study participants without chronic viral hepatitis were observed to the earliest of cLEE (elevated aminotransferase ≥6 months), death, last follow-up, or January 2, 2014. Antiretroviral treatment exposure was categorized as follows: no exposure and ongoing short- and long-term exposure (<2 or ≥2 years) after initiation. Association between development of cLEE and ART exposure was investigated using Poisson regression. Results. Among 21 485 participants observed for 105 413 person-years (PY), 6368 developed cLEE (incidence 6.04/100 PY; 95% confidence interval [CI], 5.89–6.19). Chronic liver enzyme elevation was associated with short-and long-term exposure to didanosine (<2 years rate ratio [RR] = 1.29, 95% CI, 1.11–1.49; >2 years RR = 1.26, 95% CI, 1.13–1.41); stavudine (<2 years RR = 1.51, 95% CI, 1.26–1.81; >2 years RR = 1.17, 95% CI, 1.03–1.32), and tenofovir disoproxil fumarate (<2 years RR = 1.55, 95% CI, 1.40–1.72; >2 years RR = 1.18, 95% CI, 1.05–1.32), but only short-term exposure to nevirapine (<2 years RR = 1.44, 95% CI, 1.29–1.61), efavirenz (<2 years RR = 1.14, 95% CI, 1.03–1.26), emtricitabine (<2 years RR = 1.18, 95% CI, 1.04–1.33), and atazanavir (<2 years RR = 1.20, 95% CI, 1.04–1.38). Chronic liver enzyme elevation was not associated with use of lamivudine, abacavir, and other protease inhibitors. Mortality did not differ between participants with and without cLEE. Conclusions. Although didanosine, stavudine, nevirapine, and efavirenz have been described to be hepatotoxic, we additionally observed a consistent association between tenofovir and cLEE emerging within the first 2 years after drug initiation. This novel tenofovir-cLEE signal should be

  12. Comparison of blood aminotransferase methods for assessment of myopathy and hepatopathy in Florida manatees (Trichechus manatus latirostris).

    PubMed

    Harr, Kendal E; Allison, Kathryn; Bonde, Robert K; Murphy, David; Harvey, John W

    2008-06-01

    Muscle injury is common in Florida manatees (Trichechus manatus latirostris). Plasma aspartate aminotransferase (AST) is frequently used to assess muscular damage in capture myopathy and traumatic injury. Therefore, accurate measurement of AST and alanine aminotransferase (ALT) is important in managed, free-ranging animals, as well as in those rehabilitating from injury. Activities of these enzymes, however, are usually not increased in manatees with either acute or chronic muscle damage, despite marked increases in plasma creatine kinase activity. It is hypothesized that this absence of response is due to apoenzymes in the blood not detected by commonly used veterinary assays. Addition of coenzyme pyridoxal-5-phosphate (P5P or vitamin B6) should, therefore, result in higher measured enzyme activities. The objective of this study was to determine the most accurate, precise, and diagnostically useful method for aminotransferase measurement in manatees that can be used in veterinary practices and diagnostic laboratories. Additionally, appropriate collection and storage techniques were assessed. The use of an optimized commercial wet chemical assay with 100 micromol P5P resulted in a positive bias of measured enzyme activities in a healthy population of animals. However, AST and ALT were still much lower than that typically observed in domestic animals and should not be used alone in the assessment of capture myopathy and muscular trauma. Additionally, the dry chemistry analyzer, typically used in clinics, reported significantly higher and less precise AST and ALT activities with poor correlation to those measured with wet chemical methods found in diagnostic laboratories. Therefore, these results cannot be clinically compared. Overall, the optimized wet chemical method was the most precise and diagnostically useful measurement of aminotransferase in samples. Additionally, there was a statistically significant difference between paired serum and plasma measurement

  13. Neuroprotective effects of creatine.

    PubMed

    Beal, M Flint

    2011-05-01

    There is a substantial body of literature, which has demonstrated that creatine has neuroprotective effects both in vitro and in vivo. Creatine can protect against excitotoxicity as well as against β-amyloid toxicity in vitro. We carried out studies examining the efficacy of creatine as a neuroprotective agent in vivo. We demonstrated that creatine can protect against excitotoxic lesions produced by N-methyl-D: -aspartate. We also showed that creatine is neuroprotective against lesions produced by the toxins malonate and 3-nitropropionic acid (3-NP) which are reversible and irreversible inhibitors of succinate dehydrogenase, respectively. Creatine produced dose-dependent neuroprotective effects against MPTP toxicity reducing the loss of dopamine within the striatum and the loss of dopaminergic neurons in the substantia nigra. We carried out a number of studies of the neuroprotective effects of creatine in transgenic mouse models of neurodegenerative diseases. We demonstrated that creatine produced an extension of survival, improved motor performance, and a reduction in loss of motor neurons in a transgenic mouse model of amyotrophic lateral sclerosis (ALS). Creatine produced an extension of survival, as well as improved motor function, and a reduction in striatal atrophy in the R6/2 and the N-171-82Q transgenic mouse models of Huntington's disease (HD), even when its administration was delayed until the onset of disease symptoms. We recently examined the neuroprotective effects of a combination of coenzyme Q10 (CoQ10) with creatine against both MPTP and 3-NP toxicity. We found that the combination of CoQ and creatine together produced additive neuroprotective effects in a chronic MPTP model, and it blocked the development of alpha-synuclein aggregates. In the 3-NP model of HD, CoQ and creatine produced additive neuroprotective effects against the size of the striatal lesions. In the R6/2 transgenic mouse model of HD, the combination of CoQ and creatine produced

  14. Paralogous ALT1 and ALT2 Retention and Diversification Have Generated Catalytically Active and Inactive Aminotransferases in Saccharomyces cerevisiae

    PubMed Central

    Peñalosa-Ruiz, Georgina; Aranda, Cristina; Ongay-Larios, Laura; Colon, Maritrini; Quezada, Hector; Gonzalez, Alicia

    2012-01-01

    Background Gene duplication and the subsequent divergence of paralogous pairs play a central role in the evolution of novel gene functions. S. cerevisiae possesses two paralogous genes (ALT1/ALT2) which presumably encode alanine aminotransferases. It has been previously shown that Alt1 encodes an alanine aminotransferase, involved in alanine metabolism; however the physiological role of Alt2 is not known. Here we investigate whether ALT2 encodes an active alanine aminotransferase. Principal Findings Our results show that although ALT1 and ALT2 encode 65% identical proteins, only Alt1 displays alanine aminotransferase activity; in contrast ALT2 encodes a catalytically inert protein. ALT1 and ALT2 expression is modulated by Nrg1 and by the intracellular alanine pool. ALT1 is alanine-induced showing a regulatory profile of a gene encoding an enzyme involved in amino acid catabolism, in agreement with the fact that Alt1 is the sole pathway for alanine catabolism present in S. cerevisiae. Conversely, ALT2 expression is alanine-repressed, indicating a role in alanine biosynthesis, although the encoded-protein has no alanine aminotransferase enzymatic activity. In the ancestral-like yeast L. kluyveri, the alanine aminotransferase activity was higher in the presence of alanine than in the presence of ammonium, suggesting that as for ALT1, LkALT1 expression could be alanine-induced. ALT2 retention poses the questions of whether the encoded protein plays a particular function, and if this function was present in the ancestral gene. It could be hypotesized that ALT2 diverged after duplication, through neo-functionalization or that ALT2 function was present in the ancestral gene, with a yet undiscovered function. Conclusions ALT1 and ALT2 divergence has resulted in delegation of alanine aminotransferase activity to Alt1. These genes display opposed regulatory profiles: ALT1 is alanine-induced, while ALT2 is alanine repressed. Both genes are negatively regulated by the Nrg1

  15. Elevation of Alanine Aminotransferase Activity Occurs after Activation of the Cell-Death Signaling Initiated by Pattern-Recognition Receptors ‎but before Activation of Cytolytic Effectors in NK or CD8+ T Cells in the Liver During Acute HCV Infection

    PubMed Central

    Choi, Youkyung H.; Jin, Nancy; Kelly, Fiona; Sakthivel, SenthilKumar K.; Yu, Tianwei

    2016-01-01

    Pattern-recognition receptors (PRRs) promote host defenses against HCV infection by binding to their corresponding adapter molecules leading to the initiation of innate immune responses including cell death. We investigated the expression of PRR genes, biomarkers of liver cell-death, and T cell and NK cell activation/inhibition-related genes in liver and serum obtained from three experimentally infected chimpanzees with acute HCV infection, and analyzed the correlation between gene expression levels and clinical profiles. Our results showed that expression of hepatic RIG-I, TLR3, TLR7, 2OAS1, and CXCL10 mRNAs was upregulated as early as 7 days post-inoculation and peaked 12 to 83 days post-inoculation. All of the three HCV infected chimpanzees exhibited significant elevations of serum alanine aminotransferase (ALT) activity between 70 and 95 days after inoculation. Elevated levels of serum cytokeratin 18 (CK-18) and caspases 3 and 7 activity coincided closely with the rise of ALT activity, and were preceded by significant increases in levels of caspase 3 and caspase 7 mRNAs in the liver. Particularly we found that significant positive auto-correlations were observed between RIG-I, TLR3, CXCL10, 2OAS1, and PD-L1 mRNA and ALT activity at 3 to 12 days before the peak of ALT activity. However, we observed substantial negative auto-correlations between T cell and NK cell activation/inhibition-related genes and ALT activity at 5 to 32 days after the peak of ALT activity. Our results indicated cell death signaling is preceded by early induction of RIG-I, TLR3, 2OAS1, and CXCL10 mRNAs which leads to elevation of ALT activity and this signaling pathway occurs before the activation of NK and T cells during acute HCV infection. Our study suggests that PRRs and type I IFN response may play a critical role in development of liver cell injury related to viral clearance during acute HCV infection. PMID:27788241

  16. Studies on some kinetic parameters of aminotransferases in tissues of the snail, Pila globosa (Swainson) during malathion intoxication.

    PubMed

    Sahib, I K; Rao, K R

    1988-01-01

    The aspartate and alanine aminotransferases in the tissues of the snail, Pila globosa showed high catalytic potentials (low Km and high Vmax) during malathion exposure in vivo. In vitro addition of different concentrations of malathion did not influence aminotransferase activity. The results are discussed in relation to the regulative influence of the intracellular environment of the cell.

  17. Irreversible brain creatine deficiency with elevated serum and urine creatine: a creatine transporter defect?

    PubMed

    Cecil, K M; Salomons, G S; Ball, W S; Wong, B; Chuck, G; Verhoeven, N M; Jakobs, C; DeGrauw, T J

    2001-03-01

    Recent reports highlight the utility of in vivo magnetic resonance spectroscopy (MRS) techniques to recognize creatine deficiency syndromes affecting the central nervous system (CNS). Reported cases demonstrate partial reversibility of neurologic symptoms upon restoration of CNS creatine levels with the administration of oral creatine. We describe a patient with a brain creatine deficiency syndrome detected by proton MRS that differs from published reports. Metabolic screening revealed elevated creatine in the serum and urine, with normal levels of guanidino acetic acid. Unlike the case with other reported creatine deficiency syndromes, treatment with oral creatine monohydrate demonstrated no observable increase in brain creatine with proton MRS and no improvement in clinical symptoms. In this study, we report a novel brain creatine deficiency syndrome most likely representing a creatine transporter defect.

  18. Comparison of blood aminotransferase methods for assessment of myopathy and hepatopathy in Florida manatees (Trichechus manatus latirostris)

    USGS Publications Warehouse

    Harr, K.E.; Allison, K.; Bonde, R.K.; Murphy, D.; Harvey, J.W.

    2008-01-01

    Muscle injury is common in Florida manatees (Trichechus manatus latirostris). Plasma aspartate amino-transferase (AST) is frequently used to assess muscular damage in capture myopathy and traumatic injury. Therefore, accurate measurement of AST and alanine aminotransferase (ALT) is important in managed, free-ranging animals, as well as in those rehabilitating from injury. Activities of these enzymes, however, are usually not increased in manatees with either acute or chronic muscle damage, despite marked increases in plasma creatine kinase activity. It is hypothesized that this absence of response is due to apoenzymes in the blood not detected by commonly used veterinary assays. Addition of coenzyme pyridoxal-5-phosphate (P5P or vitamin B6) should, therefore, result in higher measured enzyme activities. The objective of this study was to determine the most accurate, precise, and diagnostically useful method for aminotransferase measurement in manatees that can be used in veterinary practices and diagnostic laboratories. Additionally, appropriate collection and storage techniques were assessed. The use of an optimized commercial wet chemical assay with 100 ??mol P5P resulted in a positive bias of measured enzyme activities in a healthy population of animals. However, AST and ALT were still much lower than that typically observed in domestic animals and should not be used alone in the assessment of capture myopathy and muscular trauma. Additionally, the dry chemistry analyzer, typically used in clinics, reported significantly higher and less precise AST and ALT activities with poor correlation to those measured with wet chemical methods found in diagnostic laboratories. Therefore, these results cannot be clinically compared. Overall, the optimized wet chemical method was the most precise and diagnostically useful measurement of aminotransferase in samples. Additionally, there was a statistically significant difference between paired serum and plasma measurement

  19. Molecular cloning, expression and characterization of pyridoxamine–pyruvate aminotransferase

    PubMed Central

    Yoshikane, Yu; Yokochi, Nana; Ohnishi, Kouhei; Hayashi, Hideyuki; Yagi, Toshiharu

    2006-01-01

    Pyridoxamine–pyruvate aminotransferase is a PLP (pyridoxal 5′-phosphate) (a coenzyme form of vitamin B6)-independent aminotransferase which catalyses a reversible transamination reaction between pyridoxamine and pyruvate to form pyridoxal and L-alanine. The gene encoding the enzyme has been identified, cloned and overexpressed for the first time. The mlr6806 gene on the chromosome of a symbiotic nitrogen-fixing bacterium, Mesorhizobium loti, encoded the enzyme, which consists of 393 amino acid residues. The primary sequence was identical with those of archaeal aspartate aminotransferase and rat serine–pyruvate aminotransferase, which are PLP-dependent aminotransferases. The results of fold-type analysis and the consensus amino acid residues found around the active-site lysine residue identified in the present study showed that the enzyme could be classified into class V aminotransferases of fold type I or the AT IV subfamily of the α family of the PLP-dependent enzymes. Analyses of the absorption and CD spectra of the wild-type and point-mutated enzymes showed that Lys197 was essential for the enzyme activity, and was the active-site lysine residue that corresponded to that found in the PLP-dependent aminotransferases, as had been suggested previously [Hodsdon, Kolb, Snell and Cole (1978) Biochem. J. 169, 429–432]. The Kd value for pyridoxal determined by means of CD was 100-fold lower than the Km value for it, suggesting that Schiff base formation between pyridoxal and the active-site lysine residue is partially rate determining in the catalysis of pyridoxal. The active-site structure and evolutionary aspects of the enzyme are discussed. PMID:16545075

  20. Aminotransferase levels in relation to short-term use of acetaminophen four grams daily in postoperative cardiothoracic patients in the intensive care unit.

    PubMed

    Ahlers, S J G M; Van Gulik, L; Van Dongen, E P A; Bruins, P; Tibboel, D; Knibbe, C A J

    2011-11-01

    A volunteer study suggested that taking paracetamol 4 g daily could result in elevated alanine aminotransferase plasma levels in a substantial proportion of healthy volunteers. The safety of this dose of paracetamol for acute postoperative pain remains controversial. This study aimed to examine the incidence of alanine aminotransferase elevations after short-term use of paracetamol 4 g daily, as part of the standard pain management protocol, for 93 consecutive patients after cardiothoracic surgery. Alanine aminotransferase levels and other liver function tests were measured preoperatively as baseline and once daily after surgery during the intensive care unit stay. Preoperative alanine aminotransferase levels of more than one time the upper limit of normal (ULN >40 U/l) was observed in 11% (n=10) of the patients but none of these baseline alanine aminotransferase levels exceeded three times the ULN (>3 x ULN). The average daily dose of paracetamol administered was 50 mg/kg (SD=16) after surgery. Postoperative alanine aminotransferase levels of >1 x ULN was observed in 17% (n=16), and 4% (n=4) exceeded >3 x ULN The other liver function tests of the latter four patients, including aspartate aminotransferase (range 173 to 5590 U/l), gamma-glutamyltransferase (range 56 to 103 U/l), lactate dehydrogenase (range 376 to 3518 U/l) and the International Normalised Ratio (range 2.0 to 6.6), were all abnormal. These four patients all had right ventricular failure or cardiogenic shock during the postoperative period which could explain the significant rises in alanine aminotransferase after surgery. In conclusion, the incidence of significant alanine aminotransferase elevations after using daily paracetamol as an analgesic agent for cardiac surgery, at a dose of 4 g per day, was low and mostly due to complications after surgery. Our results, albeit still very limited, provided some reassurance about the safety of paracetamol 4 g daily, as a supplementary analgesic agent for

  1. Human, rat and chicken small intestinal Na+-Cl−-creatine transporter: functional, molecular characterization and localization

    PubMed Central

    Peral, M J; García-Delgado, M; Calonge, M L; Durán, J M; De La Horra, M C; Wallimann, T; Speer, O; Ilundáin, A A

    2002-01-01

    In spite of all the fascinating properties of oral creatine supplementation, the mechanism(s) mediating its intestinal absorption has(have) not been investigated. The purpose of this study was to characterize intestinal creatine transport. [14C]Creatine uptake was measured in chicken enterocytes and rat ileum, and expression of the creatine transporter CRT was examined in human, rat and chicken small intestine by reverse transcription-polymerase chain reaction, Northern blot, in situ hybridization, immunoblotting and immunohistochemistry. Results show that enterocytes accumulate creatine against its concentration gradient. This accumulation was electrogenic, Na+- and Cl−-dependent, with a probable stoichiometry of 2 Na+: 1 Cl−: 1 creatine, and inhibited by ouabain and iodoacetic acid. The kinetic study revealed a Km for creatine of 29 μm. [14C]Creatine uptake was efficiently antagonized by non-labelled creatine, guanidinopropionic acid and cyclocreatine. More distant structural analogues of creatine, such as GABA, choline, glycine, β-alanine, taurine and betaine, had no effect on intestinal creatine uptake, indicating a high substrate specificity of the creatine transporter. Consistent with these functional data, messenger RNA for CRT was detected only in the cells lining the intestinal villus. The sequences of partial clones, and of the full-length cDNA clone, isolated from human and rat small intestine were identical to previously cloned CRT cDNAs. Immunological analysis revealed that CRT protein was mainly associated with the apical membrane of the enterocytes. This study reports for the first time that mammalian and avian enterocytes express CRT along the villus, where it mediates high-affinity, Na+- and Cl−-dependent, apical creatine uptake. PMID:12433955

  2. Aspartate aminotransferase (AST) blood test

    MedlinePlus

    ... 2016:chap 73. Read More Acute kidney failure Acute pancreatitis Alanine transaminase (ALT) blood test ALP - blood test Burns Cardiac catheterization Enzyme Heart attack Hemolytic anemia Hepatic Liver cancer - hepatocellular carcinoma Liver ...

  3. Alanine water complexes.

    PubMed

    Vaquero, Vanesa; Sanz, M Eugenia; Peña, Isabel; Mata, Santiago; Cabezas, Carlos; López, Juan C; Alonso, José L

    2014-04-10

    Two complexes of alanine with water, alanine-(H2O)n (n = 1,2), have been generated by laser ablation of the amino acid in a supersonic jet containing water vapor and characterized using Fourier transform microwave spectroscopy. In the observed complexes, water molecules bind to the carboxylic group of alanine acting as both proton donors and acceptors. In alanine-H2O, the water molecule establishes two intermolecular hydrogen bonds forming a six-membered cycle, while in alanine-(H2O)2 the two water molecules establish three hydrogen bonds forming an eight-membered ring. In both complexes, the amino acid moiety is in its neutral form and shows the conformation observed to be the most stable for the bare molecule. The microsolvation study of alanine-(H2O)n (n = 1,2) can be taken as a first step toward understanding bulk properties at a microscopic level.

  4. The Effect of Artichoke Leaf Extract on Alanine Aminotransferase and Aspartate Aminotransferase in the Patients with Nonalcoholic Steatohepatitis

    PubMed Central

    Rangboo, Vajiheh; Noroozi, Mostafa; Zavoshy, Roza; Rezadoost, Seyed Amirmansoor; Mohammadpoorasl, Asghar

    2016-01-01

    Background. Based on recent basic and clinical investigations, the extract of artichoke (Cynara scolymus) leaf has been revealed to be used for hepatoprotective and cholesterol reducing purposes. We aimed to assess the therapeutic effects of artichoke on biochemical and liver biomarkers in patients with nonalcoholic steatohepatitis (NASH). Methods. In a randomized double blind clinical trial, 60 consecutive patients suffering NASH were randomly assigned to receive Cynara scolymus extract (as 6 tablets per day consisting of 2700 mg extract of the herb) as the intervention group or placebo as the control group for two months. Results. Comparing changes in study markers following interventions showed improvement in liver enzymes. The levels of triglycerides and cholesterol were significantly reduced in the group treated with Cynara scolymus when compared to placebo group. To compare the role of Cynara scolymus use with placebo in changes in study parameters, multivariate linear regression models were employed indicating higher improvement in liver enzymes and also lipid profile particularly triglycerides and total cholesterol following administration of Cynara scolymus in comparison with placebo use. Conclusion. This study sheds light on the potential hepatoprotective activity and hypolipidemic effect of Cynara scolymus in management of NASH. This clinical trial is registered in the IRCT, Iranian Registry of Clinical Trials, by number IRCT2014070218321N1. PMID:27293900

  5. The Effect of Artichoke Leaf Extract on Alanine Aminotransferase and Aspartate Aminotransferase in the Patients with Nonalcoholic Steatohepatitis.

    PubMed

    Rangboo, Vajiheh; Noroozi, Mostafa; Zavoshy, Roza; Rezadoost, Seyed Amirmansoor; Mohammadpoorasl, Asghar

    2016-01-01

    Background. Based on recent basic and clinical investigations, the extract of artichoke (Cynara scolymus) leaf has been revealed to be used for hepatoprotective and cholesterol reducing purposes. We aimed to assess the therapeutic effects of artichoke on biochemical and liver biomarkers in patients with nonalcoholic steatohepatitis (NASH). Methods. In a randomized double blind clinical trial, 60 consecutive patients suffering NASH were randomly assigned to receive Cynara scolymus extract (as 6 tablets per day consisting of 2700 mg extract of the herb) as the intervention group or placebo as the control group for two months. Results. Comparing changes in study markers following interventions showed improvement in liver enzymes. The levels of triglycerides and cholesterol were significantly reduced in the group treated with Cynara scolymus when compared to placebo group. To compare the role of Cynara scolymus use with placebo in changes in study parameters, multivariate linear regression models were employed indicating higher improvement in liver enzymes and also lipid profile particularly triglycerides and total cholesterol following administration of Cynara scolymus in comparison with placebo use. Conclusion. This study sheds light on the potential hepatoprotective activity and hypolipidemic effect of Cynara scolymus in management of NASH. This clinical trial is registered in the IRCT, Iranian Registry of Clinical Trials, by number IRCT2014070218321N1.

  6. Free creatine available to the creatine phosphate energy shuttle in isolated rat atria

    SciTech Connect

    Savabi, F. )

    1988-10-01

    To measure the actual percentage of intracellular free creatine participating in the process of energy transport, the incorporation of (1-{sup 14}C)creatine into the free creatine and phosphocreatine (PCr) pools in spontaneously beating isolated rat atria, under various conditions, was examined. The atria were subjected to three consecutive periods, control, anoxia, and postanoxic recover, in medium containing tracers of (1-{sup 14}C)creatine. The tissue content and specific activity of creatine and PCr were determined at the end of each period. The higher specific activity found for tissue PCr (1.87 times) than creatine, independent of the percentage of total intracellular creatine that was present as free creatine, provides evidence for the existence of two separate pools of free creatine. Analysis of the data shows that in the normal oxygenated state {approx} 9% of the total intracellular creatine is actually free to participate in the process of energy transport (shuttle pool). About 36% of the total creatine is bound to unknown intracellular components and the rest exists as PCr. The creatine that was taken up and the creatine that was released from the breakdown of PCr have much greater access to the site of phosphorylation than the rest of the intracellular creatine. A sharp increase in the specific activity of residual PCr on prolongation of anoxic time was also observed. This provides evidence for a nonhomogeneous pool of PCr, for the most recently formed (radioactive) PCr appeared to be hydrolyzed last.

  7. Augmentation of Creatine in the Heart.

    PubMed

    Zervou, Sevasti; Whittington, Hannah J; Russell, Angela J; Lygate, Craig A

    2016-01-01

    Creatine is a principle component of the creatine kinase (CK) phosphagen system common to all vertebrates. It is found in excitable cells, such as cardiomyocytes, where it plays an important role in the buffering and transport of chemical energy to ensure that supply meets the dynamic demands of the heart. Multiple components of the CK system, including intracellular creatine levels, are reduced in heart failure, while ischaemia and hypoxia represent acute crises of energy provision. Elevation of myocardial creatine levels has therefore been suggested as potentially beneficial, however, achieving this goal is not trivial. This mini-review outlines the evidence in support of creatine elevation and critically examines the pharmacological approaches that are currently available. In particular, dietary creatine-supplementation does not sufficiently elevate creatine levels in the heart due to subsequent down-regulation of the plasma membrane creatine transporter (CrT). Attempts to increase passive diffusion and bypass the CrT, e.g. via creatine esters, have yet to be tested in the heart. However, studies in mice with genetic overexpression of the CrT demonstrate proof-of-principle that elevated creatine protects the heart from ischaemia-reperfusion injury. This suggests activation of the CrT as a major unmet pharmacological target. However, translation of this finding to the clinic will require a greater understanding of CrT regulation in health and disease and the development of small molecule activators.

  8. Augmentation of Creatine in the Heart

    PubMed Central

    Zervou, Sevasti; Whittington, Hannah J.; Russell, Angela J.; Lygate, Craig A.

    2016-01-01

    Creatine is a principle component of the creatine kinase (CK) phosphagen system common to all vertebrates. It is found in excitable cells, such as cardiomyocytes, where it plays an important role in the buffering and transport of chemical energy to ensure that supply meets the dynamic demands of the heart. Multiple components of the CK system, including intracellular creatine levels, are reduced in heart failure, while ischaemia and hypoxia represent acute crises of energy provision. Elevation of myocardial creatine levels has therefore been suggested as potentially beneficial, however, achieving this goal is not trivial. This mini-review outlines the evidence in support of creatine elevation and critically examines the pharmacological approaches that are currently available. In particular, dietary creatine-supplementation does not sufficiently elevate creatine levels in the heart due to subsequent down-regulation of the plasma membrane creatine transporter (CrT). Attempts to increase passive diffusion and bypass the CrT, e.g. via creatine esters, have yet to be tested in the heart. However, studies in mice with genetic overexpression of the CrT demonstrate proof-of-principle that elevated creatine protects the heart from ischaemia-reperfusion injury. This suggests activation of the CrT as a major unmet pharmacological target. However, translation of this finding to the clinic will require a greater understanding of CrT regulation in health and disease and the development of small molecule activators. PMID:26202199

  9. Clinicopathological features of choledocholithiasis patients with high aminotransferase levels without cholangitis

    PubMed Central

    Huh, Cheal Wung; Jang, Sung Ill; Lim, Beom Jin; Kim, Hee Wook; Kim, Jae Keun; Park, Jun Sung; Kim, Ja Kyung; Lee, Se Joon; Lee, Dong Ki

    2016-01-01

    Abstract Common bile duct (CBD) stones are generally associated with greater elevations of alkaline phosphatase and gamma-glutamyl transpeptidase levels than aspartate aminotransferase and alanine aminotransferase levels. However, some patients with CBD stones show markedly increased aminotransferase levels, sometimes leading to the misdiagnosis of liver disease. Therefore, the aim of this study was to investigate the clinicopathologic features of patients with CBD stones and high aminotransferase levels. This prospective cohort study included 882 patients diagnosed with CBD stones using endoscopic retrograde cholangiopancreatography (ERCP). Among these patients, 38 (4.3%) exhibited aminotransferase levels above 400 IU/L without cholangitis (gallstone hepatitis [GSH] group), and 116 (13.2%) exhibited normal aminotransferase levels (control group). We compared groups in terms of clinical features, laboratory test results, radiologic images, and ERCP findings such as CBD diameter, CBD stone diameter and number, and periampullary diverticulum. Liver biopsy was performed for patients in the GSH group. GSH patients were younger and more likely to have gallbladder stones than control patients, implying a higher incidence of gallbladder stone migration. Also, GSH patients experienced more severe, short-lasting abdominal pain. ERCP showed narrower CBDs in GSH patients than in control patients. Histological analysis of liver tissue from GSH patients showed no abnormalities except for mild inflammation. Compared with control patients, GSH patients were younger and showed more severe, short-lasting abdominal pain, which could be due to a sudden increase of CBD pressure resulting from the migration of gallstones through narrower CBDs. These clinical features could be helpful not only for the differential diagnosis of liver disease but also for investigating the underlying mechanisms of liver damage in obstructive jaundice. Moreover, we propose a new definition of

  10. Cognitive effects of creatine ethyl ester supplementation.

    PubMed

    Ling, Jonathan; Kritikos, Minos; Tiplady, Brian

    2009-12-01

    Supplementation with creatine-based substances as a means of enhancing athletic performance has become widespread. Until recently, however, the effects of creatine supplementation on cognitive performance has been given little attention. This study used a new form of creatine--creatine ethyl ester--to investigate whether supplementation would improve performance in five cognitive tasks, using a double-blind, placebo-controlled study. Creatine dosing led to an improvement over the placebo condition on several measures. Although creatine seems to facilitate cognition on some tasks, these results require replication using objective measures of compliance. The improvement is discussed in the context of research examining the influence of brain energy capacity on cognitive performance.

  11. 21 CFR 862.1215 - Creatine phosphokinase/creatine kinase or isoenzymes test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1215 Creatine phosphokinase/creatine kinase or isoenzymes test...

  12. 21 CFR 862.1215 - Creatine phosphokinase/creatine kinase or isoenzymes test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1215 Creatine phosphokinase/creatine kinase or isoenzymes test...

  13. 21 CFR 862.1215 - Creatine phosphokinase/creatine kinase or isoenzymes test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1215 Creatine phosphokinase/creatine kinase or isoenzymes test...

  14. 21 CFR 862.1215 - Creatine phosphokinase/creatine kinase or isoenzymes test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1215 Creatine phosphokinase/creatine kinase or isoenzymes test...

  15. 21 CFR 862.1215 - Creatine phosphokinase/creatine kinase or isoenzymes test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1215 Creatine phosphokinase/creatine kinase or isoenzymes test...

  16. Structural studies of Pseudomonas and Chromobacterium ω-aminotransferases provide insights into their differing substrate specificity

    SciTech Connect

    Sayer, Christopher; Isupov, Michail N.; Westlake, Aaron; Littlechild, Jennifer A.

    2013-04-01

    The X-ray structures of two ω-aminotransferases from P. aeruginosa and C. violaceum in complex with an inhibitor offer the first detailed insight into the structural basis of the substrate specificity of these industrially important enzymes. The crystal structures and inhibitor complexes of two industrially important ω-aminotransferase enzymes from Pseudomonas aeruginosa and Chromobacterium violaceum have been determined in order to understand the differences in their substrate specificity. The two enzymes share 30% sequence identity and use the same amino acceptor, pyruvate; however, the Pseudomonas enzyme shows activity towards the amino donor β-alanine, whilst the Chromobacterium enzyme does not. Both enzymes show activity towards S-α-methylbenzylamine (MBA), with the Chromobacterium enzyme having a broader substrate range. The crystal structure of the P. aeruginosa enzyme has been solved in the holo form and with the inhibitor gabaculine bound. The C. violaceum enzyme has been solved in the apo and holo forms and with gabaculine bound. The structures of the holo forms of both enzymes are quite similar. There is little conformational difference observed between the inhibitor complex and the holoenzyme for the P. aeruginosa aminotransferase. In comparison, the crystal structure of the C. violaceum gabaculine complex shows significant structural rearrangements from the structures of both the apo and holo forms of the enzyme. It appears that the different rigidity of the protein scaffold contributes to the substrate specificity observed for the two ω-aminotransferases.

  17. Creatine supplementation influences substrate utilization at rest.

    PubMed

    Huso, M Erik; Hampl, Jeffrey S; Johnston, Carol S; Swan, Pamela D

    2002-12-01

    The influence of creatine supplementation on substrate utilization during rest was investigated using a double-blind crossover design. Ten active men participated in 12 wk of weight training and were given creatine and placebo (20 g/day for 4 days, then 2 g/day for 17 days) in two trials separated by a 4-wk washout. Body composition, substrate utilization, and strength were assessed after weeks 2, 5, 9, and 12. Maximal isometric contraction [1 repetition maximum (RM)] leg press increased significantly (P < 0.05) after both treatments, but 1-RM bench press was increased (33 +/- 8 kg, P < 0.05) only after creatine. Total body mass increased (1.6 +/- 0.5 kg, P < 0.05) after creatine but not after placebo. Significant (P < 0.05) increases in fat-free mass were found after creatine and placebo supplementation (1.9 +/- 0.8 and 2.2 +/- 0.7 kg, respectively). Fat mass did not change significantly with creatine but decreased after the placebo trial (-2.4 +/- 0.8 kg, P < 0.05). Carbohydrate oxidation was increased by creatine (8.9 +/- 4.0%, P < 0.05), whereas there was a trend for increased respiratory exchange ratio after creatine supplementation (0.03 +/- 0.01, P = 0.07). Changes in substrate oxidation may influence the inhibition of fat mass loss associated with creatine after weight training.

  18. Salivary lactate dehydrogenase and aminotransferases in diabetic patients

    PubMed Central

    Malicka, Barbara; Skoskiewicz-Malinowska, Katarzyna; Kaczmarek, Urszula

    2016-01-01

    Abstract Diabetes mellitus (DM) is a group of metabolic diseases resulting from impaired insulin secretion and/or action. DM is characterized by hyperglycemia that can lead to the dysfunction or damage of organs, including the salivary glands. The aim of this study was to compare the levels of salivary lactate dehydrogenase (LDH), aspartate aminotransferase (AST), and alanine aminotransferase (ALT) in diabetic patients. The study was approved by the Bioethics Committee of Wroclaw Medical University (Poland). The study comprised 90 adults of both sexes, aged 21 to 57 years. The patients were divided into 3 groups: type 1 diabetics (D1), type 2 diabetics (D2), and a healthy control group (C). Each group consisted of 30 age- and sex-matched subjects. Total protein (P, by Lowry method), LDH, AST, ALT (with Alpha Diagnostics kits), and salivary flow rate were measured in unstimulated mixed saliva. The level of glycosylated hemoglobin (HbA1c) was measured with DCA 2000 Reagent Kit. The obtained data were analyzed using the Mann–Whitney U test and the Spearman rank at a significance level of P < 0.05 with the use of STATISTICA 9.0 software. In comparison with C, D1 presented a significantly higher activity of LDH (P < 0.001), AST (P < 0.001), and ALT (P < 0.01), whereas D2 indicated higher levels of LDH (P < 0.001) and ALT (P < 0.05) compared with C. Comparing D1 to D2, approximately 3-fold higher activity of AST (P < 0.01) and approximately 4.5-fold higher activity of ALT (P < 0.01) was observed. Higher levels of salivary LDH, AST, and ALT in D1 compared with D2 and C confirm that salivary glands of D1 might be attributed to autoimmunological damage associated with the pathomechanism of DM. PMID:27893660

  19. The Relationships between Respiratory Virus Infection and Aminotransferase in Children

    PubMed Central

    Oh, Jun Suk; Choi, Jun Sik; Lee, Young Hyuk; Ko, Kyung Og; Lim, Jae Woo; Cheon, Eun Jung; Lee, Gyung Min

    2016-01-01

    Purpose We sought to examine the relationship between the clinical manifestations of nonspecific reactive hepatitis and respiratory virus infection in pediatric patients. Methods Patients admitted to the pediatric unit of Konyang University Hospital for lower respiratory tract disease between January 1, 2014 and December 31, 2014 and who underwent reverse transcriptase polymerase chain reaction tests were examined. The patients were divided into those with increased levels of alanine aminotransferase (ALT) or aspartate aminotransferase (AST) and those with normal ALT or AST levels. Further, patients with increased ALT and AST levels were individually compared with patients in the normal group, and the blood test results were compared according to the type of respiratory virus. Results Patients with increased ALT or AST levels had one more day of hospital stay, on average, compared with patients in the normal group (5.3±3.1 days vs. 4.4±3.0 days, p=0.019). Patients in the increased ALT level group were younger and had a longer mean hospital stay, compared with patients in the normal group (p=0.022 and 0.003, respectively). The incidences of increased ALT or AST were the highest in adenovirus infections (6/24, 25.0%), followed by enterovirus (2/11, 18.2%) and respiratory syncytial virus A (21/131, 16.0%) infections. Conclusion Nonspecific reactive hepatitis is more common among patients with adenovirus, enterovirus and respiratory syncytial virus infection, as well as among those infected at a younger age. Compared with AST levels, ALT levels are better indicators of the severity of nonspecific reactive hepatitis. PMID:28090469

  20. Health implications of creatine: can oral creatine supplementation protect against neurological and atherosclerotic disease?

    PubMed

    Wyss, Markus; Schulze, Andreas

    2002-01-01

    Major achievements made over the last several years have highlighted the important roles of creatine and the creatine kinase reaction in health and disease. Inborn errors of metabolism have been identified in the three main steps involved in creatine metabolism: arginine:glycine amidinotransferase (AGAT), S-adenosyl-L-methionine:N-guanidinoacetate methyltransferase (GAMT), and the creatine transporter. All these diseases are characterized by a lack of creatine and phosphorylcreatine in the brain, and by (severe) mental retardation. Similarly, knockout mice lacking the brain cytosolic and mitochondrial isoenzymes of creatine kinase displayed a slightly increased creatine concentration, but no phosphorylcreatine in the brain. These mice revealed decreased weight gain and reduced life expectancy, disturbed fat metabolism, behavioral abnormalities and impaired learning capacity. Oral creatine supplementation improved the clinical symptoms in both AGAT and GAMT deficiency, but not in creatine transporter deficiency. In addition, creatine supplementation displayed neuroprotective effects in several animal models of neurological disease, such as Huntington's disease, Parkinson's disease, or amyotrophic lateral sclerosis. All these findings pinpoint to a close correlation between the functional capacity of the creatine kinase/phosphorylcreatine/creatine system and proper brain function. They also offer a starting-point for novel means of delaying neurodegenerative disease, and/or for strengthening memory function and intellectual capabilities.Finally, creatine biosynthesis has been postulated as a major effector of homocysteine concentration in the plasma, which has been identified as an independent graded risk factor for atherosclerotic disease. By decreasing homocysteine production, oral creatine supplementation may, thus, also lower the risk for developing, e.g., coronary heart disease or cerebrovascular disease. Although compelling, these results require further

  1. X-linked creatine transporter deficiency: clinical aspects and pathophysiology.

    PubMed

    van de Kamp, Jiddeke M; Mancini, Grazia M; Salomons, Gajja S

    2014-09-01

    Creatine transporter deficiency was discovered in 2001 as an X-linked cause of intellectual disability characterized by cerebral creatine deficiency. This review describes the current knowledge regarding creatine metabolism, the creatine transporter and the clinical aspects of creatine transporter deficiency. The condition mainly affects the brain while other creatine requiring organs, such as the muscles, are relatively spared. Recent studies have provided strong evidence that creatine synthesis also occurs in the brain, leading to the intriguing question of why cerebral creatine is deficient in creatine transporter deficiency. The possible mechanisms explaining the cerebral creatine deficiency are discussed. The creatine transporter knockout mouse provides a good model to study the disease. Over the past years several treatment options have been explored but no treatment has been proven effective. Understanding the pathogenesis of creatine transporter deficiency is of paramount importance in the development of an effective treatment.

  2. Creatine supplementation: exploring the role of the creatine kinase/phosphocreatine system in human muscle.

    PubMed

    Hespel, P; Eijnde, B O; Derave, W; Richter, E A

    2001-01-01

    The effect of oral creatine supplementation on high-intensity exercise performance has been extensively studied over the past ten years and its ergogenic potential in young healthy subjects is now well documented. Recently, research has shifted from performance evaluation towards elucidating the mechanisms underlying enhanced muscle functional capacity after creatine supplementation. In this review, we attempt to summarise recent advances in the understanding of potential mechanisms of action of creatine supplementation at the level of skeletal muscle cells. By increasing intracellular creatine content, oral creatine ingestion conceivably stimulates operation of the creatine kinase (CK)/phosphocreatine (PCr) system, which in turn facilitates muscle relaxation. Furthermore, evidence is accumulating to suggest that creatine supplementation can beneficially impact on muscle protein and glycogen synthesis. Thus, muscle hypertrophy and glycogen supercompensation are candidate factors to explain the ergogenic potential of creatine ingestion. Additional issues discussed in this review are the fibre-type specificity of muscle creatine metabolism, the identification of responders versus non-responders to creatine intake, and the scientific background concerning potential side effects of creatine supplementation.

  3. Creatine biosynthesis and transport in health and disease.

    PubMed

    Joncquel-Chevalier Curt, Marie; Voicu, Pia-Manuela; Fontaine, Monique; Dessein, Anne-Frédérique; Porchet, Nicole; Mention-Mulliez, Karine; Dobbelaere, Dries; Soto-Ares, Gustavo; Cheillan, David; Vamecq, Joseph

    2015-12-01

    Creatine is physiologically provided equally by diet and by endogenous synthesis from arginine and glycine with successive involvements of arginine glycine amidinotransferase [AGAT] and guanidinoacetate methyl transferase [GAMT]. A specific plasma membrane transporter, creatine transporter [CRTR] (SLC6A8), further enables cells to incorporate creatine and through uptake of its precursor, guanidinoacetate, also directly contributes to creatine biosynthesis. Breakthrough in the role of creatine has arisen from studies on creatine deficiency disorders. Primary creatine disorders are inherited as autosomal recessive (mutations affecting GATM [for glycine-amidinotransferase, mitochondrial]) and GAMT genes) or X-linked (SLC6A8 gene) traits. They have highlighted the role of creatine in brain functions altered in patients (global developmental delay, intellectual disability, behavioral disorders). Creatine modulates GABAergic and glutamatergic cerebral pathways, presynaptic CRTR (SLC6A8) ensuring re-uptake of synaptic creatine. Secondary creatine disorders, addressing other genes, have stressed the extraordinary imbrication of creatine metabolism with many other cellular pathways. This high dependence on multiple pathways supports creatine as a cellular sensor, to cell methylation and energy status. Creatine biosynthesis consumes 40% of methyl groups produced as S-adenosylmethionine, and creatine uptake is controlled by AMP activated protein kinase, a ubiquitous sensor of energy depletion. Today, creatine is considered as a potential sensor of cell methylation and energy status, a neurotransmitter influencing key (GABAergic and glutamatergic) CNS neurotransmission, therapeutic agent with anaplerotic properties (towards creatine kinases [creatine-creatine phosphate cycle] and creatine neurotransmission), energetic and antioxidant compound (benefits in degenerative diseases through protection against energy depletion and oxidant species) with osmolyte behavior (retention of

  4. Creatine and guanidinoacetate content of human milk and infant formulas: implications for creatine deficiency syndromes and amino acid metabolism.

    PubMed

    Edison, Erica E; Brosnan, Margaret E; Aziz, Khalid; Brosnan, John T

    2013-09-28

    Creatine is essential for normal neural development; children with inborn errors of creatine synthesis or transport exhibit neurological symptoms such as mental retardation, speech delay and epilepsy. Creatine accretion may occur through dietary intake or de novo creatine synthesis. The objective of the present study was to determine how much creatine an infant must synthesise de novo. We have calculated how much creatine an infant needs to account for urinary creatinine excretion (creatine's breakdown product) and new muscle lay-down. To measure an infant's dietary creatine intake, we measured creatine in mother's milk and in various commercially available infant formulas. Knowing the amount of milk/formula ingested, we calculated the amount of creatine ingested. We have found that a breast-fed infant receives about 9 % of the creatine needed in the diet and that infants fed cows' milk-based formula receive up to 36 % of the creatine needed. However, infants fed a soya-based infant formula receive negligible dietary creatine and must rely solely on de novo creatine synthesis. This is the first time that it has been shown that neonatal creatine accretion is largely due to de novo synthesis and not through dietary intake of creatine. This has important implications both for infants suffering from creatine deficiency syndromes and for neonatal amino acid metabolism.

  5. Weaning Induced Hepatic Oxidative Stress, Apoptosis, and Aminotransferases through MAPK Signaling Pathways in Piglets

    PubMed Central

    Luo, Zhen; Zhu, Wei; Guo, Qi; Luo, Wenli; Zhang, Jing; Xu, Weina

    2016-01-01

    This study investigated the effects of weaning on the hepatic redox status, apoptosis, function, and the mitogen-activated protein kinase (MAPK) signaling pathways during the first week after weaning in piglets. A total of 12 litters of piglets were weaned at d 21 and divided into the weaning group (WG) and the control group (CG). Six piglets from each group were slaughtered at d 0 (d 20, referred to weaning), d 1, d 4, and d 7 after weaning. Results showed that weaning significantly increased the concentrations of hepatic free radicals H2O2 and NO, malondialdehyde (MDA), and 8-hydroxy-2′-deoxyguanosine (8-OHdG), while significantly decreasing the inhibitory hydroxyl ability (IHA) and glutathione peroxidase (GSH-Px), and altered the level of superoxide dismutase (SOD). The apoptosis results showed that weaning increased the concentrations of caspase-3, caspase-8, caspase-9 and the ratio of Bax/Bcl-2. In addition, aspartate aminotransferase transaminase (AST) and alanine aminotransferase (ALT) in liver homogenates increased after weaning. The phosphorylated JNK and ERK1/2 increased, while the activated p38 initially decreased and then increased. Our results suggested that weaning increased the hepatic oxidative stress and aminotransferases and initiated apoptosis, which may be related to the activated MAPK pathways in postweaning piglets. PMID:27807471

  6. Structural studies of Pseudomonas and Chromobacterium ω-aminotransferases provide insights into their differing substrate specificity.

    PubMed

    Sayer, Christopher; Isupov, Michail N; Westlake, Aaron; Littlechild, Jennifer A

    2013-04-01

    The crystal structures and inhibitor complexes of two industrially important ω-aminotransferase enzymes from Pseudomonas aeruginosa and Chromobacterium violaceum have been determined in order to understand the differences in their substrate specificity. The two enzymes share 30% sequence identity and use the same amino acceptor, pyruvate; however, the Pseudomonas enzyme shows activity towards the amino donor β-alanine, whilst the Chromobacterium enzyme does not. Both enzymes show activity towards S-α-methylbenzylamine (MBA), with the Chromobacterium enzyme having a broader substrate range. The crystal structure of the P. aeruginosa enzyme has been solved in the holo form and with the inhibitor gabaculine bound. The C. violaceum enzyme has been solved in the apo and holo forms and with gabaculine bound. The structures of the holo forms of both enzymes are quite similar. There is little conformational difference observed between the inhibitor complex and the holoenzyme for the P. aeruginosa aminotransferase. In comparison, the crystal structure of the C. violaceum gabaculine complex shows significant structural rearrangements from the structures of both the apo and holo forms of the enzyme. It appears that the different rigidity of the protein scaffold contributes to the substrate specificity observed for the two ω-aminotransferases.

  7. Structural studies of Pseudomonas and Chromobacterium ω-aminotransferases provide insights into their differing substrate specificity

    PubMed Central

    Sayer, Christopher; Isupov, Michail N.; Westlake, Aaron; Littlechild, Jennifer A.

    2013-01-01

    The crystal structures and inhibitor complexes of two industrially important ω-aminotransferase enzymes from Pseudomonas aeruginosa and Chromobacterium violaceum have been determined in order to understand the differences in their substrate specificity. The two enzymes share 30% sequence identity and use the same amino acceptor, pyruvate; however, the Pseudomonas enzyme shows activity towards the amino donor β-alanine, whilst the Chromobacterium enzyme does not. Both enzymes show activity towards S-α-methylbenzylamine (MBA), with the Chromobacterium enzyme having a broader substrate range. The crystal structure of the P. aeruginosa enzyme has been solved in the holo form and with the inhibitor gabaculine bound. The C. violaceum enzyme has been solved in the apo and holo forms and with gabaculine bound. The structures of the holo forms of both enzymes are quite similar. There is little conformational difference observed between the inhibitor complex and the holoenzyme for the P. aeruginosa aminotransferase. In comparison, the crystal structure of the C. violaceum gabaculine complex shows significant structural rearrangements from the structures of both the apo and holo forms of the enzyme. It appears that the different rigidity of the protein scaffold contributes to the substrate specificity observed for the two ω-­aminotransferases. PMID:23519665

  8. Creatine supplementation and swimming performance.

    PubMed

    Leenders, N M; Lamb, D R; Nelson, T E

    1999-09-01

    The purpose of this study was to determine if oral creatine (CR) ingestion, compared to a placebo (PL), would enable swimmers to maintain a higher swimming velocity across repeated interval sets over 2 weeks of supplementation. Fourteen female and 18 male university swimmers consumed a PL during a 2-week baseline period. Using a randomized, double-blind design, during the next 2 weeks subjects consumed either CR or PL. Swimming velocity was assessed twice weekly during 6 X 50-m swims and once weekly during 10 X 25-yd swims. There was no effect of CR on the 10 X 25-yd interval sets for men and women and no effect on the 6 X 50-m interval sets for women. In contrast, for men, CR significantly improved mean overall swimming velocity in the 6 X 50-m interval after 2 weeks of supplementation, whereas PL had no effect. Although ineffective in women, CR supplementation apparently enables men to maintain a faster mean overall swimming velocity during repeated swims each lasting about 30 s; however, CR was not effective for men in repeated swims each lasting about 10 - 15 s.

  9. Creatine and Caffeine: Considerations for Concurrent Supplementation.

    PubMed

    Trexler, Eric T; Smith-Ryan, Abbie E

    2015-12-01

    Nutritional supplementation is a common practice among athletes, with creatine and caffeine among the most commonly used ergogenic aids. Hundreds of studies have investigated the ergogenic potential of creatine supplementation, with consistent improvements in strength and power reported for exercise bouts of short duration (≤ 30 s) and high intensity. Caffeine has been shown to improve endurance exercise performance, but results are mixed in the context of strength and sprint performance. Further, there is conflicting evidence from studies comparing the ergogenic effects of coffee and caffeine anhydrous supplementation. Previous research has identified independent mechanisms by which creatine and caffeine may improve strength and sprint performance, leading to the formulation of multi-ingredient supplements containing both ingredients. Although scarce, research has suggested that caffeine ingestion may blunt the ergogenic effect of creatine. While a pharmacokinetic interaction is unlikely, authors have suggested that this effect may be explained by opposing effects on muscle relaxation time or gastrointestinal side effects from simultaneous consumption. The current review aims to evaluate the ergogenic potential of creatine and caffeine in the context of high-intensity exercise. Research directly comparing coffee and caffeine anhydrous is discussed, along with previous studies evaluating the concurrent supplementation of creatine and caffeine.

  10. Plasma Creatine Kinetics After Ingestion of Microencapsulated Creatine Monohydrate with Enhanced Stability in Aqueous Solutions.

    PubMed

    Hone, Michelle; Kent, Robert M; Scotto di Palumbo, Alessandro; Bleiel, Sinead B; De Vito, Giuseppe; Egan, Brendan

    2017-07-04

    Creatine monohydrate represents one of the largest sports supplement markets. Enhancing creatine (CRE) stability in aqueous solutions, such as with microencapsulation, represents innovation potential. Ten physically active male volunteers were randomly assigned in a double-blind design to either placebo (PLA) (3-g maltodextrin; n = 5) or microencapsulated CRE (3-g creatine monohydrate; n = 5) conditions. Experimental conditions involved ingestion of the samples in a 70-mL ready-to-drink format. CRE was delivered in a novel microencapsulation matrix material consisting entirely of hydrolyzed milk protein. Three hours after ingestion, plasma creatine concentrations were unchanged during PLA, and averaged ∼45 μM. During CRE, plasma creatine concentration peaked after 30 min at 101.6 ± 14.9 μM (p < 0.05), representing a 2.3-fold increase over PLA. Thereafter, plasma creatine concentration gradually trended downwards but remained significantly elevated (∼50% above resting levels) 3 hr after ingestion. These results demonstrate that the microencapsulated form of creatine monohydrate reported herein remains bioavailable when delivered in aqueous conditions, and has potential utility in ready-to-drink formulations for creatine supplementation.

  11. Whole Body Creatine and Protein Kinetics in Healthy Men and Women: Effects of creatine and amino acid supplementation

    PubMed Central

    Kalhan, Satish C; Gruca, Lourdes; Marczewski, Susan; Bennett, Carole; Kummitha, China

    2015-01-01

    Creatine kinetics were measured in young healthy subjects, eight males and seven females, age 20–30 years, after an overnight fast on creatine free diet. Whole body turnover of glycine and its appearance in creatine was quantified using [1-13C] glycine and the rate of protein turnover was quantified using L-ring [2H5] phenylalanine. The creatine pool size was estimated by the dilution of a bolus [C2H3] creatine. Studies were repeated following a five days supplement creatine 21g.day−1 and following supplement amino acids 14.3 g.day−1. Creatine caused a ten folds increase in the plasma concentration of creatine and a 50% decrease in the concentration of guanidinoacetic acid. Plasma amino acids profile showed a significant decrease in glycine, glutamine and taurine and a significant increase in citrulline, valine, lysine and cysteine. There was a significant decrease in the rate of appearance of glycine, suggesting a decrease in de-novo synthesis (p=0.006). The fractional and absolute rate of synthesis of creatine was significantly decreased by supplemental creatine. Amino acid supplement had no impact on any of the parameters. Creatine supplement caused a significant decrease in the rate of synthesis of creatine. This is the first detailed analysis of creatine kinetics and the effects of creatine supplement in healthy young men and women. These methods can be applied for the analysis of creatine kinetics in different physiological states. PMID:26480831

  12. A short review on creatine-creatine kinase system in relation to cancer and some experimental results on creatine as adjuvant in cancer therapy.

    PubMed

    Patra, Subrata; Ghosh, Alok; Roy, Soumya Sinha; Bera, Soumen; Das, Manju; Talukdar, Dipa; Ray, Subhankar; Wallimann, Theo; Ray, Manju

    2012-06-01

    The creatine/creatine kinase (CK) system plays a key role in cellular energy buffering and transport. In vertebrates, CK has four isoforms expressed in a tissue-specific manner. In the process of creatine biosynthesis several other important metabolites are formed. The anticancer effect of creatine had been reported in the past, and recent literature has reported low creatine content in several types of malignant cells. Furthermore, creatine can protect cardiac mitochondria from the deleterious effects of some anticancer compounds. Previous work from our laboratory showed progressive decrease of phosphocreatine, creatine and CK upon transformation of skeletal muscle into sarcoma. It was convincingly demonstrated that prominent expression of creatine-synthesizing enzymes L-arginine: glycine amidinotransferase and N-guanidinoacetate methyltransferase occurs in sarcoma, Ehrlich ascites carcinoma and sarcoma 180 cells; whereas, both these enzymes are virtually undetectable in skeletal muscle. Creatine transporter also remained unaltered in malignant cells. The anticancer effect of methylglyoxal had been known for a long time. The present work shows that this anticancer effect of methylglyoxal is significantly augmented in presence of creatine. On creatine supplementation the effect of methylglyoxal plus ascorbic acid was further augmented and there was no visible sign of tumor. Moreover, creatine and CK, which were very low in sarcoma tissue, were significantly elevated with the concomitant regression of tumor.

  13. Creatine supplementation and multiple sprint running performance.

    PubMed

    Glaister, Mark; Lockey, Richard A; Abraham, Corinne S; Staerck, Allan; Goodwin, Jon E; McInnes, Gillian

    2006-05-01

    The aim of this study was to examine the effects of short-term creatine monohydrate supplementation on multiple sprint running performance. Using a double-blind research design, 42 physically active men completed a series of 3 indoor multiple sprint running trials (15 x 30 m repeated at 35-second intervals). After the first 2 trials (familiarization and baseline), subjects were matched for fatigue score before being randomly assigned to 5 days of either creatine (4 x d(-1) x 5 g creatine monohydrate + 1 g maltodextrin) or placebo (4 x d(-1) x 6 g maltodextrin) supplementation. Sprint times were recorded via twin-beam photocells, and earlobe blood samples were drawn to evaluate posttest lactate concentrations. Relative to placebo, creatine supplementation resulted in a 0.7 kg increase in body mass (95% likely range: 0.02 to 1.3 kg) and a 0.4% reduction in body fat (95% likely range: -0.2 to 0.9%). There were no significant (p > 0.05) between-group differences in multiple sprint measures of fastest time, mean time, fatigue, or posttest blood lactate concentration. Despite widespread use as an ergogenic aid in sport, the results of this study suggest that creatine monohydrate supplementation conveys no benefit to multiple sprint running performance.

  14. Creatine metabolism and psychiatric disorders: Does creatine supplementation have therapeutic value?

    PubMed

    Allen, Patricia J

    2012-05-01

    Athletes, body builders, and military personnel use dietary creatine as an ergogenic aid to boost physical performance in sports involving short bursts of high-intensity muscle activity. Lesser known is the essential role creatine, a natural regulator of energy homeostasis, plays in brain function and development. Creatine supplementation has shown promise as a safe, effective, and tolerable adjunct to medication for the treatment of brain-related disorders linked with dysfunctional energy metabolism, such as Huntington's Disease and Parkinson's Disease. Impairments in creatine metabolism have also been implicated in the pathogenesis of psychiatric disorders, leaving clinicians, researchers and patients alike wondering if dietary creatine has therapeutic value for treating mental illness. The present review summarizes the neurobiology of the creatine-phosphocreatine circuit and its relation to psychological stress, schizophrenia, mood and anxiety disorders. While present knowledge of the role of creatine in cognitive and emotional processing is in its infancy, further research on this endogenous metabolite has the potential to advance our understanding of the biological bases of psychopathology and improve current therapeutic strategies.

  15. Creatine metabolism and psychiatric disorders: Does creatine supplementation have therapeutic value?

    PubMed Central

    Allen, Patricia J.

    2012-01-01

    Athletes, body builders, and military personnel use dietary creatine as an ergogenic aid to boost physical performance in sports involving short bursts of high-intensity muscle activity. Lesser known is the essential role creatine, a natural regulator of energy homeostasis, plays in brain function and development. Creatine supplementation has shown promise as a safe, effective, and tolerable adjunct to medication for the treatment of brain-related disorders linked with dysfunctional energy metabolism, such as Huntington’s Disease and Parkinson’s Disease. Impairments in creatine metabolism have also been implicated in the pathogenesis of psychiatric disorders, leaving clinicians, researchers and patients alike wondering if dietary creatine has therapeutic value for treating mental illness. The present review summarizes the neurobiology of the creatine-phosphocreatine circuit and its relation to psychological stress, schizophrenia, mood and anxiety disorders. While present knowledge of the role of creatine in cognitive and emotional processing is in its infancy, further research on this endogenous metabolite has the potential to advance our understanding of the biological bases of psychopathology and improve current therapeutic strategies. PMID:22465051

  16. Physicochemical characterization of creatine N-methylguanidinium salts.

    PubMed

    Gufford, Brandon T; Sriraghavan, Kamaraj; Miller, Nicholas J; Miller, Donald W; Gu, Xiaochen; Vennerstrom, Jonathan L; Robinson, Dennis H

    2010-09-01

    Creatine is widely used as a dietary supplement for body builders to enhance athletic performance. As the monohydrate, its low solubility in water and high dose lead to water retention and gastrointestinal discomfort. Hence, alternative creatine derivatives with enhanced water solubility and potential therapeutic advantages have been synthesized. As a zwitterionic compound, creatine can form salts at the N-methyl guanidinium or carboxylic acid functional groups. In this study, we determined the aqueous solubilities and partition coefficients of six N-methyl guanidinium salts of creatine compared to those of creatine monohydrate; two of these were new salts, namely, creatine mesylate and creatine hydrogen maleate. The aqueous solubilities of the salts were significantly more than that of creatine monohydrate with the hydrochloride and mesylate being 38 and 30 times more soluble, respectively. The partition coefficients of the creatine salts were very low indicating their relatively high polarity. Permeabilities of creatine pyruvate, citrate, and hydrochloride in Caco-2 monolayers were compared to that of creatine monohydrate. Aside from the creatine citrate salt form that had reduced permeability, there were no significant differences in permeability characteristics in Caco-2 monolayers. Typical of an amphoteric compound, creatine is least soluble in the pH region near the isoelectric point.

  17. Creatine supplementation. Its role in human performance.

    PubMed

    Kraemer, W J; Volek, J S

    1999-07-01

    Creatine supplementation is the most popular nutritional supplement today. Although many questions remain regarding the use and benefits of creatine supplementation, a fast-growing body of literature is starting to define both its acute and chronic effects on human and physiologic performance. The initial data indicate that this energetic boost of the phosphagen energy system also helps to enhance strength and power training. Few documented side effects have been demonstrated in the medical and scientific literature, but further investigation is still required as to long-term use (i.e., beyond several months).

  18. Comparison of effect of cafetière and filtered coffee on serum concentrations of liver aminotransferases and lipids: six month randomised controlled trial.

    PubMed Central

    Urgert, R.; Meyboom, S.; Kuilman, M.; Rexwinkel, H.; Vissers, M. N.; Klerk, M.; Katan, M. B.

    1996-01-01

    OBJECTIVE: To study the effects of prolonged intake of cafetière coffee, which is rich in the diterpenes cafestol and kahweol, on serum aminotransferase and lipid concentrations. DESIGN: Randomised parallel controlled trial. SUBJECTS: 46 healthy men and women aged 19 to 69. INTERVENTION: Consumption of five to six strong cups (0.9 litres) a day of either cafetière (22 subjects) or filtered coffee (24 subjects) for 24 weeks. MAIN OUTCOME MEASURES: Mean changes in serum aminotransferase and lipid concentrations. RESULTS: Cafetière coffee raised alanine aminotransferase concentration by up to 80% above baseline values relative to filtered coffee. After 24 weeks the rise was still 45% (9 U/l (95% confidence interval 3 to 15 U/l), P = 0.007). Alanine aminotransferase concentration exceeded the upper limit of normal in eight of the 22 subjects drinking cafetière coffee, being twice the upper limit of normal in three of them. Cafetière coffee raised low density lipoprotein cholesterol concentrations by 9-14%. After 24 weeks the rise was 0.26 mmol/l (0.04 to 0.47 mmol/l) (P = 0.03) relative to filtered coffee. Triglyceride concentrations initially rose by 26% with cafetière coffee but returned close to baseline values within six months. All increases were reversible after the intervention was stopped. CONCLUSIONS: Daily consumption of five to six cups of strong cafetière coffee affects the integrity of liver cells as suggested by small increases in serum alanine aminotransferase concentration. The effect does not subside with prolonged intake. High intakes of coffee brews rich in cafestol and kahweol may thus be responsible for unexplained increases in this enzyme activity in apparently healthy subjects. Cafetière coffee also raises low density lipoprotein cholesterol concentration and thus the risk of coronary heart disease. PMID:8956701

  19. [Conformation of aspartate aminotransferase in crystals].

    PubMed

    Borisov, V V; Borisova, S N; Sosfenov, N I; Dikson, Kh BF

    1983-01-01

    X-ray study of chicken cytosolic aspartate aminotransferase revealed conformational changes in the protein of two kinds: (1) a shift of the small domain adjacent to substrate-binding area due to interaction of the protein with two carboxyl groups of substrate and (2) a change in inclination of the coenzyme plane due to replacement of C = N bond of the coenzyme with Lys-258 by C = N bond with a substrate. An asymmetry in subunit behaviour is observed in both cases: the domain is shifted in one subunit and the coenzyme is rotated in other. Substrate-binding properties of each subunit are strictly dependent on the protein conformation in substrate-binding area.

  20. Neuroprotective effects of creatine administration against NMDA and malonate toxicity.

    PubMed

    Malcon, C; Kaddurah-Daouk, R; Beal, M F

    2000-03-31

    We examined whether creatine administration could exert neuroprotective effects against excitotoxicity mediated by N-methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and kainic acid. Oral administration of 1% creatine significantly attenuated striatal excitotoxic lesions produced by NMDA, but had no effect on lesions produced by AMPA or kainic acid. Both creatine and nicotinamide can exert significant protective effects against malonate-induced striatal lesions. We, therefore, examined whether nicotinamide could exert additive neuroprotective effects with creatine against malonate-induced lesions. Nicotinamide with creatine produced significantly better neuroprotection than creatine alone against malonate-induced lesions. Creatine can, therefore, produce significant neuroprotective effects against NMDA mediated excitotoxic lesions in vivo and the combination of nicotinamide with creatine exerts additive neuroprotective effects.

  1. Genetics Home Reference: X-linked creatine deficiency

    MedlinePlus

    ... instructions for making a protein that transports the compound creatine into cells. Creatine is needed for the ... j.ymgme.2014.05.011. Epub 2014 May 29. Review. Citation on PubMed Longo N, Ardon O, ...

  2. Whole body creatine and protein kinetics in healthy men and women: effects of creatine and amino acid supplementation.

    PubMed

    Kalhan, Satish C; Gruca, Lourdes; Marczewski, Susan; Bennett, Carole; Kummitha, China

    2016-03-01

    Creatine kinetics were measured in young healthy subjects, eight males and seven females, age 20-30 years, after an overnight fast on creatine-free diet. Whole body turnover of glycine and its appearance in creatine was quantified using [1-(13)C] glycine and the rate of protein turnover was quantified using L-ring [(2)H5] phenylalanine. The creatine pool size was estimated by the dilution of a bolus [C(2)H3] creatine. Studies were repeated following a five days supplement creatine 21 g.day(-1) and following supplement amino acids 14.3 g day(-1). Creatine caused a ten-fold increase in the plasma concentration of creatine and a 50 % decrease in the concentration of guanidinoacetic acid. Plasma amino acids profile showed a significant decrease in glycine, glutamine, and taurine and a significant increase in citrulline, valine, lysine, and cysteine. There was a significant decrease in the rate of appearance of glycine, suggesting a decrease in de-novo synthesis (p = 0.006). The fractional and absolute rate of synthesis of creatine was significantly decreased by supplemental creatine. Amino acid supplement had no impact on any of the parameters. This is the first detailed analysis of creatine kinetics and the effects of creatine supplement in healthy young men and women. These methods can be applied for the analysis of creatine kinetics in different physiological states.

  3. Creatine and the Male Adolescent Athlete

    ERIC Educational Resources Information Center

    Schumaker, Shauna; Eyers, Christina; Cappaert, Thomas

    2012-01-01

    As the level of competition in youth sports increases, so does athletes' vulnerability to experimenting with performance-enhancing aids (PEAs) at alarmingly young ages. One of the more commonly used PEAs is a supplement called creatine, which has the ability to generate muscular energy, allowing athletes to train at higher intensities for longer…

  4. Does brain creatine content rely on exogenous creatine in healthy youth? A proof-of-principle study.

    PubMed

    Merege-Filho, Carlos Alberto Abujabra; Otaduy, Maria Concepción Garcia; de Sá-Pinto, Ana Lúcia; de Oliveira, Maira Okada; de Souza Gonçalves, Lívia; Hayashi, Ana Paula Tanaka; Roschel, Hamilton; Pereira, Rosa Maria Rodrigues; Silva, Clovis Artur; Brucki, Sonia Maria Dozzi; da Costa Leite, Claudia; Gualano, Bruno

    2017-02-01

    It has been hypothesized that dietary creatine could influence cognitive performance by increasing brain creatine in developing individuals. This double-blind, randomized, placebo-controlled, proof-of-principle study aimed to investigate the effects of creatine supplementation on cognitive function and brain creatine content in healthy youth. The sample comprised 67 healthy participants aged 10 to 12 years. The participants were given creatine or placebo supplementation for 7 days. At baseline and after the intervention, participants undertook a battery of cognitive tests. In a random subsample of participants, brain creatine content was also assessed in the regions of left dorsolateral prefrontal cortex, left hippocampus, and occipital lobe by proton magnetic resonance spectroscopy (1H-MRS) technique. The scores obtained from verbal learning and executive functions tests did not significantly differ between groups at baseline or after the intervention (all p > 0.05). Creatine content was not significantly different between groups in left dorsolateral prefrontal cortex, left hippocampus, and occipital lobe (all p > 0.05). In conclusion, a 7-day creatine supplementation protocol did not elicit improvements in brain creatine content or cognitive performance in healthy youth, suggesting that this population mainly relies on brain creatine synthesis rather than exogenous creatine intake to maintain brain creatine homeostasis.

  5. The past and present of serum aminotransferases and the future of liver injury biomarkers

    PubMed Central

    McGill, Mitchell R.

    2016-01-01

    Laboratory testing is important in the diagnosis and monitoring of liver injury and disease. Current liver tests include plasma markers of injury (e.g. aminotransferases, γ-glutamyl transferase, and alkaline phosphatase), markers of function (e.g. prothrombin time, bilirubin), viral hepatitis serologies, and markers of proliferation (e.g. α-fetoprotein). Among the injury markers, the alanine and aspartate aminotransferases (ALT and AST, respectively) are the most commonly used. However, interpretation of ALT and AST plasma levels can be complicated. Furthermore, both have poor prognostic utility in acute liver injury and liver failure. New biomarkers of liver injury are rapidly being developed, and the US Food and Drug Administration the European Medicines Agency have recently expressed support for use of some of these biomarkers in drug trials. The purpose of this paper is to review the history of liver biomarkers, to summarize mechanisms and interpretation of ALT and AST elevation in plasma in liver injury (particularly acute liver injury), and to discuss emerging liver injury biomarkers that may complement or even replace ALT and AST in the future. PMID:28337112

  6. Preoperative Aspartate Aminotransferase to White Blood Cell Count Ratio Predicting Postoperative Outcomes of Hepatocellular Carcinoma.

    PubMed

    Liao, Weijia; Wang, Yongqin; Liao, Yan; He, Songqing; Jin, Junfei

    2016-04-01

    Effective biomarkers for predicting prognosis of hepatocellular carcinoma (HCC) patients after hepatectomy is urgently needed. The purpose of this study is to evaluate the value of the preoperative peripheral aspartate aminotransferase to white blood cell count ratio (AWR) for the prognostication of patients with HCC.Clinical data of 396 HCC patients who underwent radical hepatectomy were retrospectively analyzed. The patients were divided into the low-AWR group (AWR ≤5.2) and the high-AWR group (AWR >5.2); univariate analysis, Kaplan-Meier method analysis, and the multivariate analysis by Cox regression were conducted, respectively.The results showed that AWR was associated with alpha-fetoprotein (AFP), tumor size, Barcelona clinic liver cancer (BCLC) stage, portal vein tumor thrombus (PVTT), and alanine aminotransferase (ALT) in HCC. AWR > 5.2, AFP > 100 ng/mL, size of tumor >6 cm, number of multiple tumors, B-C of BCLC stage, PVTT, and distant metastasis were predictors of poorer disease-free survival (DFS) and overall survival (OS). Except for recurrence, which was an independent predictor for OS only, AWR >5.2, size of tumor >6 cm, and PVTT were independent predictors of both DFS and OS.We concluded that preoperative AWR > 5.2 was an adverse predictor of DFS and OS in HCC after hepatectomy, AWR might be a novel prognostic biomarker in HCC after curative resection.

  7. Elevated levels of serum creatine kinase induced by hyponatraemia.

    PubMed

    Goldenberg, I; Jonas, M; Thaler, M; Grossman, E

    1997-08-01

    Elevated serum creatine kinase levels are one of the major criteria for the diagnosis of myocardial injury. Noncardiac causes such as muscular and brain damage may also be associated with elevated serum creatine kinase levels. Hyponatremia may induce increased serum creatine kinase in association with rhabdomyolysis or with hypothyroidism. A patient is described where three episodes of hyponatraemia not associated with rhabdomyolysis or hypothyroidism induced transient elevations of serum creatine kinase levels. The association between hyponatraemia and elevated creatine kinase levels should be emphasized to prevent erroneous diagnosis of myocardial injury.

  8. AMPK and substrate availability regulate creatine transport in cultured cardiomyocytes.

    PubMed

    Darrabie, Marcus D; Arciniegas, Antonio Jose Luis; Mishra, Rajashree; Bowles, Dawn E; Jacobs, Danny O; Santacruz, Lucia

    2011-05-01

    Profound alterations in myocellular creatine and phosphocreatine levels are observed during human heart failure. To maintain its intracellular creatine stores, cardiomyocytes depend upon a cell membrane creatine transporter whose regulation is not clearly understood. Creatine transport capacity in the intact heart is modulated by substrate availability, and it is reduced in the failing myocardium, likely adding to the energy imbalance that characterizes heart failure. AMPK, a key regulator of cellular energy homeostasis, acts by switching off energy-consuming pathways in favor of processes that generate energy. Our objective was to determine the effects of substrate availability and AMPK activation on creatine transport in cardiomyocytes. We studied creatine transport in rat neonatal cardiomyocytes and HL-1 cardiac cells expressing the human creatine transporter cultured in the presence of varying creatine concentrations and the AMPK activator 5-aminoimidazole-4-carboxamide-1-β-d-ribonucleoside (AICAR). Transport was enhanced in cardiomyocytes following incubation in creatine-depleted medium or AICAR. The changes in transport were due to alterations in V(max) that correlated with changes in total and cell surface creatine transporter protein content. Our results suggest a positive role for AMPK in creatine transport modulation for cardiomyocytes in culture.

  9. Crystal Structure of Human Kynurenine Aminotransferase ll*

    SciTech Connect

    Han,Q.; Robinson, H.; Li, J.

    2008-01-01

    Human kynurenine aminotransferase II (hKAT-II) efficiently catalyzes the transamination of knunrenine to kynurenic acid (KYNA). KYNA is the only known endogenous antagonist of N-methyl-d-aspartate (NMDA) receptors and is also an antagonist of 7-nicotinic acetylcholine receptors. Abnormal concentrations of brain KYNA have been implicated in the pathogenesis and development of several neurological and psychiatric diseases in humans. Consequently, enzymes involved in the production of brain KYNA have been considered potential regulatory targets. In this article, we report a 2.16 Angstroms crystal structure of hKAT-II and a 1.95 Angstroms structure of its complex with kynurenine. The protein architecture of hKAT-II reveals that it belongs to the fold-type I pyridoxal 5-phosphate (PLP)-dependent enzymes. In comparison with all subclasses of fold-type I-PLP-dependent enzymes, we propose that hKAT-II represents a novel subclass in the fold-type I enzymes because of the unique folding of its first 65 N-terminal residues. This study provides a molecular basis for future effort in maintaining physiological concentrations of KYNA through molecular and biochemical regulation of hKAT-II.

  10. Guanidinoacetic acid versus creatine for improved brain and muscle creatine levels: a superiority pilot trial in healthy men.

    PubMed

    Ostojic, Sergej M; Ostojic, Jelena; Drid, Patrik; Vranes, Milan

    2016-09-01

    In this randomized, double-blind, crossover trial, we evaluated whether 4-week supplementation with guanidinoacetic acid (GAA) is superior to creatine in facilitating creatine levels in healthy men (n = 5). GAA (3.0 g/day) resulted in a more powerful rise (up to 16.2%) in tissue creatine levels in vastus medialis muscle, middle-cerebellar peduncle, and paracentral grey matter, as compared with creatine (P < 0.05). These results indicate that GAA as a preferred alternative to creatine for improved bioenergetics in energy-demanding tissues.

  11. [Creatine: the nutritional supplement for exercise - current concepts].

    PubMed

    Mendes, Renata Rebello; Tirapegui, Julio

    2002-06-01

    Creatine, a natural nutrient found in animal foods, is alleged to be an effective nutritional ergogenic aid to enhance sport or exercise performance. It may be formed in kidney and liver from arginina and glicina. Creatine may be delivered to the muscle, where it may combine readily with phosphate to form creatine phosphate, a high-energy phosphagen in the ATP-CP system, and is stored. The ATP-CP energy system is important for rapid energy production, such as in speed and power events. Approximately 120 g of creatine is found in a 70 kg male, 95% in the skeletal muscle. Total creatine exists in muscle as both free creatine (40%) and phosphocreatine (60%). It is only recently that a concerted effort has been undertaken to investigate its potential ergogenic effect relative to sport or exercise performance. It does appear that oral creatine monohydrate may increase muscle total creatine, including both free and phosphocreatine. Many, but not all studies suggest that creatine supplementation may enhance performance in high intensity, short-term exercise task that are dependent primarily on the ATP-CP energy system, particularly on laboratory test involving repeated exercise bouts with limited recovery time between repetitions. Short-term creatine supplementation appears to increase body mass, although the initial increase is most likely water associated with the osmotic effect of increased intramuscular total creatine. Chronic creatine supplementation in conjunction with physical training involving resistance exercise may increase muscle mass. However, confirmatory research data are needed. Creatine supplementation up to 8 weeks, with high doses, has not been associated with major health risks; with low doses, it was demonstrated that in 5 years period supplementation, there are no adverse effects. The decision to use creatine as a mean to enhance sport performance is left to the description to the individual athlete.

  12. Conversion of cysteine to 3-mercaptopyruvic acid by bacterial aminotransferases.

    PubMed

    Andreeßen, Christina; Gerlt, Vanessa; Steinbüchel, Alexander

    2017-04-01

    3-Mercaptopyruvate (3MPy), a structural analog of 3-mercaptopropionic acid, is a precursor compound for biosynthesis of polythioesters in bacteria. The cost-effectiveness and sustainability of the whole process could be greatly improved by using the cysteine degradation pathway for an intracellular supply of 3MPy. Transamination of cysteine to its corresponding α-keto acid 3MPy is catalyzed by cysteine aminotransferases (CAT). However, CAT activity has so far not been described for bacterial aminotransferases (AT), and it was unknown whether they can be applied for the conversion of cysteine to 3MPy. In this study, we selected eight bacterial aminotransferases based on sequence homology to CAT of Rattus norvegicus (Got1). The aminotransferases included four aspartate aminotransferases (AATs) and four aromatic amino acid aminotransferases (ArATs) from Advenella mimigardefordensis DPN7, Escherichia coli MG1655, Shimwellia blattae ATCC 33430, Ralstonia eutropha H16 and Paracoccus denitrificans PD1222. For a more detailed characterization, all selected AAT or ArAT encoding genes were heterologously expressed in E. coli and purified. CAT activity was detected for all aminotransferases when a novel continuous coupled enzyme assay was applied. Kinetic studies revealed the highest catalytic efficiency of 5.1mM/s for AAT from A. mimigardefordensis. Formation of 3MPy from cysteine could additionally be verified by an optimized approach using derivatization of 3MPy with the Girard T reagent and liquid chromatography-mass spectrometry analyses.

  13. Functional analysis of all aminotransferase proteins inferred from the genome sequence of Corynebacterium glutamicum.

    PubMed

    Marienhagen, Jan; Kennerknecht, Nicole; Sahm, Hermann; Eggeling, Lothar

    2005-11-01

    Twenty putative aminotransferase (AT) proteins of Corynebacterium glutamicum, or rather pyridoxal-5'-phosphate (PLP)-dependent enzymes, were isolated and assayed among others with L-glutamate, L-aspartate, and L-alanine as amino donors and a number of 2-oxo-acids as amino acceptors. One outstanding AT identified is AlaT, which has a broad amino donor specificity utilizing (in the order of preference) L-glutamate > 2-aminobutyrate > L-aspartate with pyruvate as acceptor. Another AT is AvtA, which utilizes L-alanine to aminate 2-oxo-isovalerate, the L-valine precursor, and 2-oxo-butyrate. A second AT active with the L-valine precursor and that of the other two branched-chain amino acids, too, is IlvE, and both enzyme activities overlap partially in vivo, as demonstrated by the analysis of deletion mutants. Also identified was AroT, the aromatic AT, and this and IlvE were shown to have comparable activities with phenylpyruvate, thus demonstrating the relevance of both ATs for L-phenylalanine synthesis. We also assessed the activity of two PLP-containing cysteine desulfurases, supplying a persulfide intermediate. One of them is SufS, which assists in the sulfur transfer pathway for the Fe-S cluster assembly. Together with the identification of further ATs and the additional analysis of deletion mutants, this results in an overview of the ATs within an organism that may not have been achieved thus far.

  14. Approach to asymptomatic creatine kinase elevation

    PubMed Central

    MOGHADAM-KIA, SIAMAK; ODDIS, CHESTER V.; AGGARWAL, ROHIT

    2016-01-01

    How to manage a patient who has an elevated serum creatine kinase (CK) level but no or insignificant muscle-related signs and symptoms is a clinical conundrum. The authors provide a systematic approach, including repeat testing after a period of rest, defining higher thresholds over which pursuing a diagnosis is worthwhile, and evaluating for a variety of nonneuromuscular causes. They also outline a workup for neuromuscular causes. PMID:26760521

  15. Similarities between cysteinesulphinate transaminase and aspartate aminotransferase.

    PubMed

    Recasens, M; Mandel, P

    1979-01-01

    A method for the purification of two cysteinesulphinate transaminases, A and B (EC 2.6.1), is described. These enzymes catalyse the conversion of cysteinesulphinic acid to beta-sulphinyl pyruvate. The final preparations are homogeneous by polyacrylamide gel electrophoresis, sodium dodecyl sulphate-polyacrylamide gel electrophoresis and isoelectrofocusing. The molecular weight of the subunits is 41 000 for cysteinesulphinate transaminase A and 43 400 for B. Both enzymes are unspecific, as L-asparate, L-glutamate and L-cysteic acid serve as substrates in addition to L-cysteinesulphinic acid. Cysteinesulphinate transaminase A has a Km of 9.8 mM for cysteinesulphinic acid and 0.25 mM for aspartic acid, whereas the B enzyme has a Km of 6.5 mM for cysteinesulphinic acid and 1.4 mM for aspartic acid. The Vmax values of the A and B enzymes are respectively 7.1 and 6.2 mmol h-1 mg-1 protein for aspartic acid and 45 and 9.3 mmol h-1 mg-1 protein for cysteinesulphinic acid. Both enzymes exhibit maximum activity at pH 8.6. A high specific activity is found in optimal conditions for these two transaminases, the pI values being 9.06 and 5.70 for cysteinesulphinate transaminase A and B respectively. These results have been compared with those already obtained for purified aspartate aminotransferase. Similarities in the pathways of taurine and gamma-aminobutyric acid (GABA) metabolism are discussed.

  16. Kynurenine Aminotransferase Isozyme Inhibitors: A Review

    PubMed Central

    Nematollahi, Alireza; Sun, Guanchen; Jayawickrama, Gayan S.; Church, W. Bret

    2016-01-01

    Kynurenine aminotransferase isozymes (KATs 1–4) are members of the pyridoxal-5’-phosphate (PLP)-dependent enzyme family, which catalyse the permanent conversion of l-kynurenine (l-KYN) to kynurenic acid (KYNA), a known neuroactive agent. As KATs are found in the mammalian brain and have key roles in the kynurenine pathway, involved in different categories of central nervous system (CNS) diseases, the KATs are prominent targets in the quest to treat neurodegenerative and cognitive impairment disorders. Recent studies suggest that inhibiting these enzymes would produce effects beneficial to patients with these conditions, as abnormally high levels of KYNA are observed. KAT-1 and KAT-3 share the highest sequence similarity of the isozymes in this family, and their active site pockets are also similar. Importantly, KAT-2 has the major role of kynurenic acid production (70%) in the human brain, and it is considered therefore that suitable inhibition of this isozyme would be most effective in managing major aspects of CNS diseases. Human KAT-2 inhibitors have been developed, but the most potent of them, chosen for further investigations, did not proceed in clinical studies due to the cross toxicity caused by their irreversible interaction with PLP, the required cofactor of the KAT isozymes, and any other PLP-dependent enzymes. As a consequence of the possibility of extensive undesirable adverse effects, it is also important to pursue KAT inhibitors that reversibly inhibit KATs and to include a strategy that seeks compounds likely to achieve substantial interaction with regions of the active site other than the PLP. The main purpose of this treatise is to review the recent developments with the inhibitors of KAT isozymes. This treatise also includes analyses of their crystallographic structures in complex with this enzyme family, which provides further insight for researchers in this and related studies. PMID:27314340

  17. Substrate specificity and structure of human aminoadipate aminotransferase/kynurenine aminotransferase II.

    PubMed

    Han, Qian; Cai, Tao; Tagle, Danilo A; Robinson, Howard; Li, Jianyong

    2008-08-01

    KAT (kynurenine aminotransferase) II is a primary enzyme in the brain for catalysing the transamination of kynurenine to KYNA (kynurenic acid). KYNA is the only known endogenous antagonist of the N-methyl-D-aspartate receptor. The enzyme also catalyses the transamination of aminoadipate to alpha-oxoadipate; therefore it was initially named AADAT (aminoadipate aminotransferase). As an endotoxin, aminoadipate influences various elements of glutamatergic neurotransmission and kills primary astrocytes in the brain. A number of studies dealing with the biochemical and functional characteristics of this enzyme exist in the literature, but a systematic assessment of KAT II addressing its substrate profile and kinetic properties has not been performed. The present study examines the biochemical and structural characterization of a human KAT II/AADAT. Substrate screening of human KAT II revealed that the enzyme has a very broad substrate specificity, is capable of catalysing the transamination of 16 out of 24 tested amino acids and could utilize all 16 tested alpha-oxo acids as amino-group acceptors. Kinetic analysis of human KAT II demonstrated its catalytic efficiency for individual amino-group donors and acceptors, providing information as to its preferred substrate affinity. Structural analysis of the human KAT II complex with alpha-oxoglutaric acid revealed a conformational change of an N-terminal fraction, residues 15-33, that is able to adapt to different substrate sizes, which provides a structural basis for its broad substrate specificity.

  18. Biochemical and structural characterization of mouse mitochondrial aspartate aminotransferase, a newly identified kynurenine aminotransferase-IV

    PubMed Central

    Han, Qian; Robinson, Howard; Cai, Tao; Tagle, Danilo A.; Li, Jianyong

    2010-01-01

    Synopsis Mammalian mitochondrial aspartate aminotransferase (mAspAT) is recently reported to have kynurenine aminotransferase (KAT) activity and plays a role in the biosynthesis of kynurenic acid (KYNA) in rat, mouse and human brains. This study concerns the biochemical and structural characterization of mouse mAspAT. In this study, mouse mAspAT cDNA was amplified from mouse brain first stand cDNA and its recombinant protein was expressed in an Escherichia coli expression system. Sixteen keto acids were tested for the co-substrate specificity of mouse mAspAT and fourteen of them were shown to be capable of serving as co-substrates for the enzyme. Structural analysis of mAspAT by macromolecular crystallography revealed that the cofactor binding residues of mAspAT are similar to those of other KATs. The substrate binding residues of mAspAT are slightly different from those of other KATs. Our data provide a biochemical and structural basis towards understanding the overall physiological role of mAspAT in vivo and insight into controlling the levels of endogenous KYNA through modulation of the enzyme in the mouse brain. PMID:20977429

  19. Substrate specificity and structure of human aminoadipate aminotransferase/kynurenine aminotransferase II

    PubMed Central

    HAN, Qian; CAI, Tao; TAGLE, Danilo A.; ROBINSON, Howard; LI, Jianyong

    2008-01-01

    Synopsis KAT (kynurenine aminotransferase) II is a primary enzyme in the brain for catalysing the transamination of kynurenine to KYNA (kynurenic acid). KYNA is the only known endogenous antagonist of the N-methyl-d-aspartate receptor. The enzyme also catalyses the transamination of aminoadipate to α-oxoadipate; therefore it was initially named AADAT (aminoadipate aminotransferase). As an endotoxin, aminoadipate influences various elements of glutamatergic neurotransmission and kills primary astrocytes in the brain. A number of studies dealing with the biochemical and functional characteristics of this enzyme exist in the literature, but a systematic assessment of KAT II addressing its substrate profile and kinetic properties has not been performed. The present study examines the biochemical and structural characterization of a human KAT II/AADAT. Substrate screening of human KAT II revealed that the enzyme has a very broad substrate specificity, is capable of catalysing the transamination of 16 out of 24 tested amino acids and could utilize all 16 tested α-oxo acids as amino-group acceptors. Kinetic analysis of human KAT II demonstrated its catalytic efficiency for individual amino-group donors and acceptors, providing information as to its preferred substrate affinity. Structural analysis of the human KAT II complex with α-oxoglutaric acid revealed a conformational change of an N-terminal fraction, residues 15-33, that is able to adapt to different substrate sizes, which provides a structural basis for its broad substrate specificity. PMID:18620547

  20. Biochemical and structural characterization of mouse mitochondrial aspartate aminotransferase, a newly identified kynurenine aminotransferase-IV.

    PubMed

    Han, Qian; Robinson, Howard; Cai, Tao; Tagle, Danilo A; Li, Jianyong

    2011-10-01

    Mammalian mAspAT (mitochondrial aspartate aminotransferase) is recently reported to have KAT (kynurenine aminotransferase) activity and plays a role in the biosynthesis of KYNA (kynurenic acid) in rat, mouse and human brains. This study concerns the biochemical and structural characterization of mouse mAspAT. In this study, mouse mAspAT cDNA was amplified from mouse brain first stand cDNA and its recombinant protein was expressed in an Escherichia coli expression system. Sixteen oxo acids were tested for the co-substrate specificity of mouse mAspAT and 14 of them were shown to be capable of serving as co-substrates for the enzyme. Structural analysis of mAspAT by macromolecular crystallography revealed that the cofactor-binding residues of mAspAT are similar to those of other KATs. The substrate-binding residues of mAspAT are slightly different from those of other KATs. Our results provide a biochemical and structural basis towards understanding the overall physiological role of mAspAT in vivo and insight into controlling the levels of endogenous KYNA through modulation of the enzyme in the mouse brain.

  1. Biochemical and structural characterization of mouse mitochondrial aspartate aminotransferase, a newly identified kynurenine aminotransferase-IV

    SciTech Connect

    Han, Q.; Robinson, H.; Cai, T.; Tagle, D. A.; Li, J.

    2011-10-01

    Mammalian mAspAT (mitochondrial aspartate aminotransferase) is recently reported to have KAT (kynurenine aminotransferase) activity and plays a role in the biosynthesis of KYNA (kynurenic acid) in rat, mouse and human brains. This study concerns the biochemical and structural characterization of mouse mAspAT. In this study, mouse mAspAT cDNA was amplified from mouse brain first stand cDNA and its recombinant protein was expressed in an Escherichia coli expression system. Sixteen oxo acids were tested for the co-substrate specificity of mouse mAspAT and 14 of them were shown to be capable of serving as co-substrates for the enzyme. Structural analysis of mAspAT by macromolecular crystallography revealed that the cofactor-binding residues of mAspAT are similar to those of other KATs. The substrate-binding residues of mAspAT are slightly different from those of other KATs. Our results provide a biochemical and structural basis towards understanding the overall physiological role of mAspAT in vivo and insight into controlling the levels of endogenous KYNA through modulation of the enzyme in the mouse brain.

  2. Substrate Specificity and Structure of Human aminoadipate aminotransferase/kynurenine aminotransferase II

    SciTech Connect

    Han, Q.; Cai, T; Tagle, D; Robinson, H; Li, J

    2009-01-01

    KAT (kynurenine aminotransferase) II is a primary enzyme in the brain for catalysing the transamination of kynurenine to KYNA (kynurenic acid). KYNA is the only known endogenous antagonist of the N-methyl-D-aspartate receptor. The enzyme also catalyses the transamination of aminoadipate to alpha-oxoadipate; therefore it was initially named AADAT (aminoadipate aminotransferase). As an endotoxin, aminoadipate influences various elements of glutamatergic neurotransmission and kills primary astrocytes in the brain. A number of studies dealing with the biochemical and functional characteristics of this enzyme exist in the literature, but a systematic assessment of KAT II addressing its substrate profile and kinetic properties has not been performed. The present study examines the biochemical and structural characterization of a human KAT II/AADAT. Substrate screening of human KAT II revealed that the enzyme has a very broad substrate specificity, is capable of catalysing the transamination of 16 out of 24 tested amino acids and could utilize all 16 tested alpha-oxo acids as amino-group acceptors. Kinetic analysis of human KAT II demonstrated its catalytic efficiency for individual amino-group donors and acceptors, providing information as to its preferred substrate affinity. Structural analysis of the human KAT II complex with alpha-oxoglutaric acid revealed a conformational change of an N-terminal fraction, residues 15-33, that is able to adapt to different substrate sizes, which provides a structural basis for its broad substrate specificity.

  3. Substrate Specificity and Structure of Human Aminoadipate Aminotransferase/kynurenine Aminotransferase II

    SciTech Connect

    Han,Q.; Cai, T.; Tagle, D.; Robinson, H.; Li, J.

    2008-01-01

    KAT (kynurenine aminotransferase) II is a primary enzyme in the brain for catalysing the transamination of kynurenine to KYNA (kynurenic acid). KYNA is the only known endogenous antagonist of the N-methyl-D-aspartate receptor. The enzyme also catalyses the transamination of aminoadipate to a-oxoadipate; therefore it was initially named AADAT (aminoadipate aminotransferase). As an endotoxin, aminoadipate influences various elements of glutamatergic neurotransmission and kills primary astrocytes in the brain. A number of studies dealing with the biochemical and functional characteristics of this enzyme exist in the literature, but a systematic assessment of KAT II addressing its substrate profile and kinetic properties has not been performed. The present study examines the biochemical and structural characterization of a human KAT II/AADAT. Substrate screening of human KAT II revealed that the enzyme has a very broad substrate specificity, is capable of catalysing the transamination of 16 out of 24 tested amino acids and could utilize all 16 tested a-oxo acids as amino-group acceptors. Kinetic analysis of human KAT II demonstrated its catalytic efficiency for individual amino-group donors and acceptors, providing information as to its preferred substrate affinity. Structural analysis of the human KAT II complex with a-oxoglutaric acid revealed a conformational change of an N-terminal fraction, residues 15-33, that is able to adapt to different substrate sizes, which provides a structural basis for its broad substrate specificity.

  4. Creatine metabolism: detection of creatine and guanidinoacetate in saliva of healthy subjects.

    PubMed

    Martínez, Lidia D; Bezard, Miriam; Brunotto, Mabel; Dodelson de Kremer, Raquel

    2016-04-01

    Creatine (Cr) plays an important role in storage and transmission of phosphate-bound energy. Cerebral creatine deficiency syndromes comprise three inherited defects in Cr biosynthesis and transport. The aim of this study was to investigate whether Cr and Guanidinoacetate (GAA) can be detected in saliva of healthy subjects and to establish the relationship between salivary and plasma levels of these molecules. An adapted gas chromatography (GC) method is described for the quantification of Cr and GAA biomarkers in saliva. Reference values were established for GAA and Cr in saliva. These values were age dependent (p= 0.001). No difference between genders was observed. We detected a difference between GAA and Cr concentrations in saliva and in plasma. The GC method for simultaneous determination of GAA and Cr in human saliva is fast, reliable, sensitive, non-invasive and precise to use as a biochemical approach in early detection of cerebral creatine deficiency syndromes.

  5. Synthesis and biological evaluation of new creatine fatty esters revealed dodecyl creatine ester as a promising drug candidate for the treatment of the creatine transporter deficiency.

    PubMed

    Trotier-Faurion, Alexandra; Dézard, Sophie; Taran, Frédéric; Valayannopoulos, Vassili; de Lonlay, Pascale; Mabondzo, Aloïse

    2013-06-27

    The creatine transporter deficiency is a neurological disease caused by impairment of the creatine transporter SLC6A8, resulting in mental retardation associated with a complete absence of creatine within the brain and cellular energy perturbation of neuronal cells. One of the therapeutic hypotheses was to administer lipophilic creatine derivatives which are (1) thought to have better permeability through the cell membrane and (2) would not rely on the activity of SLC6A8 to penetrate the brain. Here, we synthesized creatine fatty esters through original organic chemistry process. A screening on an in vitro rat primary cell-based blood-brain barrier model and on a rat primary neuronal cells model demonstrated interesting properties of these prodrugs to incorporate into endothelial, astroglial, and neuronal cells according to a structure-activity relationship. Dodecyl creatine ester showed then a 20-fold increase in creatine content in pathological human fibroblasts compared with the endogenous creatine content, stating that it could be a promising drug candidate.

  6. Macro creatine kinase type 1: a cause of spuriously elevated serum creatine kinase associated with leukoencephalopathy in a child.

    PubMed

    Bodensteiner, John B

    2014-07-01

    Macro creatine kinase type 1 is a complex formed by the creatine kinase isoenzyme BB and monoclonal IgG and occurs in about 1% of patients studied. First identified as a cause of spurious elevation of the total serum creatine kinase in patients suspected of myocardial infarction, the test has been largely replaced by the measurement of troponin levels. We present a child with delayed milestones and persistently elevated total serum creatine kinase measurements (∼ 1000-4000 IU) normal electromyogram and brisk myotatic reflexes. Creatine kinase isoenzymes and brain imaging showed the presence of macro creatine kinase type 1 and extensive signal abnormality of the cerebral white matter. Macro creatine kinase type 1 has been associated with several conditions though it has not been described in association with leukoencephalopathy or in patients this young. Macro creatine kinase type 1 can be a cause of elevated total creatine kinase in patients without primary muscle disease. The significance of the relationship of the macro creatine kinase to the leukoencephalopathy in this patient is unknown.

  7. Effect of a defined lacto-ovo-vegetarian diet and oral creatine monohydrate supplementation on plasma creatine concentration.

    PubMed

    Lukaszuk, Judith M; Robertson, Robert J; Arch, Judith E; Moyna, Niall M

    2005-11-01

    This study examined the effects that preceding creatine supplementation with a lacto-ovo-vegetarian diet would have on plasma creatine concentration. Twenty-six healthy moderately fit omnivorous men were assigned to either a 26-day lacto-ovo-vegetarian (LOV; n = 12) or omnivorous (Omni; n = 14) diet. On day 22, subjects were also assigned in a double-blind manner either creatine monohydrate (CM; 0.3 g.kg(-1).day(-1) + 20 g Polycose) or an equivalent dose of placebo (PL) for 5 days. Blood samples were taken on days 1, 22 and 27. Consuming a LOV diet for 21 days was effective in reducing plasma creatine concentration (p < 0.01) in the LOV group. Regardless of diet, the CM group showed an increase in plasma creatine concentrations from day 22 to 27, whereas the PL group's levels remained the same (p < 0.05). Although the LOV diet caused a deprivation effect in plasma creatine concentration relative to the Omni diet, concurrent supplementation with creatine resulted in no difference in plasma creatine concentrations between the LOV and Omni diet groups. Dietary advice should be provided to LOV athletes that supplementation with creatine may help to increase their muscle stores of creatine, and thus their ATP resynthesis capabilities, to levels similar to those of omnivores.

  8. Inhibitors of alanine racemase enzyme: a review.

    PubMed

    Azam, Mohammed Afzal; Jayaram, Unni

    2016-08-01

    Alanine racemase is a fold type III PLP-dependent amino acid racemase enzyme catalysing the conversion of l-alanine to d-alanine utilised by bacterial cell wall for peptidoglycan synthesis. As there are no known homologs in humans, it is considered as an excellent antibacterial drug target. The standard inhibitors of this enzyme include O-carbamyl-d-serine, d-cycloserine, chlorovinyl glycine, alaphosphin, etc. d-Cycloserine is indicated for pulmonary and extra pulmonary tuberculosis but therapeutic use of drug is limited due to its severe toxic effects. Toxic effects due to off-target affinities of cycloserine and other substrate analogs have prompted new research efforts to identify alanine racemase inhibitors that are not substrate analogs. In this review, an updated status of known inhibitors of alanine racemase enzyme has been provided which will serve as a rich source of structural information and will be helpful in generating selective and potent inhibitor of alanine racemase.

  9. [Raman scattering study of DL-alanine].

    PubMed

    Gong, Yan; Wang, Wen-qing

    2006-01-01

    Studies of Raman vibration spectra are useful to obtaining information on biomolecular crystals. The cell dimensions of the L- and DL-alanine crystals are nearly identical, and both structures belong to the orthorhombic system, but the space group is P2(1) 2(1) 2(1) for the L-isomer, and Pna2(1) for the racemate crystal. The Raman spectrum of L-alanine has been measured by many authors. The present work is focusing on the Raman scattering study of DL-alanine powder. Based on the analysis of the differences between DL-alanine and L-alanine Raman spectra, the authors obtained indispensable information on hydrogen bond and the motion of the molecular conformation in alanine crystals.

  10. Elevated plasma creatinine due to creatine ethyl ester use.

    PubMed

    Velema, M S; de Ronde, W

    2011-02-01

    Creatine is a nutritional supplement widely used in sport, physical fitness training and bodybuilding. It is claimed to enhance performance. We describe a case in which serum creatinine is elevated due to the use of creatine ethyl esther. One week after withdrawal, the plasma creatinine had normalised. There are two types of creatine products available: creatine ethyl esther (CEE) and creatine monohydrate (CM). Plasma creatinine is not elevated in all creatine-using subjects. CEE , but not CM, is converted into creatinine in the gastrointestinal tract. As a result the use of CEE may be associated with elevated plasma creatinine levels. Since plasma creatinine is a widely used marker for renal function, the use of CEE may lead to a false assumption of renal failure.

  11. Vibrational dynamics of crystalline L-alanine

    SciTech Connect

    Bordallo, H.N.; Eckert, J.; Barthes, M.

    1997-11-01

    The authors report a new, complete vibrational analysis of L-alanine and L-alanine-d{sub 4} which utilizes IINS intensities in addition to frequency information. The use of both isotopomers resulted in a self-consistent force field for and assignment of the molecular vibrations in L-alanine. Some details of the calculation as well as a comparison of calculated and observed IINS spectra are presented. The study clarifies a number of important issues on the vibrational dynamics of this molecule and presents a self-consistent force field for the molecular vibrations in crystalline L-alanine.

  12. On the Importance of Exchangeable NH Protons in Creatine for the Magnetic Coupling of Creatine Methyl Protons in Skeletal Muscle

    NASA Astrophysics Data System (ADS)

    Kruiskamp, M. J.; Nicolay, K.

    2001-03-01

    The methyl protons of creatine in skeletal muscle exhibit a strong off-resonance magnetization transfer effect. The mechanism of this process is unknown. We previously hypothesized that the exchangeable amide/amino protons of creatine might be involved. To test this the characteristics of the creatine magnetization transfer effect were investigated in excised rat hindleg skeletal muscle that was equilibrated in either H2O or D2O solutions containing creatine. The efficiency of off-resonance magnetization transfer to the protons of mobile creatine in excised muscle was similar to that previously reported in intact muscle in vivo. Equilibrating the isolated muscle in D2O solution had no effect on the magnetic coupling to the immobile protons. It is concluded that exchangeable protons play a negligible role in the magnetic coupling of creatine methyl protons in muscle.

  13. Catalytic Stereoinversion of L-Alanine to Deuterated D-Alanine.

    PubMed

    Moozeh, Kimia; So, Soon Mog; Chin, Jik

    2015-08-03

    A combination of an achiral pyridoxal analogue and a chiral base has been developed for catalytic deuteration of L-alanine with inversion of stereochemistry to give deuterated D-alanine under mild conditions (neutral pD and 25 °C) without the use of any protecting groups. This system can also be used for catalytic deuteration of D-alanine with retention of stereochemistry to give deuterated D-alanine. Thus a racemic mixture of alanine can be catalytically deuterated to give an enantiomeric excess of deuterated D-alanine. While catalytic deracemization of alanine is forbidden by the second law of thermodynamics, this system can be used for catalytic deracemization of alanine with deuteration. Such green and biomimetic approach to catalytic stereocontrol provides insights into efficient amino acid transformations.

  14. Elevation of creatine in red blood cells in vegetarians and nonvegetarians after creatine supplementation.

    PubMed

    Maccormick, Vanessa M; Hill, Lisa M; Macneil, Lauren; Burke, Darren G; Smith-Palmer, Truis

    2004-12-01

    The purpose of this study was to examine the effect of a 5-day creatine (CR) supplementation period on red blood cell (RBC) CR uptake in vegetarian and nonvegetarian young women. Blood samples were collected from lacto-ovo vegetarians (VG, n = 6, age 21.8 +/- 1.9 yrs) and nonvegetarians (NV, n = 6, age 21.7 +/- 1.9 yrs) before and after a 5-day CR loading period (0. 3g CR/kg lean body mass/day), and from a control group of nonvegetarians (NV, n = 5, age 22.0 +/- 0.7 yrs) who did not supplement with creatine. RBC and plasma samples were analyzed for the presence of creatine. Significant increases (p < .05) in RBC and plasma CR levels were found for vegetarians and nonvegetarians following supplementation. The initial RBC CR content was significantly lower (p < .05) in the vegetarian group. There was no significant difference between vegetarians and nonvegetarians in final RBC CR content, suggesting that a ceiling had been reached. As the uptake into both muscle and RBC is moderated by creatine transporter proteins, analysis of the uptake of CR into RBC may reflect the uptake of CR into muscle, offering an alternative to biopsies.

  15. Creatine kinase expression and creatine phosphate accumulation are developmentally regulated during differentiation of mouse and human monocytes

    PubMed Central

    1984-01-01

    We have studied the expression of creatine kinase (CK) and the accumulation of creatine phosphate during the differentiation of human and mouse peripheral blood monocytes. Mouse monocytes cultured for 24 h do not contain detectable levels of CK and creatine phosphate. However, resident tissue macrophages and inflammatory elicited macrophages obtained from the peritoneal cavities of mice have 70 and 300 mU per mg protein of CK activity and contain 3 and 6 mol of creatine phosphate per mol of ATP, respectively. The major isozyme of CK in these cells has been identified as the brain form. These findings suggest that the differentiation of monocytes into macrophages is associated with the expression of CK and the accumulation of creatine phosphate. We have found a similar pattern in human monocytes. Human blood monocytes, maintained in culture for 24 or 48 h, do not contain detectable levels of CK or creatine phosphate. Monocyte-derived macrophages (monocytes maintained in tissue cultures for 1 to 2 wk) have up to 100 mU per mg protein of CK activity and contain 0.5 mol of creatine phosphate per mol of ATP. Human macrophages express multiple isozymes of CK including the brain (BB) and possibly the mitochondrial forms of this enzyme. Thus, the expression of CK and the accumulation of creatine phosphate in human monocytes is induced by their in vitro cultivation. The induction of CK during in vitro cultivation occurs independently of the concentration of creatine in the medium. However, the size of the creatine phosphate pool varies with respect to extracellular creatine concentration. Creatine phosphate and CK are not detectable in freshly isolated human lymphocytes, polymorphonuclear leukocytes or erythrocytes, but are found in freshly isolated human platelets. PMID:6699543

  16. The effect of longer-term creatine supplementation on elite swimming performance after an acute creatine loading.

    PubMed

    Theodorou, A S; Cooke, C B; King, R F; Hood, C; Denison, T; Wainwright, B G; Havenetidis, K

    1999-11-01

    We investigated the effect of an acute creatine loading (25 g per day for 4 days) and longer-term creatine supplementation (5 g of creatine or 5 g of placebo per day for 2 months) on the performance of 22 elite swimmers during maximal interval sessions. After the acute creatine loading, the mean of the average interval swim times for all swimmers (n = 22) improved (44.3+/-16.5 s before vs. 43.7+/-16.3 s after supplementation; P<0.01). Three of the 22 swimmers did not respond positively to supplementation. After 2 months of longer-term creatine supplementation or placebo, neither group showed a significant change in swimming performance (38.7+/-13.5 s before vs. 38.7+/-14.1 s after for the creatine group; 48.7+/-18.0 s before vs. 48.7+/-18.1 s after for the placebo group). We conclude that, in elite swimmers, 4 days of acute creatine loading improves swimming performance significantly when assessed by maximal interval sessions. However, longer-term supplementation for 2 months (5 g of creatine per day) did not benefit significantly the creatine group compared with the placebo group.

  17. A potential role for creatine in drug abuse?

    PubMed

    D'Anci, Kristen E; Allen, Patricia J; Kanarek, Robin B

    2011-10-01

    Supplemental creatine has been promoted for its positive health effects and is best known for its use by athletes to increase muscle mass. In addition to its role in physical performance, creatine supplementation has protective effects on the brain in models of neuronal damage and also alters mood state and cognitive performance. Creatine is found in high protein foods, such as fish or meat, and is also produced endogenously from the biosynthesis of arginine, glycine, and methionine. Changes in brain creatine levels, as measured using magnetic resonance spectroscopy, are seen in individuals exposed to drugs of abuse and depressed individuals. These changes in brain creatine indicate that energy metabolism differs in these populations relative to healthy individuals. Recent work shows that creatine supplementation has the ability to function in a manner similar to antidepressant drugs and can offset negative consequences of stress. These observations are important in relation to addictive behaviors as addiction is influenced by psychological factors such as psychosocial stress and depression. The significance of altered brain levels of creatine in drug-exposed individuals and the role of creatine supplementation in models of drug abuse have yet to be explored and represent gaps in the current understanding of brain energetics and addiction.

  18. Does reduced creatine synthesis protect against statin myopathy?

    PubMed

    Ballard, Kevin D; Thompson, Paul D

    2013-12-03

    Statins, widely used to lower cholesterol levels, cause myopathy in some patients. Mangravite et al. (2013) show that a single nucleotide polymorphism decreasing expression of glycine amidinotransferase (GATM), the enzyme regulating creatine biosynthesis, is associated with reduced statin myopathy. Whether reduced creatine production protects against statin myopathy remains to be determined.

  19. Creatine Synthesis: An Undergraduate Organic Chemistry Laboratory Experiment

    ERIC Educational Resources Information Center

    Smith, Andri L.; Tan, Paula

    2006-01-01

    Students in introductory chemistry classes typically appreciate seeing the connection between course content and the "real world". For this reason, we have developed a synthesis of creatine monohydrate--a popular supplement used in sports requiring short bursts of energy--for introductory organic chemistry laboratory courses. Creatine monohydrate…

  20. Total-body creatine pool size and skeletal muscle mass determination by creatine-(methyl-D3) dilution in rats.

    PubMed

    Stimpson, Stephen A; Turner, Scott M; Clifton, Lisa G; Poole, James C; Mohammed, Hussein A; Shearer, Todd W; Waitt, Greg M; Hagerty, Laura L; Remlinger, Katja S; Hellerstein, Marc K; Evans, William J

    2012-06-01

    There is currently no direct, facile method to determine total-body skeletal muscle mass for the diagnosis and treatment of skeletal muscle wasting conditions such as sarcopenia, cachexia, and disuse. We tested in rats the hypothesis that the enrichment of creatinine-(methyl-d(3)) (D(3)-creatinine) in urine after a defined oral tracer dose of D(3)-creatine can be used to determine creatine pool size and skeletal muscle mass. We determined 1) an oral tracer dose of D(3)-creatine that was completely bioavailable with minimal urinary spillage and sufficient enrichment in the body creatine pool for detection of D(3)-creatine in muscle and D(3)-creatinine in urine, and 2) the time to isotopic steady state. We used cross-sectional studies to compare total creatine pool size determined by the D(3)-creatine dilution method to lean body mass determined by independent methods. The tracer dose of D(3)-creatine (<1 mg/rat) was >99% bioavailable with 0.2-1.2% urinary spillage. Isotopic steady state was achieved within 24-48 h. Creatine pool size calculated from urinary D(3)-creatinine enrichment at 72 h significantly increased with muscle accrual in rat growth, significantly decreased with dexamethasone-induced skeletal muscle atrophy, was correlated with lean body mass (r = 0.9590; P < 0.0001), and corresponded to predicted total muscle mass. Total-body creatine pool size and skeletal muscle mass can thus be accurately and precisely determined by an orally delivered dose of D(3)-creatine followed by the measurement of D(3)-creatinine enrichment in a single urine sample and is promising as a noninvasive tool for the clinical determination of skeletal muscle mass.

  1. Creatine supplementation and swim performance: a brief review.

    PubMed

    Hopwood, Melissa J; Graham, Kenneth; Rooney, Kieron B

    2006-03-01

    Nutritional supplements are popular among athletes participating in a wide variety of sports. Creatine is one of the most commonly used dietary supplements, as it has been shown to be beneficial in improving performance during repeated bouts of high-intensity anaerobic activity. This review examines the specific effects of creatine supplementation on swimming performance, and considers the effects of creatine supplementation on various measures of power development in this population. Research performed on the effect of creatine supplementation on swimming performance indicates that whilst creatine supplementation is ineffective in improving performance during a single sprint swim, dietary creatine supplementation may benefit repeated interval swim set performance. Considering the relationship between sprint swimming performance and measurements of power, the effect of creatine supplementation on power development in swimmers has also been examined. When measured on a swim bench ergometer, power development does show some improvement following a creatine supplementation regime. How this improvement in power output transfers to performance in the pool is uncertain. Although some evidence exists to suggest a gender effect on the performance improvements seen in swimmers following creatine supplementation, the majority of research indicates that male and female swimmers respond equally to supplementation. A major limitation to previous research is the lack of consideration given to the possible stroke dependant effect of creatine supplementation on swimming performance. The majority of the research conducted to date has involved examination of the freestyle swimming stroke only. The potential for performance improvements in the breaststroke and butterfly swimming strokes is discussed, with regards to the biomechanical differences and differences in efficiency between these strokes and freestyle. Key PointsCreatine supplementation does not improve single sprint

  2. Creatine supplementation reduces doxorubicin-induced cardiomyocellular injury.

    PubMed

    Santacruz, Lucia; Darrabie, Marcus D; Mantilla, Jose Gabriel; Mishra, Rajashree; Feger, Bryan J; Jacobs, Danny O

    2015-04-01

    Heart failure is a common complication of doxorubicin (DOX) therapy. Previous studies have shown that DOX adversely impacts cardiac energy metabolism, and the ensuing energy deficiencies antedate clinical manifestations of cardiac toxicity. Brief exposure of cultured cardiomyocytes to DOX significantly decreases creatine transport, which is the cell's sole source of creatine. We present the results of a study performed to determine if physiological creatine supplementation (5 mmol/L) could protect cardiomyocytes in culture from cellular injury resulting from exposure to therapeutic levels of DOX. Creatine supplementation significantly decreased cytotoxicity, apoptosis, and reactive oxygen species production caused by DOX. The protective effect was specific to creatine and depended on its transport into the cell.

  3. Complete inhibition of creatine kinase in isolated perfused rat hearts

    SciTech Connect

    Fossel, E.T.; Hoefeler, H.

    1987-01-01

    Transient exposure of an isolated isovolumic perfused rat heart to low concentrations (0.5 mM) of perfusate-born iodoacetamide resulted in complete inhibition of creatine kinase and partial inhibition of glyceraldehyde-3-phosphate dehydrogenase in the heart. At low levels of developed pressure, hearts maintained mechanical function, ATP, and creatine phosphate levels at control values. However, iodoacetamide-inhibited hearts were unable to maintain control values of end diastolic pressure or peak systolic pressure as work load increased. Global ischemia resulted in loss of all ATP without loss of creatine phosphate, indicating lack of active creatine kinase. These results indicate that isovolumic perfused rat hearts are able to maintain normal function and normal levels of high-energy phosphates without active creatine kinase at low levels of developed pressure. /sup 31/P-NMR of the heart was carried out.

  4. Structure of putrescine aminotransferase from Escherichia coli provides insights into the substrate specificity among class III aminotransferases.

    PubMed

    Cha, Hyung Jin; Jeong, Jae-Hee; Rojviriya, Catleya; Kim, Yeon-Gil

    2014-01-01

    YgjG is a putrescine aminotransferase enzyme that transfers amino groups from compounds with terminal primary amines to compounds with an aldehyde group using pyridoxal-5'-phosphate (PLP) as a cofactor. Previous biochemical data show that the enzyme prefers primary diamines, such as putrescine, over ornithine as a substrate. To better understand the enzyme's substrate specificity, crystal structures of YgjG from Escherichia coli were determined at 2.3 and 2.1 Å resolutions for the free and putrescine-bound enzymes, respectively. Sequence and structural analyses revealed that YgjG forms a dimer that adopts a class III PLP-dependent aminotransferase fold. A structural comparison between YgjG and other class III aminotransferases revealed that their structures are similar. However, YgjG has an additional N-terminal helical structure that partially contributes to a dimeric interaction with the other subunit via a helix-helix interaction. Interestingly, the YgjG substrate-binding site entrance size and charge distribution are smaller and more hydrophobic than other class III aminotransferases, which suggest that YgjG has a unique substrate binding site that could accommodate primary aliphatic diamine substrates, including putrescine. The YgjG crystal structures provide structural clues to putrescine aminotransferase substrate specificity and binding.

  5. METHANOGENS WITH PSEUDOMUREIN USE DIAMINOPIMELATE AMINOTRANSFERASE IN LYSINE BIOSYNTHESIS

    PubMed Central

    Graham, David E.; Huse, Holly K.

    2008-01-01

    Methanothermobacter thermautotrophicus uses lysine for both protein synthesis and cross-linking pseudomurein in its cell wall. A diaminopimelate aminotransferase enzyme from this methanogen (MTH0052) converts tetrahydrodipicolinate to L,L-diaminopimelate, a lysine precursor. This gene complemented an Escherichia coli diaminopimelate auxotrophy, and the purified protein catalyzed the transamination of diaminopimelate to tetrahydrodipicolinate. Phylogenetic analysis indicated this gene was recruited from anaerobic Gram-positive bacteria. These results expand the family of diaminopimelate aminotransferases to a diverse set of plant, bacterial and archaeal homologs. In contrast marine methanogens from the Methanococcales, which lack pseudomurein, appear to use a different diaminopimelate pathway for lysine biosynthesis. PMID:18371309

  6. Functional Analysis of All Aminotransferase Proteins Inferred from the Genome Sequence of Corynebacterium glutamicum

    PubMed Central

    Marienhagen, Jan; Kennerknecht, Nicole; Sahm, Hermann; Eggeling, Lothar

    2005-01-01

    Twenty putative aminotransferase (AT) proteins of Corynebacterium glutamicum, or rather pyridoxal-5′-phosphate (PLP)-dependent enzymes, were isolated and assayed among others with l-glutamate, l-aspartate, and l-alanine as amino donors and a number of 2-oxo-acids as amino acceptors. One outstanding AT identified is AlaT, which has a broad amino donor specificity utilizing (in the order of preference) l-glutamate > 2-aminobutyrate > l-aspartate with pyruvate as acceptor. Another AT is AvtA, which utilizes l-alanine to aminate 2-oxo-isovalerate, the l-valine precursor, and 2-oxo-butyrate. A second AT active with the l-valine precursor and that of the other two branched-chain amino acids, too, is IlvE, and both enzyme activities overlap partially in vivo, as demonstrated by the analysis of deletion mutants. Also identified was AroT, the aromatic AT, and this and IlvE were shown to have comparable activities with phenylpyruvate, thus demonstrating the relevance of both ATs for l-phenylalanine synthesis. We also assessed the activity of two PLP-containing cysteine desulfurases, supplying a persulfide intermediate. One of them is SufS, which assists in the sulfur transfer pathway for the Fe-S cluster assembly. Together with the identification of further ATs and the additional analysis of deletion mutants, this results in an overview of the ATs within an organism that may not have been achieved thus far. PMID:16267288

  7. Can creatine supplementation form carcinogenic heterocyclic amines in humans?

    PubMed

    Pereira, Renato Tavares dos Santos; Dörr, Felipe Augusto; Pinto, Ernani; Solis, Marina Yazigi; Artioli, Guilherme Giannini; Fernandes, Alan Lins; Murai, Igor Hisashi; Dantas, Wagner Silva; Seguro, Antônio Carlos; Santinho, Mirela Aparecida Rodrigues; Roschel, Hamilton; Carpentier, Alain; Poortmans, Jacques Remi; Gualano, Bruno

    2015-09-01

    There is a long-standing concern that creatine supplementation could be associated with cancer, possibly by facilitating the formation of carcinogenic heterocyclic amines (HCAs). This study provides compelling evidence that both low and high doses of creatine supplementation, given either acutely or chronically, does not cause a significant increase in HCA formation. HCAs detection was unrelated to creatine supplementation. Diet was likely to be the main factor responsible for HCAs formation after either placebo (n = 6) or creatine supplementation (n = 3). These results directly challenge the recently suggested biological plausibility for the association between creatine use and risk of testicular germ cell cancer. Creatine supplementation has been associated with increased cancer risk. In fact, there is evidence indicating that creatine and/or creatinine are important precursors of carcinogenic heterocyclic amines (HCAs). The present study aimed to investigate the acute and chronic effects of low- and high-dose creatine supplementation on the production of HCAs in healthy humans (i.e. 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (8-MeIQx), 2-amino-(1,6-dimethylfuro[3,2-e]imidazo[4,5-b])pyridine (IFP) and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4,8-DiMeIQx)). This was a non-counterbalanced single-blind crossover study divided into two phases, in which low- and high-dose creatine protocols were tested. After acute (1 day) and chronic supplementation (30 days), the HCAs PhIP, 8-MeIQx, IFP and 4,8-DiMeIQx were assessed through a newly developed HPLC-MS/MS method. Dietary HCA intake and blood and urinary creatinine were also evaluated. Out of 576 assessments performed (from 149 urine samples), only nine (3 from creatine and 6 from placebo) showed quantifiable levels of HCAs (8-MeIQx: n = 3; 4,8-DiMeIQx: n = 2; PhIP: n = 4). Individual analyses revealed that diet rather than creatine supplementation was

  8. Alanine increases blood pressure during hypotension

    NASA Technical Reports Server (NTRS)

    Conlay, L. A.; Maher, T. J.; Wurtman, R. J.

    1990-01-01

    The effect of L-alanine administration on blood pressure (BP) during haemorrhagic shock was investigated using anesthetized rats whose left carotid arteries were cannulated for BP measurement, blood removal, and drug administration. It was found that L-alanine, in doses of 10, 25, 50, 100, and 200 mg/kg, increased the systolic BP of hypotensive rats by 38 to 80 percent (while 100 mg/kg pyruvate increased BP by only 9.4 mmhg, not significantly different from saline). The results suggest that L-alanine might influence cardiovascular function.

  9. Creatine supplementation does not improve cognitive function in young adults.

    PubMed

    Rawson, Eric S; Lieberman, Harris R; Walsh, Talia M; Zuber, Sylwia M; Harhart, Jaclyn M; Matthews, Tracy C

    2008-09-03

    Creatine supplementation has been reported to improve certain aspects of cognitive and psychomotor function in older individuals and in young subjects following 24 and 36 h of sleep deprivation. However, the effects of creatine supplementation on cognitive processing and psychomotor performance in non-sleep deprived young adults have not been assessed with a comprehensive battery of neurocognitive tests. The primary objective of this study was to examine the effects of creatine supplementation on cognitive processing and psychomotor performance in young adults. Twenty-two subjects (21+/-2 yr) ingested creatine (0.03 g/kg/day) or placebo for 6 weeks in a double-blind placebo-controlled fashion. Subjects completed a battery of neurocognitive tests pre- and post-supplementation, including: simple reaction time (RT), code substitution (CS), code substitution delayed (CSD), logical reasoning symbolic (LRS), mathematical processing (MP), running memory (RM), and Sternberg memory recall (MR). There were no significant effects of group, no significant effects of time, and no significant group by time interactions for RT, CS, CSD, LRS, MP, RM, and MR (all p>0.05), indicating that there were no differences between creatine and placebo supplemented groups at any time. These results suggest that six weeks of creatine supplementation (0.03/g/kg/day) does not improve cognitive processing in non-sleep deprived young adults. Potentially, creatine supplementation only improves cognitive processing and psychomotor performance in individuals who have impaired cognitive processing abilities.

  10. Creatine monohydrate supplementation on body weight and percent body fat.

    PubMed

    Kutz, Matthew R; Gunter, Michael J

    2003-11-01

    Seventeen active males (age 22.9 +/- 4.9 year) participated in a study to examine the effects of creatine monohydrate supplementation on total body weight (TBW), percent body fat, body water content, and caloric intake. The TBW was measured in kilograms, percent body fat by hydrostatic weighing, body water content via bioelectrical impedance, and caloric intake by daily food log. Subjects were paired and assigned to a creatine or placebo group with a double-blind research design. Supplementation was given for 4 weeks (30 g a day for the initial 2 weeks and 15 g a day for the final 2 weeks). Subjects reported 2 days a week for supervised strength training of the lower extremity. Significant increases before and after the study were found in TBW (90.42 +/- 14.74 to 92.12 +/- 15.19 kg) and body water content (53.77 +/- 1.75 to 57.15 +/- 2.01 L) for the creatine group (p = 0.05). No significant changes were found in percent body fat or daily caloric intake in the creatine group. No significant changes were noted for the placebo group. These findings support previous research that creatine supplementation increases TBW. Mean percent body fat and caloric intake was not affected by creatine supplementation. Therefore weight gain in lieu of creatine supplementation may in part be due to water retention.

  11. Cerebral creatine deficiencies: a group of treatable intellectual developmental disorders.

    PubMed

    Stockler-Ipsiroglu, Sylvia; van Karnebeek, Clara D M

    2014-07-01

    Currently there are 91 treatable inborn errors of metabolism that cause intellectual developmental disorders. Cerebral creatine deficiencies (CDD) comprise three of these: arginine: glycine amidinotransferase [AGAT], guanidinoacetate methyltransferase [GAMT], and X-linked creatine transporter deficiency [SLC6A8]. Intellectual developmental disorder and cerebral creatine deficiency are the hallmarks of CDD. Additional clinical features include prominent speech delay, autism, epilepsy, extrapyramidal movement disorders, and signal changes in the globus pallidus. Patients with GAMT deficiency exhibit the most severe clinical spectrum. Myopathy is a distinct feature in AGAT deficiency. Guanidinoacetate (GAA) is the immediate product in the creatine biosynthetic pathway. Low GAA concentrations in urine, plasma, and cerebrospinal fluid are characteristic diagnostic markers for AGAT deficiency, while high GAA concentrations are characteristic markers for GAMT deficiency. An elevated ratio of urinary creatine /creatinine excretion serves as a diagnostic marker in males with SLC6A8 deficiency. Treatment strategies include oral supplementation of high-dose creatine-monohydrate for all three CDD. Guanidinoacetate-reducing strategies (high-dose ornithine, arginine-restricted diet) are additionally employed in GAMT deficiency. Supplementation of substrates for intracerebral creatine synthesis (arginine, glycine) has been used additionally to treat SLC6A8 deficiency. Early recognition and treatment improves outcomes. Normal outcomes in neonatally ascertained siblings from index families with AGAT and GAMT deficiency suggest a potential benefit of newborn screening for these disorders.

  12. Creatine and the Liver: Metabolism and Possible Interactions.

    PubMed

    Barcelos, R P; Stefanello, S T; Mauriz, J L; Gonzalez-Gallego, J; Soares, F A A

    2016-01-01

    The process of creatine synthesis occurs in two steps, catalyzed by L-arginine:glycine amidinotransferase (AGAT) and guanidinoacetate N-methyltransferase (GAMT), which take place mainly in kidney and liver, respectively. This molecule plays an important energy/pH buffer function in tissues, and to guarantee the maintenance of its total body pool, the lost creatine must be replaced from diet or de novo synthesis. Creatine administration is known to decrease the consumption of Sadenosyl methionine and also reduce the homocysteine production in liver, diminishing fat accumulation and resulting in beneficial effects in fatty liver and non-alcoholic liver disease. Different studies have shown that creatine supplementation could supply brain energy, presenting neuroprotective effects against the encephalopathy induced by hyperammonemia in acute liver failure. Creatine is also taken by many athletes for its ergogenic properties. However, little is known about the adverse effects of creatine supplementation, which are barely described in the literature, with reports of mainly hypothetical effects arising from a small number of scientific publications. Antioxidant effects have been found in several studies, although one of the theories regarding the potential for toxicity from creatine supplementation is that it can increase oxidative stress and potentially form carcinogenic compounds.

  13. Creatine kinase inhibits ADP-induced platelet aggregation

    PubMed Central

    Horjus, D. L.; Nieuwland, R.; Boateng, K. B.; Schaap, M. C. L.; van Montfrans, G. A.; Clark, J. F.; Sturk, A.; Brewster, L. M.

    2014-01-01

    Bleeding risk with antiplatelet therapy is an increasing clinical challenge. However, the inter-individual variation in this risk is poorly understood. We assessed whether the level of plasma creatine kinase, the enzyme that utilizes ADP and phosphocreatine to rapidly regenerate ATP, may modulate bleeding risk through a dose-dependent inhibition of ADP-induced platelet activation. Exogenous creatine kinase (500 to 4000 IU/L, phosphocreatine 5 mM) added to human plasma induced a dose-dependent reduction to complete inhibition of ADP-induced platelet aggregation. Accordingly, endogenous plasma creatine kinase, studied in 9 healthy men (mean age 27.9 y, SE 3.3; creatine kinase 115 to 859 IU/L, median 358), was associated with reduced ADP-induced platelet aggregation (Spearman's rank correlation coefficient, −0.6; p < 0.05). After exercise, at an endogenous creatine kinase level of 4664, ADP-induced platelet aggregation was undetectable, normalizing after rest, with a concomitant reduction of creatine kinase to normal values. Thus, creatine kinase reduces ADP-induced platelet activation. This may promote bleeding, in particular when patients use platelet P2Y12 ADP receptor inhibitors. PMID:25298190

  14. Strategic creatine supplementation and resistance training in healthy older adults.

    PubMed

    Candow, Darren G; Vogt, Emelie; Johannsmeyer, Sarah; Forbes, Scott C; Farthing, Jonathan P

    2015-07-01

    Creatine supplementation in close proximity to resistance training may be an important strategy for increasing muscle mass and strength; however, it is unknown whether creatine supplementation before or after resistance training is more effective for aging adults. Using a double-blind, repeated measures design, older adults (50-71 years) were randomized to 1 of 3 groups: creatine before (CR-B: n = 15; creatine (0.1 g/kg) immediately before resistance training and placebo (0.1 g/kg cornstarch maltodextrin) immediately after resistance training), creatine after (CR-A: n = 12; placebo immediately before resistance training and creatine immediately after resistance training), or placebo (PLA: n = 12; placebo immediately before and immediately after resistance training) for 32 weeks. Prior to and following the study, body composition (lean tissue, fat mass; dual-energy X-ray absorptiometry) and muscle strength (1-repetition maximum leg press and chest press) were assessed. There was an increase over time for lean tissue mass and muscle strength and a decrease in fat mass (p < 0.05). CR-A resulted in greater improvements in lean tissue mass (Δ 3.0 ± 1.9 kg) compared with PLA (Δ 0.5 ± 2.1 kg; p < 0.025). Creatine supplementation, independent of the timing of ingestion, increased muscle strength more than placebo (leg press: CR-B, Δ 36.6 ± 26.6 kg; CR-A, Δ 40.8 ± 38.4 kg; PLA, Δ 5.6 ± 35.1 kg; chest press: CR-B, Δ 15.2 ± 13.0 kg; CR-A, Δ 15.7 ± 12.5 kg; PLA, Δ 1.9 ± 14.7 kg; p < 0.025). Compared with resistance training alone, creatine supplementation improves muscle strength, with greater gains in lean tissue mass resulting from post-exercise creatine supplementation.

  15. Creatine regulation in the embryo and growing chick

    PubMed Central

    Ramírez, Oscar; Calva, Edmundo; Trejo, Augusto

    1970-01-01

    1. The absence of creatine was demonstrated enzymically in the hen's-egg yolk and in the albumin contrary to former reports. 2. A comparison of the results obtained by enzymic and colorimetric methods to measure creatine is presented. 3. Creatine phosphate was not detected in the yolk extracts. 4. The content of free arginine enzymically assayed was 15.7μmol in the yolk and 3.38μmol in the albumin. Arginine amounts to practically all of the guanidine compounds in the yolk and one-half of those in the albumin. 5. No glycine amidinotransferase activity was found in the egg-yolk homogenates. 6. The heart of the chick embryo does not receive creatine from the egg and the creatine kinase activity present in this organ starting from the 27th hour of incubation suggests that the enzyme is a constitutive one working probably as an adenosine triphosphatase in a way similar to the kinase isolated from rabbit skeletal muscle. 7. Liver glycine amidinotransferase activity appeared clearly after day 5 of incubation. The specific activity reached a maximum at day 12 and then declined; however, the activity per total mass of liver increased steadily during all the prenatal period. Concomitantly with this steady increase a rise in the creatine content of the whole embryo was observed. An analogous increasing relationship between total liver amidinotransferase activity and liver creatine content was also detected during the postnatal period. 8. Repression of amidinotransferase by creatine cannot be accepted as occurring under physiological conditions since an inverse relationship between the two parameters was not observed. 9. Repression of liver amidinotransferase is observed only when pharmacological concentrations of the exogenous creatine are present in the chick liver. PMID:5493509

  16. Production of Alanine by Fusarium moniliforme

    PubMed Central

    Carito, Sebastian L.; Pisano, Michael A.

    1966-01-01

    Fusarium moniliforme grown in a chemically defined medium in submerged culture accumulated amino acids extracellularly. Alanine and glutamic acid were present in greatest amounts, with traces of glycine, lysine, threonine, and valine detectable. Increasing the glucose and urea concentrations of the medium increased yields of alanine. Further increases in alanine production occurred with elevated levels of mineral salts in the medium, whereas the addition of a vitamin mixture proved to be inhibitory. Chemical changes resulting from the growth of F. moniliforme in the final fermentation medium disclosed maximal alanine production, mycelial weight, and glucose consumption after 72 hr of incubation at 28.5 C. Total soluble nitrogen, by contrast, was minimal at the same time period. The pH remained in the alkaline range throughout the fermentation. PMID:5914495

  17. Structural analysis and mutant growth properties reveal distinctive enzymatic and cellular roles for the three major L-alanine transaminases of Escherichia coli.

    PubMed

    Peña-Soler, Esther; Fernandez, Francisco J; López-Estepa, Miguel; Garces, Fernando; Richardson, Andrew J; Quintana, Juan F; Rudd, Kenneth E; Coll, Miquel; Vega, M Cristina

    2014-01-01

    In order to maintain proper cellular function, the metabolism of the bacterial microbiota presents several mechanisms oriented to keep a correctly balanced amino acid pool. Central components of these mechanisms are enzymes with alanine transaminase activity, pyridoxal 5'-phosphate-dependent enzymes that interconvert alanine and pyruvate, thereby allowing the precise control of alanine and glutamate concentrations, two of the most abundant amino acids in the cellular amino acid pool. Here we report the 2.11-Å crystal structure of full-length AlaA from the model organism Escherichia coli, a major bacterial alanine aminotransferase, and compare its overall structure and active site composition with detailed atomic models of two other bacterial enzymes capable of catalyzing this reaction in vivo, AlaC and valine-pyruvate aminotransferase (AvtA). Apart from a narrow entry channel to the active site, a feature of this new crystal structure is the role of an active site loop that closes in upon binding of substrate-mimicking molecules, and which has only been previously reported in a plant enzyme. Comparison of the available structures indicates that beyond superficial differences, alanine aminotransferases of diverse phylogenetic origins share a universal reaction mechanism that depends on an array of highly conserved amino acid residues and is similarly regulated by various unrelated motifs. Despite this unifying mechanism and regulation, growth competition experiments demonstrate that AlaA, AlaC and AvtA are not freely exchangeable in vivo, suggesting that their functional repertoire is not completely redundant thus providing an explanation for their independent evolutionary conservation.

  18. Solved? The reductive radiation chemistry of alanine.

    PubMed

    Pauwels, Ewald; De Cooman, Hendrik; Waroquier, Michel; Hole, Eli O; Sagstuen, Einar

    2014-02-14

    The structural changes throughout the entire reductive radiation-induced pathway of l-α-alanine are solved on an atomistic level with the aid of periodic DFT and nudged elastic band (NEB) simulations. This yields unprecedented information on the conformational changes taking place, including the protonation state of the carboxyl group in the "unstable" and "stable" alanine radicals and the internal transformation converting these two radical variants at temperatures above 220 K. The structures of all stable radicals were verified by calculating EPR properties and comparing those with experimental data. The variation of the energy throughout the full radiochemical process provides crucial insight into the reason why these structural changes and rearrangements occur. Starting from electron capture, the excess electron quickly localizes on the carbon of a carboxyl group, which pyramidalizes and receives a proton from the amino group of a neighboring alanine molecule, forming a first stable radical species (up to 150 K). In the temperature interval 150-220 K, this radical deaminates and deprotonates at the carboxyl group, the detached amino group undergoes inversion and its methyl group sustains an internal rotation. This yields the so-called "unstable alanine radical". Above 220 K, triggered by the attachment of an additional proton on the detached amino group, the radical then undergoes an internal rotation in the reverse direction, giving rise to the "stable alanine radical", which is the final stage in the reductive radiation-induced decay of alanine.

  19. Creatine supplementation and cognitive performance in elderly individuals.

    PubMed

    McMorris, Terry; Mielcarz, Gregorsz; Harris, Roger C; Swain, Jonathan P; Howard, Alan

    2007-09-01

    The purpose of this study was to examine the effect of creatine supplementation on the cognitive performance of elderly people. Participants were divided into two groups, which were tested on random number generation, forward and backward number and spatial recall, and long-term memory tasks to establish a baseline level. Group 1 (n = 15) were given 5 g four times a day of placebo for 1 week, followed by the same dosage of creatine for the second week. Group 2 (n = 17) were given placebo both weeks. Participants were retested at the end of each week. Results showed a significant effect of creatine supplementation on all tasks except backward number recall. It was concluded that creatine supplementation aids cognition in the elderly.

  20. Temperature dependent Raman and DFT study of creatine.

    PubMed

    Gangopadhyay, Debraj; Sharma, Poornima; Singh, Ranjan K

    2015-01-01

    Temperature dependent Raman spectra of creatine powder have been recorded in the temperature range 420-100K at regular intervals and different clusters of creatine have been optimized using density functional theory (DFT) in order to determine the effect of temperature on the hydrogen bonded network in the crystal structure of creatine. Vibrational assignments of all the 48 normal modes of the zwitterionic form of creatine have been done in terms of potential energy distribution obtained from DFT calculations. Precise analysis gives information about thermal motion and intermolecular interactions with respect to temperature in the crystal lattice. Formation of higher hydrogen bonded aggregates on cooling can be visualized from the spectra through clear signature of phase transition between 200K and 180K.

  1. Creatine Enhances Mitochondrial-Mediated Oligodendrocyte Survival After Demyelinating Injury

    PubMed Central

    Nanescu, Sonia E.

    2017-01-01

    Chronic oligodendrocyte loss, which occurs in the demyelinating disorder multiple sclerosis (MS), contributes to axonal dysfunction and neurodegeneration. Current therapies are able to reduce MS severity, but do not prevent transition into the progressive phase of the disease, which is characterized by chronic neurodegeneration. Therefore, pharmacological compounds that promote oligodendrocyte survival could be beneficial for neuroprotection in MS. Here, we investigated the role of creatine, an organic acid involved in adenosine triphosphate (ATP) buffering, in oligodendrocyte function. We found that creatine increased mitochondrial ATP production directly in oligodendrocyte lineage cell cultures and exerted robust protection on oligodendrocytes by preventing cell death in both naive and lipopolysaccharide-treated mixed glia. Moreover, lysolecithin-mediated demyelination in mice deficient in the creatine-synthesizing enzyme guanidinoacetate-methyltransferase (Gamt) did not affect oligodendrocyte precursor cell recruitment, but resulted in exacerbated apoptosis of regenerated oligodendrocytes in central nervous system (CNS) lesions. Remarkably, creatine administration into Gamt-deficient and wild-type mice with demyelinating injury reduced oligodendrocyte apoptosis, thereby increasing oligodendrocyte density and myelin basic protein staining in CNS lesions. We found that creatine did not affect the recruitment of macrophages/microglia into lesions, suggesting that creatine affects oligodendrocyte survival independently of inflammation. Together, our results demonstrate a novel function for creatine in promoting oligodendrocyte viability during CNS remyelination. SIGNIFICANCE STATEMENT We report that creatine enhances oligodendrocyte mitochondrial function and protects against caspase-dependent oligodendrocyte apoptosis during CNS remyelination. This work has important implications for the development of therapeutic targets for diseases characterized by

  2. Creatine supplementation and oxidative stress in rat liver

    PubMed Central

    2013-01-01

    Background The objective of this study was to determine the effects of creatine supplementation on liver biomarkers of oxidative stress in exercise-trained rats. Methods Forty 90-day-old adult male Wistar rats were assigned to four groups for the eight-week experiment. Control group (C) rats received a balanced control diet; creatine control group (CCr) rats received a balanced diet supplemented with 2% creatine; trained group (T) rats received a balanced diet and intense exercise training equivalent to the maximal lactate steady state phase; and supplemented-trained (TCr) rats were given a balanced diet supplemented with 2% creatine and subjected to intense exercise training equivalent to the maximal lactate steady state phase. At the end of the experimental period, concentrations of creatine, hydrogen peroxide (H2O2) and thiobarbituric acid reactive substances (TBARS) were measured as well as the enzyme activity of superoxide dismutase (SOD), glutathione peroxidase (GSH-GPx) and catalase (CAT). Liver tissue levels of reduced glutathione (GSH), oxidized glutathione (GSSG) and the GSH/GSSG ratio were also determined. Results Hepatic creatine levels were highest in the CCr and TCr groups with increased concentration of H2O2 observed in the T and TCr animal groups. SOD activity was decreased in the TCr group. GSH-GPx activity was increased in the T and TCr groups while CAT was elevated in the CCr and TCr groups. GSH, GGS and the GSH/GSSG ratio did not differ between all animal subsets. Conclusions Our results demonstrate that creatine supplementation acts in an additive manner to physical training to raise antioxidant enzymes in rat liver. However, because markers of liver oxidative stress were unchanged, this finding may also indicate that training-induced oxidative stress cannot be ameliorated by creatine supplementation. PMID:24325803

  3. A systems biology approach to understanding elevated serum alanine transaminase levels in a clinical trial with ximelagatran.

    PubMed

    Andersson, Ulf; Lindberg, Johan; Wang, Shunghuang; Balasubramanian, Raji; Marcusson-Ståhl, Maritha; Hannula, Mira; Zeng, Chenhui; Juhasz, Peter J; Kolmert, Johan; Bäckström, Jonas; Nord, Lars; Nilsson, Kerstin; Martin, Steve; Glinghammar, Björn; Cederbrant, Karin; Schuppe-Koistinen, Ina

    2009-12-01

    Ximelagatran was developed for the prevention and treatment of thromboembolic conditions. However, in long-term clinical trials with ximelagatran, the liver injury marker, alanine aminotransferase (ALT) increased in some patients. Analysis of plasma samples from 134 patients was carried out using proteomic and metabolomic platforms, with the aim of finding predictive biomarkers to explain the ALT elevation. Analytes that were changed after ximelagatran treatment included 3-hydroxybutyrate, pyruvic acid, CSF1R, Gc-globulin, L-glutamine, protein S and alanine, etc. Two of these analytes (pyruvic acid and CSF1R) were studied further in human cell cultures in vitro with ximelagatran. A systems biology approach applied in this study proved to be successful in generating new hypotheses for an unknown mechanism of toxicity.

  4. Creatine metabolism differs between mammals and rainbow trout (Oncorhynchus mykiss).

    PubMed

    Borchel, Andreas; Verleih, Marieke; Rebl, Alexander; Kühn, Carsten; Goldammer, Tom

    2014-01-01

    Creatine plays an important role in the cell as an energy buffer. As the energy system is a basic element of the organism it may possibly contribute to differences between rainbow trout strains selected for the traits growth and robustness, respectively. The cDNA sequences of creatine-related genes encoding glycine amidinotransferase (GATM), guanidinoacetate N-methyltransferase (GAMT), creatine kinase muscle-type (CKM) and creatine transporter 1 (CT1, encoded by gene solute carrier family 6, member 8 (SLC6A8)) were characterized in rainbow trout. Transcripts of the respective genes were quantified in kidney, liver, brain and skeletal muscle in both trout strains that had been acclimated to different temperatures. Several differences between the compared trout strains were found as well as between temperatures indicating that the energy system may contribute to differences between both strains. In addition to that, the expression data showed clear differences between the creatine system in rainbow trout and mammals, as the spatial distribution of the enzyme-encoding gene expression was clearly different from the patterns described for mammals. In rainbow trout, creatine synthesis seems to take place to a big extent in the skeletal muscle.

  5. Creatine supplementation does not improve sprint performance in competitive swimmers.

    PubMed

    Mujika, I; Chatard, J C; Lacoste, L; Barale, F; Geyssant, A

    1996-11-01

    This study was conducted to examine the effects of creatine (Cr) supplementation on sprint swimming performance and energy metabolism. Twenty highly trained swimmers (9 female, 11 male) were tested for blood ammonia and for blood lactate after the 25-, 50-, and 100-m performance in their best stroke on two occasions 7 d apart. After the first trial, subjects were evenly and randomly assigned to either a creatine (5 g creatine monohydrate 4 times per day for 5 d) or a placebo group (same dosage of a lactose placebo) in a double-blind research design. No significant differences in performance times were observed between trials. Post-exercise blood ammonia concentration decreased in the 50- and 100-m trials in the creatine group and in the 50-m trial in the placebo group. The supplementation period had no effect on post-exercise blood lactate. Therefore, creatine supplementation cannot be considered as an ergogenic aid for sprint performance in highly trained swimmers although adenine nucleotide degradation may be reduced during sprint exercise after 5 d of creatine ingestion.

  6. Inhibition of glutamine synthesis induces glutamate dehydrogenase-dependent ammonia fixation into alanine in co-cultures of astrocytes and neurons.

    PubMed

    Dadsetan, Sherry; Bak, Lasse K; Sørensen, Michael; Keiding, Susanne; Vilstrup, Hendrik; Ott, Peter; Leke, Renata; Schousboe, Arne; Waagepetersen, Helle S

    2011-09-01

    It has been previously demonstrated that ammonia exposure of neurons and astrocytes in co-culture leads to net synthesis not only of glutamine but also of alanine. The latter process involves the concerted action of glutamate dehydrogenase (GDH) and alanine aminotransferase (ALAT). In the present study it was investigated if the glutamine synthetase (GS) inhibitor methionine sulfoximine (MSO) would enhance alanine synthesis by blocking the GS-dependent ammonia scavenging process. Hence, co-cultures of neurons and astrocytes were incubated for 2.5h with [U-(13)C]glucose to monitor de novo synthesis of alanine and glutamine in the absence and presence of 5.0 mM NH(4)Cl and 10 mM MSO. Ammonia exposure led to increased incorporation of label but not to a significant increase in the amount of these amino acids. However, in the presence of MSO, glutamine synthesis was blocked and synthesis of alanine increased leading to an elevated content intra- as well as extracellularly of this amino acid. Treatment with MSO led to a dramatic decrease in glutamine content and increased the intracellular contents of glutamate and aspartate. The large increase in alanine during exposure to MSO underlines the importance of the GDH and ALAT biosynthetic pathway for ammonia fixation, and it points to the use of a GS inhibitor to ameliorate the brain toxicity and edema induced by hyperammonemia, events likely related to glutamine synthesis.

  7. Normal cardiac function in mice with supraphysiological cardiac creatine levels.

    PubMed

    Santacruz, Lucia; Hernandez, Alejandro; Nienaber, Jeffrey; Mishra, Rajashree; Pinilla, Miguel; Burchette, James; Mao, Lan; Rockman, Howard A; Jacobs, Danny O

    2014-02-01

    Creatine and phosphocreatine levels are decreased in heart failure, and reductions in myocellular phosphocreatine levels predict the severity of the disease and portend adverse outcomes. Previous studies of transgenic mouse models with increased creatine content higher than two times baseline showed the development of heart failure and shortened lifespan. Given phosphocreatine's role in buffering ATP content, we tested the hypothesis whether elevated cardiac creatine content would alter cardiac function under normal physiological conditions. Here, we report the creation of transgenic mice that overexpress the human creatine transporter (CrT) in cardiac muscle under the control of the α-myosin heavy chain promoter. Cardiac transgene expression was quantified by qRT-PCR, and human CrT protein expression was documented on Western blots and immunohistochemistry using a specific anti-CrT antibody. High-energy phosphate metabolites and cardiac function were measured in transgenic animals and compared with age-matched, wild-type controls. Adult transgenic animals showed increases of 5.7- and 4.7-fold in the content of creatine and free ADP, respectively. Phosphocreatine and ATP levels were two times as high in young transgenic animals but declined to control levels by the time the animals reached 8 wk of age. Transgenic mice appeared to be healthy and had normal life spans. Cardiac morphometry, conscious echocardiography, and pressure-volume loop studies demonstrated mild hypertrophy but normal function. Based on our characterization of the human CrT protein expression, creatine and phosphocreatine content, and cardiac morphometry and function, these transgenic mice provide an in vivo model for examining the therapeutic value of elevated creatine content for cardiac pathologies.

  8. Creatine and guanidinoacetate reference values in a French population.

    PubMed

    Joncquel-Chevalier Curt, Marie; Cheillan, David; Briand, Gilbert; Salomons, Gajja S; Mention-Mulliez, Karine; Dobbelaere, Dries; Cuisset, Jean-Marie; Lion-François, Laurence; Des Portes, Vincent; Chabli, Allel; Valayannopoulos, Vassili; Benoist, Jean-François; Pinard, Jean-Marc; Simard, Gilles; Douay, Olivier; Deiva, Kumaran; Tardieu, Marc; Afenjar, Alexandra; Héron, Delphine; Rivier, François; Chabrol, Brigitte; Prieur, Fabienne; Cartault, François; Pitelet, Gaëlle; Goldenberg, Alice; Bekri, Soumeya; Gerard, Marion; Delorme, Richard; Porchet, Nicole; Vianey-Saban, Christine; Vamecq, Joseph

    2013-11-01

    Creatine and guanidinoacetate are biomarkers of creatine metabolism. Their assays in body fluids may be used for detecting patients with primary creatine deficiency disorders (PCDD), a class of inherited diseases. Their laboratory values in blood and urine may vary with age, requiring that reference normal values are given within the age range. Despite the long known role of creatine for muscle physiology, muscle signs are not necessarily the major complaint expressed by PCDD patients. These disorders drastically affect brain function inducing, in patients, intellectual disability, autistic behavior and other neurological signs (delays in speech and language, epilepsy, ataxia, dystonia and choreoathetosis), being a common feature the drop in brain creatine content. For this reason, screening of PCDD patients has been repeatedly carried out in populations with neurological signs. This report is aimed at providing reference laboratory values and related age ranges found for a large scale population of patients with neurological signs (more than 6 thousand patients) previously serving as a background population for screening French patients with PCDD. These reference laboratory values and age ranges compare rather favorably with literature values for healthy populations. Some differences are also observed, and female participants are discriminated from male participants as regards to urine but not blood values including creatine on creatinine ratio and guanidinoacetate on creatinine ratio values. Such gender differences were previously observed in healthy populations; they might be explained by literature differential effects of testosterone and estrogen in adolescents and adults, and by estrogen effects in prepubertal age on SLC6A8 function. Finally, though they were acquired on a population with neurological signs, the present data might reasonably serve as reference laboratory values in any future medical study exploring abnormalities of creatine metabolism and

  9. Observed activities of serum creatine kinase: total and B subunit activity and other enzymes in young persons abusing solvents.

    PubMed Central

    Spooner, R J; Birrell, R C; McLelland, A S; Baird, J A; Sourindhrin, I

    1984-01-01

    Serum enzymes (aspartate transaminase, alanine transaminase, alkaline phosphatase (ALP), gamma-glutamyltransferase, and creatine kinase (CK] were measured in 296 young persons who admitted to recent inhalation of solvents, usually toluene based glues. In general, results fell within expected adult reference ranges except for ALP and CK. About 60% of subjects had CK activities above the upper reference limit and these activities were investigated in terms of their isoenzyme composition. CK B subunit activity was measured in 90 subjects with raised total CK activities. In five instances the CK B subunit activity was judged abnormal and in two subjects the presence of CK BB was confirmed. These two subjects were thought to have a circulating macro CK, type 1. It is concluded that the increased total CK activity found in this group of solvent abusers was due to physical activity, but a contribution from specific muscle toxicity by solvents cannot be excluded. PMID:6142904

  10. Can creatine supplementation form carcinogenic heterocyclic amines in humans?

    PubMed Central

    Pereira, Renato Tavares dos Santos; Dörr, Felipe Augusto; Pinto, Ernani; Solis, Marina Yazigi; Artioli, Guilherme Giannini; Fernandes, Alan Lins; Murai, Igor Hisashi; Dantas, Wagner Silva; Seguro, Antônio Carlos; Santinho, Mirela Aparecida Rodrigues; Roschel, Hamilton; Carpentier, Alain; Poortmans, Jacques Remi; Gualano, Bruno

    2015-01-01

    Abstract Creatine supplementation has been associated with increased cancer risk. In fact, there is evidence indicating that creatine and/or creatinine are important precursors of carcinogenic heterocyclic amines (HCAs). The present study aimed to investigate the acute and chronic effects of low- and high-dose creatine supplementation on the production of HCAs in healthy humans (i.e. 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (8-MeIQx),  2-amino-(1,6-dimethylfuro[3,2-e]imidazo[4,5-b])pyridine (IFP) and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4,8-DiMeIQx)). This was a non-counterbalanced single-blind crossover study divided into two phases, in which low- and high-dose creatine protocols were tested. After acute (1 day) and chronic supplementation (30 days), the HCAs PhIP, 8-MeIQx, IFP and 4,8-DiMeIQx were assessed through a newly developed HPLC–MS/MS method. Dietary HCA intake and blood and urinary creatinine were also evaluated. Out of 576 assessments performed (from 149 urine samples), only nine (3 from creatine and 6 from placebo) showed quantifiable levels of HCAs (8-MeIQx: n = 3; 4,8-DiMeIQx: n = 2; PhIP: n = 4). Individual analyses revealed that diet rather than creatine supplementation was the main responsible factor for HCA formation in these cases. This study provides compelling evidence that both low and high doses of creatine supplementation, given either acutely or chronically, did not cause increases in the carcinogenic HCAs PhIP, 8-MeIQx, IFP and 4,8-DiMeIQx in healthy subjects. These findings challenge the long-existing notion that creatine supplementation could potentially increase the risk of cancer by stimulating the formation of these mutagens. Key points There is a long-standing concern that creatine supplementation could be associated with cancer, possibly by facilitating the formation of carcinogenic heterocyclic amines (HCAs). This study provides compelling evidence

  11. Macro-creatine kinase: a neglected cause of elevated creatine kinase.

    PubMed

    Aljuani, F; Tournadre, A; Cecchetti, S; Soubrier, M; Dubost, J J

    2015-04-01

    Macro-creatine kinase (macro-CK) is a neglected cause of raised CK. Over a 10-year period, we observed five cases. Three patients had macro-CK type 1. One patient with fibromyalgia underwent several explorations to find a muscular pathology; another, who had elevated CK-MB (muscle-brain fraction) activity, was referred to a cardiologist, and statin therapy was erroneously discontinued in two patients. Two patients had macro-CK type 2: a man with a neuroendocrine carcinoma and a woman with rheumatoid arthritis. Diagnosis of type 1 obviates the need to carry out pointless and expensive investigations seeking a neuromuscular or cardiac pathology, and also, the unwarranted discontinuation of statin therapy. Type 2 must prompt investigations for a neoplasm.

  12. 21 CFR 582.5118 - Alanine.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Alanine. 582.5118 Section 582.5118 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  13. 21 CFR 582.5118 - Alanine.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Alanine. 582.5118 Section 582.5118 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  14. 21 CFR 582.5118 - Alanine.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Alanine. 582.5118 Section 582.5118 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  15. 21 CFR 582.5118 - Alanine.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Alanine. 582.5118 Section 582.5118 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  16. 21 CFR 582.5118 - Alanine.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Alanine. 582.5118 Section 582.5118 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  17. Meta-analysis of Creatine for neuroprotection against Parkinson's disease.

    PubMed

    Attia, Attia; Ahmed, Hussien; Gadelkarim, Mohamed; Morsi, Mahmoud; Awad, Kamal; Elnenny, Mohamed; Ghanem, Esraa; El-Jafaary, Shaimaa; Negida, Ahmed

    2016-11-04

    Background Creatine is an antioxidant agent that showed neuroprotective effects in animal models of Parkinson's disease (PD). Creatine was selected by the National Institute of Neurological Disorders and Stroke as a possible disease modifying agent for Parkinson's disease. Therefore, many clinical trials evaluated the efficacy of creatine for patients with PD. The aim of this systematic review and meta-analysis is to synthesize evidence from published randomized controlled trials (RCTs) about the efficacy of Creatine for patients with PD. Methods We followed PRISMA statement guidelines during the preparation of this systematic review and meta-analysis. A computer literature search for PubMed, EBSCO, web of science and Ovid Midline was carried out. We included RCTs comparing creatine with placebo in terms of motor functions and quality of life. Outcomes of total Unified Parkinson's Disease Rating Scale (UPDRS), UPDRS I, UPDRS II, and UPDRS III were pooled as mean difference (MD) between two groups from baseline to the endpoint. Statistical heterogeneity was assessed by visual inspection of the forest plot and measured by chi-square and I square tests. Results Three RCTs (n=1935) were included in this study. The overall effect did not favor either of the two groups in terms of: UPDRS total score (MD 1.07, 95% CI [3.38 to 1.25], UPDRS III (MD 0.62, 95% CI [2.27 to 1.02]), UPDRS II (MD 0.03, 95% CI [0.81 to 0.86], or UPDRS I (MD 0.03, 95% CI [0.33 to 0.28]). Conclusion Current evidence does not support the use of creatine for neuroprotection against PD. Future well-designed, randomized controlled trials are needed.

  18. Creatine supplementation, sleep deprivation, cortisol, melatonin and behavior.

    PubMed

    McMorris, T; Harris, R C; Howard, A N; Langridge, G; Hall, B; Corbett, J; Dicks, M; Hodgson, C

    2007-01-30

    The effect of creatine supplementation and sleep deprivation, with intermittent moderate-intensity exercise, on cognitive and psychomotor performance, mood state, effort and salivary concentrations of cortisol and melatonin were examined. Subjects were divided into a creatine supplementation group and a placebo group. They took 5 g of creatine monohydrate or a placebo, dependent on their group, four times a day for 7 days immediately prior to the experiment. They undertook tests examining central executive functioning, short-term memory, choice reaction time, balance, mood state and effort at baseline and following 18-, 24- and 36-h sleep deprivation, with moderate intermittent exercise. Saliva samples were taken prior to each set of tests. A group x time analysis of covariance, with baseline performance the covariate, showed that the creatine group performed significantly (p < 0.05) better than the placebo group on the central executive task but only at 36 h. The creatine group demonstrated a significant (p < 0.01) linear improvement in performance of the central executive task throughout the experiment, while the placebo group showed no significant effects. There were no significant differences between the groups for any of the other variables. A significant (p < 0.001) main effect of time was found for the balance test with a linear improvement being registered. Cortisol concentrations on Day 1 were significantly (p < 0.01) higher than on Day 2. Mood significantly (p < 0.001) deteriorated up to 24 h with no change from 24 to 36 h. Effort at baseline was significantly (p < 0.01) lower than in the other conditions. It was concluded that, during sleep deprivation with moderate-intensity exercise, creatine supplementation only affects performance of complex central executive tasks.

  19. Creatine kinase MB isoenzyme in dermatomyositis: a noncardiac source

    SciTech Connect

    Larca, L.J.; Coppola, J.T.; Honig, S.

    1981-03-01

    Three patients with polymyositis had elevated serum levels of creatine kinase MB isoenzyme. The presence of this isoenzyme is used extensively to diagnose myocardial infarction, but the isoenzyme is also found in sera of patients with primary muscular and neuromuscular disorders. Researchers studied cardiac function in two of our patients with electrocardiograms, technetium stannous pyrophosphate scanning, and technetium 99m-labeled erythrocyte gated blood pool imaging and in the third patient by postmortem examination. There was no evidence of myocardial involvement to account for the high serum levels of isoenzyme. Creatine kinase MB in the sera of patients with polymyositis does not necessarily indicate myocardial necrosis.

  20. Biocatalytic potential of vanillin aminotransferase from Capsicum chinense

    PubMed Central

    2014-01-01

    Background The conversion of vanillin to vanillylamine is a key step in the biosynthetic route towards capsaicinoids in pungent cultivars of Capsicum sp. The reaction has previously been annotated to be catalysed by PAMT (putative aminotransferase; [GenBank: AAC78480.1, Swiss-Prot: O82521]), however, the enzyme has previously not been biochemically characterised in vitro. Results The biochemical activity of the transaminase was confirmed by direct measurement of the reaction with purified recombinant enzyme. The enzyme accepted pyruvate, and oxaloacetate but not 2-oxoglutarate as co-substrate, which is in accordance with other characterised transaminases from the plant kingdom. The enzyme was also able to convert (S)-1-phenylethylamine into acetophenone with high stereo-selectivity. Additionally, it was shown to be active at a broad pH range. Conclusions We suggest PAMT to be renamed to VAMT (vanillin aminotransferase, abbreviation used in this study) as formation of vanillin from vanillylamine could be demonstrated. Furthermore, due to high stereoselectivity and activity at physiological pH, VAMT is a suitable candidate for biocatalytic transamination in a recombinant whole-cell system. PMID:24712445

  1. Brain alanine formation as an ammonia-scavenging pathway during hyperammonemia: effects of glutamine synthetase inhibition in rats and astrocyte-neuron co-cultures.

    PubMed

    Dadsetan, Sherry; Kukolj, Eva; Bak, Lasse K; Sørensen, Michael; Ott, Peter; Vilstrup, Hendrik; Schousboe, Arne; Keiding, Susanne; Waagepetersen, Helle S

    2013-08-01

    Hyperammonemia is a major etiological toxic factor in the development of hepatic encephalopathy. Brain ammonia detoxification occurs primarily in astrocytes by glutamine synthetase (GS), and it has been proposed that elevated glutamine levels during hyperammonemia lead to astrocyte swelling and cerebral edema. However, ammonia may also be detoxified by the concerted action of glutamate dehydrogenase (GDH) and alanine aminotransferase (ALAT) leading to trapping of ammonia in alanine, which in vivo likely leaves the brain. Our aim was to investigate whether the GS inhibitor methionine sulfoximine (MSO) enhances incorporation of (15)NH4(+) in alanine during acute hyperammonemia. We observed a fourfold increased amount of (15)NH4 incorporation in brain alanine in rats treated with MSO. Furthermore, co-cultures of neurons and astrocytes exposed to (15)NH4Cl in the absence or presence of MSO demonstrated a dose-dependent incorporation of (15)NH4 into alanine together with increased (15)N incorporation in glutamate. These findings provide evidence that ammonia is detoxified by the concerted action of GDH and ALAT both in vivo and in vitro, a mechanism that is accelerated in the presence of MSO thereby reducing the glutamine level in brain. Thus, GS could be a potential drug target in the treatment of hyperammonemia in patients with hepatic encephalopathy.

  2. A creatine-driven substrate cycle enhances energy expenditure and thermogenesis in beige fat.

    PubMed

    Kazak, Lawrence; Chouchani, Edward T; Jedrychowski, Mark P; Erickson, Brian K; Shinoda, Kosaku; Cohen, Paul; Vetrivelan, Ramalingam; Lu, Gina Z; Laznik-Bogoslavski, Dina; Hasenfuss, Sebastian C; Kajimura, Shingo; Gygi, Steve P; Spiegelman, Bruce M

    2015-10-22

    Thermogenic brown and beige adipose tissues dissipate chemical energy as heat, and their thermogenic activities can combat obesity and diabetes. Herein the functional adaptations to cold of brown and beige adipose depots are examined using quantitative mitochondrial proteomics. We identify arginine/creatine metabolism as a beige adipose signature and demonstrate that creatine enhances respiration in beige-fat mitochondria when ADP is limiting. In murine beige fat, cold exposure stimulates mitochondrial creatine kinase activity and induces coordinated expression of genes associated with creatine metabolism. Pharmacological reduction of creatine levels decreases whole-body energy expenditure after administration of a β3-agonist and reduces beige and brown adipose metabolic rate. Genes of creatine metabolism are compensatorily induced when UCP1-dependent thermogenesis is ablated, and creatine reduction in Ucp1-deficient mice reduces core body temperature. These findings link a futile cycle of creatine metabolism to adipose tissue energy expenditure and thermal homeostasis. PAPERCLIP.

  3. 21 CFR 172.540 - DL-Alanine.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true DL-Alanine. 172.540 Section 172.540 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN... Agents and Related Substances § 172.540 DL-Alanine. DL-Alanine (a racemic mixture of D- and...

  4. On the existence of ‘L-alanine cadmium bromide'

    NASA Astrophysics Data System (ADS)

    Srinivasan, Bikshandarkoil R.

    2013-12-01

    It is argued that the recently reported nonlinear optical crystal L-alanine cadmium bromide, grown by slow solvent evaporation method at room temperature [P. Ilayabarathi, J. Chandrasekaran, Spectrochim. Acta 96A (2012) 684-689] is the well-known L-alanine crystal. The isolation of L-alanine crystal is explained due to fractional crystallization.

  5. On the existence of 'L-alanine cadmium bromide'.

    PubMed

    Srinivasan, Bikshandarkoil R

    2013-12-01

    It is argued that the recently reported nonlinear optical crystal L-alanine cadmium bromide, grown by slow solvent evaporation method at room temperature [P. Ilayabarathi, J. Chandrasekaran, Spectrochim. Acta 96A (2012) 684-689] is the well-known L-alanine crystal. The isolation of L-alanine crystal is explained due to fractional crystallization.

  6. Prevalence and Predictors of Elevated Aspartate Aminotransferase-to-Platelet Ratio Index in Latin American Perinatally HIV-infected Children

    PubMed Central

    Siberry, George K.; Cohen, Rachel A.; Harris, D. Robert; Cruz, Maria Leticia Santos; Oliveira, Ricardo; Peixoto, Mario F.; Cervi, Maria Celia; Hazra, Rohan; Pinto, Jorge A.

    2013-01-01

    Background Chronic liver disease has emerged as an important problem in adults with longstanding HIV infection, but data are lacking for children. We characterized elevated aspartate aminotransferase (AST)-to-platelet ratio index (APRI ), a marker of possible liver fibrosis, in perinatally HIV-infected children. Methods NISDI [NICHD (National Institute of Child Health and Human Development) International Site Development Initiative] enrolled HIV-infected children (ages 0.1-20.1 years) from five Latin American countries in an observational cohort from 2002–2009. Twice yearly visits included medical history, physical examination and laboratory evaluations. The prevalence (95% confidence interval [CI]) of APRI>1.5 was calculated and associations with demographic, HIV-related and liver-related variables were investigated in bivariate analyses. Results APRI was available for 1012 of 1032 children. APRI was >1.5 in 32 (3.2%, 95% CI: 2.2%-4.4%) including 2 of 4 participants with hepatitis B (HBV) infection. Factors significantly associated with APRI>1.5 (p<0.01 compared to APRI≤1.5) included country, younger age, past or current HBV, higher alanine aminotransferase, lower total cholesterol, higher log10 current viral load, lower current CD4 count, lower nadir CD4 count, use of hepatotoxic non-antiretroviral (ARV) medications, and no prior ARV use. Rates of APRI>1.5 varied significantly by current ARV regimen (p=0.0002), from 8.0% for no ARV to 3.2% for non-protease inhibitor (PI) regimens to 1.5% for PI-based regimens. Conclusions Elevated APRI occurred in approximately 3% of perinatally HIV-infected children. PI-based ARVs appeared protective while inadequate HIV control appeared to increase risk of elevated APRI. Additional investigations are needed to better assess potential subclinical, chronic liver disease in HIV-infected children. PMID:23799515

  7. Concerted modulation of alanine and glutamate metabolism in young Medicago truncatula seedlings under hypoxic stress.

    PubMed

    Limami, Anis M; Glévarec, Gaëlle; Ricoult, Claudie; Cliquet, Jean-Bernard; Planchet, Elisabeth

    2008-01-01

    The modulation of primary nitrogen metabolism by hypoxic stress was studied in young Medicago truncatula seedlings. Hypoxic seedlings were characterized by the up-regulation of glutamate dehydrogenase 1 (GDH1) and mitochondrial alanine aminotransferase (mAlaAT), and down-regulation of glutamine synthetase 1b (GS1b), NADH-glutamate synthase (NADH-GOGAT), glutamate dehydrogenase 3 (GDH3), and isocitrate dehydrogenase (ICDH) gene expression. Hypoxic stress severely inhibited GS activity and stimulated NADH-GOGAT activity. GDH activity was lower in hypoxic seedlings than in the control, however, under either normoxia or hypoxia, the in vivo activity was directed towards glutamate deamination. (15)NH(4) labelling showed for the first time that the adaptive reaction of the plant to hypoxia consisted of a concerted modulation of nitrogen flux through the pathways of both alanine and glutamate synthesis. In hypoxic seedlings, newly synthesized (15)N-alanine increased and accumulated as the major amino acid, asparagine synthesis was inhibited, while (15)N-glutamate was synthesized at a similar rate to that in the control. A discrepancy between the up-regulation of GDH1 expression and the down-regulation of GDH activity by hypoxic stress highlighted for the first time the complex regulation of this enzyme by hypoxia. Higher rates of glycolysis and ethanol fermentation are known to cause the fast depletion of sugar stores and carbon stress. It is proposed that the expression of GDH1 was stimulated by hypoxia-induced carbon stress, while the enzyme protein might be involved during post-hypoxic stress contributing to the regeneration of 2-oxoglutarate via the GDH shunt.

  8. Radioimmunoassay measurement of creatine kinase bb in the serum of schizophrenic patients

    SciTech Connect

    Lerner, M.H.; Friedhoff, A.J.

    1980-03-03

    Brain type creatine kinase (BB) isoenzyme was measured using a highly sensitive and specific radioimmunoassay procedure in two schizophrenic populations. The data would indicate that in the schizophrenic populations examined there is insufficient tissue disruption to cause abnormal build-up of brain creatine kinase levels. However the possibility of a rapid removal of creatine kinase BB from the circulation exists. The elevated creatine kinase reported in acute schizophrenics is most likely not of brain origin.

  9. Influence of Asymptomatic Pneumonia on the Response to Hemorrhage and Resuscitation in Swine

    DTIC Science & Technology

    2010-01-01

    dehydrogenase (LDH), aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatine kinase (CK), amylase and lactate (Vitros Chemistry System...CK. Creatinine increased at 15 min in both groups and remained elevated throughout the study. Mean total protein, amylase and ALT decreased similarly

  10. Creatine Loading, Resistance Exercise Performance, and Muscle Mechanics.

    ERIC Educational Resources Information Center

    Stevenson, Scott W.; Dudley, Gary A.

    2001-01-01

    Examined whether creatine (CR) monohydrate loading would alter resistance exercise performance, isometric strength, or in vivo contractile properties of the quadriceps femoris muscle compared with placebo loading in resistance-trained athletes. Overall, CR loading did not provide an ergogenic benefit for the unilateral dynamic knee extension…

  11. 21 CFR 862.1210 - Creatine test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Creatine test system. 862.1210 Section 862.1210 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 Systems §...

  12. 21 CFR 862.1210 - Creatine test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Creatine test system. 862.1210 Section 862.1210 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 Systems §...

  13. 21 CFR 862.1210 - Creatine test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Creatine test system. 862.1210 Section 862.1210 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 Systems §...

  14. 21 CFR 862.1210 - Creatine test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Creatine test system. 862.1210 Section 862.1210 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 Systems §...

  15. 21 CFR 862.1210 - Creatine test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Creatine test system. 862.1210 Section 862.1210 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 Systems §...

  16. Creatine Revealed Anticonvulsant Properties on Chemically and Electrically Induced Seizures in Mice.

    PubMed

    Shafaroodi, Hamed; Shahbek, Farnaz; Faizi, Mehrdad; Ebrahimi, Farzad; Moezi, Leila

    2016-01-01

    Creatine exerts beneficial effects on a variety of pathologies in which energy metabolism and oxidative stress play an etiological role. Creatine supplements have shown beneficial effects on neurological disorders including Parkinson׳s disease, Huntington›s disease, amyotrophic lateral sclerosis, as well as Alzheimer›s disease and stroke. However, the potential benefits of creatine for patients with convulsive disorders remain poorly defined. While some authors did not suggest any anti- or pro-convulsant roles for creatine treatment, others suggest that creatine may be an anticonvulsant agent. In this study, we investigated the effects of creatine on seizures in mice. Three models were used to explore the role of creatine on seizures in mice including intravenous pentylenetetrazole (PTZ), intraperitoneal PTZ, and electroshock models. Acute creatine treatment (10, 20, 40 and 80 mg/Kg) significantly increased the clonic seizure threshold in the intravenous PTZ model. Sub-chronic administration of creatine (10 and 20 mg/Kg) revealed a significant anticonvulsant effect in intravenous PTZ model. Acute creatine administration (10, 20 and 40 mg/Kg) significantly decreased the frequency of clonic seizures in the intraperitoneal PTZ model. Besides, acute creatine (40 and 80 mg/Kg) decreased the incidence of tonic seizures after electroshock. In conclusion, creatine exerts anticonvulsant effects in three seizure models; therefore, it may act as a potential drug to help patients with convulsions. However, further investigations should be done to clarify these results more.

  17. Creatine Revealed Anticonvulsant Properties on Chemically and Electrically Induced Seizures in Mice

    PubMed Central

    Shafaroodi, Hamed; Shahbek, Farnaz; Faizi, Mehrdad; Ebrahimi, Farzad; Moezi, Leila

    2016-01-01

    Creatine exerts beneficial effects on a variety of pathologies in which energy metabolism and oxidative stress play an etiological role. Creatine supplements have shown beneficial effects on neurological disorders including Parkinson׳s disease, Huntington›s disease, amyotrophic lateral sclerosis, as well as Alzheimer›s disease and stroke. However, the potential benefits of creatine for patients with convulsive disorders remain poorly defined. While some authors did not suggest any anti- or pro-convulsant roles for creatine treatment, others suggest that creatine may be an anticonvulsant agent. In this study, we investigated the effects of creatine on seizures in mice. Three models were used to explore the role of creatine on seizures in mice including intravenous pentylenetetrazole (PTZ), intraperitoneal PTZ, and electroshock models. Acute creatine treatment (10, 20, 40 and 80 mg/Kg) significantly increased the clonic seizure threshold in the intravenous PTZ model. Sub-chronic administration of creatine (10 and 20 mg/Kg) revealed a significant anticonvulsant effect in intravenous PTZ model. Acute creatine administration (10, 20 and 40 mg/Kg) significantly decreased the frequency of clonic seizures in the intraperitoneal PTZ model. Besides, acute creatine (40 and 80 mg/Kg) decreased the incidence of tonic seizures after electroshock. In conclusion, creatine exerts anticonvulsant effects in three seizure models; therefore, it may act as a potential drug to help patients with convulsions. However, further investigations should be done to clarify these results more. PMID:28243281

  18. Characterization of dehydration behavior of untreated and pulverized creatine monohydrate powders.

    PubMed

    Sakata, Yukoh; Shiraishi, Sumihiro; Otsuka, Makoto

    2004-06-01

    Creatine, which is well known as an important substance for muscular activity, is synthesized from amino acids such as glycine, arginine and ornithine in liver and kidney. It then accumulates in skeletal muscle as creatine phosphoric acid. The aim of this study was to understand the dehydration behavior of untreated and pulverized creatine monohydrate at various temperatures. The removal of crystal water was investigated by using differential scanning calorimetry (DSC), X-ray powder diffraction and scanning electron microscopy (SEM). The X-ray diffraction pattern of untreated and pulverized creatine monohydrate agreed with reported data for creatine monohydrate. However, the diffraction peaks of the (100), (200) and (300) planes of pulverized creatine monohydrate were much stronger than those of untreated creatine monohydrate. On the other hand, the diffraction peaks of the (012) and (013) planes of untreated creatine monohydrate were much stronger than those of pulverized creatine monohydrate. The dehydration of untreated and pulverized creatine monohydrate was investigated at various storage temperatures, and the results indicated that untreated and pulverized creatine monohydrate were transformed into the anhydrate at more than 30 degrees C. After dehydration, the particles of untreated and pulverized creatine anhydrate had many cracks. The dehydration kinetics of untreated and pulverized creatine monohydrate were analyzed by the Hancock-Sharp equation on the basis of the isothermal DSC data. The dehydrations of untreated and pulverized creatine monohydrate both followed a zero-order mechanism (Polany-Winger equation). However, the transition rate constant, calculated from the slope of the straight line, was about 2.2-7.7 times higher for pulverized creatine monohydrate than for untreated creatine monohydrate. The Arrhenius plots (natural logarithm of the dehydration rate constant versus the reciprocal of absolute temperature) of the isothermal DSC data for

  19. Simultaneous determination of creatine phosphate, creatine and 12 nucleotides in rat heart by LC-MS/MS.

    PubMed

    Wang, Jun-mei; Chu, Yang; Li, Wei; Wang, Xiang-yang; Guo, Jia-hua; Yan, Lu-lu; Ma, Xiao-hui; Ma, Ying-li; Yin, Qi-hui; Liu, Chang-xiao

    2014-05-01

    A simple, rapid and sensitive LC-MS/MS method was developed and validated for simultaneous determination of creatine phosphate (CP), creatine (Cr) and 12 nucleotides in rat heart. The analytes, ATP, ADP, AMP, GTP, GDP, GMP, CTP, CDP, CMP, UTP, UDP, UMP, CP, Cr, were extracted from heart tissue with pre-cooled (0°C) methanol/water (1:1, v/v) and separated on a Hypersil Gold AQ C18 column (150mm×4.6mm, 3μm) using an isocratic elution with a mobile phase consisting of 2mmol/L ammonium acetate in water (pH 10.0, adjusted with ammonia). The detection was performed by negative ion electrospray ionization in selective reaction monitoring mode (SRM). In the assay, all the analytes showed good linearity over the investigated concentration range (r>0.99). The accuracy was between 80.7% and 120.6% and the precision expressed in RSD was less than 15.6%. This method was successfully applied to measure the concentrations of the 12 nucleotides, creatine phosphate and creatine in rat heart for the first time.

  20. Synthesis and evaluation of novel heteroaromatic substrates of GABA aminotransferase

    PubMed Central

    Hawker, Dustin D.; Silverman, Richard B.

    2012-01-01

    Two principal neurotransmitters are involved in the regulation of mammalian neuronal activity, namely, γ-aminobutyric acid (GABA), an inhibitory neurotransmitter, and L-glutamic acid, an excitatory neurotransmitter. Low GABA levels in the brain have been implicated in epilepsy and several other neurological diseases. Because of GABA’s poor ability to cross the blood-brain barrier (BBB), a successful strategy to raise brain GABA concentrations is the use of a compound that does cross the BBB and inhibits or inactivates GABA aminotransferase (GABA-AT), the enzyme responsible for GABA catabolism. Vigabatrin, a mechanism-based inactivator of GABA-AT, is currently a successful therapeutic for epilepsy, but has harmful side effects, leaving a need for improved GABA-AT inactivators. Here, we report the synthesis and evaluation of a series of heteroaromatic GABA analogues as substrates of GABA-AT, which will be used as the basis for the design of novel enzyme inactivators. PMID:22944334

  1. Ambient Ionization Mass Spectrometry Measurement of Aminotransferase Activity

    NASA Astrophysics Data System (ADS)

    Yan, Xin; Li, Xin; Zhang, Chengsen; Xu, Yang; Cooks, R. Graham

    2017-01-01

    A change in enzyme activity has been used as a clinical biomarker for diagnosis and is useful in evaluating patient prognosis. Current laboratory measurements of enzyme activity involve multi-step derivatization of the reaction products followed by quantitative analysis of these derivatives. This study simplified the reaction systems by using only the target enzymatic reaction and directly detecting its product. A protocol using paper spray mass spectrometry for identifying and quantifying the reaction product has been developed. Evaluation of the activity of aspartate aminotransferase (AST) was chosen as a proof-of-principle. The volume of sample needed is greatly reduced compared with the traditional method. Paper spray has a desalting effect that avoids sprayer clogging problems seen when examining serum samples by nanoESI. This very simple method does not require sample pretreatment and additional derivatization reactions, yet it gives high quality kinetic data, excellent limits of detection (60 ppb from serum), and coefficients of variation <10% in quantitation.

  2. Contribution of creatine kinase MB mass concentration at admission to early diagnosis of acute myocardial infarction.

    PubMed Central

    Bakker, A J; Gorgels, J P; van Vlies, B; Koelemay, M J; Smits, R; Tijssen, J G; Haagen, F D

    1994-01-01

    OBJECTIVE--To assess the diagnostic value at admission of creatine kinase MB mass concentration, alone or in combination with electrocardiographic changes, in suspected myocardial infarction. DESIGN--Prospective study of all consecutive patients admitted within 12 hours after onset of chest pain to a coronary care unit for evaluation of suspected myocardial infarction. SETTING--Large regional hospital. PATIENTS--In 297 patients creatine kinase and creatine kinase MB activities and creatine kinase MB mass concentration were determined. Myocardial infarction according to the criteria of the World Health Organisation was diagnosed in 154 patients and excluded in 143 patients (including 70 with unstable angina pectoris). RESULTS--Sensitivity/specificity for creatine kinase MB mass concentration in patients admitted within 4 hours and 4-12 hours after onset of chest pain were 45%/94% and 76%/79% respectively. Corresponding values for creatine kinase activity were 20%/89% and 59%/83%, and for creatine kinase MB activity 16%/87% and 53%/87%. Raised creatine kinase MB mass concentration was seen in 17% of patients with unstable angina pectoris. Stepwise logistic regression analysis showed that independent predictors of acute myocardial infarction in patients admitted within 4 hours after onset of chest pain were electrocardiographic changes and creatine kinase MB mass concentration on admission; in patients admitted 4-12 hours after the onset of pain independent predictors were electrocardiographic changes and creatine kinase MB mass concentration and activity. CONCLUSION--Creatine kinase MB mass concentration is a more sensitive marker for myocardial infarction than the activity of creatine kinase and its MB isoenzyme. Electrocardiographic changes on admission in combination with creatine kinase MB mass concentration (instead of creatine kinase and creatine kinase MB activities) are best in diagnosing myocardial infarction. PMID:7917680

  3. Unique substrate specificity of ornithine aminotransferase from Toxoplasma gondii.

    PubMed

    Astegno, Alessandra; Maresi, Elena; Bertoldi, Mariarita; La Verde, Valentina; Paiardini, Alessandro; Dominici, Paola

    2017-03-07

    Toxoplasma gondii is a protozoan parasite of medical and veterinary relevance responsible for toxoplasmosis in humans. As an efficacious vaccine remains a challenge, chemotherapy is still the most effective way to combat the disease. In search of novel druggable targets, we performed a thorough characterization of the putative pyridoxal 5'-phosphate (PLP)-dependent enzyme ornithine aminotransferase from T. gondii ME49 (TgOAT). We overexpressed the protein in Escherichia coli and analysed its molecular and kinetic properties by UV-visible absorbance, fluorescence and CD spectroscopy, in addition to kinetic studies of both the steady state and pre-steady state. TgOAT is largely similar to OATs from other species regarding its general transamination mechanism and spectral properties of PLP; however, it does not show a specific ornithine aminotransferase activity like its human homologue, but exhibits both N-acetylornithine and γ-aminobutyric acid (GABA) transaminase activity in vitro, suggesting a role in both arginine and GABA metabolism in vivo The presence of Val79 in the active site of TgOAT in place of Tyr, as in its human counterpart, provides the necessary room to accommodate N-acetylornithine and GABA, resembling the active site arrangement of GABA transaminases. Moreover, mutation of Val79 to Tyr results in a change of substrate preference between GABA, N-acetylornithine and L-ornithine, suggesting a key role of Val79 in defining substrate specificity. The findings that TgOAT possesses parasite-specific structural features as well as differing substrate specificity from its human homologue make it an attractive target for anti-toxoplasmosis inhibitor design that can be exploited for chemotherapeutic intervention.

  4. Effect of pulverization on hydration kinetic behaviors of creatine anhydrate powders.

    PubMed

    Sakata, Yukoh; Shiraishi, Sumihiro; Otsuka, Makoto

    2004-12-25

    The crystal orientation of creatine monohydrate varies significantly with tableting performance and pulverizing mechanism. Furthermore, the X-ray diffraction patterns of anhydrous forms of untreated creatine monohydrate and of pulverized creatine monohydrate exhibit different crystal orientations. However, hygroscopic forms of unpulverized creatine anhydrate and pulverized creatine anhydrate was exhibit the same diffraction peak pattern. The hygroscopicity of unpulverized and pulverized creatine anhydrate has been investigated by hydration kinetic methods using isothermal differential scanning calorimetry data. Testing of the hygroscopicity of unpulverized and pulverized creatine anhydrate at various levels of relative humidity (RH) at 25 degrees C revealed that the anhydrate was stable at less than 33% RH, but was transformed into the monohydrate at more than 52% RH. Hydration data of unpulverized and pulverized creatine anhydrate at 60% and 75% RH were calculated to determine hydration kinetics using various solid-state kinetic models. The hydration type of unpulverized and pulverized creatine anhydrate powder follows the zero-order mechanism (Polany-Winger equation) R1. The transition rate constant of pulverized creatine anhydrate, calculated from the slope of the straight line, was about 1.34-1.36 times higher than that of unpulverized creatine anhydrate.

  5. Creatine protects against mitochondrial dysfunction associated with HIV-1 Tat-induced neuronal injury

    PubMed Central

    Stevens, Patrick R.; Gawryluk, Jeremy W.; Hui, Liang; Chen, Xuesong; Geiger, Jonathan D.

    2015-01-01

    HIV-1 infected individuals are living longer but experiencing a prevalence rate of over 50% for HIV-1 associated neurocognitive disorders (HAND) for which no effective treatment is available. Viral and cellular factors secreted by HIV-1 infected cells leads to neuronal injury and HIV-1 Tat continues to be implicated in the pathogenesis of HAND. Here we tested the hypothesis that creatine protected against HIV-1 Tat-induced neuronal injury by preventing mitochondrial bioenergetic crisis and/or redox catastrophe. Creatine blocked HIV-1 Tat1-72-induced increases in neuron cell death and synaptic area loss. Creatine protected against HIV-1 Tat-induced decreases in ATP. Creatine and creatine plus HIV-1 Tat increased cellular levels of creatine, and creatine plus HIV-1 Tat further decreased ratios of phosphocreatine to creatine observed with creatine or HIV-1 Tat treatments alone. Additionally, creatine protected against HIV-1 Tat-induced mitochondrial hypopolarization and HIV-1 Tat-induced mitochondrial permeability transition pore opening. Thus, creatine may be a useful adjunctive therapy against HAND. PMID:25613139

  6. NQR in Alanine and Lysine Iodates

    NASA Astrophysics Data System (ADS)

    Petrosyan, A. M.; Burbelo, V. M.; Tamazyan, R. A.; Karapetyan, H. A.; Sukiasyan, R. P.

    2000-02-01

    The structure o f iodates of α- and β-alanine ( Ala) (2(β-Ala • HIO3) • H2O , β-Ala-2HIO3 , D L-Ala• HIO3 • 2H2O, L-Ala • HIO3) and L-lysine (L-Lys) (L-Lys • HIO3, L-Lys • 2HIO3,L-Lys • 3HIO3, L-Lys • 6HIO3) have been investigated by means of iodine-127 NQR, IR spectroscopy and X-ray diffraction

  7. Swelling-activated taurine and creatine effluxes from rat cortical astrocytes are pharmacologically distinct.

    PubMed

    Bothwell, J H; Styles, P; Bhakoo, K K

    2002-01-15

    Primary cultures of rat cortical astrocytes undergo a swelling-activated loss of taurine and creatine. In this study, the pharmacological characteristics of the taurine and creatine efflux pathways were compared, and significant differences were shown to exist between the two. Both taurine and creatine effluxes were rapidly activated upon exposure of astrocytes to hypo-osmotic media, and rapidly inactivated upon their return to iso-osmotic media. The relative rates of taurine and creatine efflux depended upon the magnitude of the hypo-osmotic shock. Anion-transport inhibitors strongly inhibited taurine efflux, with the order of potency being NPPB > DIDS > niflumic acid. DIDS and NPPB had less of an inhibitory effect on creatine efflux, whereas tamoxifen and niflumic acid actually stimulated creatine efflux. These data are consistent with separate pathways for taurine and creatine loss during astrocyte swelling.

  8. Characterisation of Potential Antimicrobial Targets in Bacillus spp. I. Aminotransferases and Methionine Regeneration in Bacillus subtilis

    DTIC Science & Technology

    2002-07-01

    targets in Bacillus spp. I. Aminotransferases and methionine regeneration in Bacillus subtilis. Bradley J. Berger and Marvin H. Knodel Defence R&D...Characterisation of potential antimicrobial targets in Bacillus spp. I. Aminotransferases and methionine regeneration in Bacillus subtilis. Bradley J...examined in the gram-positive bacterium Bacillus subtilis. Homogenates of this bacterium were able to convert ketomethiobutyrate to methionine, utilising

  9. Short and longer-term effects of creatine supplementation on exercise induced muscle damage.

    PubMed

    Rosene, John; Matthews, Tracey; Ryan, Christine; Belmore, Keith; Bergsten, Alisa; Blaisdell, Jill; Gaylord, James; Love, Rebecca; Marrone, Michael; Ward, Kristine; Wilson, Eric

    2009-01-01

    The purpose of this investigation was to determine if creatine supplementation assisted with reducing the amount of exercise induced muscle damage and if creatine supplementation aided in recovery from exercise induced muscle damage. Two groups of subjects (group 1 = creatine; group 2 = placebo) participated in an eccentric exercise protocol following 7 and 30 days of creatine or placebo supplementation (20 g.d(-1) for 7 d followed by 6g.d(-1) for 23 d = 30 d). Prior to the supplementation period, measurements were obtained for maximal dynamic strength, maximal isometric force, knee range of motion, muscle soreness, and serum levels of creatine kinase (CK) and lactate dehydrogenase (LDH). Following 7 days of creatine supplementation, on day 8, subjects began consuming 6 g.d(-1) of creatine for 23 days. Additionally on days 8 and 31, subjects performed an eccentric exercise protocol using the knee extensors to induce muscle damage. Indirect markers of muscle damage, including maximal isometric force, knee range of motion, muscle soreness, and serum levels of CK and LDH, were collected at 12, 24, and 48 hours following each exercise bout. The results indicated that acute bouts of creatine have no effect on indirect markers of muscle damage for the acute (7 days) bout. However, maximal isometric force was greater for the creatine group versus placebo for the chronic (30 days) bout. This suggests that the ergogenic effect of creatine following 30 days of supplementation may have a positive impact on exercise induced muscle damage. Key pointsEccentric muscle actions highly associated with exercise induced muscle damage.Creatine supplementation has ergogenic effect to increase protein synthesis.Creatine supplementation does not attenuate exercise induced muscle damage with short term supplementation (7 days).Increased maximal isometric force seen with creatine supplementation after 30 days following exercise induced muscle damage.Ergogenic effect of creatine

  10. Short and longer-term effects of creatine supplementation on exercise induced muscle damage

    PubMed Central

    Rosene, John; Matthews, Tracey; Ryan, Christine; Belmore, Keith; Bergsten, Alisa; Blaisdell, Jill; Gaylord, James; Love, Rebecca; Marrone, Michael; Ward, Kristine; Wilson, Eric

    2009-01-01

    The purpose of this investigation was to determine if creatine supplementation assisted with reducing the amount of exercise induced muscle damage and if creatine supplementation aided in recovery from exercise induced muscle damage. Two groups of subjects (group 1 = creatine; group 2 = placebo) participated in an eccentric exercise protocol following 7 and 30 days of creatine or placebo supplementation (20 g.d-1 for 7 d followed by 6g.d-1 for 23 d = 30 d). Prior to the supplementation period, measurements were obtained for maximal dynamic strength, maximal isometric force, knee range of motion, muscle soreness, and serum levels of creatine kinase (CK) and lactate dehydrogenase (LDH). Following 7 days of creatine supplementation, on day 8, subjects began consuming 6 g.d-1 of creatine for 23 days. Additionally on days 8 and 31, subjects performed an eccentric exercise protocol using the knee extensors to induce muscle damage. Indirect markers of muscle damage, including maximal isometric force, knee range of motion, muscle soreness, and serum levels of CK and LDH, were collected at 12, 24, and 48 hours following each exercise bout. The results indicated that acute bouts of creatine have no effect on indirect markers of muscle damage for the acute (7 days) bout. However, maximal isometric force was greater for the creatine group versus placebo for the chronic (30 days) bout. This suggests that the ergogenic effect of creatine following 30 days of supplementation may have a positive impact on exercise induced muscle damage. Key points Eccentric muscle actions highly associated with exercise induced muscle damage. Creatine supplementation has ergogenic effect to increase protein synthesis. Creatine supplementation does not attenuate exercise induced muscle damage with short term supplementation (7 days). Increased maximal isometric force seen with creatine supplementation after 30 days following exercise induced muscle damage. Ergogenic effect of creatine

  11. [Association between occupational stress and aminotransferase activity in patients with metabolic syndrome].

    PubMed

    Zhao, H; Song, L; Qiang, Y; Liu, H R; Qiu, F Y; Li, X Z; Song, H

    2016-12-20

    Objective: To investigate the association between occupational stress and activity of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in patients with metabolic syndrome. Methods: A case-control study was performed. According to inclusion and exclusion criteria, among the staff members of enterprises and public institutions aged 20~60 years who underwent physical examination in The Affiliated Hospital of Ningxia Medical University and The People's Hospital of Wuzhong from October 2011 to October 2012, 622 patients with metabolic syndrome who did not have a blood relationship with each other were enrolled as case group, and 600 healthy staff members who also did not have a blood relationshipwith each otherwere enrolled as control group. Questionnaire investigation, chronic occupational stress investigation, physical examination, and laboratory tests were performed for all subjects. Results: Compared with the control group, the case group had significantly higher serum levels and abnormal rates of AST and ALT (t=-4.338 and-5.485, χ(2)=11.168 and 34.302, all P<0.05) . There were no significantdifferences in the serum level and abnormal rate of AST between the subgroups with different occupational stresses in both groups (F=2.192 and 2.567, χ(2)=2.694 and 5.402, all P>0.05) , but there were significant differencesbetween the subgroups in all subjects (F=5.005, χ(2)=6.398, all P<0.05) . There were no significant differences in the serum level and abnormal rate of ALT between thesubgroups with different occupational stresses in the case group, the control group, and all subjects (F=0.845, 0.450, and 1.416, χ(2)=2.564, 1.344, and 3.147, all P>0.05) . The partial correlation analysis showed that the total score of occupational stress was positively correlated withthe serum level of AST (r=0.071, P<0.05) and was not correlated with the serum level of ALT (r=-0.044, P>0.05) , and that the serum level of AST was positively correlated with that of ALT

  12. Diagnostic methods and recommendations for the cerebral creatine deficiency syndromes.

    PubMed

    Clark, Joseph F; Cecil, Kim M

    2015-03-01

    Primary care pediatricians and a variety of specialist physicians strive to define an accurate diagnosis for children presenting with impairment of expressive speech and delay in achieving developmental milestones. Within the past two decades, a group of disorders featuring this presentation have been identified as cerebral creatine deficiency syndromes (CCDS). Patients with these disorders were initially discerned using proton magnetic resonance spectroscopy of the brain within a magnetic resonance imaging (MRI) examination. The objective of this review is to provide the clinician with an overview of the current information available on identifying and treating these conditions. We explain the salient features of creatine metabolism, synthesis, and transport required for normal development. We propose diagnostic approaches for confirming a CCDS diagnosis. Finally, we describe treatment approaches for managing patients with these conditions.

  13. [Acute myocardial infarct and the kinetics of creatine kinase].

    PubMed

    Sochman, J; Fabiían, J; Englis, M; Belán, A

    1989-10-01

    The authors criticize contemporary views on creatine kinase kinetics in relation to the patency or occlusion of the coronary artery in the area of the infarction focus. In the investigation proper the time needed to achieve the peak plasma creatine kinase activity after the onset of infarction pain in patients with necroses in different areas of the left ventricle is assessed. Although the interpretation of the observed phenomenon is not clear so far, this finding makes the informative value of the hitherto used time parameter of the kinetics of this enzyme doubtful, in particular in thrombolytic treatment of myocardial infarction. In practice it is thus not possible to evaluate the restored patency of the artery to the necrotic focus on the basis of the above parameter.

  14. Elevated creatine kinase and transaminases in asymptomatic SBMA.

    PubMed

    Sorenson, Eric J; Klein, Christopher J

    2007-02-01

    X-linked spinal and bulbar muscular atrophy (SBMA or Kennedy's disease) has a variable prognosis. Most male carriers are affected by their fourth or fifth decade of life, while some remain asymptomatic lifelong. Elevations of serum creatine kinase are well known to occur in clinically manifesting SBMA patients. Elevations prior to the onset of the clinical syndrome have not been reported. Here we report two cases of SBMA presenting with 'idiopathic' elevations of serum transaminases and creatine kinase a decade in advance of their symptomatic onset. These cases emphasize the need to consider SBMA and genetic testing for the androgen receptor trinucleotide CAG expansion in males otherwise healthy with 'idiopathic' elevated creatinine kinase.

  15. Crystallization and preliminary crystallographic analysis of d-alanine-d-alanine ligase from Streptococcus mutans

    SciTech Connect

    Lu, Yong-Zhi; Sheng, Yu; Li, Lan-Fen; Tang, De-Wei; Liu, Xiang-Yu; Zhao, Xiaojun; Liang, Yu-He Su, Xiao-Dong

    2007-09-01

    A potential target for antibiotic drug design, d-alanine-d-alanine ligase from S. mutans, was expressed in E. coli, purified and crystallized. Diffraction data were collected to 2.4 Å resolution. d-Alanine-d-alanine ligase is encoded by the gene ddl (SMU-599) in Streptococcus mutans. This ligase plays a very important role in cell-wall biosynthesis and may be a potential target for drug design. To study the structure and function of this ligase, the gene ddl was amplified from S. mutans genomic DNA and cloned into the expression vector pET28a. The protein was expressed in soluble form in Escherichia coli strain BL21 (DE3). Homogeneous protein was obtained using a two-step procedure consisting of Ni{sup 2+}-chelating and size-exclusion chromatography. Purified protein was crystallized and the cube-shaped crystal diffracted to 2.4 Å. The crystal belongs to space group P3{sub 1}21 or P3{sub 2}21, with unit-cell parameters a = b = 79.50, c = 108.97 Å. There is one molecule per asymmetric unit.

  16. Racemization of alanine by the alanine racemases from Salmonella typhimurium and Bacillus stearothermophilus: energetic reaction profiles

    SciTech Connect

    Faraci, W.S.; Walsh, C.T.

    1988-05-03

    Alanine racemases are bacterial pyridoxal 5'-phosphate (PLP) dependent enzymes providing D-alanine as an essential building block for biosynthesis of the peptidoglycan layer of the cell wall. Two isozymic alanine racemases, encoded by the dadB gene and the alr gene, from the Gram-negative mesophilic Salmonella typhimurium and one from the Gram-positive thermophilic Bacillus stearothermophilus have been examined for the racemization mechanism. Substrate deuterium isotope effects and solvent deuterium isotope effects have been measured in both L ..-->.. D and D..-->.. L directions for all three enzymes to assess the degree to which abstraction of the ..cap alpha..-proton or protonation of substrate PLP carbanion is limiting in catalysis. Additionally, experiments measuring internal return of ..cap alpha..-/sup 3/H from substrate to product and solvent exchange/substrate conversion experiments in /sup 3/H/sub 2/O have been used with each enzyme to examine the partitioning of substrate PLP carbanion intermediates and to obtain the relative heights of kinetically significant energy barriers in alanine racemase catalysis.

  17. Creatine supplementation improves muscular performance in older women.

    PubMed

    Gotshalk, Lincoln A; Kraemer, William J; Mendonca, Mario A G; Vingren, Jakob L; Kenny, Anne M; Spiering, Barry A; Hatfield, Disa L; Fragala, Maren S; Volek, Jeff S

    2008-01-01

    Muscle power and strength decrease with age leading to reduced independence and increased health risk from falls. Creatine supplementation can increase muscle power and strength. The purpose of this study was to examine the effects of 7 days of creatine supplementation on body composition, muscular strength, and lower-body motor functional performance in older women. Thirty 58-71 year old women performed three test sessions (T1-T3) each separated by one week. Each session consisted of one repetition maximum tests for bench press and leg press, and isometric hand-grip, tandem gait, upper-body ergometer, and lower-body ergometer tests. Following T2, subjects were assigned to a creatine monohydrate (0.3 g kg body mass(-1) for 7 days) (CR: 63.31 +/- 1.22 year, 160.00 +/- 1.58 cm, 67.11 +/- 4.38 kg) or a placebo (PL: 62.98 +/- 1.11 year, 162.25 +/- 2.09 cm, 67.84 +/- 3.90 kg) supplementation group. CR significantly (P < 0.05) increased bench press (1.7 +/- 0.4 kg), leg press (5.2 +/- 1.8 kg), body mass (0.49 +/- 0.04 kg) and fat free mass (0.52 +/- 0.05) and decreased completion time on the functional tandem gait tests from T2-T3. No significant changes were found for PL on any of the measured variables. No adverse side-effects were reported by either group. Short-term creatine supplementation resulted in an increase in strength, power, and lower-body motor functional performance in older women without any adverse side effects.

  18. Combined creatine and sodium bicarbonate supplementation enhances interval swimming.

    PubMed

    Mero, Antti A; Keskinen, Kari L; Malvela, Marko T; Sallinen, Janne M

    2004-05-01

    This study examined the effect of simultaneous supplementation of creatine and sodium bicarbonate on consecutive maximal swims. Sixteen competitive male and female swimmers completed, in a randomized order, 2 different treatments (placebo and a combination of creatine and sodium bicarbonate) with 30 days of washout period between treatments in a double-blind crossover procedure. Both treatments consisted of placebo or creatine supplementation (20 g per day) in 6 days. In the morning of the seventh day, there was placebo or sodium bicarbonate supplementation (0.3 g per kg body weight) during 2 hours before a warm-up for 2 maximal 100-m freestyle swims that were performed with a passive recovery of 10 minutes in between. The first swims were similar, but the increase in time of the second versus the first 100-m swimming time was 0.9 seconds less (p < 0.05) in the combination group than in placebo. Mean blood pH was higher (p < 0.01-0.001) in the combination group than in placebo after supplementation on the test day. Mean blood pH decreased (p < 0.05) similarly during the swims in both groups. Mean blood lactate increased (p < 0.001) during the swims, but there were no differences in peak blood lactate between the combination group (14.9 +/- 0.9 mmol.L(-1)) and placebo (13.4 +/- 1.0 mmol.L(-1)). The data indicate that simultaneous supplementation of creatine and sodium bicarbonate enhances performance in consecutive maximal swims.

  19. Meta-analysis of the influence of TM6SF2 E167K variant on Plasma Concentration of Aminotransferases across different Populations and Diverse Liver Phenotypes

    PubMed Central

    Sookoian, Silvia; Pirola, Carlos J.

    2016-01-01

    A nonsynonymous E167K (rs58542926 C/T) variant in TM6SF2 gene was recently associated with nonalcoholic fatty liver disease (NAFLD). We explored the association between E167K and plasma concentrations of alanine (ALT) and aspartate (AST) aminotransferases through a meta-analysis. We also estimated the strength of the effect across diverse liver phenotypes, including NAFLD and chronic viral hepatitis; fourteen studies were included. We found that ALT (p = 3.2 × 10−6, n = 94,414) and AST (p = 0007, n = 93,809) levels were significantly associated with rs58542926 in NAFLD. By contrast, rs58542926 was not associated with either ALT (p = 0.24, n = 4187) or AST (p = 0.17, n = 2678) levels in four studies on chronic hepatitis. In conclusion, the results of the pooled estimates in patients with NAFLD showed that carriers of the T allele (EK + KK), when compared with homozygous subjects for the C allele (EE genotype) have increased levels of aminotransferases; however, this increase represents –2.5 (9.8%) and 1.2 (5%) IU/L of ALT and AST respectively, which is fairly small compared with the large effect of PNPLA3- rs738409-G allele that is associated with a –28% increase in serum ALT. PMID:27278285

  20. Assignment of the creatine transporter gene (SLC6A8) to human chromosome Xq28 telomeric to G6PD

    SciTech Connect

    Gregor, P.; Nash, S.R.; Caron, M.G.

    1995-01-01

    The creatine-phosphocreatine shuttle has important functions in the temporal and spatial maintenance of the energy supply to skeletal and cardiac muscle. Muscle cells do not synthesize creatine, but take it up via a specific sodium-dependent transporter - the creatine transporter. Thus, the creatine transporter has an important role in muscular physiology. Furthermore, inhibition of creatine transport in experimental animals causes muscle weakness. Recently, creatine transporter cDNAs have been isolated and characterized from rabbit and human. In this communication we report mapping of the creatine transporter gene to human chromosome Xq28. 12 refs., 1 fig.

  1. Improved radioimmunoassay for creatine kinase isoenzymes in plasma

    SciTech Connect

    Ritter, C.S.; Mumm, S.R.; Roberts, R.

    1981-11-01

    We describe convenient and relatively rapid procedures for purifying creatine kinase isoenzymes MM, BB, and MB, and their use in an improved radioimmunoassay for creatine kinase isoenzymes in plasma. The modifications include use of: (a) BB with a specific activity of 400 kU/G, which can be labeled with a specific radioactivity of 20 Ci/g; (b) albumin-free purified MB as inhibitor; (c) antiserum to MB creatine kinase; and (d) a second-antibody technique that necessitates only a 15-min incubation. The radioimmunoassay for MB has a sensitivity of 0.2 ..mu..g/L (80 mU/L) and a CV of <5%. Plasma MB average 22 (SD 12) ..mu..g/L in 200 normal subjects; 24 (SD 12) ..mu..g/L in 200 patients with chest pain without infarction; and 23 (SD 7) ..mu..g/L in 43 patients with renal disease, whether measured before or after dialysis. Peak values for plasma MB averaged 191 (SD 86) ..mu..g/L in 325 patients with documented myocardial infarction; BB was negligible. Extensive clinical experience indicates the radioimmunoassay to be suitably rapid, highly sensitive, and reliable as a diagnostic assay for MB on plasma.

  2. Ergolytic/ergogenic effects of creatine on aerobic power.

    PubMed

    Smith, A E; Fukuda, D H; Ryan, E D; Kendall, K L; Cramer, J T; Stout, J

    2011-12-01

    This study evaluated the effects of creatine (Cr) loading and sex differences on aerobic running performance. 27 men (mean±SD; age: 22.2±3.1 years, ht: 179.5±8.7 cm, wt: 78.0±9.8 kg) and 28 women (age: 21.2±2.1 years, ht: 166.0±5.8 cm, wt: 63.4±8.9 kg) were randomly assigned to either creatine (Cr, di-creatine citrate; n=27) or a placebo (PL; n=28) group, ingesting 1 packet 4 times daily (total of 20 g/day) for 5 days. Aerobic power (maximal oxygen consumption: VO2max) was assessed before and after supplementation using open circuit spirometry (Parvo-Medics) during graded exercise tests on a treadmill. 4 high-speed runs to exhaustion were conducted at 110, 105, 100, and 90% of peak velocity to determine critical velocity (CV). Distances achieved were plotted over times-to-exhaustion and linear regression was used to determine the slopes (critical velocity, CV) assessing aerobic performance. The results indicated that Cr loading did not positively or negatively influence VO2max, CV, time to exhaustion or body mass (p>0.05). These results suggest Cr supplementation may be used in aerobic running activities without detriments to performance.

  3. Estimation of skeletal muscle mass from body creatine content

    NASA Technical Reports Server (NTRS)

    Pace, N.; Rahlmann, D. F.

    1982-01-01

    Procedures have been developed for studying the effect of changes in gravitational loading on skeletal muscle mass through measurements of the body creatine content. These procedures were developed for studies of gravitational scale effects in a four-species model, comprising the hamster, rat, guinea pig, and rabbit, which provides a sufficient range of body size for assessment of allometric parameters. Since intracellular muscle creatine concentration varies among species, and with age within a given species, the concentration values for metabolically mature individuals of these four species were established. The creatine content of the carcass, skin, viscera, smooth muscle, and skeletal muscle was determined for each species. In addition, the skeletal muscle mass of the major body components was determined, as well as the total and fat-free masses of the body and carcass, and the percent skeletal muscle in each. It is concluded that these procedures are particularly useful for studying the effect of gravitational loading on the skeletal muscle content of the animal carcass, which is the principal weight-bearing organ of the body.

  4. Macro creatine kinase: determination and differentiation of two types by their activation energies

    SciTech Connect

    Stein, W.; Bohner, J.; Steinhart, R.; Eggstein, M.

    1982-01-01

    Determination of the MB isoenzyme of creatine kinase in patients with acute myocardial infarction may be disturbed by the presence of macro creatine kinase. The relative molecular mass of this form of creatine kinase in human serum is at least threefold that of the ordinary enzyme, and it is more thermostable. Here we describe our method for determination of macro creatine kinases and an easy-to-perform test for differentiating two forms of macro creatine kinase, based on their distinct activation energies. The activation energies of serum enzymes are mostly in the range of 40-65 kJ/mol of substrate. Unlike normal cytoplasmatic creatine kinases and IgG-linked CK-BB (macro creatine kinase type 1) a second form of macro creatine kinase (macro creatine kinase type 2) shows activation energies greater than 80 kJ/mol of substrate. The exact composition of macro creatine kinase type 2 is still unknown, but there is good reason to believe that it is of mitochondrial origin.

  5. Creatine supplementation with specific view to exercise/sports performance: an update.

    PubMed

    Cooper, Robert; Naclerio, Fernando; Allgrove, Judith; Jimenez, Alfonso

    2012-07-20

    Creatine is one of the most popular and widely researched natural supplements. The majority of studies have focused on the effects of creatine monohydrate on performance and health; however, many other forms of creatine exist and are commercially available in the sports nutrition/supplement market. Regardless of the form, supplementation with creatine has regularly shown to increase strength, fat free mass, and muscle morphology with concurrent heavy resistance training more than resistance training alone. Creatine may be of benefit in other modes of exercise such as high-intensity sprints or endurance training. However, it appears that the effects of creatine diminish as the length of time spent exercising increases. Even though not all individuals respond similarly to creatine supplementation, it is generally accepted that its supplementation increases creatine storage and promotes a faster regeneration of adenosine triphosphate between high intensity exercises. These improved outcomes will increase performance and promote greater training adaptations. More recent research suggests that creatine supplementation in amounts of 0.1 g/kg of body weight combined with resistance training improves training adaptations at a cellular and sub-cellular level. Finally, although presently ingesting creatine as an oral supplement is considered safe and ethical, the perception of safety cannot be guaranteed, especially when administered for long period of time to different populations (athletes, sedentary, patient, active, young or elderly).

  6. Creatine supplementation with specific view to exercise/sports performance: an update

    PubMed Central

    2012-01-01

    Creatine is one of the most popular and widely researched natural supplements. The majority of studies have focused on the effects of creatine monohydrate on performance and health; however, many other forms of creatine exist and are commercially available in the sports nutrition/supplement market. Regardless of the form, supplementation with creatine has regularly shown to increase strength, fat free mass, and muscle morphology with concurrent heavy resistance training more than resistance training alone. Creatine may be of benefit in other modes of exercise such as high-intensity sprints or endurance training. However, it appears that the effects of creatine diminish as the length of time spent exercising increases. Even though not all individuals respond similarly to creatine supplementation, it is generally accepted that its supplementation increases creatine storage and promotes a faster regeneration of adenosine triphosphate between high intensity exercises. These improved outcomes will increase performance and promote greater training adaptations. More recent research suggests that creatine supplementation in amounts of 0.1 g/kg of body weight combined with resistance training improves training adaptations at a cellular and sub-cellular level. Finally, although presently ingesting creatine as an oral supplement is considered safe and ethical, the perception of safety cannot be guaranteed, especially when administered for long period of time to different populations (athletes, sedentary, patient, active, young or elderly). PMID:22817979

  7. Ornithine-δ-Aminotransferase Inhibits Neurogenesis During Xenopus Embryonic Development

    PubMed Central

    Peng, Ying; Cooper, Sandra K.; Li, Yi; Mei, Jay M.; Qiu, Shuwei; Borchert, Gregory L.; Donald, Steven P.; Kung, Hsiang-fu; Phang, James M.

    2015-01-01

    Purpose. In humans, deficiency of ornithine-δ-aminotransferase (OAT) results in progressive degeneration of the neural retina (gyrate atrophy) with blindness in the fourth decade. In this study, we used the Xenopus embryonic developmental model to study functions of the OAT gene on embryonic development. Methods. We cloned and sequenced full-length OAT cDNA from Xenopus oocytes (X-OAT) and determined X-OAT expression in various developmental stages of Xenopus embryos and in a variety of adult tissues. The phenotype, gene expression of neural developmental markers, and enzymatic activity were detected by gain-of-function and loss-of-function manipulations. Results. We showed that X-OAT is essential for Xenopus embryonic development, and overexpression of X-OAT produces a ventralized phenotype characterized by a small head, lack of axial structure, and defective expression of neural developmental markers. Using X-OAT mutants based on mutations identified in humans, we found that substitution of both Arg 180 and Leu 402 abrogated both X-OAT enzymatic activity and ability to modulate the developmental phenotype. Neurogenesis is inhibited by X-OAT during Xenopus embryonic development. Conclusions. Neurogenesis is inhibited by X-OAT during Xenopus embryonic development, but it is essential for Xenopus embryonic development. The Arg 180 and Leu 402 are crucial for these effects of the OAT molecule in development. PMID:25783604

  8. Tyrosine aminotransferase from Leishmania infantum: A new drug target candidate

    PubMed Central

    Moreno, Miguel Angel; Alonso, Ana; Alcolea, Pedro Jose; Abramov, Ariel; de Lacoba, Mario García; Abendroth, Jan; Zhang, Sunny; Edwards, Thomas; Lorimer, Don; Myler, Peter John; Larraga, Vicente

    2014-01-01

    Leishmania infantum is the etiological agent of zoonotic visceral leishmaniasis in the Mediterranean basin. The disease is fatal without treatment, which has been based on antimonial pentavalents for more than 60 years. Due to resistances, relapses and toxicity to current treatment, the development of new drugs is required. The structure of the L. infantum tyrosine aminotransferase (LiTAT) has been recently solved showing important differences with the mammalian orthologue. The characterization of LiTAT is reported herein. This enzyme is cytoplasmic and is over-expressed in the more infective stages and nitric oxide resistant parasites. Unlike the mammalian TAT, LiTAT is able to use ketomethiobutyrate as co-substrate. The pharmacophore model of LiTAT with this specific co-substrate is described herein. This may allow the identification of new inhibitors present in the databases. All the data obtained support that LiTAT is a good target candidate for the development of new anti-leishmanial drugs. PMID:25516846

  9. Design and Mechanism of Tetrahydrothiophene-based GABA Aminotransferase Inactivators

    PubMed Central

    Le, Hoang V.; Hawker, Dustin D.; Wu, Rui; Doud, Emma; Widom, Julia; Sanishvili, Ruslan; Liu, Dali; Kelleher, Neil L.; Silverman, Richard B.

    2015-01-01

    Low levels of γ-aminobutyric acid (GABA), one of two major neurotransmitters that regulate brain neuronal activity, are associated with many neurological disorders, such as epilepsy, Parkinson’s disease, Alzheimer’s disease, Huntington’s disease, and cocaine addiction. One of the main methods to raise the GABA level in human brain is to use small molecules that cross the blood-brain barrier and inhibit the activity of γ-aminobutyric acid aminotransferase (GABA-AT), the enzyme that degrades GABA. We have designed a series of conformationally-restricted, tetrahydrothiophene-based GABA analogs with a properly-positioned leaving group that could facilitate a ring-opening mechanism, leading to inactivation of GABA-AT. One compound in the series is eight times more efficient an inactivator of GABA-AT than vigabatrin, the only FDA-approved inactivator of GABA-AT. Our mechanistic studies show that the compound inactivates GABA-AT by a new mechanism. The metabolite resulting from inactivation does not covalently bind to amino acid residues of GABA-AT but stays in the active site via H-bond interactions with Arg-192, a π-π interaction with Phe-189, and a weak nonbonded S···O=C interaction with Glu-270, thereby inactivating the enzyme. PMID:25781189

  10. Cloning and expression of human tyrosine aminotransferase cDNA.

    PubMed

    Séralini, G E; Luu-Thé, V; Labrie, F

    1995-01-02

    Complementary DNA clones encoding human tyrosine aminotransferase (TAT) were isolated by screening a normal adult woman liver lambda gt11 library with rat TAT cDNA. The largest isolated cDNA is 2051 bp long (EMBL accession number X55675). This cDNA was subcloned downstream of the cytomegalovirus promoter in the pCMV vector for transfection into human cervical carcinoma HeLa cells. Expression of the TAT cDNA resulted in the synthesis of a protein with a molecular mass of approximately 50 kDa, as assessed by Western analysis, a value which is in close agreement with the predicted molecular weight of 50,399, for a deduced sequence of 454 amino acids. The expressed protein catalyzed specifically the conversion of L-[14C]tyrosine into p-[14C]hydroxyphenylpyruvate. The availability of a functional TAT cDNA provides a useful tool for detailed study of the structure-function relationship of the enzyme and its mutated derivatives.

  11. International society of sports nutrition position stand: Beta-Alanine.

    PubMed

    Trexler, Eric T; Smith-Ryan, Abbie E; Stout, Jeffrey R; Hoffman, Jay R; Wilborn, Colin D; Sale, Craig; Kreider, Richard B; Jäger, Ralf; Earnest, Conrad P; Bannock, Laurent; Campbell, Bill; Kalman, Douglas; Ziegenfuss, Tim N; Antonio, Jose

    2015-01-01

    The International Society of Sports Nutrition (ISSN) provides an objective and critical review of the mechanisms and use of beta-alanine supplementation. Based on the current available literature, the conclusions of the ISSN are as follows: 1) Four weeks of beta-alanine supplementation (4-6 g daily) significantly augments muscle carnosine concentrations, thereby acting as an intracellular pH buffer; 2) Beta-alanine supplementation currently appears to be safe in healthy populations at recommended doses; 3) The only reported side effect is paraesthesia (tingling), but studies indicate this can be attenuated by using divided lower doses (1.6 g) or using a sustained-release formula; 4) Daily supplementation with 4 to 6 g of beta-alanine for at least 2 to 4 weeks has been shown to improve exercise performance, with more pronounced effects in open end-point tasks/time trials lasting 1 to 4 min in duration; 5) Beta-alanine attenuates neuromuscular fatigue, particularly in older subjects, and preliminary evidence indicates that beta-alanine may improve tactical performance; 6) Combining beta-alanine with other single or multi-ingredient supplements may be advantageous when supplementation of beta-alanine is high enough (4-6 g daily) and long enough (minimum 4 weeks); 7) More research is needed to determine the effects of beta-alanine on strength, endurance performance beyond 25 min in duration, and other health-related benefits associated with carnosine.

  12. Expression and processing of human ornithine-delta-aminotransferase in Saccharomyces cerevisiae.

    PubMed

    Dougherty, K M; Swanson, D A; Brody, L C; Valle, D

    1993-11-01

    Ornithine-delta-aminotransferase catalyzes the conversion of ornithine to glutamate-gamma-semialdehyde. In humans, deficiency of this mitochondrial matrix enzyme results in the progressive blinding disorder, gyrate atrophy of the choroid and retina. To explore yeast as an expression system, we introduced a cDNA encoding human ornithine-delta-aminotransferase into an ornithine aminotransferase-deficient strain of Saccharomyces cerevisiae. The human enzyme was expressed at high levels, with activity 20-fold greater than that of wild-type yeast and 10-fold higher than in human fibroblasts. Although the normal location of ornithine-delta-aminotransferase in S. cerevisiae is cytosolic, human ornithine-delta-aminotransferase expressed in S. cerevisiae was localized to the mitochondrial matrix with correct proteolytic processing of its mitochondrial leader sequence. Despite this anomalous location in yeast, human ornithine-delta-aminotransferase complemented the phenotype of the mutant strain, restoring its ability to utilize ornithine as a sole nitrogen source. We also expressed a vitamin B6-responsive missense allele of ornithine-delta-aminotransferase (V332M) and showed that the biochemical phenotype of this allele is easily demonstrated confirming the usefulness of this system for examining mutations causing gyrate atrophy.

  13. Crystal structure of the Apo form of D-Alanine:D-Alanine ligase (DDl) from Streptococcus mutans.

    PubMed

    Lu, Yongzhi; Xu, Hongyan; Zhao, Xiaojun

    2010-08-01

    D-Alanine:D-Alanine ligase (DDl) catalyzes the formation of D-Alanine:D-Alanine dipeptide and is an essential enzyme in bacterial cell wall biosynthesis.. This enzyme does not have a human ortholog, making it an attractive target for developing new antibiotic drugs. We determined the crystal structure at 2.23 A resolution of DDl from Streptococcus mutans (SmDDl), the principal aetiological agent of human dental caries. This structure reveals that SmDDl is a dimer and has a disordered omega-loop region.

  14. Sensitive non-radioactive determination of aminotransferase stereospecificity for C-4' hydrogen transfer on the coenzyme.

    PubMed

    Jomrit, Juntratip; Summpunn, Pijug; Meevootisom, Vithaya; Wiyakrutta, Suthep

    2011-02-25

    A sensitive non-radioactive method for determination of the stereospecificity of the C-4' hydrogen transfer on the coenzymes (pyridoxal phosphate, PLP; and pyridoxamine phosphate, PMP) of aminotransferases has been developed. Aminotransferase of unknown stereospecificity in its PLP form was incubated in (2)H(2)O with a substrate amino acid resulted in PMP labeled with deuterium at C-4' in the pro-S or pro-R configuration according to the stereospecificity of the aminotransferase tested. The [4'-(2)H]PMP was isolated from the enzyme protein and divided into two portions. The first portion was incubated in aqueous buffer with apo-aspartate aminotransferase (a reference si-face specific enzyme), and the other was incubated with apo-branched-chain amino acid aminotransferase (a reference re-face specific enzyme) in the presence of a substrate 2-oxo acid. The (2)H at C-4' is retained with the PLP if the aminotransferase in question transfers C-4' hydrogen on the opposite face of the coenzyme compared with the reference aminotransferase, but the (2)H is removed if the test and reference aminotransferases catalyze hydrogen transfer on the same face. PLP formed in the final reactions was analyzed by LC-MS/MS for the presence or absence of (2)H. The method was highly sensitive that for the aminotransferase with ca. 50 kDa subunit molecular weight, only 2mg of the enzyme was sufficient for the whole test. With this method, the use of radioactive substances could be avoided without compromising the sensitivity of the assay.

  15. Sensitive non-radioactive determination of aminotransferase stereospecificity for C-4' hydrogen transfer on the coenzyme

    SciTech Connect

    Jomrit, Juntratip; Summpunn, Pijug; Meevootisom, Vithaya; Wiyakrutta, Suthep

    2011-02-25

    Research highlights: {yields} Stereochemical mechanism of PLP enzymes is important but difficult to determine. {yields} This new method is significantly less complicated than the previous ones. {yields} This assay is as sensitive as the radioactive based method. {yields} LC-MS/MS positively identify the analyte coenzyme. {yields} The method can be used with enzyme whose apo form is unstable. -- Abstract: A sensitive non-radioactive method for determination of the stereospecificity of the C-4' hydrogen transfer on the coenzymes (pyridoxal phosphate, PLP; and pyridoxamine phosphate, PMP) of aminotransferases has been developed. Aminotransferase of unknown stereospecificity in its PLP form was incubated in {sup 2}H{sub 2}O with a substrate amino acid resulted in PMP labeled with deuterium at C-4' in the pro-S or pro-R configuration according to the stereospecificity of the aminotransferase tested. The [4'-{sup 2}H]PMP was isolated from the enzyme protein and divided into two portions. The first portion was incubated in aqueous buffer with apo-aspartate aminotransferase (a reference si-face specific enzyme), and the other was incubated with apo-branched-chain amino acid aminotransferase (a reference re-face specific enzyme) in the presence of a substrate 2-oxo acid. The {sup 2}H at C-4' is retained with the PLP if the aminotransferase in question transfers C-4' hydrogen on the opposite face of the coenzyme compared with the reference aminotransferase, but the {sup 2}H is removed if the test and reference aminotransferases catalyze hydrogen transfer on the same face. PLP formed in the final reactions was analyzed by LC-MS/MS for the presence or absence of {sup 2}H. The method was highly sensitive that for the aminotransferase with ca. 50 kDa subunit molecular weight, only 2 mg of the enzyme was sufficient for the whole test. With this method, the use of radioactive substances could be avoided without compromising the sensitivity of the assay.

  16. Engineering of alanine dehydrogenase from Bacillus subtilis for novel cofactor specificity.

    PubMed

    Lerchner, Alexandra; Jarasch, Alexander; Skerra, Arne

    2016-09-01

    The l-alanine dehydrogenase of Bacillus subtilis (BasAlaDH), which is strictly dependent on NADH as redox cofactor, efficiently catalyzes the reductive amination of pyruvate to l-alanine using ammonia as amino group donor. To enable application of BasAlaDH as regenerating enzyme in coupled reactions with NADPH-dependent alcohol dehydrogenases, we alterated its cofactor specificity from NADH to NADPH via protein engineering. By introducing two amino acid exchanges, D196A and L197R, high catalytic efficiency for NADPH was achieved, with kcat /KM  = 54.1 µM(-1)  Min(-1) (KM  = 32 ± 3 µM; kcat  = 1,730 ± 39 Min(-1) ), almost the same as the wild-type enzyme for NADH (kcat /KM  = 59.9 µM(-1)  Min(-1) ; KM  = 14 ± 2 µM; kcat  = 838 ± 21 Min(-1) ). Conversely, recognition of NADH was much diminished in the mutated enzyme (kcat /KM  = 3 µM(-1)  Min(-1) ). BasAlaDH(D196A/L197R) was applied in a coupled oxidation/transamination reaction of the chiral dicyclic dialcohol isosorbide to its diamines, catalyzed by Ralstonia sp. alcohol dehydrogenase and Paracoccus denitrificans ω-aminotransferase, thus allowing recycling of the two cosubstrates NADP(+) and l-Ala. An excellent cofactor regeneration with recycling factors of 33 for NADP(+) and 13 for l-Ala was observed with the engineered BasAlaDH in a small-scale biocatalysis experiment. This opens a biocatalytic route to novel building blocks for industrial high-performance polymers.

  17. Effects of creatine supplementation on the performance and body composition of competitive swimmers.

    PubMed

    Mendes, Renata Rebello; Pires, Ivanir; Oliveira, Althair; Tirapegui, Julio

    2004-08-01

    The objective of this study was to determine the effect of creatine supplementation on performance and body composition of swimmers. Eighteen swimmers were evaluated in terms of post-performance lactate accumulation, body composition, creatine and creatinine excretion, and serum creatinine concentrations before and after creatine or placebo supplementation. No significant differences were observed in the marks obtained in swimming tests after supplementation, although lactate concentrations were higher in placebo group during this period. In the creatine-supplemented group, urinary creatine, creatinine, and body mass, lean mass and body water were significantly increased, but no significant difference in muscle or bone mass was observed. These results suggest that creatine supplementation cannot be considered to be an ergogenic supplement ensuring improved performance and muscle mass gain in swimmers.

  18. Biochemical and Structural Properties of Mouse Kynurenine Aminotransferase III

    SciTech Connect

    Han, Q.; Robinson, H; Cai, T; Tagle, D; Li, J

    2009-01-01

    Kynurenine aminotransferase III (KAT III) has been considered to be involved in the production of mammalian brain kynurenic acid (KYNA), which plays an important role in protecting neurons from overstimulation by excitatory neurotransmitters. The enzyme was identified based on its high sequence identity with mammalian KAT I, but its activity toward kynurenine and its structural characteristics have not been established. In this study, the biochemical and structural properties of mouse KAT III (mKAT III) were determined. Specifically, mKAT III cDNA was amplified from a mouse brain cDNA library, and its recombinant protein was expressed in an insect cell protein expression system. We established that mKAT III is able to efficiently catalyze the transamination of kynurenine to KYNA and has optimum activity at relatively basic conditions of around pH 9.0 and at relatively high temperatures of 50 to 60C. In addition, mKAT III is active toward a number of other amino acids. Its activity toward kynurenine is significantly decreased in the presence of methionine, histidine, glutamine, leucine, cysteine, and 3-hydroxykynurenine. Through macromolecular crystallography, we determined the mKAT III crystal structure and its structures in complex with kynurenine and glutamine. Structural analysis revealed the overall architecture of mKAT III and its cofactor binding site and active center residues. This is the first report concerning the biochemical characteristics and crystal structures of KAT III enzymes and provides a basis toward understanding the overall physiological role of mammalian KAT III in vivo and insight into regulating the levels of endogenous KYNA through modulation of the enzyme in the mouse brain.

  19. Biochemical and structural properties of mouse kynurenine aminotransferase III.

    PubMed

    Han, Qian; Robinson, Howard; Cai, Tao; Tagle, Danilo A; Li, Jianyong

    2009-02-01

    Kynurenine aminotransferase III (KAT III) has been considered to be involved in the production of mammalian brain kynurenic acid (KYNA), which plays an important role in protecting neurons from overstimulation by excitatory neurotransmitters. The enzyme was identified based on its high sequence identity with mammalian KAT I, but its activity toward kynurenine and its structural characteristics have not been established. In this study, the biochemical and structural properties of mouse KAT III (mKAT III) were determined. Specifically, mKAT III cDNA was amplified from a mouse brain cDNA library, and its recombinant protein was expressed in an insect cell protein expression system. We established that mKAT III is able to efficiently catalyze the transamination of kynurenine to KYNA and has optimum activity at relatively basic conditions of around pH 9.0 and at relatively high temperatures of 50 to 60 degrees C. In addition, mKAT III is active toward a number of other amino acids. Its activity toward kynurenine is significantly decreased in the presence of methionine, histidine, glutamine, leucine, cysteine, and 3-hydroxykynurenine. Through macromolecular crystallography, we determined the mKAT III crystal structure and its structures in complex with kynurenine and glutamine. Structural analysis revealed the overall architecture of mKAT III and its cofactor binding site and active center residues. This is the first report concerning the biochemical characteristics and crystal structures of KAT III enzymes and provides a basis toward understanding the overall physiological role of mammalian KAT III in vivo and insight into regulating the levels of endogenous KYNA through modulation of the enzyme in the mouse brain.

  20. Biochemical and Structural Properties of Mouse Kynurenine Aminotransferase III▿

    PubMed Central

    Han, Qian; Robinson, Howard; Cai, Tao; Tagle, Danilo A.; Li, Jianyong

    2009-01-01

    Kynurenine aminotransferase III (KAT III) has been considered to be involved in the production of mammalian brain kynurenic acid (KYNA), which plays an important role in protecting neurons from overstimulation by excitatory neurotransmitters. The enzyme was identified based on its high sequence identity with mammalian KAT I, but its activity toward kynurenine and its structural characteristics have not been established. In this study, the biochemical and structural properties of mouse KAT III (mKAT III) were determined. Specifically, mKAT III cDNA was amplified from a mouse brain cDNA library, and its recombinant protein was expressed in an insect cell protein expression system. We established that mKAT III is able to efficiently catalyze the transamination of kynurenine to KYNA and has optimum activity at relatively basic conditions of around pH 9.0 and at relatively high temperatures of 50 to 60°C. In addition, mKAT III is active toward a number of other amino acids. Its activity toward kynurenine is significantly decreased in the presence of methionine, histidine, glutamine, leucine, cysteine, and 3-hydroxykynurenine. Through macromolecular crystallography, we determined the mKAT III crystal structure and its structures in complex with kynurenine and glutamine. Structural analysis revealed the overall architecture of mKAT III and its cofactor binding site and active center residues. This is the first report concerning the biochemical characteristics and crystal structures of KAT III enzymes and provides a basis toward understanding the overall physiological role of mammalian KAT III in vivo and insight into regulating the levels of endogenous KYNA through modulation of the enzyme in the mouse brain. PMID:19029248

  1. Tyrosine aminotransferase: biochemical and structural properties and molecular dynamics simulations

    SciTech Connect

    Mehere, P.; Robinson, H.; Han, Q.; Lemkul, J. A.; Vavricka, C. J.; Bevan, D. R.; Li, J.

    2010-11-01

    Tyrosine aminotransferase (TAT) catalyzes the transamination of tyrosine and other aromatic amino acids. The enzyme is thought to play a role in tyrosinemia type II, hepatitis and hepatic carcinoma recovery. The objective of this study is to investigate its biochemical and structural characteristics and substrate specificity in order to provide insight regarding its involvement in these diseases. Mouse TAT (mTAT) was cloned from a mouse cDNA library, and its recombinant protein was produced using Escherichia coli cells and purified using various chromatographic techniques. The recombinant mTAT is able to catalyze the transamination of tyrosine using {alpha}-ketoglutaric acid as an amino group acceptor at neutral pH. The enzyme also can use glutamate and phenylalanine as amino group donors and p-hydroxy-phenylpyruvate, phenylpyruvate and alpha-ketocaproic acid as amino group acceptors. Through macromolecular crystallography we have determined the mTAT crystal structure at 2.9 {angstrom} resolution. The crystal structure revealed the interaction between the pyridoxal-5'-phosphate cofactor and the enzyme, as well as the formation of a disulphide bond. The detection of disulphide bond provides some rational explanation regarding previously observed TAT inactivation under oxidative conditions and reactivation of the inactive TAT in the presence of a reducing agent. Molecular dynamics simulations using the crystal structures of Trypanosoma cruzi TAT and human TAT provided further insight regarding the substrate-enzyme interactions and substrate specificity. The biochemical and structural properties of TAT and the binding of its cofactor and the substrate may help in elucidation of the mechanism of TAT inhibition and activation.

  2. Tyrosine Aminotransferase: Biochemical and Structural Properties and Molecular Dynamics Simulations

    SciTech Connect

    P Mehere; Q Han; J Lemkul; C Vavricka; H Robinson; D Bevan; J Li

    2011-12-31

    Tyrosine aminotransferase (TAT) catalyzes the transamination of tyrosine and other aromatic amino acids. The enzyme is thought to play a role in tyrosinemia type II, hepatitis and hepatic carcinoma recovery. The objective of this study is to investigate its biochemical and structural characteristics and substrate specificity in order to provide insight regarding its involvement in these diseases. Mouse TAT (mTAT) was cloned from a mouse cDNA library, and its recombinant protein was produced using Escherichia coli cells and purified using various chromatographic techniques. The recombinant mTAT is able to catalyze the transamination of tyrosine using {alpha}-ketoglutaric acid as an amino group acceptor at neutral pH. The enzyme also can use glutamate and phenylalanine as amino group donors and p-hydroxy-phenylpyruvate, phenylpyruvate and alpha-ketocaproic acid as amino group acceptors. Through macromolecular crystallography we have determined the mTAT crystal structure at 2.9 {angstrom} resolution. The crystal structure revealed the interaction between the pyridoxal-5'-phosphate cofactor and the enzyme, as well as the formation of a disulphide bond. The detection of disulphide bond provides some rational explanation regarding previously observed TAT inactivation under oxidative conditions and reactivation of the inactive TAT in the presence of a reducing agent. Molecular dynamics simulations using the crystal structures of Trypanosoma cruzi TAT and human TAT provided further insight regarding the substrate-enzyme interactions and substrate specificity. The biochemical and structural properties of TAT and the binding of its cofactor and the substrate may help in elucidation of the mechanism of TAT inhibition and activation.

  3. Tyrosine aminotransferase contributes to benzylisoquinoline alkaloid biosynthesis in opium poppy.

    PubMed

    Lee, Eun-Jeong; Facchini, Peter J

    2011-11-01

    Tyrosine aminotransferase (TyrAT) catalyzes the transamination of L-Tyr and α-ketoglutarate, yielding 4-hydroxyphenylpyruvic acid and L-glutamate. The decarboxylation product of 4-hydroxyphenylpyruvic acid, 4-hydroxyphenylacetaldehyde, is a precursor to a large and diverse group of natural products known collectively as benzylisoquinoline alkaloids (BIAs). We have isolated and characterized a TyrAT cDNA from opium poppy (Papaver somniferum), which remains the only commercial source for several pharmaceutical BIAs, including codeine, morphine, and noscapine. TyrAT belongs to group I pyridoxal 5'-phosphate (PLP)-dependent enzymes wherein Schiff base formation occurs between PLP and a specific Lys residue. The amino acid sequence of TyrAT showed considerable homology to other putative plant TyrATs, although few of these have been functionally characterized. Purified, recombinant TyrAT displayed a molecular mass of approximately 46 kD and a substrate preference for L-Tyr and α-ketoglutarate, with apparent K(m) values of 1.82 and 0.35 mm, respectively. No specific requirement for PLP was detected in vitro. Liquid chromatography-tandem mass spectrometry confirmed the conversion of L-Tyr to 4-hydroxyphenylpyruvate. TyrAT gene transcripts were most abundant in roots and stems of mature opium poppy plants. Virus-induced gene silencing was used to evaluate the contribution of TyrAT to BIA metabolism in opium poppy. TyrAT transcript levels were reduced by at least 80% in silenced plants compared with controls and showed a moderate reduction in total alkaloid content. The modest correlation between transcript levels and BIA accumulation in opium poppy supports a role for TyrAT in the generation of alkaloid precursors, but it also suggests the occurrence of other sources for 4-hydroxyphenylacetaldehyde.

  4. Purification and properties of 7, 8-diaminopelargonic acid aminotransferase.

    PubMed

    Stoner, G L; Eisenberg, M A

    1975-06-10

    The enzyme 7, 8-diaminopelargonic acid aminotransferase utilizes S-adenosyl-L-methionine to transaminate the biotin precurson 7-keto-8-aminopelargonic acid and form the next intermediate in the pathway, 7, 8-diaminopelargonic acid. The enzyme has been purified nearly 1000-fold from an extract of a regulatory mutant of Escherichia coli which is derepressed for the enzymes of the biotin operon. The extract was treated with protamine sulfate, ammonium sulfate, and subjected to acid and heat treatments. Subsequently, the enzyme was chromatographed on columns of DEAE-cellulose, phosphocellulose, hydroxylapatite, and two Sephadex G-100. The resulting purified preparation was judged 86% homogeneous by the scanning of of a stained disc gel. The enzymatic activity was associated with the major band in gels run at two different gel concentrations and two different pH values. The cofactor, pyridoxal phosphate, can be resolved from the enzyme in the presence of phosphate buffer after incubation with the amino donor, S-adenosyl-L-methionine. A molecular weight estimation of 94,000 plus or minus 10, 000 has been obtained by gel filtration and sucrose gradient sedimentation studies. Gel electrophoresis in the presence of sodium dodecyl sulfate, shows a single subunit with a molecular weight of 47, 000 plus or minus 3, 000 indicating a dimeric enzyme. A neutral compound was detected in the acidified reaction mixture which was derived from the methionine moiety of S-adenosyl-L-methionine and was present in amounts equivalent to the 7, 8-diaminopelargonic acid produced in the reaction mixture. It is suggested that the keto product of the reaction, i.e. S-adenosyl-2-oxo-4-methylthiobutyric acid, may decompose nonenzymatically under the conditions of the reaction to form 5'-methylthioadenosine and the neutral compound, 2-oxo-3-butenoic acid.

  5. Protective effect of creatine against inhibition by methylglyoxal of mitochondrial respiration of cardiac cells.

    PubMed

    Roy, Soumya Sinha; Biswas, Swati; Ray, Manju; Ray, Subhankar

    2003-06-01

    Previous publications from our laboratory have shown that methylglyoxal inhibits mitochondrial respiration of malignant and cardiac cells, but it has no effect on mitochondrial respiration of other normal cells [Biswas, Ray, Misra, Dutta and Ray (1997) Biochem. J. 323, 343-348; Ray, Biswas and Ray (1997) Mol. Cell. Biochem. 171, 95-103]. However, this inhibitory effect of methylglyoxal is not significant in cardiac tissue slices. Moreover, post-mitochondrial supernatant (PMS) of cardiac cells could almost completely protect the mitochondrial respiration against the inhibitory effect of methylglyoxal. A systematic search indicated that creatine present in cardiac cells is responsible for this protective effect. Glutathione has also some protective effect. However, creatine phosphate, creatinine, urea, glutathione disulphide and beta-mercaptoethanol have no protective effect. The inhibitory and protective effects of methylglyoxal and creatine respectively on cardiac mitochondrial respiration were studied with various concentrations of both methylglyoxal and creatine. Interestingly, neither creatine nor glutathione have any protective effect on the inhibition by methylglyoxal on the mitochondrial respiration of Ehrlich ascites carcinoma cells. The creatine and glutathione contents of several PMS, which were tested for the possible protective effect, were measured. The activities of two important enzymes, namely glyoxalase I and creatine kinase, which act upon glutathione plus methylglyoxal and creatine respectively, were also measured in different PMS. Whether mitochondrial creatine kinase had any role in the protective effect of creatine had also been investigated using 1-fluoro-2,4-dinitrobenzene, an inhibitor of creatine kinase. The differential effect of creatine on mitochondria of cardiac and malignant cells has been discussed with reference to the therapeutic potential of methylglyoxal.

  6. Dietary Creatine as a Possible Novel Treatment for Crohn’s Ileitis

    PubMed Central

    Lee, David

    2016-01-01

    Creatine, a commonly used dietary supplement, plays an important role in maintaining gut barrier function. Given that dysregulation of the intestinal epithelial barrier is a hallmark of inflammatory bowel disease, it is plausible that creatine supplementation may attenuate disease severity. We present a patient with Crohn’s ileitis who responded to creatine supplementation with both symptomatic and endoscopic improvement in disease activity. PMID:28008406

  7. Creatine supplementation enhances corticomotor excitability and cognitive performance during oxygen deprivation.

    PubMed

    Turner, Clare E; Byblow, Winston D; Gant, Nicholas

    2015-01-28

    Impairment or interruption of oxygen supply compromises brain function and plays a role in neurological and neurodegenerative conditions. Creatine is a naturally occurring compound involved in the buffering, transport, and regulation of cellular energy, with the potential to replenish cellular adenosine triphosphate without oxygen. Creatine is also neuroprotective in vitro against anoxic/hypoxic damage. Dietary creatine supplementation has been associated with improved symptoms in neurological disorders defined by impaired neural energy provision. Here we investigate, for the first time in humans, the utility of creatine as a dietary supplement to protect against energetic insult. The aim of this study was to assess the influence of oral creatine supplementation on the neurophysiological and neuropsychological function of healthy young adults during acute oxygen deprivation. Fifteen healthy adults were supplemented with creatine and placebo treatments for 7 d, which increased brain creatine on average by 9.2%. A hypoxic gas mixture (10% oxygen) was administered for 90 min, causing global oxygen deficit and impairing a range of neuropsychological processes. Hypoxia-induced decrements in cognitive performance, specifically attentional capacity, were restored when participants were creatine supplemented, and corticomotor excitability increased. A neuromodulatory effect of creatine via increased energy availability is presumed to be a contributing factor of the restoration, perhaps by supporting the maintenance of appropriate neuronal membrane potentials. Dietary creatine monohydrate supplementation augments neural creatine, increases corticomotor excitability, and prevents the decline in attention that occurs during severe oxygen deficit. This is the first demonstration of creatine's utility as a neuroprotective supplement when cellular energy provision is compromised.

  8. Alanine repeats influence protein localization in splicing speckles and paraspeckles.

    PubMed

    Chang, Shuo-Hsiu; Chang, Wei-Lun; Lu, Chia-Chen; Tarn, Woan-Yuh

    2014-12-16

    Mammalian splicing regulatory protein RNA-binding motif protein 4 (RBM4) has an alanine repeat-containing C-terminal domain (CAD) that confers both nuclear- and splicing speckle-targeting activities. Alanine-repeat expansion has pathological potential. Here we show that the alanine-repeat tracts influence the subnuclear targeting properties of the RBM4 CAD in cultured human cells. Notably, truncation of the alanine tracts redistributed a portion of RBM4 to paraspeckles. The alanine-deficient CAD was sufficient for paraspeckle targeting. On the other hand, alanine-repeat expansion reduced the mobility of RBM4 and impaired its splicing activity. We further took advantage of the putative coactivator activator (CoAA)-RBM4 conjoined splicing factor, CoAZ, to investigate the function of the CAD in subnuclear targeting. Transiently expressed CoAZ formed discrete nuclear foci that emerged and subsequently separated-fully or partially-from paraspeckles. Alanine-repeat expansion appeared to prevent CoAZ separation from paraspeckles, resulting in their complete colocalization. CoAZ foci were dynamic but, unlike paraspeckles, were resistant to RNase treatment. Our results indicate that the alanine-rich CAD, in conjunction with its conjoined RNA-binding domain(s), differentially influences the subnuclear localization and biogenesis of RBM4 and CoAZ.

  9. The influence of creatine supplementation on the cognitive functioning of vegetarians and omnivores.

    PubMed

    Benton, David; Donohoe, Rachel

    2011-04-01

    Creatine when combined with P forms phosphocreatine that acts as a reserve of high-energy phosphate. Creatine is found mostly in meat, fish and other animal products, and the levels of muscle creatine are known to be lower in vegetarians. Creatine supplementation influences brain functioning as indicated by imaging studies and the measurement of oxygenated Hb. Given the key role played by creatine in the provision of energy, the influence of its supplementation on cognitive functioning was examined, contrasting the effect in omnivores and vegetarians. Young adult females (n 128) were separated into those who were and were not vegetarian. Randomly and under a double-blind procedure, subjects consumed either a placebo or 20 g of creatine supplement for 5 d. Creatine supplementation did not influence measures of verbal fluency and vigilance. However, in vegetarians rather than in those who consume meat, creatine supplementation resulted in better memory. Irrespective of dietary style, the supplementation of creatine decreased the variability in the responses to a choice reaction-time task.

  10. Novel creatine biosensors based on all solid-state contact ammonium-selective membrane electrodes.

    PubMed

    Altikatoglu, Melda; Karakus, Emine; Erci, Vildan; Pekyardımcı, Sule; Isildak, Ibrahim

    2013-04-01

    Novel creatine bienzymatic potentiometric biosensors were prepared by immobilizing urease and creatinase on all solid-state contact PVC-containing palmitic acid and carboxylated PVC matrix membrane ammonium-selective electrodes without inner reference solution. Potentiometric characteristics of biosensors were examined in physiological model solutions at different creatine concentrations. The linear working range and long-term sensitivity of the biosensors were also determined. The creatine biosensors prepared by using the carboxylated PVC membrane electrodes showed more effective performance than those of the PVC containing palmitic acid membrane electrodes. Creatine assay in serum samples was successfully carried out by using the standard addition method.

  11. Creatine inhibits adipogenesis by downregulating insulin-induced activation of the phosphatidylinositol 3-kinase signaling pathway.

    PubMed

    Lee, Nayeon; Kim, Inhee; Park, Soojeong; Han, Dasol; Ha, Soobong; Kwon, Mookwang; Kim, Juwan; Byun, Sung-Hyun; Oh, Wonil; Jeon, Hong Bae; Kweon, Dae-Hyuk; Cho, Jae Youl; Yoon, Keejung

    2015-04-15

    Creatine is a nitrogenous organic acid known to function in adenosine triphosphate (ATP) metabolism. Recent evidence indicates that creatine regulates the differentiation of mesenchymal stem cells (MSCs) in processes such as osteogenesis and myogenesis. In this study, we show that creatine also has a negative regulatory effect on fat cell formation. Creatine inhibits the accumulation of cytoplasmic triglycerides in a dose-dependent manner irrespective of the adipogenic cell models used, including a C3H10T1/2 MSC line, 3T3-L1 preadipocytes, and primary human MSCs. Consistently, a dramatic reduction in mRNA expression of adipogenic transcription factors, peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer-binding protein α (C/EBPα), glucose transporters, 1 and 4 (Glut1, Glut4), and adipocyte markers, aP2 and adipsin, was observed in the presence of creatine. Creatine appears to exert its inhibitory effects on adipogenesis during early differentiation, but not late differentiation, or proliferation stages through inhibition of the PI3K-Akt-PPARγ signaling pathway. In an in vivo model, administration of creatine into mice resulted in body mass increase without fat accumulation. In summary, our results indicate that creatine downregulates adipogenesis through inhibition of phosphatidylinositol 3-kinase (PI3K) activation and imply the potent therapeutic value of creatine in treating obesity and obesity-related metabolic disorders.

  12. Maternal creatine supplementation affects the morpho-functional development of hippocampal neurons in rat offspring.

    PubMed

    Sartini, S; Lattanzi, D; Ambrogini, P; Di Palma, M; Galati, C; Savelli, D; Polidori, E; Calcabrini, C; Rocchi, M B L; Sestili, P; Cuppini, R

    2016-01-15

    Creatine supplementation has been shown to protect neurons from oxidative damage due to its antioxidant and ergogenic functions. These features have led to the hypothesis of creatine supplementation use during pregnancy as prophylactic treatment to prevent CNS damage, such as hypoxic-ischemic encephalopathy. Unfortunately, very little is known on the effects of creatine supplementation during neuron differentiation, while in vitro studies revealed an influence on neuron excitability, leaving the possibility of creatine supplementation during the CNS development an open question. Using a multiple approach, we studied the hippocampal neuron morphological and functional development in neonatal rats born by dams supplemented with 1% creatine in drinking water during pregnancy. CA1 pyramidal neurons of supplemented newborn rats showed enhanced dendritic tree development, increased LTP maintenance, larger evoked-synaptic responses, and higher intrinsic excitability in comparison to controls. Moreover, a faster repolarizing phase of action potential with the appearance of a hyperpolarization were recorded in neurons of the creatine-treated group. Consistently, CA1 neurons of creatine exposed pups exhibited a higher maximum firing frequency than controls. In summary, we found that creatine supplementation during pregnancy positively affects morphological and electrophysiological development of CA1 neurons in offspring rats, increasing neuronal excitability. Altogether, these findings emphasize the need to evaluate the benefits and the safety of maternal intake of creatine in humans.

  13. Age dependence of myosin heavy chain transitions induced by creatine depletion in rat skeletal muscle

    NASA Technical Reports Server (NTRS)

    Adams, Gregory R.; Baldwin, Kenneth M.

    1995-01-01

    This study was designed to test the hypothesis that myosin heavy chain (MHC) plasticity resulting from creatine depletion is an age-dependent process. At weaning (age 28 days), rat pups were placed on either standard rat chow (normal diet juvenile group) or the same chow supplemented with 1% wt/wt of the creatine analogue beta-guanidinopropionic acid (creatine depletion juvenile (CDJ) group). Two groups of adult rats (age approximately 8 wk) were placed on the same diet regimens (normal diet adult and creatine depletion adult (CDA) groups). After 40 days (CDJ and normal diet juvenile groups) and 60 days (CDA and normal diet adult groups), animals were killed and several skeletal muscles were removed for analysis of creatine content or MHC ditribution. In the CDJ group, creatine depletion (78%) was accompanied by significant shifts toward expression of slower MHC isoforms in two slow and three fast skeletal muscles. In contrast, creatine depletion in adult animals did not result in similar shifts toward slow MHC isoform expression in either muscle type. The results of this study indicate that there is a differential effect of creatine depletion on MHC tranitions that appears to be age dependent. These results strongly suggest that investigators contemplating experimental designs involving the use of the creatine analogue beta-guanidinopropionic acid should consider the age of the animals to be used.

  14. Novel sensory surface for creatine kinase electrochemical detection.

    PubMed

    Moreira, Felismina T C; Dutra, Rosa A F; Noronha, João P; Sales, M Goreti F

    2014-06-15

    This work describes a novel concept of biosensor for quantifying enzymes, where the substrate is immobilized directly over the working area of a screen printed electrode (Au-SPE). This concept is applied here to creatine kinase isoenzyme (CK-MB), a cardiac biomarker in ischemic conditions. It acts as a phospho-transferase on creatine (Crea), requiring the presence of phosphate. So, the phosphorylated form of creatine (Pcrea) was immobilized on the Au/SPE previously aminated with cysteamine (Cys) by self-assembling monolayer technique. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) studies were used to follow the chemical modifications in the Au-SPE. Since Pcrea is an electroactive species at low potential, its consumption over the platform by the enzyme changed the electrical response of the biosensor. So, CK-MB determination has been achieved in mediator free-conditions due the redox proprieties of the Pcrea. The analytical features of the resulting biosensor were studied by square wave voltammetry (SWV). The limit of detection was 0.11 µg/mL and the slope was -0.029(± 0.0035) µA × mL/µg. The interference effect of troponin T (TnT), bovine serum albumin (BSA) and myoglobin (Myo) in the performance of the sensor was tested and good selectivity was observed. The biosensor was successfully applied to biological fluids, showing good stability at room temperature and excellent sensitivity and selectivity. This new concept of biosensor is especially useful for point of care (POC) applications, due to the low cost and small size of the final device.

  15. Production of D-Alanine by Corynebacterium fascians

    PubMed Central

    Yamada, Shigeki; Maeshima, Haruko; Wada, Mitsuru; Chibata, Ichiro

    1973-01-01

    A strain identified as Corynebacterium fascians was found to accumulate extracellular D-alanine from glycerol. Cultural conditions for the accumulation of D-alanine were investigated and, as a result, a yield of 7 g of D-alanine per liter was obtained after a 96-h incubation in a medium containing 5% glycerol, 4% (NH4)2HPO4, and 0.3% corn steep liquor. Optical purity of D-alanine was dependent upon the concentration of corn steep liquor. At the optimal condition, almost optically pure D-alanine was formed and readily isolated (5 g/liter) from the fermentation broth. The product was not contaminated with any detectable amount of other amino acids, except for glycine which was present at a concentration of less than 1 percent. PMID:4699220

  16. The structure of alanine racemase from Acinetobacter baumannii.

    PubMed

    Davis, Emily; Scaletti-Hutchinson, Emma; Opel-Reading, Helen; Nakatani, Yoshio; Krause, Kurt L

    2014-09-01

    Acinetobacter baumannii is an opportunistic Gram-negative bacterium which is a common cause of hospital-acquired infections. Numerous antibiotic-resistant strains exist, emphasizing the need for the development of new antimicrobials. Alanine racemase (Alr) is a pyridoxal 5'-phosphate dependent enzyme that is responsible for racemization between enantiomers of alanine. As D-alanine is an essential component of the bacterial cell wall, its inhibition is lethal to prokaryotes, making it an excellent antibiotic drug target. The crystal structure of A. baumannii alanine racemase (AlrAba) from the highly antibiotic-resistant NCTC13302 strain has been solved to 1.9 Å resolution. Comparison of AlrAba with alanine racemases from closely related bacteria demonstrates a conserved overall fold. The substrate entryway and active site of the enzymes were shown to be highly conserved. The structure of AlrAba will provide the template required for future structure-based drug-design studies.

  17. Chronotypic induction of tyrosine aminotransferase by. cap alpha. -methyl-p-tyrosine. [Rat liver

    SciTech Connect

    Cahill, A.L.; Ferguson, S.M.; Ehret, C.F.

    1981-04-06

    ..cap alpha..Methyl-p-tyrosine induced hepatic tyrosine aminotransferase activity to different extents depending upon the time of day of administration of the drug. Maximal induction occurred when ..cap alpha..-methyl-p-tyrosine (100 mg/kg) was injected intraperitoneally during the first several hours of the light phase of the daily cycle, but the magnitude of the induction depended on the nutritional state of the animal. Induction was 4- to 5-fold greater in fasting rats. The effect of ..cap alpha..-methyl-ptyrosine on hepatic tyrosine aminotransferase is believed to be mediated by decreases in hypothalamic norepinephrine. This hypothesis was supported by the demonstration that decreasing levels of hypothalamic norepinephrine at times of day when hypothalamic turnover of norepinephrine was greatest resulted in the greatest induction of tyrosine aminotransferase, while lowering hypothalamic norepinephrine at times when turnover was minimal resulted in minimal induction of tyrosine aminotransferase.

  18. An alternate method for demonstration of erythrocytic aminotransferases on starch gels

    PubMed Central

    Scott, Edward M.; Wright, Rita C.

    1981-01-01

    A two-stage procedure using MTT tetrazolium for the demonstration of aminotransferases (GPT and GOT) either singly or together was developed. Identification of phenotypes was unequivocal in over 99% of the individuals studied. ImagesFig. 1 PMID:7258187

  19. Plastidic aspartate aminotransferases and the biosynthesis of essential amino acids in plants.

    PubMed

    de la Torre, Fernando; Cañas, Rafael A; Pascual, M Belén; Avila, Concepción; Cánovas, Francisco M

    2014-10-01

    In the chloroplasts and in non-green plastids of plants, aspartate is the precursor for the biosynthesis of different amino acids and derived metabolites that play distinct and important roles in plant growth, reproduction, development or defence. Aspartate biosynthesis is mediated by the enzyme aspartate aminotransferase (EC 2.6.1.1), which catalyses the reversible transamination between glutamate and oxaloacetate to generate aspartate and 2-oxoglutarate. Plastids contain two aspartate aminotransferases: a eukaryotic-type and a prokaryotic-type bifunctional enzyme displaying aspartate and prephenate aminotransferase activities. A general overview of the biochemistry, regulation, functional significance, and phylogenetic origin of both enzymes is presented. The roles of these plastidic aminotransferases in the biosynthesis of essential amino acids are discussed.

  20. Comparison of EPR response of alanine and Gd₂O₃-alanine dosimeters exposed to TRIGA Mainz reactor.

    PubMed

    Marrale, M; Schmitz, T; Gallo, S; Hampel, G; Longo, A; Panzeca, S; Tranchina, L

    2015-12-01

    In this work we report some preliminary results regarding the analysis of electron paramagnetic resonance (EPR) response of alanine pellets and alanine pellets added with gadolinium used for dosimetry at the TRIGA research reactor in Mainz, Germany. Two set-ups were evaluated: irradiation inside PMMA phantom and irradiation inside boric acid phantom. We observed that the presence of Gd2O3 inside alanine pellets increases the EPR signal by a factor of 3.45 and 1.24 in case of PMMA and boric acid phantoms, respectively. We can conclude that in the case of neutron beam with a predominant thermal neutron component the addition of gadolinium oxide can significantly improve neutron sensitivity of alanine pellets. Monte Carlo (MC) simulations of both response of alanine and Gd-added alanine pellets with FLUKA code were performed and a good agreement was achieved for pure alanine dosimeters. For Gd2O3-alanine deviations between MC simulations and experimental data were observed and discussed.

  1. l-glutamine and l-alanine supplementation increase glutamine-glutathione axis and muscle HSP-27 in rats trained using a progressive high-intensity resistance exercise.

    PubMed

    Leite, Jaqueline Santos Moreira; Raizel, Raquel; Hypólito, Thaís Menezes; Rosa, Thiago Dos Santos; Cruzat, Vinicius Fernandes; Tirapegui, Julio

    2016-08-01

    In this study we investigated the chronic effects of oral l-glutamine and l-alanine supplementation, either in their free or dipeptide form, on glutamine-glutathione (GLN-GSH) axis and cytoprotection mediated by HSP-27 in rats submitted to resistance exercise (RE). Forty Wistar rats were distributed into 5 groups: sedentary; trained (CTRL); and trained supplemented with l-alanyl-l-glutamine, l-glutamine and l-alanine in their free form (GLN+ALA), or free l-alanine (ALA). All trained animals were submitted to a 6-week ladder-climbing protocol. Supplementations were offered in a 4% drinking water solution for 21 days prior to euthanasia. Plasma glutamine, creatine kinase (CK), myoglobin (MYO), and erythrocyte concentration of reduced GSH and glutathione disulfide (GSSG) were measured. In tibialis anterior skeletal muscle, GLN-GSH axis, thiobarbituric acid reactive substances (TBARS), and the expression of heat shock factor 1 (HSF-1), 27-kDa heat shock protein (HSP-27), and glutamine synthetase were determined. In CRTL animals, high-intensity RE reduced muscle glutamine levels and increased GSSG/GSH rate and TBARS, as well as augmented plasma CK and MYO levels. Conversely, l-glutamine-supplemented animals showed an increase in plasma and muscle levels of glutamine, with a reduction in GSSG/GSH rate, TBARS, and CK. Free l-alanine administration increased plasma glutamine concentration and lowered muscle TBARS. HSF-1 and HSP-27 were high in all supplemented groups when compared with CTRL (p < 0.05). The results presented herein demonstrate that l-glutamine supplemented with l-alanine, in both a free or dipeptide form, improve the GLN-GSH axis and promote cytoprotective effects in rats submitted to high-intensity RE training.

  2. Guanidinoacetate is more effective than creatine at enhancing tissue creatine stores while consequently limiting methionine availability in Yucatan miniature pigs.

    PubMed

    McBreairty, Laura E; Robinson, Jason L; Furlong, Kayla R; Brunton, Janet A; Bertolo, Robert F

    2015-01-01

    Creatine (Cr) is an important high-energy phosphate buffer in tissues with a high energy demand such as muscle and brain and is consequently a highly consumed nutritional supplement. Creatine is synthesized via the S-adenosylmethionine (SAM) dependent methylation of guanidinoacetate (GAA) which is not regulated by a feedback mechanism. The first objective of this study was to determine the effectiveness of GAA at increasing tissue Cr stores. Because SAM is required for other methylation reactions, we also wanted to determine whether an increased creatine synthesis would lead to a lower availability of methyl groups for other methylated products. Three month-old pigs (n = 18) were fed control, GAA- or Cr-supplemented diets twice daily. On day 18 or 19, anesthesia was induced 1-3 hours post feeding and a bolus of [methyl-3H]methionine was intravenously infused. After 30 minutes, the liver was analyzed for methyl-3H incorporation into protein, Cr, phosphatidylcholine (PC) and DNA. Although both Cr and GAA led to higher hepatic Cr concentration, only supplementation with GAA led to higher levels of muscle Cr (P < 0.05). Only GAA supplementation resulted in lower methyl-3H incorporation into PC and protein as well as lower hepatic SAM concentration compared to the controls, suggesting that Cr synthesis resulted in a limited methyl supply for PC and protein synthesis (P < 0.05). Although GAA is more effective than Cr at supporting muscle Cr accretion, further research should be conducted into the long term consequences of a limited methyl supply and its effects on protein and PC homeostasis.

  3. Guanidinoacetate Is More Effective than Creatine at Enhancing Tissue Creatine Stores while Consequently Limiting Methionine Availability in Yucatan Miniature Pigs

    PubMed Central

    McBreairty, Laura E.; Robinson, Jason L.; Furlong, Kayla R.; Brunton, Janet A.; Bertolo, Robert F.

    2015-01-01

    Creatine (Cr) is an important high-energy phosphate buffer in tissues with a high energy demand such as muscle and brain and is consequently a highly consumed nutritional supplement. Creatine is synthesized via the S-adenosylmethionine (SAM) dependent methylation of guanidinoacetate (GAA) which is not regulated by a feedback mechanism. The first objective of this study was to determine the effectiveness of GAA at increasing tissue Cr stores. Because SAM is required for other methylation reactions, we also wanted to determine whether an increased creatine synthesis would lead to a lower availability of methyl groups for other methylated products. Three month-old pigs (n = 18) were fed control, GAA- or Cr-supplemented diets twice daily. On day 18 or 19, anesthesia was induced 1–3 hours post feeding and a bolus of [methyl-3H]methionine was intravenously infused. After 30 minutes, the liver was analyzed for methyl-3H incorporation into protein, Cr, phosphatidylcholine (PC) and DNA. Although both Cr and GAA led to higher hepatic Cr concentration, only supplementation with GAA led to higher levels of muscle Cr (P < 0.05). Only GAA supplementation resulted in lower methyl-3H incorporation into PC and protein as well as lower hepatic SAM concentration compared to the controls, suggesting that Cr synthesis resulted in a limited methyl supply for PC and protein synthesis (P < 0.05). Although GAA is more effective than Cr at supporting muscle Cr accretion, further research should be conducted into the long term consequences of a limited methyl supply and its effects on protein and PC homeostasis. PMID:26110793

  4. Primary structure of a key enzyme in plant tetrapyrrole synthesis: glutamate 1-semialdehyde aminotransferase.

    PubMed Central

    Grimm, B

    1990-01-01

    The formation of delta-aminolevulinate from glutamate 1-semialdehyde (GSA) is catalyzed by glutamate 1-semialdehyde aminotransferase (EC 5.4.3.8). The active form of the barley enzyme appears to be a dimer of identical subunits with a molecular mass of 46 kDa. From the purified enzyme, amino acid sequences of the N-terminal ends of the mature protein as well as an internal peptide were determined. DNA primers deduced from these peptide sequences were used to amplify with the polymerase chain reaction a cDNA sequence encoding part of the enzyme. Screening a cDNA library with this DNA fragment identified a full-length clone encoding the 49,540-Da precursor of the GSA aminotransferase. The transit peptide for chloroplast import consists of 34 amino acids. GSA aminotransferase and a precursor form were expressed on a multicopy plasmid in Escherichia coli. Both recombinant gene products reacted with an antibody against the barley GSA aminotransferase. Active barley GSA aminotransferase expressed in E. coli was shown to be active in assays of bacterial cell extracts. As a gene symbol for barley GSA aminotransferase, Gsa is proposed. Images PMID:2349227

  5. Systems bioenergetics of creatine kinase networks: physiological roles of creatine and phosphocreatine in regulation of cardiac cell function.

    PubMed

    Guzun, R; Timohhina, N; Tepp, K; Gonzalez-Granillo, M; Shevchuk, I; Chekulayev, V; Kuznetsov, A V; Kaambre, T; Saks, V A

    2011-05-01

    Physiological role of creatine (Cr) became first evident in the experiments of Belitzer and Tsybakova in 1939, who showed that oxygen consumption in a well-washed skeletal muscle homogenate increases strongly in the presence of creatine and with this results in phosphocreatine (PCr) production with PCr/O(2) ratio of about 5-6. This was the beginning of quantitative analysis in bioenergetics. It was also observed in many physiological experiments that the contractile force changes in parallel with the alteration in the PCr content. On the other hand, it was shown that when heart function is governed by Frank-Starling law, work performance and oxygen consumption rate increase in parallel without any changes in PCr and ATP tissue contents (metabolic homeostasis). Studies of cellular mechanisms of all these important phenomena helped in shaping new approach to bioenergetics, Molecular System Bioenergetics, a part of Systems Biology. This approach takes into consideration intracellular interactions that lead to novel mechanisms of regulation of energy fluxes. In particular, interactions between mitochondria and cytoskeleton resulting in selective restriction of permeability of outer mitochondrial membrane anion channel (VDAC) for adenine nucleotides and thus their recycling in mitochondria coupled to effective synthesis of PCr by mitochondrial creatine kinase, MtCK. Therefore, Cr concentration and the PCr/Cr ratio became important kinetic parameters in the regulation of respiration and energy fluxes in muscle cells. Decrease in the intracellular contents of Cr and PCr results in a hypodynamic state of muscle and muscle pathology. Many experimental studies have revealed that PCr may play two important roles in the regulation of muscle energetics: first by maintaining local ATP pools via compartmentalized creatine kinase reactions, and secondly by stabilizing cellular membranes due to electrostatic interactions with phospholipids. The second mechanism decreases the

  6. Enzymes of creatine biosynthesis, arginine and methionine metabolism in normal and malignant cells.

    PubMed

    Bera, Soumen; Wallimann, Theo; Ray, Subhankar; Ray, Manju

    2008-12-01

    The creatine/creatine kinase system decreases drastically in sarcoma. In the present study, an investigation of catalytic activities, western blot and mRNA expression unambiguously demonstrates the prominent expression of the creatine-synthesizing enzymes l-arginine:glycine amidinotransferase and N-guanidinoacetate methyltransferase in sarcoma, Ehrlich ascites carcinoma and Sarcoma 180 cells, whereas both enzymes were virtually undetectable in normal muscle. Compared to that of normal animals, these enzymes remained unaffected in the kidney or liver of sarcoma-bearing mice. High activity and expression of mitochondrial arginase II in sarcoma indicated increased ornithine formation. Slightly or moderately higher levels of ornithine, guanidinoacetate and creatinine were observed in sarcoma compared to muscle. Despite the intrinsically low level of creatine in Ehrlich ascites carcinoma and Sarcoma 180 cells, these cells could significantly take up and release creatine, suggesting a functional creatine transport, as verified by measuring mRNA levels of creatine transporter. Transcript levels of arginase II, ornithine-decarboxylase, S-adenosyl-homocysteine hydrolase and methionine-synthase were significantly upregulated in sarcoma and in Ehrlich ascites carcinoma and Sarcoma 180 cells. Overall, the enzymes related to creatine and arginine/methionine metabolism were found to be significantly upregulated in malignant cells. However, the low levels of creatine kinase in the same malignant cells do not appear to be sufficient for the building up of an effective creatine/phosphocreatine pool. Instead of supporting creatine biosynthesis, l-arginine:glycine amidinotransferase and N-guanidinoacetate methyltransferase appear to be geared to support cancer cell metabolism in the direction of polyamine and methionine synthesis because both these compounds are in high demand in proliferating cancer cells.

  7. Myocardial Creatine Levels Do Not Influence Response to Acute Oxidative Stress in Isolated Perfused Heart

    PubMed Central

    Aksentijević, Dunja; Zervou, Sevasti; Faller, Kiterie M. E.; McAndrew, Debra J.; Schneider, Jurgen E.; Neubauer, Stefan; Lygate, Craig A.

    2014-01-01

    Background Multiple studies suggest creatine mediates anti-oxidant activity in addition to its established role in cellular energy metabolism. The functional significance for the heart has yet to be established, but antioxidant activity could contribute to the cardioprotective effect of creatine in ischaemia/reperfusion injury. Objectives To determine whether intracellular creatine levels influence responses to acute reactive oxygen species (ROS) exposure in the intact beating heart. We hypothesised that mice with elevated creatine due to over-expression of the creatine transporter (CrT-OE) would be relatively protected, while mice with creatine-deficiency (GAMT KO) would fare worse. Methods and Results CrT-OE mice were pre-selected for creatine levels 20–100% above wild-type using in vivo 1H–MRS. Hearts were perfused in isovolumic Langendorff mode and cardiac function monitored throughout. After 20 min equilibration, hearts were perfused with either H2O2 0.5 µM (30 min), or the anti-neoplastic drug doxorubicin 15 µM (100 min). Protein carbonylation, creatine kinase isoenzyme activities and phospho-PKCδ expression were quantified in perfused hearts as markers of oxidative damage and apoptotic signalling. Wild-type hearts responded to ROS challenge with a profound decline in contractile function that was ameliorated by co-administration of catalase or dexrazoxane as positive controls. In contrast, the functional deterioration in CrT-OE and GAMT KO hearts was indistinguishable from wild-type controls, as was the extent of oxidative damage and apoptosis. Exogenous creatine supplementation also failed to protect hearts from doxorubicin-induced dysfunction. Conclusions Intracellular creatine levels do not influence the response to acute ROS challenge in the intact beating heart, arguing against creatine exerting (patho-)physiologically relevant anti-oxidant activity. PMID:25272153

  8. Abnormalities in Human Brain Creatine Metabolism in Gulf War Illness Probed with MRS

    DTIC Science & Technology

    2014-12-01

    TYPE Final 3. DATES COVERED 30 Sep 2012 - 29 Sep 2014 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Abnormalities in Human Brain Creatine Metabolism in...1H transverse relaxation times (T2s) of the methyl peaks of the molecules phosphocreatine (PCr) and free creatine (Cr) in brains of ill and well

  9. Structural correlates of the creatine transporter function regulation: the undiscovered country.

    PubMed

    Santacruz, Lucia; Jacobs, Danny O

    2016-08-01

    Creatine (Cr) and phosphocreatine constitute an energy shuttle that links ATP production in mitochondria to subcellular locations of ATP consumption. Cells in tissues that are reliant on this energy shuttle, such as myocytes and neurons, appear to have very limited ability to synthesize creatine. Therefore, these cells depend on Cr uptake across the cell membrane by a specialized creatine transporter (CrT solute carrier SLC6A8) in order to maintain intracellular creatine levels. Cr supplementation has been shown to have a beneficial effect in numerous in vitro and in vivo models, particularly in cases of oxidative stress, and is also widely used by athletes as a performance enhancement nutraceutical. Intracellular creatine content is maintained within narrow limits. However, the physiological and cellular mechanisms that mediate Cr transport during health and disease (such as cardiac failure) are not understood. In this narrative mini-review, we summarize the last three decades of research on CrT structure, function and regulation.

  10. Focally Elevated Creatine Detected in Amyloid Precursor Protein (APP) Transgenic Mice and Alzheimer Disease Brain Tissue

    SciTech Connect

    Gallant,M.; Rak, M.; Szeghalmi, A.; Del Bigio, M.; Westaway, D.; Yang, J.; Julian, R.; Gough, K.

    2006-01-01

    The creatine/phosphocreatine system, regulated by creatine kinase, plays an important role in maintaining energy balance in the brain. Energy metabolism and the function of creatine kinase are known to be affected in Alzheimer diseased brain and in cells exposed to the {beta}-amyloid peptide. We used infrared microspectroscopy to examine hippocampal, cortical, and caudal tissue from 21-89-week-old transgenic mice expressing doubly mutant (K670N/M671L and V717F) amyloid precursor protein and displaying robust pathology from an early age. Microcrystalline deposits of creatine, suggestive of perturbed energetic status, were detected by infrared microspectroscopy in all animals with advanced plaque pathology. Relatively large creatine deposits were also found in hippocampal sections from post-mortem Alzheimer diseased human brain, compared with hippocampus from non-demented brain. We therefore speculate that this molecule is a marker of the disease process.

  11. Function of the D-alanine:D-alanine ligase lid loop: a molecular modeling and bioactivity study.

    PubMed

    Hrast, Martina; Vehar, Blaž; Turk, Samo; Konc, Janez; Gobec, Stanislav; Janežič, Dušanka

    2012-08-09

    D-Alanine:D-alanine ligase (Ddl) is an essential ATP-dependent bacterial enzyme involved in peptidoglycan biosynthesis. Discovery of Ddl inhibitors not competitive with ATP has proven to be difficult because the Ddl bimolecular d-alanine binding pocket is very restricted, as is accessibility to the active site for larger molecules in the catalytically active closed conformation of Ddl. A molecular dynamics study of the opening and closing of the Ddl lid loop informs future structure-based design efforts that allow for the flexibility of Ddl. A virtual screen on generated enzyme conformations yielded some hit inhibitors whose bioactivity was determined.

  12. Cerebral energetic effects of creatine supplementation in humans.

    PubMed

    Pan, J W; Takahashi, K

    2007-04-01

    There has been considerable interest in the use of creatine (Cr) supplementation to treat neurological disorders. However, in contrast to muscle physiology, there are relatively few studies of creatine supplementation in the brain. In this report, we use high-field MR (31)P and (1)H spectroscopic imaging of human brain with a 7-day protocol of oral Cr supplementation to examine its effects on cerebral energetics (phosphocreatine, PCr; ATP) and mitochondrial metabolism (N-acetyl aspartate, NAA; and Cr). We find an increased ratio of PCr/ATP (day 0, 0.80 +/- 0.10; day 7, 0.85 +/- 09), with this change largely due to decreased ATP, from 2.7 +/- 0.3 mM to 2.5 +/- 0.3 mM. The ratio of NAA/Cr also decreased (day 0, 1.32 +/- 0.17; day 7 1.18 +/- 0.13), primarily from increased Cr (9.6 +/- 1.9 to 10.1 +/- 2.0 mM). The Cr-induced changes significantly correlated with the basal state, with the fractional increase in PCr/ATP negatively correlating with the basal PCr/ATP value (R = -0.74, P < 0.001). As NAA is a measure of mitochondrial function, there was also a significant negative correlation between basal NAA concentrations with the fractional change in PCr and ATP. Thus healthy human brain energetics is malleable and shifts with 7 days of Cr supplementation, with the regions of initially low PCr showing the largest increments in PCr. Overall, Cr supplementation appears to improve high-energy phosphate turnover in healthy brain and can result in either a decrease or an increase in high-energy phosphate concentrations.

  13. Impaired Brain Creatine Kinase Activity in Huntington's Disease

    PubMed Central

    Zhang, S.F.; Hennessey, T.; Yang, L.; Starkova, N.N.; Beal, M.F.; Starkov, A.A.

    2011-01-01

    Background Huntington's disease (HD) is associated with impaired energy metabolism in the brain. Creatine kinase (CK) catalyzes ATP-dependent phosphorylation of creatine (Cr) into phosphocreatine (PCr), thereby serving as readily available high-capacity spatial and temporal ATP buffering. Objective: Substantial evidence supports a specific role of the Cr/PCr system in neurodegenerative diseases. In the brain, the Cr/PCr ATP-buffering system is established by a concerted operation of the brain-specific cytosolic enzyme BB-CK and ubiquitous mitochondrial uMt-CK. It is not yet established whether the activity of these CK isoenzymes is impaired in HD. Methods We measured PCr, Cr, ATP and ADP in brain extracts of 3 mouse models of HD – R6/2 mice, N171-82Q and HdhQ111 mice – and the activity of CK in cytosolic and mitochondrial brain fractions from the same mice. Results The PCr was significantly increased in mouse HD brain extracts as compared to nontransgenic littermates. We also found an approximately 27% decrease in CK activity in both cytosolic and mitochondrial fractions of R6/2 and N171-82Q mice, and an approximately 25% decrease in the mitochondria from HdhQ111 mice. Moreover, uMt-CK and BB-CK activities were approximately 63% lower in HD human brain samples as compared to nondiseased controls. Conclusion Our findings lend strong support to the role of impaired energy metabolism in HD, and point out the potential importance of impairment of the CK-catalyzed ATP-buffering system in the etiology of HD. PMID:21124007

  14. [The effect of diet ratio of polyunsaturated fatty acids of omega-3 and omega-6 families on activity of aminotransferases and gamma-glutamyltransferase in rat blood serum].

    PubMed

    Ketsa, O V; Marchenko, M M

    2014-01-01

    The effect of diet fat compositions with various ratio of omega-3 and omega-6 polyunsaturated fatty acids (PUFAs) on alanine aminotransferase (ALT), aspartate aminotransferase (AST) and gamma-glutamyltransferase (GGT) activities in blood serum of 45 white mongrel rats weighing 90-110 g (9 animals in group) has been investigated. Fat components in the semi-synthetic diet, compiled on the basis of AIN-93 diet, and sources of omega-6 and omega-3 PUFA were presented by sunflower oil, soybean oil and fish oil. It has been shown that four-week inclusion of linoleic acid (LA) and alpha-linolenic acid (alpha-LNA) in a ratio of 7:1 into the diet (soybean oil) as well as use of only omega-6 PUFA (sunflower oil) has lead to an increase in the activity of ALT and GGT in rat blood serum compared to control animals treated with the complex of linolenic, eicosapentaenoic (EPA) and docosahexaenoic (DHA) acid through the mixture of sunflower oil and fish oil (9:1) with the ratio of omega-6 and omega-3 PUFA 7:1. Along with this, the AST:ALT ratio (de Ritis ratio) was lower (p < 0.05) as compared with the control group of rat, amounting respectively 0.92 +/- 0.08 and 0.79 +/- 0.12 vs 1.26 +/- 0.10. The use of high doses of omega-3 fatty acids (600 mg EPA and 400 mg DHA per kg of animal weight per day coming through fish oil) did not affect the activity of ALT and GGT, but increased AST serum activity (0.47 +/- 0.04 micromoles/min per mg protein) and the de Ritis ratio (2.53 +/- 0.23). The diet deprived with fat increased enzyme activity of ALT, AST and GGT in rat blood serum.

  15. Relationship Between Hepatic Steatosis and the Elevation of Aminotransferases in HBV-Infected Patients With HBe-Antigen Negativity and a Low Viral Load

    PubMed Central

    Enomoto, Hirayuki; Aizawa, Nobuhiro; Nishikawa, Hiroki; Ikeda, Naoto; Sakai, Yoshiyuki; Takata, Ryo; Hasegawa, Kunihiro; Nakano, Chikage; Nishimura, Takashi; Yoh, Kazunori; Ishii, Akio; Takashima, Tomoyuki; Iwata, Yoshinori; Iijima, Hiroko; Nishiguchi, Shuhei

    2016-01-01

    Abstract Nonalcoholic fatty liver disease has been suggested to be associated with alanine aminotransferase (ALT) elevation in hepatitis B virus (HBV)-infected patients with HBe antigen (HBeAg)-negativity and a low HBV-DNA level. However, few studies have evaluated the association according to histological findings of the liver. Among a total of 198 HBV-infected patients who received a percutaneous liver biopsy, we studied the histological and laboratory findings of HBeAg-negative patients without receiving nucleoside/nucleotide analogues treatment (N = 70) in order to evaluate whether hepatic steatosis and its related metabolic disorders were associated with an elevation in ALT levels in HBeAg-negative patients. In HBeAg-negative patients with a high serum HBV-DNA level (≥2000 IU/mL), the level of HBV-DNA was the only significant factor related to ALT elevation. However, in HBeAg-negative patients with a low HBV-DNA level, the serum ferritin level, and histologically observed hepatic steatosis were significantly associated factors with ALT elevation. When we evaluated 2 metabolic variables (serum ferritin and fasting insulin) that are suggested to be relevant to the presence of progressive disease in Japanese patients, we found that the rate of metabolic disorders was significantly higher among patients with a high ALT level and a low HBV-DNA level than it was among those with other conditions. The triglyceride level and the frequency of moderate or severe hepatic steatosis were significantly higher in patients with a low HBV-DNA level than in those with a high HBV-DNA level. Histologically proven hepatic steatosis and its related metabolic disorders are suggested to be involved in the elevation of aminotransferases of HBeAg-negative patients, particularly those with low HBV-DNA levels. PMID:27124068

  16. D-Amino acid dipeptide production utilizing D-alanine-D-alanine ligases with novel substrate specificity.

    PubMed

    Sato, Masaru; Kirimura, Kohtaro; Kino, Kuniki

    2005-06-01

    D-Alanine-D-alanine ligase (Ddl) is an important enzyme in the synthesis of bacterial peptidoglycan. The genes encoding Ddls from Escherichia coli K12 (EcDdlB), Oceanobacillus iheyensis JCM 11309 (OiDdl), Synechocystis sp. PCC 6803 (SsDdl) and Thermotoga maritima ATCC 43589 (TmDdl), the genomic DNA sequences of which have been determined, were cloned and the substrate specificities of these recombinant Ddls were investigated. Although OiDdl had a high substrate specificity for D-alanine; EcDdlB, SsDdl and TmDdl showed broad substrate specificities for D-serine, D-threonine, D-cysteine and glycine, in addition to D-alanine. Four D-amino acid dipeptides were produced using EcDdlB, and D-amino acid homo-dipeptides were successfully produced at high yields except for D-threonyl-D-threonine.

  17. Immunoglobulin-associated creatine kinase masquerading as macro-creatine kinase type 2 in a statin user.

    PubMed

    Loh, Tze Ping; Ang, Yan Hoon; Neo, Siew Fong; Yin, Cecilia; Wong, Moh Sim; Leong, Sai Mun; Saw, Sharon; Sethi, Sunil K

    2012-01-01

    Macro-creatine kinase (CK) is a cause of falsely elevated CK. Macro-CK type 1 is immunoglobulin-associated CK; type 2 is polymeric mitochondrial-CK. An elderly asymptomatic lady had an elevated CK level after receiving statin therapy. Her CK gel electrophoresis analysis demonstrated coexisting macro-CK type 1 and type 2 patterns. Further analysis by immunofixation and mixing this patient's serum with CK control material revealed an IgG-associated macro-CK that mimicked the electrophoretic pattern of macro-CK type 2. This highly unusual discovery suggests the possibility of the misinterpretation of macro-CK type 1 as macro-CK type 2. Falsely elevated CK is still common despite modern laboratory instrumentation and should be investigated.

  18. Noncovalent and covalent functionalization of a (5, 0) single-walled carbon nanotube with alanine and alanine radicals.

    PubMed

    Rajarajeswari, Muthusivarajan; Iyakutti, Kombiah; Kawazoe, Yoshiyuki

    2012-02-01

    We have systematically investigated the noncovalent and covalent adsorption of alanine and alanine radicals, respectively, onto a (5, 0) single-walled carbon nanotube using first-principles calculation. It was found that XH···π (X = N, O, C) interactions play a crucial role in the non-ovalent adsorption and that the functional group close to the carbon nanotube exhibits a significant influence on the binding strength. Noncovalent functionalization of the carbon nanotube with alanine enhances the conductivity of the metallic (5, 0) nanotube. In the covalent adsorption of each alanine radical onto a carbon nanotube, the binding energy depends on the adsorption site on CNT and the electronegative atom that binds with the CNT. The strongest complex is formed when the alanine radical interacts with a (5, 0) carbon nanotube through the amine group. In some cases, the covalent interaction of the alanine radical introduces a half-filled band at the Fermi level due to the local sp (3) hybridization, which modifies the conductivity of the tube.

  19. Structure of D-alanine-D-alanine ligase from Yersinia pestis: nucleotide phosphate recognition by the serine loop.

    PubMed

    Tran, Huyen Thi; Hong, Myoung Ki; Ngo, Ho Phuong Thuy; Huynh, Kim Hung; Ahn, Yeh Jin; Wang, Zhong; Kang, Lin Woo

    2016-01-01

    D-Alanyl-D-alanine is an essential precursor of bacterial peptidoglycan and is synthesized by D-alanine-D-alanine ligase (DDL) with hydrolysis of ATP; this reaction makes DDL an important drug target for the development of antibacterial agents. Five crystal structures of DDL from Yersinia pestis (YpDDL) were determined at 1.7-2.5 Å resolution: apo, AMP-bound, ADP-bound, adenosine 5'-(β,γ-imido)triphosphate-bound, and D-alanyl-D-alanine- and ADP-bound structures. YpDDL consists of three domains, in which four loops, loop 1, loop 2 (the serine loop), loop 3 (the ω-loop) and loop 4, constitute the binding sites for two D-alanine molecules and one ATP molecule. Some of them, especially the serine loop and the ω-loop, show flexible conformations, and the serine loop is mainly responsible for the conformational change in substrate nucleotide phosphates. Enzyme-kinetics assays were carried out for both the D-alanine and ATP substrates and a substrate-binding mechanism was proposed for YpDDL involving conformational changes of the loops.

  20. Creatine transporter (SLC6A8) knockout mice display an increased capacity for in vitro creatine biosynthesis in skeletal muscle.

    PubMed

    Russell, Aaron P; Ghobrial, Lobna; Wright, Craig R; Lamon, Séverine; Brown, Erin L; Kon, Michihiro; Skelton, Matthew R; Snow, Rodney J

    2014-01-01

    The present study aimed to investigate whether skeletal muscle from whole body creatine transporter (CrT; SLC6A8) knockout mice (CrT(-/y)) actually contained creatine (Cr) and if so, whether this Cr could result from an up regulation of muscle Cr biosynthesis. Gastrocnemius muscle from CrT(-/y) and wild type (CrT(+/y)) mice were analyzed for ATP, Cr, Cr phosphate (CrP), and total Cr (TCr) content. Muscle protein and gene expression of the enzymes responsible for Cr biosynthesis L-arginine:glycine amidotransferase (AGAT) and guanidinoacetate methyltransferase (GAMT) were also determined as were the rates of in vitro Cr biosynthesis. CrT(-/y) mice muscle contained measurable (22.3 ± 4.3 mmol.kg(-1) dry mass), but markedly reduced (P < 0.05) TCr levels compared with CrT(+/y) mice (125.0 ± 3.3 mmol.kg(-1) dry mass). AGAT gene and protein expression were higher (~3 fold; P < 0.05) in CrT(-/y) mice muscle, however GAMT gene and protein expression remained unchanged. The in vitro rate of Cr biosynthesis was elevated 1.5 fold (P < 0.05) in CrT(-/y) mice muscle. These data clearly demonstrate that in the absence of CrT protein, skeletal muscle has reduced, but not absent, levels of Cr. This presence of Cr may be at least partly due to an up regulation of muscle Cr biosynthesis as evidenced by an increased AGAT protein expression and in vitro Cr biosynthesis rates in CrT(-/y) mice. Of note, the up regulation of Cr biosynthesis in CrT(-/y) mice muscle was unable to fully restore Cr levels to that found in wild type muscle.

  1. Use of creatine in the elderly and evidence for effects on cognitive function in young and old.

    PubMed

    Rawson, Eric S; Venezia, Andrew C

    2011-05-01

    The ingestion of the dietary supplement creatine (about 20 g/day for 5 days or about 2 g/day for 30 days) results in increased skeletal muscle creatine and phosphocreatine. Subsequently, the performance of high-intensity exercise tasks, which rely heavily on the creatine-phosphocreatine energy system, is enhanced. The well documented benefits of creatine supplementation in young adults, including increased lean body mass, increased strength, and enhanced fatigue resistance are particularly important to older adults. With aging and reduced physical activity, there are decreases in muscle creatine, muscle mass, bone density, and strength. However, there is evidence that creatine ingestion may reverse these changes, and subsequently improve activities of daily living. Several groups have demonstrated that in older adults, short-term high-dose creatine supplementation, independent of exercise training, increases body mass, enhances fatigue resistance, increases muscle strength, and improves the performance of activities of daily living. Similarly, in older adults, concurrent creatine supplementation and resistance training increase lean body mass, enhance fatigue resistance, increase muscle strength, and improve performance of activities of daily living to a greater extent than resistance training alone. Additionally, creatine supplementation plus resistance training results in a greater increase in bone mineral density than resistance training alone. Higher brain creatine is associated with improved neuropsychological performance, and recently, creatine supplementation has been shown to increase brain creatine and phosphocreatine. Subsequent studies have demonstrated that cognitive processing, that is either experimentally (following sleep deprivation) or naturally (due to aging) impaired, can be improved with creatine supplementation. Creatine is an inexpensive and safe dietary supplement that has both peripheral and central effects. The benefits afforded to

  2. The Effects of Creatine Supplementation on Explosive Performance and Optimal Individual Postactivation Potentiation Time.

    PubMed

    Wang, Chia-Chi; Yang, Ming-Ta; Lu, Kang-Hao; Chan, Kuei-Hui

    2016-03-04

    Creatine plays an important role in muscle energy metabolism. Postactivation potentiation (PAP) is a phenomenon that can acutely increase muscle power, but it is an individualized process that is influenced by muscle fatigue. This study examined the effects of creatine supplementation on explosive performance and the optimal individual PAP time during a set of complex training bouts. Thirty explosive athletes performed tests of back squat for one repetition maximum (1RM) strength and complex training bouts for determining the individual optimal timing of PAP, height and peak power of a counter movement jump before and after the supplementation. Subjects were assigned to a creatine or placebo group and then consumed 20 g of creatine or carboxymethyl cellulose per day for six days. After the supplementation, the 1RM strength in the creatine group significantly increased (p < 0.05). The optimal individual PAP time in the creatine group was also significant earlier than the pre-supplementation and post-supplementation of the placebo group (p < 0.05). There was no significant difference in jump performance between the groups. This study demonstrates that creatine supplementation improves maximal muscle strength and the optimal individual PAP time of complex training but has no effect on explosive performance.

  3. The effect of creatine supplementation on mass and performance of rat skeletal muscle.

    PubMed

    Young, Robert E; Young, John C

    2007-08-09

    This study investigated the effect of dietary creatine supplementation on hypertrophy and performance of rat skeletal muscle. Male Sprague-Dawley rats underwent either tibialis anterior ablation or partial ablation of the plantaris/gastrocnemius to induce compensatory hypertrophy of the extensor digitorum longus (EDL) or soleus respectively, or sham surgery. Creatine (300 mg/kg) was administered to one half of each group for 5 weeks, after which force production was measured. With the leg fixed at the knee and ankle, the distal tendon of the EDL or soleus was attached to a force transducer and the muscle was electrically stimulated via the sciatic nerve. Synergist ablation resulted in a significant increase in EDL mass and in soleus mass relative to control muscles. However, no effect of creatine supplementation on muscle mass or performance was found between control and either group of creatine-treated rats. Despite an apparent increase in muscle creatine content, creatine supplementation did not augment muscle hypertrophy or force production in rat EDL or soleus muscle, providing evidence that the potential benefits of creatine supplementation are not due to a direct effect on muscle but rather to an enhanced ability to train.

  4. The Effects of Creatine Supplementation on Explosive Performance and Optimal Individual Postactivation Potentiation Time

    PubMed Central

    Wang, Chia-Chi; Yang, Ming-Ta; Lu, Kang-Hao; Chan, Kuei-Hui

    2016-01-01

    Creatine plays an important role in muscle energy metabolism. Postactivation potentiation (PAP) is a phenomenon that can acutely increase muscle power, but it is an individualized process that is influenced by muscle fatigue. This study examined the effects of creatine supplementation on explosive performance and the optimal individual PAP time during a set of complex training bouts. Thirty explosive athletes performed tests of back squat for one repetition maximum (1RM) strength and complex training bouts for determining the individual optimal timing of PAP, height and peak power of a counter movement jump before and after the supplementation. Subjects were assigned to a creatine or placebo group and then consumed 20 g of creatine or carboxymethyl cellulose per day for six days. After the supplementation, the 1RM strength in the creatine group significantly increased (p < 0.05). The optimal individual PAP time in the creatine group was also significant earlier than the pre-supplementation and post-supplementation of the placebo group (p < 0.05). There was no significant difference in jump performance between the groups. This study demonstrates that creatine supplementation improves maximal muscle strength and the optimal individual PAP time of complex training but has no effect on explosive performance. PMID:26959056

  5. Caffeine and Creatine Content of Dietary Supplements Consumed by Brazilian Soccer Players.

    PubMed

    Inácio, Suelen Galante; de Oliveira, Gustavo Vieira; Alvares, Thiago Silveira

    2016-08-01

    Caffeine and creatine are ingredients in the most popular dietary supplements consumed by soccer players. However, some products may not contain the disclosed amounts of the ingredients listed on the label, compromising the safe usage and the effectiveness of these supplements. Therefore, the aim of this study was to evaluate the content of caffeine and creatine in dietary supplements consumed by Brazilian soccer players. The results obtained were compared with the caffeine content listed on the product label. Two batches of the supplement brands consumed by ≥ 50% of the players were considered for analysis. The quantification of caffeine and creatine in the supplements was determined by a high-performance liquid chromatography system with UV detector. Nine supplements of caffeine and 7 supplements of creatine met the inclusion criteria for analysis. Eight brands of caffeine and five brands of creatine showed significantly different values (p < .05) as compared with the values stated on the label. There were no significant differences between the two batches of supplements analyzed, except for one caffeine supplement. It can be concluded that caffeine and creatine dietary supplements consumed by Brazilian soccer players present inaccurate values listed on the label, although most presented no difference among batches. To ensure consumer safety and product efficacy, accurate information on caffeine and creatine content should be provided on all dietary supplement labels.

  6. A Pilot Clinical Trial of Creatine and Minocycline in Early Parkinson Disease: 18-Month Results

    PubMed Central

    2015-01-01

    Objective To report an 18-month follow-up on creatine and minocycline futility study, the Neuroprotective Exploratory Trials in Parkinson Disease, Futility Study 1 (NET-PD FS-1). Background The NET-PD FS-1 futility study on creatine and minocycline found neither agent futile in slowing down the progression of disability in Parkinson disease (PD) at 12 months using the prespecified futility threshold. An additional 6 months of follow-up aimed to assess safety and potential interactions of the study interventions with anti-parkinsonian therapy. Methods Additional 6 months of follow-up in randomized, blinded phase II trial of creatine (dosage, 10 g/d) and minocycline (dosage, 200 mg/d) in subjects with early PD. Results By 18 months, symptomatic treatment of PD symptoms was required in 61% of creatine, 62% of minocycline, and 60% of placebo-treated subjects. Study treatment was prematurely discontinued in 9%, 23%, and 6% of subjects in the creatine, minocycline, and placebo arms, respectively. Creatine and minocycline did not seem to adversely influence the response to symptomatic therapy nor increase adverse events. Conclusions Data from this small, 18-month phase II trial of creatine and minocycline do not demonstrate safety concerns that would preclude a large, phase III efficacy trial, although the decreased tolerability of minocycline is a concern. PMID:18520981

  7. Effect of creatine and pioglitazone on Hk-2 cell line cisplatin nephrotoxicity.

    PubMed

    Genc, Gurkan; Kilinc, Veli; Bedir, Abdulkerim; Ozkaya, Ozan

    2014-08-01

    Cisplatin is a chemotherapeutic agent, which is used in the treatment of various solid organ cancers, and its main dose limiting side effect of cisplatin is nephrotoxicity. The aim of this study is to investigate the role of pioglitazone and creatine on cisplatin nephrotoxicity in vitro. Real-time cell analyzer system (RTCA) was used for real-time and time-dependent analysis of the cellular response of HK-2 cells following incubation with cisplatin and combination with creatine or pioglitazone hydrochloride. First, half-maximal inhibitory concentrations (IC50) of cisplatin, creatine and pioglitazone were calculated by RTCA system. Afterwards creatine and pioglitazone was administered with serial dilutions under RTCA system. IC50 dose for cisplatin was 7.69 M × 10(-5) at 24th hour and 3.93 M × 10(-6) at 48th hour. IC50 dose for pioglitazone was 1.61 M × 10(-3) at 24th hour and 2.85 M × 10(-4) at 48th hour. Although cells were treated the dose of 40,225 mM creatine, IC50 dose could not been reached. Neither pioglitazone nor creatine had additional protective effect in any dose. Consequently, beneficial effect of creatine and pioglitazone on cisplatin-induced cell death could not be found. Further studies and clinical trials are needed to evaluate the effect of different doses of these drugs in cisplatin-induced nephrotoxicity.

  8. Creatine as a compatible osmolyte in muscle cells exposed to hypertonic stress

    PubMed Central

    Alfieri, Roberta R; Bonelli, Mara A; Cavazzoni, Andrea; Brigotti, Maurizio; Fumarola, Claudia; Sestili, Piero; Mozzoni, Paola; De Palma, Giuseppe; Mutti, Antonio; Carnicelli, Domenica; Vacondio, Federica; Silva, Claudia; Borghetti, Angelo F; Wheeler, Kenneth P; Petronini, Pier Giorgio

    2006-01-01

    Exposure of C2C12 muscle cells to hypertonic stress induced an increase in cell content of creatine transporter mRNA and of creatine transport activity, which peaked after about 24 h incubation at 0.45 osmol (kg H2O)−1. This induction of transport activity was prevented by addition of either cycloheximide, to inhibit protein synthesis, or of actinomycin D, to inhibit RNA synthesis. Creatine uptake by these cells is largely Na+ dependent and kinetic analysis revealed that its increase under hypertonic conditions resulted from an increase in Vmax of the Na+-dependent component, with no significant change in the Km value of about 75 μmol l−1. Quantitative real-time PCR revealed a more than threefold increase in the expression of creatine transporter mRNA in cells exposed to hypertonicity. Creatine supplementation significantly enhanced survival of C2C12 cells incubated under hypertonic conditions and its effect was similar to that obtained with the well known compatible osmolytes, betaine, taurine and myo-inositol. This effect seemed not to be linked to the energy status of the C2C12 cells because hypertonic incubation caused a decrease in their ATP content, with or without the addition of creatine at 20 mmol l−1 to the medium. This induction of creatine transport activity by hypertonicity is not confined to muscle cells: a similar induction was shown in porcine endothelial cells. PMID:16873409

  9. Total body skeletal muscle mass: estimation by creatine (methyl-d3) dilution in humans.

    PubMed

    Clark, Richard V; Walker, Ann C; O'Connor-Semmes, Robin L; Leonard, Michael S; Miller, Ram R; Stimpson, Stephen A; Turner, Scott M; Ravussin, Eric; Cefalu, William T; Hellerstein, Marc K; Evans, William J

    2014-06-15

    Current methods for clinical estimation of total body skeletal muscle mass have significant limitations. We tested the hypothesis that creatine (methyl-d3) dilution (D3-creatine) measured by enrichment of urine D3-creatinine reveals total body creatine pool size, providing an accurate estimate of total body skeletal muscle mass. Healthy subjects with different muscle masses [n = 35: 20 men (19-30 yr, 70-84 yr), 15 postmenopausal women (51-62 yr, 70-84 yr)] were housed for 5 days. Optimal tracer dose was explored with single oral doses of 30, 60, or 100 mg D3-creatine given on day 1. Serial plasma samples were collected for D3-creatine pharmacokinetics. All urine was collected through day 5. Creatine and creatinine (deuterated and unlabeled) were measured by liquid chromatography mass spectrometry. Total body creatine pool size and muscle mass were calculated from D3-creatinine enrichment in urine. Muscle mass was also measured by magnetic resonance imaging (MRI), dual-energy x-ray absorptiometry (DXA), and traditional 24-h urine creatinine. D3-creatine was rapidly absorbed and cleared with variable urinary excretion. Isotopic steady-state of D3-creatinine enrichment in the urine was achieved by 30.7 ± 11.2 h. Mean steady-state enrichment in urine provided muscle mass estimates that correlated well with MRI estimates for all subjects (r = 0.868, P < 0.0001), with less bias compared with lean body mass assessment by DXA, which overestimated muscle mass compared with MRI. The dilution of an oral D3-creatine dose determined by urine D3-creatinine enrichment provides an estimate of total body muscle mass strongly correlated with estimates from serial MRI with less bias than total lean body mass assessment by DXA.

  10. The effects of creatine monohydrate supplementation with and without D-pinitol on resistance training adaptations.

    PubMed

    Kerksick, Chad M; Wilborn, Colin D; Campbell, William I; Harvey, Travis M; Marcello, Brandon M; Roberts, Mike D; Parker, Adam G; Byars, Allyn G; Greenwood, Lori D; Almada, Anthony L; Kreider, Richard B; Greenwood, Mike

    2009-12-01

    Coingestion of D-pinitol with creatine (CR) has been reported to enhance creatine uptake. The purpose of this study was to evaluate whether adding D-pinitol to CR affects training adaptations, body composition, whole-body creatine retention, and/or blood safety markers when compared to CR ingestion alone after 4 weeks of resistance training. Twenty-four resistance trained males were randomly assigned in a double-blind manner to creatine + pinitol (CRP) or creatine monohydrate (CR) prior to beginning a supervised 4-week resistance training program. Subjects ingested a typical loading phase (i.e., 20 g/d-1 for 5 days) before ingesting 5 g/d-1 the remaining 23 days. Performance measures were assessed at baseline (T0), week 1 (T1), and week 4 (T2) and included 1 repetition maximum (1RM) bench press (BP), 1RM leg press (LP), isokinetic knee extension, and a 30-second Wingate anaerobic capacity test. Fasting blood and body composition using dual-energy x-ray absorptiometry (DEXA) were determined at T1 and T3. Data were analyzed by repeated measures analysis of variance (ANOVA). Creatine retention increased (p < 0.001) in both groups as a result of supplementation but was not different between groups (p > 0.05). Significant improvements in upper- and lower-body strength and body composition occurred in both groups. However, significantly greater increases in lean mass and fat-free mass occurred in the CR group when compared to CRP (p <0.05). Adding D-pinitol to creatine monohydrate does not appear to facilitate further physiological adaptations while resistance training. Creatine monohydrate supplementation helps to improve strength and body composition while resistance training. Data from this study assist in determining the potential role the addition of D-pinitol to creatine may aid in facilitating training adaptations to exercise.

  11. Creatine affords protection against glutamate-induced nitrosative and oxidative stress.

    PubMed

    Cunha, Mauricio P; Lieberknecht, Vicente; Ramos-Hryb, Ana Belén; Olescowicz, Gislaine; Ludka, Fabiana K; Tasca, Carla I; Gabilan, Nelson H; Rodrigues, Ana Lúcia S

    2016-05-01

    Creatine has been reported to exert beneficial effects in several neurodegenerative diseases in which glutamatergic excitotoxicity and oxidative stress play an etiological role. The purpose of this study was to investigate the protective effects of creatine, as compared to the N-Methyl-d-Aspartate (NMDA) receptor antagonist dizocilpine (MK-801), against glutamate or hydrogen peroxide (H2O2)-induced injury in human neuroblastoma SH-SY5Y cells. Exposure of cells to glutamate (60-80 mM) or H2O2 (200-300 μM) for 24 h decreased cellular viability and increased dichlorofluorescein (DCF) fluorescence (indicative of increased reactive oxygen species, ROS) and nitric oxide (NO) production (assessed by mono-nitrogen oxides, NOx, levels). Creatine (1-10 mM) or MK-801 (0.1-10 μM) reduced glutamate- and H2O2-induced toxicity. The protective effect of creatine against glutamate-induced toxicity involves its antioxidant effect, since creatine, similar to MK-801, prevented the increase on DCF fluorescence induced by glutamate or H2O2. Furthermore, creatine or MK-801 blocked glutamate- and H2O2-induced increases in NOx levels. In another set of experiments, the repeated, but not acute, administration of creatine (300 mg/kg, po) in mice prevented the decreases on cellular viability and mitochondrial membrane potential (assessed by tetramethylrhodamine ethyl ester, TMRE, probe) of hippocampal slices incubated with glutamate (10 mM). Creatine concentration-dependent decreased the amount of nitrite formed in the reaction of oxygen with NO produced from sodium nitroprusside solution, suggesting that its protective effect against glutamate or H2O2-induced toxicity might be due to its scavenger activity. Overall, the results suggest that creatine may be useful as adjuvant therapy for neurodegenerative disease treatments.

  12. Creatine and Its Potential Therapeutic Value for Targeting Cellular Energy Impairment in Neurodegenerative Diseases

    PubMed Central

    Adhihetty, Peter J.

    2010-01-01

    Substantial evidence indicates bioenergetic dysfunction and mitochondrial impairment contribute either directly and/or indirectly to the pathogenesis of numerous neurodegenerative disorders. Treatment paradigms aimed at ameliorating this cellular energy deficit and/or improving mitochondrial function in these neurodegenerative disorders may prove to be useful as a therapeutic intervention. Creatine is a molecule that is produced both endogenously, and acquired exogenously through diet, and is an extremely important molecule that participates in buffering intracellular energy stores. Once creatine is transported into cells, creatine kinase catalyzes the reversible transphosphorylation of creatine via ATP to enhance the phosphocreatine energy pool. Creatine kinase enzymes are located at strategic intracellular sites to couple areas of high energy expenditure to the efficient regeneration of ATP. Thus, the creatinekinase/phosphocreatine system plays an integral role in energy buffering and overall cellular bioenergetics. Originally, exogenous creatine supplementation was widely used only as an ergogenic aid to increase the phosphocreatine pool within muscle to bolster athletic performance. However, the potential therapeutic value of creatine supplementation has recently been investigated with respect to various neurodegenerative disorders that have been associated with bioenergetic deficits as playing a role in disease etiology and/or progression which include; Alzheimer’s, Parkinson’s, amyotrophic lateral sclerosis (ALS), and Huntington’s disease. This review discusses the contribution of mitochondria and bioenergetics to the progression of these neurodegenerative diseases and investigates the potential neuroprotective value of creatine supplementation in each of these neurological diseases. In summary, current literature suggests that exogenous creatine supplementation is most efficacious as a treatment paradigm in Huntington’s and Parkinson’s disease but

  13. Chronic Creatine Supplementation Alters Depression-like Behavior in Rodents in a Sex-Dependent Manner

    PubMed Central

    Allen, Patricia J; D'Anci, Kristen E; Kanarek, Robin B; Renshaw, Perry F

    2010-01-01

    Impairments in bioenergetic function, cellular resiliency, and structural plasticity are associated with the pathogenesis of mood disorders. Preliminary evidence suggests that creatine, an ergogenic compound known to promote cell survival and influence the production and usage of energy in the brain, can improve mood in treatment-resistant patients. This study examined the effects of chronic creatine supplementation using the forced swim test (FST), an animal model selectively sensitive to antidepressants with clinical efficacy in human beings. Thirty male (experiment 1) and 36 female (experiment 2) Sprague–Dawley rats were maintained on either chow alone or chow blended with either 2% w/w creatine monohydrate or 4% w/w creatine monohydrate for 5 weeks before the FST. Open field exploration and wire suspension tests were used to rule out general psychostimulant effects. Male rats maintained on 4% creatine displayed increased immobility in the FST as compared with controls with no differences by diet in the open field test, whereas female rats maintained on 4% creatine displayed decreased immobility in the FST and less anxiety in the open field test compared with controls. Open field and wire suspension tests confirmed that creatine supplementation did not produce differences in physical ability or motor function. The present findings suggest that creatine supplementation alters depression-like behavior in the FST in a sex-dependent manner in rodents, with female rats displaying an antidepressant-like response. Although the mechanisms of action are unclear, sex differences in creatine metabolism and the hormonal milieu are likely involved. PMID:19829292

  14. Biochemical properties and crystal structure of a β-phenylalanine aminotransferase from Variovorax paradoxus.

    PubMed

    Crismaru, Ciprian G; Wybenga, Gjalt G; Szymanski, Wiktor; Wijma, Hein J; Wu, Bian; Bartsch, Sebastian; de Wildeman, Stefaan; Poelarends, Gerrit J; Feringa, Ben L; Dijkstra, Bauke W; Janssen, Dick B

    2013-01-01

    By selective enrichment, we isolated a bacterium that can use β-phenylalanine as a sole nitrogen source. It was identified by 16S rRNA gene sequencing as a strain of Variovorax paradoxus. Enzyme assays revealed an aminotransferase activity. Partial genome sequencing and screening of a cosmid DNA library resulted in the identification of a 1,302-bp aminotransferase gene, which encodes a 46,416-Da protein. The gene was cloned and overexpressed in Escherichia coli. The recombinant enzyme was purified and showed a specific activity of 17.5 U mg(-1) for (S)-β-phenylalanine at 30°C and 33 U mg(-1) at the optimum temperature of 55°C. The β-specific aminotransferase exhibits a broad substrate range, accepting ortho-, meta-, and para-substituted β-phenylalanine derivatives as amino donors and 2-oxoglutarate and pyruvate as amino acceptors. The enzyme is highly enantioselective toward (S)-β-phenylalanine (enantioselectivity [E], >100) and derivatives thereof with different substituents on the phenyl ring, allowing the kinetic resolution of various racemic β-amino acids to yield (R)-β-amino acids with >95% enantiomeric excess (ee). The crystal structures of the holoenzyme and of the enzyme in complex with the inhibitor 2-aminooxyacetate revealed structural similarity to the β-phenylalanine aminotransferase from Mesorhizobium sp. strain LUK. The crystal structure was used to rationalize the stereo- and regioselectivity of V. paradoxus aminotransferase and to define a sequence motif with which new aromatic β-amino acid-converting aminotransferases may be identified.

  15. The polyproline II conformation in short alanine peptides is noncooperative.

    PubMed

    Chen, Kang; Liu, Zhigang; Kallenbach, Neville R

    2004-10-26

    The finding that short alanine peptides possess a high fraction of polyproline II (PII) structure (Phi=-75 degrees, Psi=+145 degrees ) at low temperature has broad implications for unfolded states of proteins. An important question concerns whether or not this structure is locally determined or cooperative. We have monitored the conformation of alanine in a series of model peptides AcGGAnGGNH2 (n=1-3) over a temperature range from -10 degrees C to +80 degrees C. Use of 15N-labeled alanine substitutions makes it possible to measure 3JalphaN coupling constants accurately over the full temperature range. Based on a 1D next-neighbor model, the cooperative parameter sigma of PII nucleation is evaluated from the coupling constant data. The finding that sigma is close to unity (1 +/- 0.2) indicates a noncooperative role for alanine in PII structure formation, consistent with statistical surveys of the Protein Data Bank that suggest that most PII structure occurs in isolated residues. Lack of cooperativity in these models implies that hydration effects that influence PII conformation in water are highly localized. Using a nuclear Overhauser effect ratio strategy to define the alanine Psi angle, we estimate that, at 40 degrees C, the time-averaged alanine conformation (Phi=-80 degrees, Psi=+170 degrees ) deviates from canonical PII structure, indicating that PII melts at high temperature. Thus, the high-temperature state of short alanine peptides seems to be an unfolded ensemble with higher distribution in the extended beta structure basin, but not a coil.

  16. EPR/alanine dosimetry for two therapeutic proton beams

    NASA Astrophysics Data System (ADS)

    Marrale, Maurizio; Carlino, Antonio; Gallo, Salvatore; Longo, Anna; Panzeca, Salvatore; Bolsi, Alessandra; Hrbacek, Jan; Lomax, Tony

    2016-02-01

    In this work the analysis of the electron paramagnetic resonance (EPR) response of alanine pellets exposed to two different clinical proton beams employed for radiotherapy is performed. One beam is characterized by a passive delivery technique and is dedicated to the eyes treatment (OPTIS2 beam line). Alanine pellets were irradiated with a 70 MeV proton beam corresponding to 35 mm range in eye tissue. We investigated how collimators with different sizes and shape used to conform the dose to the planned target volume influence the delivered dose. For this purpose we performed measurements with varying the collimator size (Output Factor) and the results were compared with those obtained with other dosimetric techniques (such as Markus chamber and diode detector). This analysis showed that the dosimeter response is independent of collimator diameter if this is larger than or equal to 10 mm. The other beam is characterized by an active spot-scanning technique, the Gantry1 beam line (maximum energy 230 MeV), and is used to treat deep-seated tumors. The dose linearity of alanine response in the clinical dose range was tested and the alanine dose response at selected locations in depth was measured and compared with the TPS planned dose in a quasi-clinical scenario. The alanine response was found to be linear in the dose in the clinical explored range (from 10 to 70 Gy). Furthermore, a depth dose profile in a quasi-clinical scenario was measured and compared to the dose computed by the Treatment Planning System PSIPLAN. The comparison of calibrated proton alanine measurements and TPS dose shows a difference under 1% in the SOBP and a "quenching" effect up to 4% in the distal part of SOBP. The positive dosimetric characteristics of the alanine pellets confirm the feasibility to use these detectors for "in vivo" dosimetry in clinical proton beams.

  17. Biochemical and Structural Characterization of a Ureidoglycine Aminotransferase in the Klebsiella pneumoniae Uric Acid Catabolic Pathway

    SciTech Connect

    French, Jarrod B.; Ealick, Steven E.

    2010-09-03

    Many plants, fungi, and bacteria catabolize allantoin as a mechanism for nitrogen assimilation. Recent reports have shown that in plants and some bacteria the product of hydrolysis of allantoin by allantoinase is the unstable intermediate ureidoglycine. While this molecule can spontaneously decay, genetic analysis of some bacterial genomes indicates that an aminotransferase may be present in the pathway. Here we present evidence that Klebsiella pneumoniae HpxJ is an aminotransferase that preferentially converts ureidoglycine and an {alpha}-keto acid into oxalurate and the corresponding amino acid. We determined the crystal structure of HpxJ, allowing us to present an explanation for substrate specificity.

  18. Three-step preparation and purification of phosphorus-33-labeled creatine phosphate of high specific activity

    SciTech Connect

    Savabi, F.; Geiger, P.J.; Bessman, S.P.

    1984-03-01

    Rabbit heart mitochondria were used as a source of enzymes for the synthesis of phosphorus-labeled creatine phosphate. This method is based on the coupled reaction between mitochondrial oxidative phosphorylation and mitochondrial-bound creatine kinase. It is possible to convert more than 90% of the inorganic phosphate (P/sub i/) to creatine phosphate. The method used only small amounts of adenine nucleotides which led to a product with only slight nucleotide contamination. This could be removed by activated charcoal extraction. For further purification, a method for the removal of residual P/sub i/ is described. 20 references.

  19. Creatine as a booster for human brain function. How might it work?

    PubMed

    Rae, Caroline D; Bröer, Stefan

    2015-10-01

    Creatine, a naturally occurring nitrogenous organic acid found in animal tissues, has been found to play key roles in the brain including buffering energy supply, improving mitochondrial efficiency, directly acting as an anti-oxidant and acting as a neuroprotectant. Much of the evidence for these roles has been established in vitro or in pre-clinical studies. Here, we examine the roles of creatine and explore the current status of translation of this research into use in humans and the clinic. Some further possibilities for use of creatine in humans are also discussed.

  20. Effect of adrenergic agonists and antagonists on alanine amino transferase, fructose-1:6-bisphosphatase and glucose production in hepatocytes.

    PubMed

    Begum, N A; Datta, A G

    1992-08-18

    Using rat hepatocytes we confirmed our previous results that glucagon and beta-adrenergic agonists increased the enzyme activity of alanine aminotransferase (AAT) and propranolol abolished their effects. Only the enzyme activity was measured and other parameters like quantity of the enzyme or activation due to modification were not looked for. As in perfusion experiment phenylephrine and phenoxybenzamine (alpha-agonist and alpha-antagonist respectively) also alpha-antagonist respectively) also increased the AAT activity in isolated rat hepatocytes and propranolol reversed these effects. The additive effect of glucagon and phenoxybenzamine on AAT was also persistent in hepatocyte system. Fructose-1:6-bisphosphatase (Fru-P2-ase), another key enzyme in gluconeogenic pathway, was elevated by glucagon and other beta-adrenergic agonists both in liver perfusion and isolated hepatocyte experiments and was brought back to the normal level by propranolol. In this case also only the enzyme activity was measured and no other parameters were looked for. Unlike AAT this enzyme was not stimulated by phenylephrine or phenoxybenzamine. But AAT and Fru-P2-ase activities were increased significantly by adenylate cyclase activators like fluoride or forskolin. Thus, it appears that the regulation of fru-P2-ase by glucagon is purely a b-receptor mediated process whereas AAT activation shows a mixed type of regulation where some well known alpha-agonist and antagonists are behaving as beta-agonists. Results further indicate the presence of phosphodiesterase in hepatocyte membrane which was stimulated by glucagon and brought back to the normal level by propranolol. The different adrenergic compounds stated above, not only modified the activity of the above two enzymes but also stimulated glucose production by hepatocytes from alanine which was in turn abolished by propranolol as well as amino oxyacetate (AOA), a highly specified inhibitor of AAT. This confirm the participation of AAT in

  1. Method of empirical dependences in estimation and prediction of activity of creatine kinase isoenzymes in cerebral ischemia

    NASA Astrophysics Data System (ADS)

    Sergeeva, Tatiana F.; Moshkova, Albina N.; Erlykina, Elena I.; Khvatova, Elena M.

    2016-04-01

    Creatine kinase is a key enzyme of energy metabolism in the brain. There are known cytoplasmic and mitochondrial creatine kinase isoenzymes. Mitochondrial creatine kinase exists as a mixture of two oligomeric forms - dimer and octamer. The aim of investigation was to study catalytic properties of cytoplasmic and mitochondrial creatine kinase and using of the method of empirical dependences for the possible prediction of the activity of these enzymes in cerebral ischemia. Ischemia was revealed to be accompanied with the changes of the activity of creatine kinase isoenzymes and oligomeric state of mitochondrial isoform. There were made the models of multiple regression that permit to study the activity of creatine kinase system in cerebral ischemia using a calculating method. Therefore, the mathematical method of empirical dependences can be applied for estimation and prediction of the functional state of the brain by the activity of creatine kinase isoenzymes in cerebral ischemia.

  2. β-Alanine as an Ethylene Precursor. Investigations Towards Preparation, and Properties, of a Soluble Enzyme System From a Subcellular Particulate Fraction of Bean Cotyledons 1

    PubMed Central

    Stinson, Robert A.; Spencer, Mary

    1969-01-01

    A method is described for the preparation, from a subcellular particulate fraction of wax bean cotyledons, of a soluble enzyme system that is capable of converting β-alanine to ethylene. In the presence of ATP, CoA, thiamine pyrophosphate, MgSO4, and pyridoxal phosphate, ethylene production is maximum at a 0.5 mm concentration of β-alanine. The system exhibits a pH optimum at 7.0 but when the pH is raised above 8, evolution of the volatile again increases and continues to do so up to pH 12. The enzyme system is stimulated by either NADPH or NADH; the concentration of NADPH necessary to obtain maximum activity is twice that of NADH. The requirement for a reducing agent is in agreement with the proposal that malonate semialdehyde, formed by an aminotransferase reaction from β-alanine, is reduced to β-hydroxypropionate. Both malonate semialdehyde and β-hydroxypropionate are better stimulators of production of the volatile in the soluble system than is β-alanine, and β-hydroxypropionate is a better stimulator than malonate semialdehyde. This system is also able to incorporate tritium from tritiated water into ethylene; this supports the proposal that ethylene is formed by the decarboxylation of acrylate, the latter being formed from β-hydroxypropionate. Experiments with both cold and labeled malonate suggest that this compound stimulates ethylene production by acting as an end product inhibitor that prevents the loss of malonate semialdehyde from the pathway. Malonate does not appear to serve as a precursor. Addition of cytoplasmic enzymes to the `soluble system' (prepared from particulate enzymes) results in a considerable boost in ethylene production, but the specific activity (mμ1 / mg protein) is lowered from that of the particulate enzymes alone. PMID:16657194

  3. Distribution of creatine, guanidinoacetate and the enzymes for their biosynthesis in the animal kingdom. Implications for phylogeny.

    PubMed

    Van Pilsum, J F; Stephens, G C; Taylor, D

    1972-01-01

    1. The distribution of creatine and the creatine-synthesizing enzymes in the animal kingdom has been investigated. Creatine was found in tissues of all vertebrates examined, and in various invertebrates from phyla Annelida, Echinodermata, Hemichordata and Chordata, subphylum Cephalochordata. The activities of the creatine-synthesizing enzymes, arginine-glycine transamidinase and guanidinoacetate methylpherase, were not detected in the hagfish or in any of the invertebrates, including those in which creatine was found, with the exception that transamidinase activities were detected in the amphioxus and salt water clam; however, these activities are considered to be artifacts for reasons mentioned in the text. Additional evidence that the hagfish and various creatine-containing invertebrates could not synthesize creatine was the observation that these animals did not convert one or the other of the likely precursors of creatine (arginine and glycine) into creatine, in vivo. Further, the inability of these animals to synthesize creatine is correlated with the observations that all animals tested were able to abstract creatine from their aqueous environment. 2. The activities of the creatine-synthesizing enzymes were detected in the sea lamprey and in all but a few of the other vertebrates examined. Neither activity could be detected in the sharks and rays (cartilaginous fish), buffalo fish (bony fish) or the snapping turtle. Transamidinase or guanidinoacetate methylpherase activity could not be found in the salamander or garter snake, respectively. 3. The results obtained with the lamprey are in direct contrast with those obtained with the hagfish (both subphylum Agnatha, class Cyclostomata). The lamprey had the ability to synthesize creatine and did not abstract creatine from lake water. The hagfish did not have any apparent ability to synthesize creatine and did abstract creatine from sea water. The present report thus supports the theory that the myxinoid (hagfish

  4. Distribution of creatine, guanidinoacetate and the enzymes for their biosynthesis in the animal kingdom. Implications for phylogeny

    PubMed Central

    Van Pilsum, John F.; Stephens, Grover C.; Taylor, Dorris

    1972-01-01

    1. The distribution of creatine and the creatine-synthesizing enzymes in the animal kingdom has been investigated. Creatine was found in tissues of all vertebrates examined, and in various invertebrates from phyla Annelida, Echinodermata, Hemichordata and Chordata, subphylum Cephalochordata. The activities of the creatine-synthesizing enzymes, arginine–glycine transamidinase and guanidinoacetate methylpherase, were not detected in the hagfish or in any of the invertebrates, including those in which creatine was found, with the exception that transamidinase activities were detected in the amphioxus and salt water clam; however, these activities are considered to be artifacts for reasons mentioned in the text. Additional evidence that the hagfish and various creatine-containing invertebrates could not synthesize creatine was the observation that these animals did not convert one or the other of the likely precursors of creatine (arginine and glycine) into creatine, in vivo. Further, the inability of these animals to synthesize creatine is correlated with the observations that all animals tested were able to abstract creatine from their aqueous environment. 2. The activities of the creatine-synthesizing enzymes were detected in the sea lamprey and in all but a few of the other vertebrates examined. Neither activity could be detected in the sharks and rays (cartilaginous fish), buffalo fish (bony fish) or the snapping turtle. Transamidinase or guanidinoacetate methylpherase activity could not be found in the salamander or garter snake, respectively. 3. The results obtained with the lamprey are in direct contrast with those obtained with the hagfish (both subphylum Agnatha, class Cyclostomata). The lamprey had the ability to synthesize creatine and did not abstract creatine from lake water. The hagfish did not have any apparent ability to synthesize creatine and did abstract creatine from sea water. The present report thus supports the theory that the myxinoid

  5. On the existence of ``l-threonine formate'', ``l-alanine lithium chloride'' and ``bis l-alanine lithium chloride'' crystals

    NASA Astrophysics Data System (ADS)

    Petrosyan, A. M.; Ghazaryan, V. V.; Fleck, M.

    2013-03-01

    We argue that the recently reported crystals "L-threonine formate" as well as "L-alanine lithium chloride" and "bis L-alanine lithium chloride" actually are the well-known crystals L-threonine and L-alanine, respectively.

  6. Abnormal N-Glycosylation of a Novel Missense Creatine Transporter Mutant, G561R, Associated with Cerebral Creatine Deficiency Syndromes Alters Transporter Activity and Localization.

    PubMed

    Uemura, Tatsuki; Ito, Shingo; Ohta, Yusuke; Tachikawa, Masanori; Wada, Takahito; Terasaki, Tetsuya; Ohtsuki, Sumio

    2017-01-01

    Cerebral creatine deficiency syndromes (CCDSs) are caused by loss-of-function mutations in creatine transporter (CRT, SLC6A8), which transports creatine at the blood-brain barrier and into neurons of the central nervous system (CNS). This results in low cerebral creatine levels, and patients exhibit mental retardation, poor language skills and epilepsy. We identified a novel human CRT gene missense mutation (c.1681 G>C, G561R) in Japanese CCDSs patients. The purpose of the present study was to evaluate the reduction of creatine transport in G561R-mutant CRT-expressing 293 cells, and to clarify the mechanism of its functional attenuation. G561R-mutant CRT exhibited greatly reduced creatine transport activity compared to wild-type CRT (WT-CRT) when expressed in 293 cells. Also, the mutant protein is localized mainly in intracellular membrane fraction, while WT-CRT is localized in plasma membrane. Western blot analysis revealed a 68 kDa band of WT-CRT protein in plasma membrane fraction, while G561R-mutant CRT protein predominantly showed bands at 55, 110 and 165 kDa in crude membrane fraction. The bands of both WT-CRT and G561R-mutant CRT were shifted to 50 kDa by N-glycosidase treatment. Our results suggest that the functional impairment of G561R-mutant CRT was probably caused by incomplete N-linked glycosylation due to misfolding during protein maturation, leading to oligomer formation and changes of cellular localization.

  7. Effect of Preexercise Creatine Ingestion on Muscle Performance in Healthy Aging Males.

    PubMed

    Baker, Taylor P; Candow, Darren G; Farthing, Jonathan P

    2016-06-01

    Preexercise creatine supplementation may have a beneficial effect on aging muscle performance. Using a double-blind, repeated measures, crossover design, healthy males (N = 9, 54.8 ± 4.3 years; 92.9 ± 11.5 kg; 179.2 ± 11.1 cm) were randomized to consume creatine (20 g) and placebo (20 g corn starch maltodextrin), on 2 separate occasions (7 days apart), 3 hours before performing leg press and chest press repetitions to muscle fatigue (3 sets at 70% 1-repetition maximum; 1 minute rest between sets). There was a set main effect (p ≤ 0.05) for the leg press and chest press with the number of repetitions performed decreasing similarly for creatine and placebo. These results suggest that a bolus ingestion of creatine consumed 3 hours before resistance exercise has no effect on upper or lower-body muscle performance in healthy aging males.

  8. CINRG randomized controlled trial of creatine and glutamine in Duchenne muscular dystrophy.

    PubMed

    Escolar, Diana M; Buyse, Gunnar; Henricson, Erik; Leshner, Robert; Florence, Julaine; Mayhew, Jill; Tesi-Rocha, Carolina; Gorni, Ksenija; Pasquali, Livia; Patel, Kantilal M; McCarter, Robert; Huang, Jennifer; Mayhew, Thomas; Bertorini, Tulio; Carlo, Jose; Connolly, Anne M; Clemens, Paula R; Goemans, Nathalie; Iannaccone, Susan T; Igarashi, Masanori; Nevo, Yoram; Pestronk, Alan; Subramony, S H; Vedanarayanan, V V; Wessel, Henry

    2005-07-01

    We tested the efficacy and safety of glutamine (0.6 gm/kg/day) and creatine (5 gm/day) in 50 ambulant boys with Duchenne muscular dystrophy in a 6-month, double-blind, placebo-controlled clinical trial. Drug efficacy was tested by measuring muscle strength manually (34 muscle groups) and quantitatively (10 muscle groups). Timed functional tests, functional parameters, and pulmonary function tests were secondary outcome measures. Although there was no statistically significant effect of either therapy based on manual and quantitative measurements of muscle strength, a disease-modifying effect of creatine in older Duchenne muscular dystrophy and creatine and glutamine in younger Duchenne muscular dystrophy cannot be excluded. Creatine and glutamine were well tolerated.

  9. [Effects of ß-alanine supplementation on athletic performance].

    PubMed

    Domínguez, Raúl; Hernández Lougedo, Juan; Maté-Muñoz, José Luis; Garnacho-Castaño, Manuel Vicente

    2014-10-06

    Carnosine, dipeptide formed by amino acids ß-alanine and L-histidine, has important physiological functions among which its antioxidant and related memory and learning. However, in connection with the exercise, the most important functions would be associated with muscle contractility, improving calcium sensitivity in muscle fibers, and the regulatory function of pH. Thus, it is proposed that carnosine is the major intracellular buffer, but could contribute to 7-10% in buffer or buffer capacity. Since carnosine synthesis seems to be limited by the availability of ß-alanine supplementation with this compound has been gaining increasing popularity among the athlete population. Therefore, the objective of this study literature review was to examine all those research works have shown the effect of ß-alanine supplementation on athletic performance. Moreover, it also has attempted to establish a specific dosage that maximizing the potential benefits, minimize paresthesia, the main side effect presented in response to supplementation.

  10. First-principles studies of pure and fluorine substituted alanines

    NASA Astrophysics Data System (ADS)

    Ahmad, Sardar; Vaizie, Hamide; Rahnamaye Aliabad, H. A.; Ahmad, Rashid; Khan, Imad; Ali, Zahid; Jalali-Asadabadi, S.; Ahmad, Iftikhar; Khan, Amir Abdullah

    2016-05-01

    This paper communicates the structural, electronic and optical properties of L-alanine, monofluoro and difluoro substituted alanines using density functional calculations. These compounds exist in orthorhombic crystal structure and the calculated structural parameters such as lattice constants, bond angles and bond lengths are in agreement with the experimental results. L-alanine is an indirect band gap insulator, while its fluorine substituted compounds (monofluoroalanine and difluoroalanine) are direct band gap insulators. The substitution causes reduction in the band gap and hence these optically tailored direct wide band gap materials have enhanced optical properties in the ultraviolet (UV) region of electromagnetic spectrum. Therefore, optical properties like dielectric function, refractive index, reflectivity and energy loss function are also investigated. These compounds have almost isotropic nature in the lower frequency range while at higher energies, they have a significant anisotropic nature.

  11. The effects of oral creatine supplementation on performance in single and repeated sprint swimming.

    PubMed

    Peyrebrune, M C; Nevill, M E; Donaldson, F J; Cosford, D J

    1998-04-01

    We studied the effects of oral creatine supplementation on sprint swimming performance in 14 elite competitive male swimmers. The subjects performed a single sprint (1 x 50 yards [45.72 m]) and repeated sprint set (8 x 50 yards at intervals of 1 min 30 s) before and after a 5 day period of either creatine (9 g creatine + 4.5 g maltodextrin + 4.5 g glucose day(-1)) or placebo (18 g glucose day(-1); double-blind protocol) supplementation. Venous and capillary blood samples were taken for the determination of plasma ammonia, blood pH and lactate. Mean times recorded for the single 50 yard sprint were unchanged as a result of supplementation (creatine vs control, N.S.). During the repeated sprint test, mean times increased (P< 0.01, main effect time) during all trials, but performance was improved as a result of creatine supplementation (sprints 1-8: control pre-, 23.35+/-0.68 to 26.32+/-1.34 s; control post-, 23.59+/-0.66 to 26.19+/-1.48 s; creatine pre-, 23.20+/-0.67 to 26.85+/-0.42 s; creatine post-, 23.39+/-0.54 to 25.73+/-0.26 s; P < 0.03, group x trial interaction). Thus the percentage decline in performance times was reduced after creatine supplementation (control, 12.7+/-5.7% vs 11.0+/-5.5%; creatine, 15.7+/-4.3% vs 10.0+/-2.5%; P< 0.05, group x trial interaction). The metabolic response was similar before and after supplementation, with no differences in the blood lactate or pH response. Plasma ammonia was lower on the second trial (P< 0.05, main effect trial), but this could not be attributed to the effect of supplementation (group x trial interaction, N.S.). A further urinary analysis study supported these findings by demonstrating an approximately 67% (approximately 26 g) retention of the administered creatine in this group of swimmers after an identical supplementation regimen. In summary, our results suggest that ingesting 9 g creatine per day for 5 days can improve swimming performance in elite competitors during repeated sprints, but appears to have no

  12. Structure, expression, and function of kynurenine aminotransferases in human and rodent brains.

    PubMed

    Han, Qian; Cai, Tao; Tagle, Danilo A; Li, Jianyong

    2010-02-01

    Kynurenine aminotransferases (KATs) catalyze the synthesis of kynurenic acid (KYNA), an endogenous antagonist of N-methyl-D: -aspartate and alpha 7-nicotinic acetylcholine receptors. Abnormal KYNA levels in human brains are implicated in the pathophysiology of schizophrenia, Alzheimer's disease, and other neurological disorders. Four KATs have been reported in mammalian brains, KAT I/glutamine transaminase K/cysteine conjugate beta-lyase 1, KAT II/aminoadipate aminotransferase, KAT III/cysteine conjugate beta-lyase 2, and KAT IV/glutamic-oxaloacetic transaminase 2/mitochondrial aspartate aminotransferase. KAT II has a striking tertiary structure in N-terminal part and forms a new subgroup in fold type I aminotransferases, which has been classified as subgroup Iepsilon. Knowledge regarding KATs is vast and complex; therefore, this review is focused on recent important progress of their gene characterization, physiological and biochemical function, and structural properties. The biochemical differences of four KATs, specific enzyme activity assays, and the structural insights into the mechanism of catalysis and inhibition of these enzymes are discussed.

  13. Structure, expression, and function of kynurenine aminotransferases in human and rodent brains

    PubMed Central

    Han, Qian; Cai, Tao; Tagle, Danilo A.

    2010-01-01

    Kynurenine aminotransferases (KATs) catalyze the synthesis of kynurenic acid (KYNA), an endogenous antagonist of N-methyl-d-aspartate and alpha 7-nicotinic acetylcholine receptors. Abnormal KYNA levels in human brains are implicated in the pathophysiology of schizophrenia, Alzheimer's disease, and other neurological disorders. Four KATs have been reported in mammalian brains, KAT I/glutamine transaminase K/cysteine conjugate beta-lyase 1, KAT II/aminoadipate aminotransferase, KAT III/cysteine conjugate beta-lyase 2, and KAT IV/glutamic-oxaloacetic transaminase 2/mitochondrial aspartate aminotransferase. KAT II has a striking tertiary structure in N-terminal part and forms a new subgroup in fold type I aminotransferases, which has been classified as subgroup Iε. Knowledge regarding KATs is vast and complex; therefore, this review is focused on recent important progress of their gene characterization, physiological and biochemical function, and structural properties. The biochemical differences of four KATs, specific enzyme activity assays, and the structural insights into the mechanism of catalysis and inhibition of these enzymes are discussed. PMID:19826765

  14. Structural Insights into a Novel Class of Aspartate Aminotransferase from Corynebacterium glutamicum

    PubMed Central

    Son, Hyeoncheol Francis; Kim, Kyung-Jin

    2016-01-01

    Aspartate aminotransferase from Corynebacterium glutamicum (CgAspAT) is a PLP-dependent enzyme that catalyzes the production of L-aspartate and α-ketoglutarate from L-glutamate and oxaloacetate in L-lysine biosynthesis. In order to understand the molecular mechanism of CgAspAT and compare it with those of other aspartate aminotransferases (AspATs) from the aminotransferase class I, we determined the crystal structure of CgAspAT. CgAspAT functions as a dimer, and the CgAspAT monomer consists of two domains, the core domain and the auxiliary domain. The PLP cofactor is found to be bound to CgAspAT and stabilized through unique residues. In our current structure, a citrate molecule is bound at the active site of one molecule and mimics binding of the glutamate substrate. The residues involved in binding of the PLP cofactor and the glutamate substrate were confirmed by site-directed mutagenesis. Interestingly, compared with other AspATs from aminotransferase subgroup Ia and Ib, CgAspAT exhibited unique binding sites for both cofactor and substrate; moreover, it was found to have unusual structural features in the auxiliary domain. Based on these structural differences, we propose that CgAspAT does not belong to either subgroup Ia or Ib, and can be categorized into a subgroup Ic. The phylogenetic tree and RMSD analysis also indicates that CgAspAT is located in an independent AspAT subgroup. PMID:27355211

  15. Structure Expression and Function of kynurenine Aminotransferases in Human and Rodent Brains

    SciTech Connect

    Q Han; T Cai; D Tagle; J Li

    2011-12-31

    Kynurenine aminotransferases (KATs) catalyze the synthesis of kynurenic acid (KYNA), an endogenous antagonist of N-methyl-D: -aspartate and alpha 7-nicotinic acetylcholine receptors. Abnormal KYNA levels in human brains are implicated in the pathophysiology of schizophrenia, Alzheimer's disease, and other neurological disorders. Four KATs have been reported in mammalian brains, KAT I/glutamine transaminase K/cysteine conjugate beta-lyase 1, KAT II/aminoadipate aminotransferase, KAT III/cysteine conjugate beta-lyase 2, and KAT IV/glutamic-oxaloacetic transaminase 2/mitochondrial aspartate aminotransferase. KAT II has a striking tertiary structure in N-terminal part and forms a new subgroup in fold type I aminotransferases, which has been classified as subgroup Iepsilon. Knowledge regarding KATs is vast and complex; therefore, this review is focused on recent important progress of their gene characterization, physiological and biochemical function, and structural properties. The biochemical differences of four KATs, specific enzyme activity assays, and the structural insights into the mechanism of catalysis and inhibition of these enzymes are discussed.

  16. Creatine supplementation enhances endurance performance in trained rats.

    PubMed

    Malin, Steven K; Cotugna, Nancy

    2008-01-01

    Minimal evidence has shown creatine (Cr) supplementation to enhance endurance performance in either humans or rats. The purpose of this study was to examine the effects of Cr supplementation on endurance performance during high-intensity exercise in trained male rats. Endurance performance was defined as the distance run. Sixteen days of running were performed over 28 days. A cycle of 7 days consisted of 2 days of training, 1 day off, 2 days of training then 2 days off and this was repeated over a total of 28 days. Cr was administered on all 28 days. Treatment rats (n = 7) drank water containing Cr while the control rats drank water with no supplement (n = 6). The Cr group's average distance run increased significantly from baseline to exercise day 16 (baseline = 128.91 m ± 18.23 vs. exercise day 16 = 217.11m ± 18.11; p < 0.005), while the control groups did not (baseline = 137.24 m ± 10.14, exercise day 16 = 101.04 m ± 14.97; p > 0.05). Over the course of the study, the treatment group's running endurance improved by 81% compared to baseline (p < 0.001) and we conclude that Cr supplementation provided rats an increased ability to run farther demonstrating possible implications for improving endurance athletes' performances.

  17. Influence of local vibration on plasma creatine phosphokinase (CPK) activity.

    PubMed Central

    Okada, A; Okuda, H; Inaba, R; Ariizumi, M

    1985-01-01

    This study was designed to obtain basic information about the mechanism of the occurrence of muscular disorders after exposure to vibration. The hind legs of rats were exposed to acute and chronic local vibration at frequencies of 30, 60, 120, 240, 480, and 960 Hz with a constant acceleration of 50 m/sec2. The exposure time was four hours for acute, and four hours a day for two weeks continuously for chronic exposure. Blood was collected after exposure to measure plasma creatine phosphokinase (CPK) activity. In both exposure groups the activity of plasma CPK was significantly higher at 30, 60, 120, 240, and 480 Hz compared with the control group and was especially high at 30 Hz; there was no significant change at 960 Hz. As a result of an analysis of the CPK isoenzymes, the increase in plasma CPK activity was shown to be due to the activity of the plasma CPK-MM fraction, originating in the skeletal muscle. Plasma CPK activity showed a tendency to decrease gradually with the increase in vibration frequency during acute exposure but showed no such tendency during chronic exposure. There was no remarkable pathohistological change in muscle preparations from the hind legs, hence it was presumed that the increase in plasma CPK activity was caused not by the morphological changes of muscle but by other mechanisms, such as an increase in the permeability of the cell membrane. Images PMID:4041385

  18. Swim performance following creatine supplementation in Division III athletes.

    PubMed

    Selsby, Joshua T; Beckett, Keith D; Kern, Michael; Devor, Steven T

    2003-08-01

    Creatine (Cr) supplementation has yielded inconsistent results when applied to competitive swimming. To further define the role of Cr, we tested the hypothesis that a Cr supplementation group of Division III swimmers would demonstrate enhanced performance when compared with placebo. In order to test this hypothesis, 8 male and 7 female collegiate Division III swimmers were assigned in a random, double-blind manner into either a Cr supplementation group (0.3 g Cr.kg(-1) body mass) or a placebo group. Loading was maintained for 5 days followed by a 9-day period where Cr-supplemented subjects consumed 2.25 g Cr regardless of body weight. A 50- and 100-yd sprint was performed prior to and following the supplementation regimens. The Cr supplementation group decreased their finish times in both the 50- and 100-yd sprints. Support of the hypothesis suggests that Cr supplementation for swimming events is effective for singular effort sprints of 50 and 100 yd in Division III athletes.

  19. Diagnostic value of creatine kinase activity in canine cerebrospinal fluid

    PubMed Central

    Ferreira, Alexandra

    2016-01-01

    This study aimed to determine whether creatine kinase (CK) activity in cerebrospinal fluid (CSF) has diagnostic value for various groups of neurological conditions or for different anatomical areas of the nervous system (NS). The age, breed, results of CSF analysis, and diagnosis of 578 canine patients presenting with various neurological conditions between January 2009 and February 2015 were retrospectively collected. The cases were divided according to anatomical areas of the nervous system, i.e., brain, spinal cord, and peripheral nervous system, and into groups according to the nature of the condition diagnosed: vascular, immune/inflammatory/infectious, traumatic, toxic, anomalous, metabolic, idiopathic, neoplastic, and degenerative. Statistical analysis showed that CSF-CK alone cannot be used as a diagnostic tool and that total proteins in the CSF and red blood cells (RBCs) do not have a significant relationship with the CSF-CK activity. CSF-CK did not have a diagnostic value for different disease groups or anatomical areas of the nervous system. PMID:27708448

  20. Does exercise training alter myocardial creatine kinase MB isoenzyme content?

    PubMed

    Miller, T D; Rogers, P J; Bauer, B A; O'Brien, J F; Squires, R W; Bailey, K R; Bove, A A

    1989-08-01

    Skeletal muscle biopsies from highly trained endurance athletes have been shown to contain an increased percentage of the creatine kinase MB (CK-MB) isoenzyme, which has been attributed to continuous regeneration of the skeletal muscle fibers in response to exercise-induced injury. The purpose of this study was to determine whether myocardium undergoes a similar degenerative-regenerative process as a result of exercise training. Fifteen mongrel dogs underwent a 12-wk period of training (N = 8) or cage confinement (N = 7). The animals were then sacrificed, and samples of left and right ventricular myocardium were analyzed for total CK activity and CK-MB isoenzyme content. Percentages of CK-MB were slightly but insignificantly higher from both ventricles of exercise-trained as compared with cage-confined dogs: left ventricle, 4.6 +/- 0.6% vs 3.3 +/- 0.6%, respectively (P = 0.15); right ventricle, 4.0 +/- 0.4% vs 3.0 +/- 0.8%, respectively (P = 0.29). We conclude that chronic exercise training does not induce physiologically important degenerative changes in myocardium.

  1. The genomic organization of a human creatine transporter (CRTR) gene located in Xq28

    SciTech Connect

    Sandoval, N.; Bauer, D.; Brenner, V.

    1996-07-15

    During the course of a large-scale sequencing project in Xq28, a human creatine transporter (CRTR) gene was discovered. The gene is located approximately 36 kb centromeric to ALD. The gene contains 13 exons and spans about 8.5 kb of genomic DNA. Since the creatine transporter has a prominent function in muscular physiology, it is a candidate gene for Barth syndrome and infantile cardiomyopathy mapped to Xq28. 19 refs., 1 fig., 1 tab.

  2. Effects of creatine supplementation and resistance training on muscle strength and weightlifting performance.

    PubMed

    Rawson, Eric S; Volek, Jeff S

    2003-11-01

    Creatine monohydrate has become the supplement of choice for many athletes striving to improve sports performance. Recent data indicate that athletes may not be using creatine as a sports performance booster per se but instead use creatine chronically as a training aid to augment intense resistance training workouts. Although several studies have evaluated the combined effects of creatine supplementation and resistance training on muscle strength and weightlifting performance, these data have not been analyzed collectively. The purpose of this review is to evaluate the effects of creatine supplementation on muscle strength and weightlifting performance when ingested concomitant with resistance training. The effects of gender, interindividual variability, training status, and possible mechanisms of action are discussed. Of the 22 studies reviewed, the average increase in muscle strength (1, 3, or 10 repetition maximum [RM]) following creatine supplementation plus resistance training was 8% greater than the average increase in muscle strength following placebo ingestion during resistance training (20 vs. 12%). Similarly, the average increase in weightlifting performance (maximal repetitions at a given percent of maximal strength) following creatine supplementation plus resistance training was 14% greater than the average increase in weightlifting performance following placebo ingestion during resistance training (26 vs. 12%). The increase in bench press 1RM ranged from 3 to 45%, and the improvement in weightlifting performance in the bench press ranged from 16 to 43%. Thus there is substantial evidence to indicate that creatine supplementation during resistance training is more effective at increasing muscle strength and weightlifting performance than resistance training alone, although the response is highly variable.

  3. Creatine supplementation augments skeletal muscle carnosine content in senescence-accelerated mice (SAMP8).

    PubMed

    Derave, Wim; Jones, Glenys; Hespel, Peter; Harris, Roger C

    2008-06-01

    The histidine-containing dipeptides (HCD) carnosine and anserine are found in high concentrations in mammalian skeletal muscle. Given its versatile biologic properties, such as antioxidative, antiglycation, and pH buffering capacity, carnosine has been implicated as a protective factor in the aging process. The present study aimed to systematically explore age-related changes in skeletal muscles HCD content in a murine model of accelerated aging. Additionally, we investigated the effect of lifelong creatine supplementation on muscle HCD content and contractile fatiguability. Male senescence-accelerated mice (SAMP8) were fed control or creatine-supplemented (2% of food intake) diet from the age of 10 to 60 weeks. At week 10, 25, and 60, tibialis anterior muscles were dissected and analysed for HCD and taurine content by HPLC. Soleus and EDL muscles were tested for in vitro contractile fatigue and recovery. From 10 to 60 weeks of age, muscular carnosine (-45%), taurine (-24%), and total creatine (-42%) concentrations gradually and significantly decreased. At 25 but not at 60 weeks, oral creatine supplementation significantly increased carnosine (+88%) and anserine (+40%) content compared to age-matched control-fed animals. Taurine and total creatine content were not affected by creatine supplementation at any age. Creatine-treated mice showed attenuated muscle fatigue (soleus) and enhanced force recovery (m. extensor digitorum longus [EDL]) compared to controls at 25 weeks, but not at 60 weeks. From the present study, we can conclude that skeletal muscle tissue exhibits a significant decline in HCD content at old age. Oral creatine supplementation is able to transiently but potently increase muscle carnosine and anserine content, which coincides with improved resistance to contractile fatigue.

  4. Homocysteine induces energy imbalance in rat skeletal muscle: is creatine a protector?

    PubMed

    Kolling, Janaína; Scherer, Emilene B S; Siebert, Cassiana; Hansen, Fernanda; Torres, Felipe V; Scaini, Giselli; Ferreira, Gabriela; de Andrade, Rodrigo B; Gonçalves, Carlos A S; Streck, Emílio L; Wannmacher, Clovis M D; Wyse, Angela T S

    2013-10-01

    Homocystinuria is a neurometabolic disease caused by a severe deficiency of cystathionine beta-synthase activity, resulting in severe hyperhomocysteinemia. Affected patients present several symptoms including a variable degree of motor dysfunction. In this study, we investigated the effect of chronic hyperhomocysteinemia on the cell viability of the mitochondrion, as well as on some parameters of energy metabolism, such as glucose oxidation and activities of pyruvate kinase, citrate synthase, isocitrate dehydrogenase, malate dehydrogenase, respiratory chain complexes and creatine kinase in gastrocnemius rat skeletal muscle. We also evaluated the effect of creatine on biochemical alterations elicited by hyperhomocysteinemia. Wistar rats received daily subcutaneous injections of homocysteine (0.3-0.6 µmol/g body weight) and/or creatine (50 mg/kg body weight) from the 6th to the 28th days of age. The animals were decapitated 12 h after the last injection. Homocysteine decreased the cell viability of the mitochondrion and the activities of pyruvate kinase and creatine kinase. Succinate dehydrogenase was increased other evaluated parameters were not changed by this amino acid. Creatine, when combined with homocysteine, prevented or caused a synergistic effect on some changes provoked by this amino acid. Creatine per se or creatine plus homocysteine altered glucose oxidation. These findings provide insights into the mechanisms by which homocysteine exerts its effects on skeletal muscle function, more studies are needed to elucidate them. Although creatine prevents some alterations caused by homocysteine, it should be used with caution, mainly in healthy individuals because it could change the homeostasis of normal physiological functions.

  5. Purification and characterization of creatine kinase isozymes from the nurse shark Ginglymostoma cirratum.

    PubMed

    Gray, K A; Grossman, S H; Summers, D D

    1986-01-01

    Creatine kinase from nurse shark brain and muscle has been purified to apparent homogeneity. In contrast to creatine kinases from most other vertebrate species, the muscle isozyme and the brain isozyme from nurse shark migrate closely in electrophoresis and, unusually, the muscle isozyme is anodal to the brain isozyme. The isoelectric points are 5.3 and 6.2 for the muscle and brain isozymes, respectively. The purified brain preparation also contains a second active protein with pI 6.0. The amino acid content of the muscle isozyme is compared with other isozymes of creatine kinase using the Metzger Difference Index as an estimation of compositional relatedness. All comparisons show a high degree of compositional similarity including arginine kinase from lobster muscle. The muscle isozyme is marginally more resistant to temperature inactivation than the brain isozyme; the muscle protein does not exhibit unusual stability towards high concentrations of urea. Kinetic analysis of the muscle isozyme reveals Michaelis constants of 1.6 mM MgATP, 12 mM creatine, 1.2 mM MgADP and 50 mM creatine phosphate. Dissociation constants for the same substrate from the binary and ternary enzyme-substrate complex do not differ significantly, indicating limited cooperatively in substrate binding. Enzyme activity is inhibited by small planar anions, most severely by nitrate. Shark muscle creatine kinase hybridizes in vitro with rabbit muscle or monkey brain creatine kinase; shark brain isozyme hybridizes with monkey brain or rabbit brain creatine kinase. Shark muscle and shark brain isozymes, under a wide range of conditions, failed to produce a detectable hybrid.

  6. Atomic Layer Deposition of L-Alanine Polypeptide

    SciTech Connect

    Fu, Yaqin; Li, Binsong; Jiang, Ying-Bing; Dunphy, Darren R.; Tsai, Andy; Tam, Siu-Yue; Fan, Hongyou Y.; Zhang, Hongxia; Rogers, David; Rempe, Susan; Atanassov, Plamen; Cecchi, Joseph L.; Brinker, C. Jeffrey

    2014-10-30

    L-Alanine polypeptide thin films were synthesized via atomic layer deposition (ALD). Rather, instead of using an amino acid monomer as the precursor, an L-alanine amino acid derivatized with a protecting group was used to prevent self-polymerization, increase the vapor pressure, and allow linear cycle-by-cycle growth emblematic of ALD. Moreover, the successful deposition of a conformal polypeptide film has been confirmed by FTIR, TEM, and Mass Spectrometry, and the ALD process has been extended to polyvaline.

  7. Incorrect calculation of power outputs masks the ergogenic capacity of creatine supplementation.

    PubMed

    Havenetidis, Konstadinos; Cooke, Carlton B; Butterly, Ron; King, Roderick F G J

    2006-10-01

    This study assessed the effect of incorrect calculation of power output measurement on the ergogenic properties of creatine. Fifteen males performed repeated Wingate anaerobic tests, under baseline, placebo, and creatine conditions. Statistics showed significant differences (p < 0.05) following creatine-supplemented conditions compared with placebo conditions, whereas no significant differences existed between the baseline and placebo conditions. However, the performance enhancement effect of creatine became significant only when the corrected (for the inertia of the flywheel) method was employed for measuring peak and minimum power. Mean (+/- SD) values across all cycle sprints for placebo versus creatine were 1033 +/- 100 W versus 1130 +/- 95 W for peak power and 385 +/- 78 W versus 427 +/- 70 W for minimum power. No significant differences were shown using the uncorrected method for peak power (756 +/- 97 W versus 786 +/- 88 W) and minimum power 440 +/- 64 W pre versus 452 +/- 65 W post). In conclusion, the present study suggests that the potentiating effect of creatine might be underestimated if the inertial effects of the flywheel are not considered in power output determination.

  8. Proton transfer pathways, energy landscape, and kinetics in creatine-water systems.

    PubMed

    Ivchenko, Olga; Whittleston, Chris S; Carr, Joanne M; Imhof, Petra; Goerke, Steffen; Bachert, Peter; Wales, David J

    2014-02-27

    We study the exchange processes of the metabolite creatine, which is present in both tumorous and normal tissues and has NH2 and NH groups that can transfer protons to water. Creatine produces chemical exchange saturation transfer (CEST) contrast in magnetic resonance imaging (MRI). The proton transfer pathway from zwitterionic creatine to water is examined using a kinetic transition network constructed from the discrete path sampling approach and an approximate quantum-chemical energy function, employing the self-consistent-charge density-functional tight-binding (SCC-DFTB) method. The resulting potential energy surface is visualized by constructing disconnectivity graphs. The energy landscape consists of two distinct regions corresponding to the zwitterionic creatine structures and deprotonated creatine. The activation energy that characterizes the proton transfer from the creatine NH2 group to water was determined from an Arrhenius fit of rate constants as a function of temperature, obtained from harmonic transition state theory. The result is in reasonable agreement with values obtained in water exchange spectroscopy (WEX) experiments.

  9. Creatine Supplementation Increases Total Body Water in Soccer Players: a Deuterium Oxide Dilution Study.

    PubMed

    Deminice, R; Rosa, F T; Pfrimer, K; Ferrioli, E; Jordao, A A; Freitas, E

    2016-02-01

    This study aimed to evaluate changes in total body water (TBW) in soccer athletes using a deuterium oxide dilution method and bioelectrical impedance (BIA) formulas after 7 days of creatine supplementation. In a double-blind controlled manner, 13 healthy (under-20) soccer players were divided randomly in 2 supplementation groups: Placebo (Pla, n=6) and creatine supplementation (CR, n=7). Before and after the supplementation period (0.3 g/kg/d during 7 days), TBW was determined by deuterium oxide dilution and BIA methods. 7 days of creatine supplementation lead to a large increase in TBW (2.3±1.0 L) determined by deuterium oxide dilution, and a small but significant increase in total body weight (1.0±0.4 kg) in Cr group compared to Pla. The Pla group did not experience any significant changes in TBW or body weight. Although 5 of 6 BIA equations were sensitive to determine TBW changes induced by creatine supplementation, the Kushner et al. 16 method presented the best concordance levels when compared to deuterium dilution method. In conclusion, 7-days of creatine supplementation increased TBW determined by deuterium oxide dilution or BIA formulas. BIA can be useful to determine TBW changes promoted by creatine supplementation in soccer athletes, with special concern for formula choice.

  10. Crystal structure of Trypanosoma cruzi tyrosine aminotransferase: substrate specificity is influenced by cofactor binding mode.

    PubMed Central

    Blankenfeldt, W.; Nowicki, C.; Montemartini-Kalisz, M.; Kalisz, H. M.; Hecht, H. J.

    1999-01-01

    The crystal structure of tyrosine aminotransferase (TAT) from the parasitic protozoan Trypanosoma cruzi, which belongs to the aminotransferase subfamily Igamma, has been determined at 2.5 A resolution with the R-value R = 15.1%. T. cruzi TAT shares less than 15% sequence identity with aminotransferases of subfamily Ialpha but shows only two larger topological differences to the aspartate aminotransferases (AspATs). First, TAT contains a loop protruding from the enzyme surface in the larger cofactor-binding domain, where the AspATs have a kinked alpha-helix. Second, in the smaller substrate-binding domain, TAT has a four-stranded antiparallel beta-sheet instead of the two-stranded beta-sheet in the AspATs. The position of the aromatic ring of the pyridoxal-5'-phosphate cofactor is very similar to the AspATs but the phosphate group, in contrast, is closer to the substrate-binding site with one of its oxygen atoms pointing toward the substrate. Differences in substrate specificities of T. cruzi TAT and subfamily Ialpha aminotransferases can be attributed by modeling of substrate complexes mainly to this different position of the cofactor-phosphate group. Absence of the arginine, which in the AspATs fixes the substrate side-chain carboxylate group by a salt bridge, contributes to the inability of T. cruzi TAT to transaminate acidic amino acids. The preference of TAT for tyrosine is probably related to the ability of Asn17 in TAT to form a hydrogen bond to the tyrosine side-chain hydroxyl group. PMID:10595543

  11. Isolation and characterization of a gene coding for a novel aspartate aminotransferase from Rhizobium meliloti.

    PubMed Central

    Alfano, J R; Kahn, M L

    1993-01-01

    Aspartate aminotransferase (AAT) is an important enzyme in aspartate catabolism and biosynthesis and, by converting tricarboxylic acid cycle intermediates to amino acids, AAT is also significant in linking carbon metabolism with nitrogen metabolism. To examine the role of AAT in symbiotic nitrogen fixation further, plasmids encoding three different aminotransferases from Rhizobium meliloti 104A14 were isolated by complementation of an Escherichia coli auxotroph that lacks three aminotransferases. pJA10 contained a gene, aatB, that coded for a previously undescribed AAT, AatB. pJA30 encoded an aromatic aminotransferase, TatA, that had significant AAT activity, and pJA20 encoded a branched-chain aminotransferase designated BatA. Genes for the latter two enzymes, tatA and batA, were previously isolated from R. meliloti. aatB is distinct from but hybridizes to aatA, which codes for AatA, a protein required for symbiotic nitrogen fixation. The DNA sequence of aatB contained an open reading frame that could encode a protein 410 amino acids long and with a monomer molecular mass of 45,100 Da. The amino acid sequence of aatB is unusual, and AatB appears to be a member of a newly described class of AATs. AatB expressed in E. coli has a Km for aspartate of 5.3 mM and a Km for 2-oxoglutarate of 0.87 mM. Its pH optimum is between 8.0 and 8.5. Mutations were constructed in aatB and tatA and transferred to the genome of R. meliloti 104A14. Both mutants were prototrophs and were able to carry out symbiotic nitrogen fixation. Images PMID:8320232

  12. Stereoselective aminoacylation of a dinucleoside monophosphate by the imidazolides of DL-alanine and N-(tert-butoxycarbonyl)-DL-alanine

    NASA Technical Reports Server (NTRS)

    Profy, A. T.; Usher, D. A.

    1984-01-01

    The aminoacylation of diinosine monophosphate was studied experimentally. When the acylating agent was the imidazolide of N-(tert-butoxycarbonyl)-DL-alanine, a 40 percent enantiomeric excess of the isomer was incorporated at the 2' site and the positions of equilibrium for the reversible 2'-3' migration reaction differed for the D and L enantiomers. The reactivity of the nucleoside hydroxyl groups was found to decrease on the order 2'(3') less than internal 2' and less than 5', and the extent of the reaction was affected by the concentration of the imidazole buffer. Reaction of IpI with imidazolide of unprotected DL-alanine, by contrast, led to an excess of the D isomer at the internal 2' site. Finally, reaction with the N-carboxy anhydride of DL-alanine occurred without stereoselection. These results are found to be relevant to the study of the evolution of optical chemical activity and the origin of genetically directed protein synthesis.

  13. Attenuation by creatine of myocardial metabolic stress in Brattleboro rats caused by chronic inhibition of nitric oxide synthase.

    PubMed Central

    Constantin-Teodosiu, D.; Greenhaff, P. L.; Gardiner, S. M.; Randall, M. D.; March, J. E.; Bennett, T.

    1995-01-01

    1. The present experiment was undertaken to investigate: (a) the effect of nitric oxide synthase (NOS) inhibition, mediated by oral supplementation of the NOS inhibitor, NG-nitro-L-arginine methyl ester (L-NAME), on measures of myocardial energy metabolism and function: (b) the effect of oral creatine supplementation on these variables, in the absence and presence of L-NAME. 2. In one series of experiments, 4 weeks oral administration of L-NAME (0.05 mg ml-1 day-1 in the drinking water) to Brattleboro rats caused significant reductions in myocardial ATP, creatine, and total creatine concentrations and an accumulation of tissue lactate when compared with control animals. Administration of creatine (0.63 mg ml-1 day-1 in the drinking water) for 4 weeks elevated myocardial creatine and total creatine concentrations and reduced lactate accumulation, but did not significantly affect ATP or phosphocreatine (PCr). Concurrent treatment with creatine and L-NAME prevented the reduction in creatine and total creatine concentrations, and significantly attenuated the accumulation of lactate and the reduction in ATP seen with L-NAME alone. 3. In a second series of experiments, 4 weeks treatment with L-NAME and creatine plus L-NAME increased mean arterial blood pressure in conscious Brattleboro rats. Hearts isolated from these animals showed decreased coronary flow and left ventricular developed pressure (LVDP), and total mechanical performance. Treatment with creatine alone had no measurable effect on either mean arterial blood pressure or coronary flow in isolated hearts. However, there was an increase in LVDP, but not in total mechanical performance, because there was a bradycardia. 4. These results indicate that creatine supplementation can attenuate the metabolic stress associated with L-NAME administration and that this effect occurs as a consequence of the action of creatine on myocardial energy metabolism. PMID:8719809

  14. [Regulation of key enzymes of L-alanine biosynthesis by Brevibacterium flavum producer strains].

    PubMed

    Melkonian, L O; Avetisova, G E; Ambartsumian, A A; Chakhalian, A Kh; Sagian, A S

    2013-01-01

    The mechanisms of L-alanine overproduction by Brevibacterium flavum producer strains were studied. It was shown that beta-CI-L-alanine is an inhibitor of some key enzymes involved in the synthesis of L-alanine, including alanine transaminase and valine-pyruvate transaminase. Two highly active B. flavum GL1 and GL1 8 producer strains, which are resistant to the inhibitory effect of beta-Cl-L-alanine, were obtained using a parental B. flavum AA5 producer strain, characterized by a reduced activity of alanine racemase (>or=98%). It was demonstrated that the increased L-alanine synthesis efficiency observed in the producer strains developed in this work is associated with the absence of inhibition of alanine transaminase by the end product of the biosynthesis reaction, as well as with the effect of derepression of both alanine transaminase and valine-pyruvate transaminase synthesis by the studied compound.

  15. Creatine supplementation with methylglyoxal: a potent therapy for cancer in experimental models.

    PubMed

    Pal, Aparajita; Roy, Anirban; Ray, Manju

    2016-08-01

    The anti-cancer effect of methylglyoxal (MG) is now well established in the literature. The main aim of this study was to investigate the effect of creatine as a supplement in combination with MG both in vitro and in vivo. In case of the in vitro studies, two different cell lines, namely MCF-7 (human breast cancer cell line) and C2C12 (mouse myoblast cell line) were chosen. MG in combination with creatine showed enhanced apoptosis as well as higher cytotoxicity in the breast cancer MCF-7 cell line, compared to MG alone. Pre-treatment of well-differentiated C2C12 myotubes with cancerogenic 3-methylcholanthrene (3MC) induced a dedifferentiation of these myotubes towards cancerous cells (that mimic the effect of 3MC observed in solid fibro-sarcoma animal models) and subsequent exposure of these induced cancer cells with MG proved to be cytotoxic. Thus, creatine plus ascorbic acid enhanced the anti-cancer effects of MG. In contrast, when normal C2C12 muscle cells or myotubes (mouse normal myoblast cell line) were treated with MG or MG plus creatine and ascorbic acid, no detrimental effects were seen. This indicated that cytotoxic effects of MG are specifically limited towards cancer cells and are further enhanced when MG is used in combination with creatine and ascorbic acid. For the in vivo studies, tumors were induced by injecting Sarcoma-180 cells (2 × 10(6) cells/mouse) in the left hind leg. After 7 days of tumor inoculation, treatments were started with MG (20 mg/kg body wt/day, via the intravenous route), with or without creatine (150 mg/kg body wt/day, fed orally) and ascorbic acid (50 mg/kg body wt/day, fed orally) and continued for 10 consecutive days. Significant regression of tumor size was observed when Sarcoma-180 tumor-bearing mice were treated with MG and even more so with the aforesaid combination. The creatine-supplemented group demonstrated better overall survival in comparison with tumor-bearing mice without creatine. In conclusion, it may be

  16. Formation of {gamma}-alumina nanorods in presence of alanine

    SciTech Connect

    Dabbagh, Hossein A.; Rasti, Elham; Yalfani, Mohammad S.; Medina, Francesc

    2011-02-15

    Graphical abstract: Nanorod aluminas with a possible hexagonal symmetry, high surface area and relatively narrow pore size distribution were obtained. Research highlights: {yields} Research highlights {yields} Boehmite was prepared using a green sol-gel process in the presence of alanine. {yields} Nanorod aluminas with a high surface area were obtained. {yields} Addition of alanine would shape the size of the holes and crevices. {yields} The morphologies of the nanorods were revealed by transmission electron microscope. -- Abstract: Boehmite and alumina nanostructures were prepared using a simple green sol-gel process in the presence of alanine in water medium at room temperature. The uncalcined (dried at 200 {sup o}C) and the calcined materials (at 500, 600 and 700 {sup o}C for 4 h) were characterized using XRD, TEM, SEM, N{sub 2} physisorption and TGA. Nanorod aluminas with a possible hexagonal symmetry, high surface area and relatively narrow pore size distribution were obtained. The surface area was enhanced and crystallization was retarded as the alanine content increased. The morphologies of the nanoparticles and nanorods were revealed by a transmission electron microscope (TEM).

  17. A theoretical study of alanine dipeptide and analogs

    SciTech Connect

    Head-Gordon, T.; Head-Gordon, M.; Brooks, C. III; Pople, J. ); Frisch, M.J. )

    1989-01-01

    We Present a preliminary report on the conformational and energetic analysis of the molecule (S)-2-acetylamino-N-methylpropanamide (alanine dipeptide) and an analog molecule, (S)-{alpha}-formylaminopropanamide, using high-quality ab initio methods. Alanine dipeptide and its analogs are of interest since they incorporate many of the structural features found in proteins, such as intramolecular hydrogen bonds, conformational flexibility, and a variety of chemical functionality. One purpose of this study is to provide a useful benchmark calculation, MP2/6-31+G{sup **}//HF/6-31+G{sup *}, for a number of conformations of the alanine system. Based on the comparison of these benchmark calculations with lower-level basis sets, HF/3-21G was chosen to generate a fully relaxed {phi}, {psi} dihedral map. These calculations are the first of their kind on systems of this size. Features of the {phi},{psi} alanine dipeptide map that are discussed include the energetically accessible conformations and possible pathways for their interconversion. In addition, we illustrate the importance of fully optimized geometries and the proper evaluation of correlation energies,

  18. Spectrophotometric readout for an alanine dosimeter for food irradiation applications

    NASA Astrophysics Data System (ADS)

    Ebraheem, S.; Beshir, W. B.; Eid, S.; Sobhy, R.; Kovács, A.

    2003-06-01

    The alanine-electron spin resonance (EPR) readout system is well known as a reference and transfer dosimetry system for the evaluation of high doses in radiation processing. The high cost of an EPR/alanine dosimetry system is a serious handicap for large-scale routine application in irradiation facilities. In this study, the use of a complex produced by dissolving irradiated L-alanine in 1,4-phenyl diammonium dichloride solution was investigated for dosimetry purposes. This complex—having a purple colour—has an increasing absorbance with increasing dose in the range of 1-20 kGy. The applicability of spectrophotometric evaluation was studied by measuring the absorbance intensity of this complex at 360 and 505 nm, respectively. Fluorimetric evaluation was also investigated by measuring the emission of the complex at 435 nm as a function of dose. The present method is easy for routine application. The effect of the dye concentration as well as the suitable amount of irradiated alanine has been studied. With respect to routine application, the stability of the product complex after its formation was also investigated.

  19. Computational alanine scanning with linear scaling semiempirical quantum mechanical methods.

    PubMed

    Diller, David J; Humblet, Christine; Zhang, Xiaohua; Westerhoff, Lance M

    2010-08-01

    Alanine scanning is a powerful experimental tool for understanding the key interactions in protein-protein interfaces. Linear scaling semiempirical quantum mechanical calculations are now sufficiently fast and robust to allow meaningful calculations on large systems such as proteins, RNA and DNA. In particular, they have proven useful in understanding protein-ligand interactions. Here we ask the question: can these linear scaling quantum mechanical methods developed for protein-ligand scoring be useful for computational alanine scanning? To answer this question, we assembled 15 protein-protein complexes with available crystal structures and sufficient alanine scanning data. In all, the data set contains Delta Delta Gs for 400 single point alanine mutations of these 15 complexes. We show that with only one adjusted parameter the quantum mechanics-based methods outperform both buried accessible surface area and a potential of mean force and compare favorably to a variety of published empirical methods. Finally, we closely examined the outliers in the data set and discuss some of the challenges that arise from this examination.

  20. [A family with creatine transporter deficiency diagnosed with urinary creatine/creatinine ratio and the family history: the third Japanese familial case].

    PubMed

    Nozaki, Fumihito; Kumada, Tomohiro; Shibata, Minoru; Fujii, Tatsuya; Wada, Takahito; Osaka, Hitoshi

    2015-01-01

    Creatine transporter deficiency (CRTR-D) is an X-linked disorder characterized by hypotonia, developmental delay, and seizures. We report the third Japanese family with CRTR-D. The proband was an 8-year-old boy who presented with hypotonia, severe intellectual disability and two episodes of seizures associated with/without fever. Among 7 siblings (4 males, 3 females), the eldest brother had severe intellectual disability, epilepsy, and sudden death at 17 years of age, while 18-year-old third elder brother had severe intellectual disability, autism, and drug-resistant epilepsy. The proband's urinary creatine/creatinine ratio was increased. A reduced creatine peak on brain magnetic resonance spectroscopy and a known pathogenic mutation in the SLC6A8 gene (c.1661 C > T;p.Pro554Leu) confirmed the diagnosis of CRTR-D. The same mutation was found in the third elder brother. Their mother was a heterozygote. Symptoms of CRTR-D are non-specific. Urinary creatine/creatinine ratio should be measured in patients with hypotonia, developmental delay, seizure and autism whose family history indicates an X-linked inheritance.

  1. Effects of Coffee and Caffeine Anhydrous Intake During Creatine Loading.

    PubMed

    Trexler, Eric T; Smith-Ryan, Abbie E; Roelofs, Erica J; Hirsch, Katie R; Persky, Adam M; Mock, Meredith G

    2016-05-01

    The purpose of this study was to determine the effect of 5 days of creatine (CRE) loading alone or in combination with caffeine anhydrous (CAF) or coffee (COF) on upper-body and lower-body strength and sprint performance. Physically active males (n = 54; mean ± SD; age = 20.1 ± 2.1 years; weight = 78.8 ± 8.8 kg) completed baseline testing, consisting of 1 repetition maximum (1RM) and repetitions to fatigue with 80% 1RM for bench press and leg press, followed by a repeated sprint test of five, 10-second sprints separated by 60-second rest on a cycle ergometer to determine peak power (PP) and total power (TP). At least 72 hours later, subjects were randomly assigned to supplement with CRE (5 g of CRE monohydrate, 4 times per day; n = 14), CRE + CAF (CRE +300 mg·d of CAF; n = 13), CRE + COF (CRE +8.9 g of COF, yielding 303 mg of CAF; n = 13), or placebo (PLA; n = 14) for 5 days. Serum creatinine (CRN) was measured before and after supplementation, and on day 6, participants repeated pretesting procedures. Strength measures were improved in all groups (p ≤ 0.05), with no significant time × treatment interactions. No significant interaction or main effects were observed for PP. For TP, a time × sprint interaction was observed (p ≤ 0.05), with no significant interactions among treatment groups. A time × treatment interaction was observed for serum CRN values (p ≤ 0.05) that showed increases in all groups except PLA. Four subjects reported mild gastrointestinal discomfort with CRE + CAF, with no side effects reported in other groups. These findings suggest that neither CRE alone nor in combination with CAF or COF significantly affected performance compared with PLA.

  2. Reference intervals for serum creatine kinase in athletes

    PubMed Central

    Mougios, Vassilis

    2007-01-01

    Background The serum concentration of creatine kinase (CK) is used widely as an index of skeletal muscle fibre damage in sport and exercise. Since athletes have higher CK values than non‐athletes, comparing the values of athletes to the normal values established in non‐athletes is pointless. The purpose of this study was to introduce reference intervals for CK in athletes. Method CK was assayed in serum samples from 483 male athletes and 245 female athletes, aged 7–44. Samples had been obtained throughout the training and competition period. For comparison, CK was also assayed in a smaller number of non‐athletes. Reference intervals (2.5th to 97.5th percentile) were calculated by the non‐parametric method. Results The reference intervals were 82–1083 U/L (37°C) in male and 47–513 U/L in female athletes. The upper reference limits were twice the limits reported for moderately active non‐athletes in the literature or calculated in the non‐athletes in this study. The upper limits were up to six times higher than the limits reported for inactive individuals in the literature. When reference intervals were calculated specifically in male football (soccer) players and swimmers, a threefold difference in the upper reference limit was found (1492 vs 523 U/L, respectively), probably resulting from the different training and competition demands of the two sports. Conclusion Sport training and competition have profound effects on the reference intervals for serum CK. Introducing sport‐specific reference intervals may help to avoid misinterpretation of high values and to optimise training. PMID:17526622

  3. The unresolved puzzle why alanine extensions cause disease.

    PubMed

    Winter, Reno; Liebold, Jens; Schwarz, Elisabeth

    2013-08-01

    The prospective increase in life expectancy will be accompanied by a rise in the number of elderly people who suffer from ill health caused by old age. Many diseases caused by aging are protein misfolding diseases. The molecular mechanisms underlying these disorders receive constant scientific interest. In addition to old age, mutations also cause congenital protein misfolding disorders. Chorea Huntington, one of the most well-known examples, is caused by triplet extensions that can lead to more than 100 glutamines in the N-terminal region of huntingtin, accompanied by huntingtin aggregation. So far, nine disease-associated triplet extensions have also been described for alanine codons. The extensions lead primarily to skeletal malformations. Eight of these proteins represent transcription factors, while the nuclear poly-adenylate binding protein 1, PABPN1, is an RNA binding protein. Additional alanines in PABPN1 lead to the disease oculopharyngeal muscular dystrophy (OPMD). The alanine extension affects the N-terminal domain of the protein, which has been shown to lack tertiary contacts. Biochemical analyses of the N-terminal domain revealed an alanine-dependent fibril formation. However, fibril formation of full-length protein did not recapitulate the findings of the N-terminal domain. Fibril formation of intact PABPN1 was independent of the alanine segment, and the fibrils displayed biochemical properties that were completely different from those of the N-terminal domain. Although intranuclear inclusions have been shown to represent the histochemical hallmark of OPMD, their role in pathogenesis is currently unclear. Several cell culture and animal models have been generated to study the molecular processes involved in OPMD. These studies revealed a number of promising future therapeutic strategies that could one day improve the quality of life for the patients.

  4. Crystal structures of d-alanine-d-alanine ligase from Xanthomonas oryzae pv. oryzae alone and in complex with nucleotides.

    PubMed

    Doan, Thanh Thi Ngoc; Kim, Jin-Kwang; Ngo, Ho-Phuong-Thuy; Tran, Huyen-Thi; Cha, Sun-Shin; Min Chung, Kyung; Huynh, Kim-Hung; Ahn, Yeh-Jin; Kang, Lin-Woo

    2014-03-01

    D-Alanine-D-alanine ligase (DDL) catalyzes the biosynthesis of d-alanyl-d-alanine, an essential bacterial peptidoglycan precursor, and is an important drug target for the development of antibacterials. We determined four different crystal structures of DDL from Xanthomonas oryzae pv. oryzae (Xoo) causing Bacteria Blight (BB), which include apo, ADP-bound, ATP-bound, and AMPPNP-bound structures at the resolution between 2.3 and 2.0 Å. Similarly with other DDLs, the active site of XoDDL is formed by three loops from three domains at the center of enzyme. Compared with d-alanyl-d-alanine and ATP-bound TtDDL structure, the γ-phosphate of ATP in XoDDL structure was shifted outside toward solution. We swapped the ω-loop (loop3) of XoDDL with those of Escherichia coli and Helicobacter pylori DDLs, and measured the enzymatic kinetics of wild-type XoDDL and two mutant XoDDLs with the swapped ω-loops. Results showed that the direct interactions between ω-loop and other two loops are essential for the active ATP conformation for D-ala-phosphate formation.

  5. Diminution of Oxidative Damage to Human Erythrocytes and Lymphocytes by Creatine: Possible Role of Creatine in Blood.

    PubMed

    Qasim, Neha; Mahmood, Riaz

    2015-01-01

    Creatine (Cr) is naturally produced in the body and stored in muscles where it is involved in energy generation. It is widely used, especially by athletes, as a staple supplement for improving physical performance. Recent reports have shown that Cr displays antioxidant activity which could explain its beneficial cellular effects. We have evaluated the ability of Cr to protect human erythrocytes and lymphocytes against oxidative damage. Erythrocytes were challenged with model oxidants, 2, 2'-azobis(2-amidinopropane) dihydrochloride (AAPH) and hydrogen peroxide (H2O2) in the presence and absence of Cr. Incubation of erythrocytes with oxidant alone increased hemolysis, methemoglobin levels, lipid peroxidation and protein carbonyl content. This was accompanied by decrease in glutathione levels. Antioxidant enzymes and antioxidant power of the cell were compromised while the activity of membrane bound enzyme was lowered. This suggests induction of oxidative stress in erythrocytes by AAPH and H2O2. However, Cr protected the erythrocytes by ameliorating the AAPH and H2O2 induced changes in these parameters. This protective effect was confirmed by electron microscopic analysis which showed that oxidant-induced cell damage was attenuated by Cr. No cellular alterations were induced by Cr alone even at 20 mM, the highest concentration used. Creatinine, a by-product of Cr metabolism, was also shown to exert protective effects, although it was slightly less effective than Cr. Human lymphocytes were similarly treated with H2O2 in absence and presence of different concentrations of Cr. Lymphocytes incubated with oxidant alone had alterations in various biochemical and antioxidant parameters including decrease in cell viability and induction of DNA damage. The presence of Cr attenuated all these H2O2-induced changes in lymphocytes. Thus, Cr can function as a blood antioxidant, protecting cells from oxidative damage, genotoxicity and can potentially increase their lifespan.

  6. Diminution of Oxidative Damage to Human Erythrocytes and Lymphocytes by Creatine: Possible Role of Creatine in Blood

    PubMed Central

    Qasim, Neha; Mahmood, Riaz

    2015-01-01

    Creatine (Cr) is naturally produced in the body and stored in muscles where it is involved in energy generation. It is widely used, especially by athletes, as a staple supplement for improving physical performance. Recent reports have shown that Cr displays antioxidant activity which could explain its beneficial cellular effects. We have evaluated the ability of Cr to protect human erythrocytes and lymphocytes against oxidative damage. Erythrocytes were challenged with model oxidants, 2, 2'-azobis(2-amidinopropane) dihydrochloride (AAPH) and hydrogen peroxide (H2O2) in the presence and absence of Cr. Incubation of erythrocytes with oxidant alone increased hemolysis, methemoglobin levels, lipid peroxidation and protein carbonyl content. This was accompanied by decrease in glutathione levels. Antioxidant enzymes and antioxidant power of the cell were compromised while the activity of membrane bound enzyme was lowered. This suggests induction of oxidative stress in erythrocytes by AAPH and H2O2. However, Cr protected the erythrocytes by ameliorating the AAPH and H2O2 induced changes in these parameters. This protective effect was confirmed by electron microscopic analysis which showed that oxidant-induced cell damage was attenuated by Cr. No cellular alterations were induced by Cr alone even at 20 mM, the highest concentration used. Creatinine, a by-product of Cr metabolism, was also shown to exert protective effects, although it was slightly less effective than Cr. Human lymphocytes were similarly treated with H2O2 in absence and presence of different concentrations of Cr. Lymphocytes incubated with oxidant alone had alterations in various biochemical and antioxidant parameters including decrease in cell viability and induction of DNA damage. The presence of Cr attenuated all these H2O2-induced changes in lymphocytes. Thus, Cr can function as a blood antioxidant, protecting cells from oxidative damage, genotoxicity and can potentially increase their lifespan. PMID

  7. Formation of simple biomolecules from alanine in ocean by impacts

    NASA Astrophysics Data System (ADS)

    Umeda, Y.; Sekine, T.; Furukawa, Y.; Kakegawa, T.; Kobayashi, T.

    2013-12-01

    The biomolecules on the Earth are thought either to have originated from the extraterrestrial parts carried with flying meteorites or to have been formed from the inorganic materials on the Earth through given energy. From the standpoint to address the importance of impact energy, it is required to simulate experimentally the chemical reactions during impacts, because violent impacts may have occurred 3.8-4.0 Gyr ago to create biomolecules initially. It has been demonstrated that shock reactions among ocean (H2O), atmospheric nitrogen, and meteoritic constitution (Fe) can induce locally reduction environment to form simple bioorganic molecules such as ammonia and amino acid (Nakazawa et al., 2005; Furukawa et al., 2009). We need to know possible processes for alanine how chemical reactions proceed during repeated impacts and how complicated biomolecules are formed. Alanine can be formed from glycine (Umeda et al., in preparation). In this study, we carried out shock recovery experiments at pressures of 4.4-5.7 GPa to investigate the chemical reactions of alanine. Experiments were carried out with a propellant gun. Stainless steel containers (30 mm in diameter, 30 mm long) with 13C-labeled alanine aqueous solution immersed in olivine or hematite powders were used as targets. Air gap was present in the sample room (18 mm in diameter, 2 mm thick) behind the sample. The powder, solution, and air represent meteorite, ocean, and atmosphere on early Earth, respectively. Two powders of olivine and hematite help to keep the oxygen fugacity low and high during experiments, respectively in order to investigate the effect of oxygen fugacity on chemical processes of alanine. The recovered containers, after cleaned completely, were immersed into liquid nitrogen to freeze sample solution and then we drilled on the impact surface to extract water-soluble run products using pure water. Thus obtained products were analyzed by LC/MS for four amino acids (glycine, alanine, valine, and

  8. Participation of cysteine and cystine in inactivation of tyrosine aminotransferase in rat liver homogenates.

    PubMed Central

    Buckley, W T; Milligan, L P

    1978-01-01

    1. Inactivation of tyrosine aminotransferase was studied in rat liver homogenates. Under an O2 atmosphere with cysteine added, inactivation was rapid after a lag period of approx. 1h, whereas a N2 atmosphere extended the lag period to approx. 3h. 2. Replacement of cysteine with cystine resulted in rapid inactivation both aerobically and anaerobically. 3. Removal of the particulate fraction by centrifuging rat liver homogenates at 13,000g for 9min resulted in an aerobic lag period of 0.5h in the presence of cystine and approx. 3h in the presence of cysteine. 4. It is proposed that the stimulatory effect of cysteine on tyrosine aminotransferase inactivation occurs largely as a result of oxidation to cystine, which appears to be a more directly effective agent. PMID:33669

  9. Kinetic mechanism and inhibition of Mycobacterium tuberculosis D-alanine:D-alanine ligase by the antibiotic D-cycloserine.

    PubMed

    Prosser, Gareth A; de Carvalho, Luiz Pedro S

    2013-02-01

    D-cycloserine (DCS) is an antibiotic that is currently used in second-line treatment of tuberculosis. DCS is a structural analogue of D-alanine, and targets two enzymes involved in the cytosolic stages of peptidoglycan synthesis: alanine racemase (Alr) and D-alanine:D-alanine ligase (Ddl). The mechanisms of inhibition of DCS have been well-assessed using Alr and Ddl enzymes from various bacterial species, but little is known regarding the interactions of DCS with the mycobacterial orthologues of these enzymes. We have over-expressed and purified recombinant Mycobacterium tuberculosis Ddl (MtDdl; Rv2981c), and report a kinetic examination of the enzyme with both its native substrate and DCS. MtDdl is activated by K(+), follows an ordered ter ter mechanism and displays distinct affinities for D-Ala at each D-Ala binding site (K(m,D-Ala1) = 0.075 mm, K(m,D-Ala2) = 3.6 mm). ATP is the first substrate to bind and is necessary for subsequent binding of D-alanine or DCS. The pH dependence of MtDdl kinetic parameters indicate that general base chemistry is involved in the catalytic step. DCS was found to competitively inhibit D-Ala binding at both MtDdl D-Ala sites with equal affinity (K(i,DCS1) = 14 μm, K(i,DCS2) = 25 μm); however, each enzyme active site can only accommodate a single DCS molecule at a given time. The pH dependence of K(i,DCS2) revealed a loss of DCS binding affinity at high pH (pK(a) = 7.5), suggesting that DCS binds optimally in the zwitterionic form. The results of this study may assist in the design and development of novel Ddl-specific inhibitors for use as anti-mycobacterial agents.

  10. Structure of the Mycobacterium tuberculosis D-Alanine:D-Alanine Ligase, a Target of the Antituberculosis Drug D-Cycloserine

    SciTech Connect

    Bruning, John B.; Murillo, Ana C.; Chacon, Ofelia; Barletta, Raúl G.; Sacchettini, James C.

    2011-09-28

    D-Alanine:D-alanine ligase (EC 6.3.2.4; Ddl) catalyzes the ATP-driven ligation of two D-alanine (D-Ala) molecules to form the D-alanyl:D-alanine dipeptide. This molecule is a key building block in peptidoglycan biosynthesis, making Ddl an attractive target for drug development. D-Cycloserine (DCS), an analog of D-Ala and a prototype Ddl inhibitor, has shown promise for the treatment of tuberculosis. Here, we report the crystal structure of Mycobacterium tuberculosis Ddl at a resolution of 2.1 {angstrom}. This structure indicates that Ddl is a dimer and consists of three discrete domains; the ligand binding cavity is at the intersection of all three domains and conjoined by several loop regions. The M. tuberculosis apo Ddl structure shows a novel conformation that has not yet been observed in Ddl enzymes from other species. The nucleotide and D-alanine binding pockets are flexible, requiring significant structural rearrangement of the bordering regions for entry and binding of both ATP and D-Ala molecules. Solution affinity and kinetic studies showed that DCS interacts with Ddl in a manner similar to that observed for D-Ala. Each ligand binds to two binding sites that have significant differences in affinity, with the first binding site exhibiting high affinity. DCS inhibits the enzyme, with a 50% inhibitory concentration (IC{sub 50}) of 0.37 mM under standard assay conditions, implicating a preferential and weak inhibition at the second, lower-affinity binding site. Moreover, DCS binding is tighter at higher ATP concentrations. The crystal structure illustrates potential drugable sites that may result in the development of more-effective Ddl inhibitors.

  11. Deciphering the role of aspartate and prephenate aminotransferase activities in plastid nitrogen metabolism.

    PubMed

    de la Torre, Fernando; El-Azaz, Jorge; Avila, Concepción; Cánovas, Francisco M

    2014-01-01

    Chloroplasts and plastids of nonphotosynthetic plant cells contain two aspartate (Asp) aminotransferases: a eukaryotic type (Asp5) and a prokaryotic-type bifunctional enzyme displaying Asp and prephenate aminotransferase activities (PAT). We have identified the entire Asp aminotransferase gene family in Nicotiana benthamiana and isolated and cloned the genes encoding the isoenzymes with plastidic localization: NbAsp5 and NbPAT. Using a virus-induced gene silencing approach, we obtained N. benthamiana plants silenced for NbAsp5 and/or NbPAT. Phenotypic and metabolic analyses were conducted in silenced plants to investigate the specific roles of these enzymes in the biosynthesis of essential amino acids within the plastid. The NbAsp5 silenced plants had no changes in phenotype, exhibiting similar levels of free Asp and glutamate as control plants, but contained diminished levels of asparagine and much higher levels of lysine. In contrast, the suppression of NbPAT led to a severe reduction in growth and strong chlorosis symptoms. NbPAT silenced plants exhibited extremely reduced levels of asparagine and were greatly affected in their phenylalanine metabolism and lignin deposition. Furthermore, NbPAT suppression triggered a transcriptional reprogramming in plastid nitrogen metabolism. Taken together, our results indicate that NbPAT has an overlapping role with NbAsp5 in the biosynthesis of Asp and a key role in the production of phenylalanine for the biosynthesis of phenylpropanoids. The analysis of NbAsp5/NbPAT cosilenced plants highlights the central role of both plastidic aminotransferases in nitrogen metabolism; however, only NbPAT is essential for plant growth and development.

  12. Determination of the anti-inflammatory and cytoprotective effects of l-glutamine and l-alanine, or dipeptide, supplementation in rats submitted to resistance exercise.

    PubMed

    Raizel, Raquel; Leite, Jaqueline Santos Moreira; Hypólito, Thaís Menezes; Coqueiro, Audrey Yule; Newsholme, Philip; Cruzat, Vinicius Fernandes; Tirapegui, Julio

    2016-08-01

    We evaluated the effects of chronic oral supplementation with l-glutamine and l-alanine in their free form or as the dipeptide l-alanyl-l-glutamine (DIP) on muscle damage, inflammation and cytoprotection, in rats submitted to progressive resistance exercise (RE). Wistar rats (n 8/group) were submitted to 8-week RE, which consisted of climbing a ladder with progressive loads. In the final 21 d before euthanasia, supplements were delivered in a 4 % solution in drinking water. Glutamine, creatine kinase (CK), lactate dehydrogenase (LDH), TNF-α, specific IL (IL-1β, IL-6 and IL-10) and monocyte chemoattractant protein-1 (MCP-1) levels were evaluated in plasma. The concentrations of glutamine, TNF-α, IL-6 and IL-10, as well as NF-κB activation, were determined in extensor digitorum longus (EDL) skeletal muscle. HSP70 level was assayed in EDL and peripheral blood mononuclear cells (PBMC). RE reduced glutamine concentration in plasma and EDL (P<0·05 v. sedentary group). However, l-glutamine supplements (l-alanine plus l-glutamine (GLN+ALA) and DIP groups) restored glutamine levels in plasma (by 40 and 58 %, respectively) and muscle (by 93 and 105 %, respectively). GLN+ALA and DIP groups also exhibited increased level of HSP70 in EDL and PBMC, consistent with the reduction of NF-κB p65 activation and cytokines in EDL. Muscle protection was also indicated by attenuation in plasma levels of CK, LDH, TNF-α and IL-1β, as well as an increase in IL-6, IL-10 and MCP-1. Our study demonstrates that chronic oral l-glutamine treatment (given with l-alanine or as dipeptide) following progressive RE induces cyprotective effects mediated by HSP70-associated responses to muscle damage and inflammation.

  13. Dietary supplementation of creatine monohydrate reduces the human fMRI BOLD signal.

    PubMed

    Hammett, Stephen T; Wall, Matthew B; Edwards, Thomas C; Smith, Andrew T

    2010-08-02

    Creatine monohydrate is an organic acid that plays a key role in ATP re-synthesis. Creatine levels in the human brain vary considerably and dietary supplementation has been found to enhance cognitive performance in healthy individuals. To explore the possibility that the fMRI Blood Oxygen Level Dependent (BOLD) response is influenced by creatine levels, BOLD responses to visual stimuli were measured in visual cortex before and after a week of creatine administration in healthy human volunteers. The magnitude of the BOLD response decreased by 16% following creatine supplementation of a similar dose to that previously shown to increase cerebral levels of phosphocreatine. We also confirmed that cognitive performance (memory span) is increased. These changes were not found in a placebo group. Possible mechanisms of BOLD change are considered. The results offer potential for insight into the coupling between neural activity and the BOLD response and the more immediate possibility of accounting for an important source of variability during fMRI analysis in clinical studies and other investigations where between-subjects variance is an issue.

  14. Effects of creatine supplementation on biomarkers of hepatic and renal function in young trained rats.

    PubMed

    Souza, William Marciel; Heck, Thiago Gomes; Wronski, Evanio Castor; Ulbrich, Anderson Zampier; Boff, Everton

    2013-11-01

    Creatine supplementation has been widely used by athletes and young physical exercise practioneers in order of increasing muscle mass and enhancing athletic performance, but their use/overuse may represent a health risk on hepatic and renal impaired function. In this study, we evaluated the effects of 40 days of oral creatine supplementation on hepatic and renal function biomarkers in a young animal model. Wistar rats (5 weeks old) were divided in five groups (n = 7): control (CONTR), oral creatine supplementation (CREAT), moderate exercise training (EXERC), moderate exercise training plus oral creatine supplementation (EXERC + CREAT) and pathological group (positive control for liver and kidney injury) by the administration of rifampicin (RIFAMPICIN). Exercise groups were submitted to 60 min/day of swimming exercise session with a 4% of body weight workload for six weeks. The EXERC + CREAT showed the higher body weight at the end of the training protocol. The CREAT and EXERC + CREAT group showed an increase in hepatic (Aspartate transaminase and gamma-glutamyl transpeptidase) and renal (urea and creatinine) biomarkers levels (p < 0.05). Our study showed that the oral creatine supplementation promoted hepatic and renal function challenge in young rats submitted to moderate exercise training.

  15. Low dose creatine supplementation enhances sprint phase of 400 meters swimming performance.

    PubMed

    Anomasiri, Wilai; Sanguanrungsirikul, Sompol; Saichandee, Pisut

    2004-09-01

    This study demonstrated the effect of low dose creatine supplement (10 g. per day) on the sprinting time in the last 50 meters of 400 meters swimming competition, as well as the effect on exertion. Nineteen swimmers in the experimental group received creatine monohydrate 5 g with orange solution 15 g, twice per day for 7 days and nineteen swimmers in the control group received the same quantity of orange solution. The results showed that the swimmers who received creatine supplement lessened the sprinting time in the last 50 meters of 400 meters swimming competition than the control group. (p<0.05). The results of Wingate test (anaerobic power, anaerobic capacity and fatigue index) compared between pre and post supplementation. There was significant difference at p<0.05 in the control group from training effect whereas there was significant difference at p<0.000 from training effect and creatine supplement in the experiment group. Therefore, the creatine supplement in amateur swimmers in the present study enhanced the physical performance up to the maximum capacity.

  16. ALTERATIONS IN BRAIN CREATINE CONCENTRATIONS UNDER LONG-TERM SOCIAL ISOLATION (EXPERIMENTAL STUDY).

    PubMed

    Koshoridze, N; Kuchukashvili, Z; Menabde, K; Lekiashvili, Sh; Koshoridze, M

    2016-02-01

    Stress represents one of the main problems of modern humanity. This study was done for understanding more clearly alterations in creatine content of the brain under psycho-emotional stress induced by long-term social isolation. It was shown that under 30 days social isolation creatine amount in the brain was arisen, while decreasing concentrations of synthesizing enzymes (AGAT, GAMT) and creatine transporter protein (CrT). Another important point was that such changes were accompanied by down-regulation of creatine kinase (CK), therefore the enzyme's concentration was lowered. In addition, it was observed that content of phosphocreatine (PCr) and ATP were also reduced, thus indicating down-regulation of energy metabolism of brain that is really a crucial point for its normal functioning. To sum up the results it can be underlined that long-term social isolation has negative influence on energy metabolism of brain; and as a result reduce ATP content, while increase of free creatine concentration, supposedly maintaining maximal balance for ATP amount, but here must be also noted that up-regulated oxidative pathways might have impact on blood brain barrier, resulting on its permeability.

  17. [Analysis of creatine kinase variation in some members of the family Acipenseridae].

    PubMed

    Kuz'min, E V

    2008-04-01

    Creatine kinase (E.C. 2.7.3.2) was examined in stellate sturgeon Acipenser stellatus Pallas, Russian sturgeon A. gueldenstaedtii Brandt, European sterlet A. ruthenus L., Siberian sterlet A. ruthenus marsiglii Brandt, and great sturgeon (beluga) Huso huso L., using polyacrylamide gel electrophoresis. Two loci for creatine kinase were identified: CK-A* in white skeletal muscle and CK-C* in stomach wall muscle. Most species proved to be monomorphic at the CK-A* locus, showing the same phenotype represented by a single band. Heterogeneity and polymorphism in creatine kinase, determined by the CK-A* locus, were found only in Russian sturgeon. Based on the results of densitometric analysis of band staining intensity, we have advanced a hypothesis that synthesis of subunits of the CK-A* product in this species was controlled by eight genes. However, the genotype frequencies in the sample were significantly different from those theoretically expected upon free and independent gene recombination. The results of this study support the hypothesis on the absence of heterodimeric creatine kinase molecules in the skeletal muscle of Russian sturgeon. Locus CK-C* in sterlet was revealed as a single, intensely stained, rapidly migrating fraction, whereas in Russian sturgeon, the enzyme activity in this zone was very weak. No creatine kinase was found in liver, kidneys, spleen, heart, and intestine mucous tunic.

  18. Degradation of glycine and alanine on irradiated quartz.

    PubMed

    Pawlikowski, Maciej; Benko, Aleksandra; Wróbel, Tomasz P

    2013-04-01

    Recent researches suggest participation of minerals in the formation of life under primordial conditions. Among all of the minerals, quartz seems to be one of the most probable to take part in such processes. However, an external source of energy is needed, e.g. electric discharge. A device simulating the proposed conditions was designed and was used to simulate prebiotic conditions. Investigation of processes occurring during the stimulation of quartz with electric discharge was studied by means of Ultraviolet-visible (UV-VIS) spectroscopy, in order to monitor the generation kinetics of free radicals. Additionally, infrared spectroscopy was applied to identify chemical reaction products created in a solution of alanine or glycine, in the presence of quartz treated with electric discharge. Formation of increased amounts of free radicals, compared to experiments performed without quartz and/or amino acid, is reported, along with identification of possible degradation products of alanine. No synthetic reactions were observed.

  19. Clinical applications of alanine/electron spin resonance dosimetry.

    PubMed

    Baffa, Oswaldo; Kinoshita, Angela

    2014-05-01

    This paper discusses the clinical applications of electron spin resonance (ESR) dosimetry focusing on the ESR/alanine system. A review of few past studies in this area is presented offering a critical overview of the challenges and opportunities for extending this system into clinical applications. Alanine/ESR dosimetry fulfills many of the required properties for several clinical applications such as water-equivalent composition, independence of the sensitivity for the energy range used in therapy and high precision. Improvements in sensitivity and the development of minidosimeters coupled with the use of a spectrometer of higher microwave frequency expanded the possibilities for clinical applications to the new modalities of radiotherapy (intensity-modulated radiation therapy and radiosurgery) and to the detection of low doses such as those present in some radiological image procedures.

  20. Physiological hypercortisolemia increases proteolysis, glutamine, and alanine production

    SciTech Connect

    Darmaun, D.; Matthews, D.E.; Bier, D.M. Cornell Univ. Medical College, New York, NY )

    1988-09-01

    Physiological elevations of plasma cortisol levels, as are encountered in stress and severe trauma, were produced in six normal subjects by infusing them with hydrocortisone for 64 h. Amino acid kinetics were measured in the postabsorptive state using three 4-h infusions of L-(1-{sup 13}C)leucine, L-phenyl({sup 2}H{sub 5})phenylalanine, L-(2-{sup 15}N)glutamine, and L-(1-{sup 13}C)alanine tracers (1) before, (2) at 12 h, and (3) at 60 h of cortisol infusion. Before and throughout the study, the subjects ate a normal diet of adequate protein and energy intake. The cortisol infusion raised plasma cortisol levels significantly from 10 {plus minus} 1 to 32 {plus minus} 4 {mu}g/dl, leucine flux from 83 {plus minus} 3 to 97 {plus minus} 3 {mu}mol{center dot}kg{sup {minus}1}{center dot}h{sup {minus}1}, and phenylalanine flux from 34 {plus minus} 1 to 39 {plus minus} 1 (SE) {mu}mol{center dot}kg{sup {minus}1}{center dot}h{sup {minus}1} after 12 h of cortisol infusion. These increases were maintained until the cortisol infusion was terminated. These nearly identical 15% increases in two different essential amino acid appearance rates are reflective of increased whole body protein breakdown. Glutamine flux rose by 12 h of cortisol infusion and remained elevated at the same level at 64 h. The increase in flux was primarily due to a 55% increase in glutamine de novo synthesis. Alanine flux increased with acute hypercortisolemia and increased further at 60 h of cortisol infusion, a result primarily of increased alanine de novo synthesis. Insulin, alanine, and lactate plasma levels responded similarly with significant rises between the acute and chronic periods of cortisol infusion. Thus hypercortisolemia increases both protein breakdown and the turnover of important nonessential amino acids for periods of up to 64 h.

  1. NcoI and TaqI RFLPs for human M creatine kinase (CKM)

    SciTech Connect

    Perryman, M.B.; Hejtmancik, J.F.; Ashizawa, Tetsuo; Armstrong, R.; Lin, Sunchiang; Roberts, R.; Epstein, H.F. )

    1988-09-12

    Probe pHMCKUT contains a 135 bp cDNA fragment inserted into pGEM 3. The probe corresponds to nucleotides 1,201 to 1,336 located in the 3{prime} untranslated region of human M creatine kinase. The probe is specific for human M creatine kinase and does not hybridize to human B cretine kinase sequences. NcoI identifies a two allele polymorphism of a band at either 2.5 kb or 3.6 kb. TaqI identifies a two allele polymorphism at either 3.8 kb or 4.5 kb. Human M creatine has been localized to chromosome 19q. Autosomal co-dominant inheritance was shown in six informative Caucasian families.

  2. In sickness and in health: the widespread application of creatine supplementation.

    PubMed

    Gualano, Bruno; Roschel, Hamilton; Lancha-Jr, Antonio Herbert; Brightbill, Charles E; Rawson, Eric S

    2012-08-01

    There is an extensive and still growing body of the literature supporting the efficacy of creatine (Cr) supplementation. In sports, creatine has been recognized as the most effective nutritional supplement in enhancing exercise tolerance, muscle strength and lean body mass. From a clinical perspective, the application of Cr supplementation is indeed exciting. Evidences of benefits from this supplement have been reported in a broad range of diseases, including myopathies, neurodegenerative disorders, cancer, rheumatic diseases, and type 2 diabetes. In addition, after hundreds of published studies and millions of exposures creatine supplementation maintains an excellent safety profile. Thus, we contend that the widespread application of this supplement may benefit athletes, elderly people and various patient populations. In this narrative review, we aimed to summarize both the ergogenic and therapeutic effects of Cr supplementation. Furthermore, we reviewed the impact of Cr supplementation on kidney function.

  3. Insights into the Phosphoryl Transfer Mechanism of Human Ubiquitous Mitochondrial Creatine Kinase.

    PubMed

    Li, Quanjie; Fan, Shuai; Li, Xiaoyu; Jin, Yuanyuan; He, Weiqing; Zhou, Jinming; Cen, Shan; Yang, ZhaoYong

    2016-12-02

    Human ubiquitous mitochondrial creatine kinase (uMtCK) is responsible for the regulation of cellular energy metabolism. To investigate the phosphoryl-transfer mechanism catalyzed by human uMtCK, in this work, molecular dynamic simulations of uMtCK∙ATP-Mg(2+)∙creatine complex and quantum mechanism calculations were performed to make clear the puzzle. The theoretical studies hereof revealed that human uMtCK utilizes a two-step dissociative mechanism, in which the E227 residue of uMtCK acts as the catalytic base to accept the creatine guanidinium proton. This catalytic role of E227 was further confirmed by our assay on the phosphatase activity. Moreover, the roles of active site residues in phosphoryl transfer reaction were also identified by site directed mutagenesis. This study reveals the structural basis of biochemical activity of uMtCK and gets insights into its phosphoryl transfer mechanism.

  4. Insights into the Phosphoryl Transfer Mechanism of Human Ubiquitous Mitochondrial Creatine Kinase

    PubMed Central

    Li, Quanjie; Fan, Shuai; Li, Xiaoyu; Jin, Yuanyuan; He, Weiqing; Zhou, Jinming; Cen, Shan; Yang, ZhaoYong

    2016-01-01

    Human ubiquitous mitochondrial creatine kinase (uMtCK) is responsible for the regulation of cellular energy metabolism. To investigate the phosphoryl-transfer mechanism catalyzed by human uMtCK, in this work, molecular dynamic simulations of uMtCK∙ATP-Mg2+∙creatine complex and quantum mechanism calculations were performed to make clear the puzzle. The theoretical studies hereof revealed that human uMtCK utilizes a two-step dissociative mechanism, in which the E227 residue of uMtCK acts as the catalytic base to accept the creatine guanidinium proton. This catalytic role of E227 was further confirmed by our assay on the phosphatase activity. Moreover, the roles of active site residues in phosphoryl transfer reaction were also identified by site directed mutagenesis. This study reveals the structural basis of biochemical activity of uMtCK and gets insights into its phosphoryl transfer mechanism. PMID:27909311

  5. Guanidinoacetate methyltransferase deficiency: the first inborn error of creatine metabolism in man.

    PubMed Central

    Stöckler, S.; Isbrandt, D.; Hanefeld, F.; Schmidt, B.; von Figura, K.

    1996-01-01

    In two children with an accumulation of guanidinoacetate in brain and a deficiency of creatine in blood, a severe deficiency of guanidinoacetate methyltransferase (GAMT) activity was detected in the liver. Two mutant GAMT alleles were identified that carried a single base substitution within a 5' splice site or a 13-nt insertion and gave rise to four mutant transcripts. Three of the transcripts encode truncated polypeptides that lack a residue known to be critical for catalytic activity of GAMT. Deficiency of GAMT is the first inborn error of creatine metabolism. It causes a severe developmental delay and extrapyramidal symptoms in early infancy and is treatable by oral substitution with creatine. Images Figure 2 PMID:8651275

  6. Selected Cytokines Serve as Potential Biomarkers for Predicting Liver Inflammation and Fibrosis in Chronic Hepatitis B Patients With Normal to Mildly Elevated Aminotransferases.

    PubMed

    Deng, Yong-Qiong; Zhao, Hong; Ma, An-Lin; Zhou, Ji-Yuan; Xie, Shi-Bin; Zhang, Xu-Qing; Zhang, Da-Zhi; Xie, Qing; Zhang, Guo; Shang, Jia; Cheng, Jun; Zhao, Wei-Feng; Zou, Zhi-Qiang; Zhang, Ming-Xiang; Wang, Gui-Qiang

    2015-11-01

    Previous studies of small cohorts have implicated several circulating cytokines with progression of chronic hepatitis B (CHB). However, to date there have been no reliable biomarkers for assessing histological liver damage in CHB patients with normal or mildly elevated alanine aminotransferase (ALT). The aim of the present study was to investigate the association between circulating cytokines and histological liver damage in a large cohort. Also, this study was designed to assess the utility of circulating cytokines in diagnosing liver inflammation and fibrosis in CHB patients with ALT less than 2 times the upper limit of normal range (ULN). A total of 227 CHB patients were prospectively enrolled. All patients underwent liver biopsy and staging by Ishak system. Patients with at least moderate inflammation showed significantly higher levels of CXCL-11, CXCL-10, and interleukin (IL)-2 receptor (R) than patients with less than moderate inflammation (P < 0.001). Patients with significant fibrosis had higher levels of IL-8 (P = 0.027), transforming growth factor alpha (TGF-α) (P = 0.011), IL-2R (P = 0.002), and CXCL-11 (P = 0.032) than the group without significant fibrosis. In addition, 31.8% and 29.1% of 151 patients with ALT < 2 × ULN had at least moderate inflammation and significant fibrosis, respectively. Multivariate analysis demonstrated that CXCL-11 was independently associated with at least moderate inflammation, and TGF-α and IL-2R independently correlated with significant fibrosis in patients with ALT < 2 × ULN. Based on certain cytokines and clinical parameters, an inflammation-index and fib-index were developed, which showed areas under the receiver operating characteristics curve (AUROC) of 0.75 (95% CI 0.66-0.84) for at least moderate inflammation and 0.82 (95% CI 0.75-0.90) for significant fibrosis, correspondingly. Compared to existing scores, fib-index was significantly superior to aspartate aminotransferase

  7. Selected Cytokines Serve as Potential Biomarkers for Predicting Liver Inflammation and Fibrosis in Chronic Hepatitis B Patients With Normal to Mildly Elevated Aminotransferases

    PubMed Central

    Deng, Yong-Qiong; Zhao, Hong; Ma, An-Lin; Zhou, Ji-Yuan; Xie, Shi-Bin; Zhang, Xu-Qing; Zhang, Da-Zhi; Xie, Qing; Zhang, Guo; Shang, Jia; Cheng, Jun; Zhao, Wei-Feng; Zou, Zhi-Qiang; Zhang, Ming-Xiang; Wang, Gui-Qiang

    2015-01-01

    Abstract Previous studies of small cohorts have implicated several circulating cytokines with progression of chronic hepatitis B (CHB). However, to date there have been no reliable biomarkers for assessing histological liver damage in CHB patients with normal or mildly elevated alanine aminotransferase (ALT). The aim of the present study was to investigate the association between circulating cytokines and histological liver damage in a large cohort. Also, this study was designed to assess the utility of circulating cytokines in diagnosing liver inflammation and fibrosis in CHB patients with ALT less than 2 times the upper limit of normal range (ULN). A total of 227 CHB patients were prospectively enrolled. All patients underwent liver biopsy and staging by Ishak system. Patients with at least moderate inflammation showed significantly higher levels of CXCL-11, CXCL-10, and interleukin (IL)-2 receptor (R) than patients with less than moderate inflammation (P < 0.001). Patients with significant fibrosis had higher levels of IL-8 (P = 0.027), transforming growth factor alpha (TGF-α) (P = 0.011), IL-2R (P = 0.002), and CXCL-11 (P = 0.032) than the group without significant fibrosis. In addition, 31.8% and 29.1% of 151 patients with ALT < 2 × ULN had at least moderate inflammation and significant fibrosis, respectively. Multivariate analysis demonstrated that CXCL-11 was independently associated with at least moderate inflammation, and TGF-α and IL-2R independently correlated with significant fibrosis in patients with ALT < 2 × ULN. Based on certain cytokines and clinical parameters, an inflammation-index and fib-index were developed, which showed areas under the receiver operating characteristics curve (AUROC) of 0.75 (95% CI 0.66–0.84) for at least moderate inflammation and 0.82 (95% CI 0.75–0.90) for significant fibrosis, correspondingly. Compared to existing scores, fib-index was significantly superior to aspartate

  8. Cellular compartmentation of energy metabolism: creatine kinase microcompartments and recruitment of B-type creatine kinase to specific subcellular sites.

    PubMed

    Schlattner, Uwe; Klaus, Anna; Ramirez Rios, Sacnicte; Guzun, Rita; Kay, Laurence; Tokarska-Schlattner, Malgorzata

    2016-08-01

    There is an increasing body of evidence for local circuits of ATP generation and consumption that are largely independent of global cellular ATP levels. These are mostly based on the formation of multiprotein(-lipid) complexes and diffusion limitations existing in cells at different levels of organization, e.g., due to the viscosity of the cytosolic medium, macromolecular crowding, multiple and bulky intracellular structures, or controlled permeability across membranes. Enzymes generating ATP or GTP are found associated with ATPases and GTPases enabling the direct fueling of these energy-dependent processes, and thereby implying that it is the local and not the global concentration of high-energy metabolites that is functionally relevant. A paradigm for such microcompartmentation is creatine kinase (CK). Cytosolic and mitochondrial isoforms of CK constitute a well established energy buffering and shuttling system whose functions are very much based on local association of CK isoforms with ATP-providing and ATP-consuming processes. Here we review current knowledge on the subcellular localization and direct protein and lipid interactions of CK isoforms, in particular about cytosolic brain-type CK (BCK) much less is known compared to muscle-type CK (MCK). We further present novel data on BCK, based on three different experimental approaches: (1) co-purification experiments, suggesting association of BCK with membrane structures such as synaptic vesicles and mitochondria, involving hydrophobic and electrostatic interactions, respectively; (2) yeast-two-hybrid analysis using cytosolic split-protein assays and the identifying membrane proteins VAMP2, VAMP3 and JWA as putative BCK interaction partners; and (3) phosphorylation experiments, showing that the cellular energy sensor AMP-activated protein kinase (AMPK) is able to phosphorylate BCK at serine 6 to trigger BCK localization at the ER, in close vicinity of the highly energy-demanding Ca(2+) ATPase pump. Thus

  9. Pancreatic stellate cells support tumour metabolism through autophagic alanine secretion.

    PubMed

    Sousa, Cristovão M; Biancur, Douglas E; Wang, Xiaoxu; Halbrook, Christopher J; Sherman, Mara H; Zhang, Li; Kremer, Daniel; Hwang, Rosa F; Witkiewicz, Agnes K; Ying, Haoqiang; Asara, John M; Evans, Ronald M; Cantley, Lewis C; Lyssiotis, Costas A; Kimmelman, Alec C

    2016-08-25

    Pancreatic ductal adenocarcinoma (PDAC) is an aggressive disease characterized by an intense fibrotic stromal response and deregulated metabolism. The role of the stroma in PDAC biology is complex and it has been shown to play critical roles that differ depending on the biological context. The stromal reaction also impairs the vasculature, leading to a highly hypoxic, nutrient-poor environment. As such, these tumours must alter how they capture and use nutrients to support their metabolic needs. Here we show that stroma-associated pancreatic stellate cells (PSCs) are critical for PDAC metabolism through the secretion of non-essential amino acids (NEAA). Specifically, we uncover a previously undescribed role for alanine, which outcompetes glucose and glutamine-derived carbon in PDAC to fuel the tricarboxylic acid (TCA) cycle, and thus NEAA and lipid biosynthesis. This shift in fuel source decreases the tumour’s dependence on glucose and serum-derived nutrients, which are limited in the pancreatic tumour microenvironment. Moreover, we demonstrate that alanine secretion by PSCs is dependent on PSC autophagy, a process that is stimulated by cancer cells. Thus, our results demonstrate a novel metabolic interaction between PSCs and cancer cells, in which PSC-derived alanine acts as an alternative carbon source. This finding highlights a previously unappreciated metabolic network within pancreatic tumours in which diverse fuel sources are used to promote growth in an austere tumour microenvironment.

  10. Characterization of psychrophilic alanine racemase from Bacillus psychrosaccharolyticus.

    PubMed

    Okubo, Y; Yokoigawa, K; Esaki, N; Soda, K; Kawai, H

    1999-03-16

    A psychrophilic alanine racemase gene from Bacillus psychrosaccharolyticus was cloned and expressed in Escherichia coli SOLR with a plasmid pYOK3. The gene starting with the unusual initiation codon GTG showed higher preference for codons ending in A or T. The enzyme purified to homogeneity showed the high catalytic activity even at 0 degrees C and was extremely labile over 35 degrees C. The enzyme was found to have a markedly large Km value (5.0 microM) for the pyridoxal 5'-phosphate (PLP) cofactor in comparison with other reported alanine racemases, and was stabilized up to 50 degrees C in the presence of excess amounts of PLP. The low affinity of the enzyme for PLP may be related to the thermolability, and may be related to the high catalytic activity, initiated by the transaldimination reaction, at low temperature. The enzyme has a distinguishing hydrophilic region around the residue no. 150 in the deduced amino acid sequence (383 residues), whereas the corresponding regions of other Bacillus alanine racemases are hydrophobic. The position of the region in the three dimensional structure of C atoms of the enzyme was predicted to be in a surface loop surrounding the active site. The region may interact with solvent and reduce the compactness of the active site.

  11. Probing alanine transaminase catalysis with hyperpolarized 13CD3-pyruvate

    PubMed Central

    Barb, A.W.; Hekmatyar, S.K.; Glushka, J.N.; Prestegard, J.H.

    2013-01-01

    Hyperpolarized metabolites offer a tremendous sensitivity advantage (>104 fold) when measuring flux and enzyme activity in living tissues by magnetic resonance methods. These sensitivity gains can also be applied to mechanistic studies that impose time and metabolite concentration limitations. Here we explore the use of hyperpolarization by dissolution dynamic nuclear polarization (DNP) in mechanistic studies of alanine transaminase (ALT), a well-established biomarker of liver disease and cancer that converts pyruvate to alanine using glutamate as a nitrogen donor. A specific deuterated, 13C-enriched analog of pyruvic acid, 13C3D3-pyruvic acid, is demonstrated to have advantages in terms of detection by both direct 13C observation and indirect observation through methyl protons introduced by ALT-catalyzed H–D exchange. Exchange on injecting hyperpolarized 13C3D3-pyruvate into ALT dissolved in buffered 1H2O, combined with an experimental approach to measure proton incorporation, provided information on mechanistic details of transaminase action on a 1.5 s timescale. ALT introduced, on average, 0.8 new protons into the methyl group of the alanine produced, indicating the presence of an off-pathway enamine intermediate. The opportunities for exploiting mechanism-dependent molecular signatures as well as indirect detection of hyperpolarized 13C3-pyruvate and products in imaging applications are discussed. PMID:23357427

  12. Pressure-induced phase transitions in L-alanine, revisited.

    PubMed

    Tumanov, N A; Boldyreva, E V; Kolesov, B A; Kurnosov, A V; Quesada Cabrera, R

    2010-08-01

    The effect of pressure on L-alanine has been studied by X-ray powder diffraction (up to 12.3 GPa), single-crystal X-ray diffraction, Raman spectroscopy and optical microscopy (up to approximately 6 GPa). No structural phase transitions have been observed. At approximately 2 GPa the cell parameters a and b become accidentally equal to each other, but without a change in space-group symmetry. Neither of two transitions reported by others (to a tetragonal phase at approximately 2 GPa and to a monoclinic phase at approximately 9 GPa) was observed. The changes in cell parameters were continuous up to the highest measured pressures and the cells remained orthorhombic. Some important changes in the intermolecular interactions occur, which also manifest themselves in the Raman spectra. Two new orthorhombic phases could be crystallized from a MeOH/EtOH/H(2)O pressure-transmitting mixture in the pressure range 0.8-4.7 GPa, but only if the sample was kept at these pressures for at least 1-2 d. The new phases converted back to L-alanine on decompression. Judging from the Raman spectra and cell parameters, the new phases are most probably not L-alanine but its solvates.

  13. The effects of polyethylene glycosylated creatine supplementation on anaerobic performance measures and body composition.

    PubMed

    Camic, Clayton L; Housh, Terry J; Zuniga, Jorge M; Traylor, Daniel A; Bergstrom, Haley C; Schmidt, Richard J; Johnson, Glen O; Housh, Dona J

    2014-03-01

    The purpose of this study was to examine the effects of 28 days of polyethylene glycosylated creatine (PEG-creatine) supplementation (1.25 and 2.50 g·d) on anaerobic performance measures (vertical and broad jumps, 40-yard dash, 20-yard shuttle run, and 3-cone drill), upper- and lower-body muscular strength and endurance (bench press and leg extension), and body composition. This study used a randomized, double-blind, placebo-controlled parallel design. Seventy-seven adult men (mean age ± SD, 22.1 ± 2.5 years; body mass, 81.7 ± 10.8 kg) volunteered to participate and were randomly assigned to a placebo (n = 23), 1.25 g·d of PEG-creatine (n = 27), or 2.50 g·d of PEG-creatine (n = 27) group. The subjects performed anaerobic performance measures, muscular strength (one-repetition maximum [1RM]), and endurance (80% 1RM) tests for bench press and leg extension, and underwater weighing for the determination of body composition at day 0 (baseline), day 14, and day 28. The results indicated that there were improvements (p < 0.0167) in vertical jump, 20-yard shuttle run, 3-cone drill, muscular endurance for bench press, and body mass for at least one of the PEG-creatine groups without changes for the placebo group. Thus, the present results demonstrated that PEG-creatine supplementation at 1.25 or 2.50 g·d had an ergogenic effect on lower-body vertical power, agility, change-of-direction ability, upper-body muscular endurance, and body mass.

  14. Creatine and pyruvate prevent behavioral and oxidative stress alterations caused by hypertryptophanemia in rats.

    PubMed

    Andrade, Vivian Strassburger; Rojas, Denise Bertin; Oliveira, Lenise; Nunes, Mychely Lopes; de Castro, Fernanda Luz; Garcia, Cristina; Gemelli, Tanise; de Andrade, Rodrigo Binkowski; Wannmacher, Clóvis Milton Duval

    2012-03-01

    It is known that the accumulation of tryptophan and its metabolites is related to brain damage associated with both hypertryptophanemia and neurodegenerative diseases. In this study, we investigated the effect of tryptophan administration on various parameters of behavior in the open-field task and oxidative stress, and the effects of creatine and pyruvate, on the effect of tryptophan. Forty, 60-day-old male Wistar rats, were randomly divided into four groups: saline, tryptophan, pyruvate + creatine, tryptophan + pyruvate + creatine. Animals received three subcutaneous injections of tryptophan (2 μmol/g body weight each one at 3 h of intervals) and/or pyruvate (200 μg/g body weight 1 h before tryptophan), and/or creatine (400 μg/g body weight twice a day for 5 days before tryptophan twice a day for 5 days before training); controls received saline solution (NaCl 0.85%) at the same volumes (30 μl/g body weight) than the other substances. Results showed that tryptophan increased the activity of the animals, suggesting a reduction in the ability of habituation to the environment. Tryptophan induced increase of TBA-RS and total sulfhydryls. The effects of tryptophan in the open field, and in oxidative stress were fully prevented by the combination of creatine plus pyruvate. In case these findings also occur in humans affected by hypertryptophanemia or other neurodegenerative disease in which tryptophan accumulates, it is feasible that oxidative stress may be involved in the mechanisms leading to the brain injury, suggesting that creatine and pyruvate supplementation could benefit patients affected by these disorders.

  15. Umbrella sampling of proton transfer in a creatine-water system

    NASA Astrophysics Data System (ADS)

    Ivchenko, Olga; Bachert, Peter; Imhof, Petra

    2014-04-01

    Proton transfer reactions are among the most common processes in chemistry and biology. Proton transfer between creatine and surrounding solvent water is underlying the chemical exchange saturation transfer used as a contrast in magnetic resonance imaging. The free energy barrier, determined by first-principles umbrella sampling simulations (EaDFT 3 kcal/mol) is in the same order of magnitude as the experimentally obtained activation energy. The underlying mechanism is a first proton transfer from the guanidinium group to the water pool, followed by a second transition where a proton is "transferred back" from the nearest water molecule to the deprotonated nitrogen atom of creatine.

  16. Radiation inactivation method provides evidence that membrane-bound mitochondrial creatine kinase is an oligomer

    SciTech Connect

    Quemeneur, E.; Eichenberger, D.; Goldschmidt, D.; Vial, C.; Beauregard, G.; Potier, M.

    1988-06-30

    Lyophilized suspensions of rabbit heart mitochondria have been irradiated with varying doses of gamma rays. Mitochondrial creatine kinase activity was inactivated exponentially with a radiation inactivation size of 352 or 377 kDa depending upon the initial medium. These values are in good agreement with the molecular mass previously deduced from by permeation experiments: 357 kDa. This is the first direct evidence showing that the native form of mitochondrial creatine kinase is associated to the inner membrane as an oligomer, very likely an octamer.

  17. Raman spectroscopic approach to monitor the in vitro cyclization of creatine → creatinine

    NASA Astrophysics Data System (ADS)

    Gangopadhyay, Debraj; Sharma, Poornima; Singh, Sachin Kumar; Singh, Pushkar; Tarcea, Nicolae; Deckert, Volker; Popp, Jürgen; Singh, Ranjan K.

    2015-01-01

    The creatine → creatinine cyclization, an important metabolic phenomenon has been initiated in vitro at acidic pH and studied through Raman spectroscopic and DFT approach. The equilibrium composition of neutral, zwitterionic and protonated microspecies of creatine has been monitored with time as the reaction proceeds. Time series Raman spectra show clear signature of creatinine formation at pH 3 after ∼240 min at room temperature and reaction is faster at higher temperature. The spectra at pH 1 and pH 5 do not show such signature up to 270 min implying faster reaction rate at pH 3.

  18. A Pilot Study of Creatine as a Novel Treatment for Depression in Methamphetamine Using Females

    PubMed Central

    Hellem, Tracy L.; Sung, Young-Hoon; Shi, Xian-Feng; Pett, Marjorie A.; Latendresse, Gwen; Morgan, Jubel; Huber, Rebekah S.; Kuykendall, Danielle; Lundberg, Kelly J.; Renshaw, Perry F.

    2015-01-01

    Objective Depression among methamphetamine users is more prevalent in females than males, but gender specific treatment options for this comorbidity have not been described. Reduced brain phosphocreatine levels have been shown to be lower in female methamphetamine users compared to males, and, of relevance, studies have demonstrated an association between treatment resistant depression and reduced brain phosphocreatine concentrations. The nutritional supplement creatine monohydrate has been reported to reduce symptoms of depression in female adolescents and adults taking antidepressants, as well as to increase brain phosphocreatine in healthy volunteers. Therefore, the purpose of this pilot study was to investigate creatine monohydrate as a treatment for depression in female methamphetamine users. Methods Fourteen females with depression and comorbid methamphetamine dependence were enrolled in an 8 week open label trial of 5 grams of daily creatine monohydrate and of these 14, eleven females completed the study. Depression was measured using the Hamilton Depression Rating Scale (HAMD) and brain phosphocreatine levels were measured using phosphorus magnetic resonance spectroscopy pre- and post-creatine treatment. Secondary outcome measures included anxiety symptoms, measured with the Beck Anxiety Inventory (BAI), as well as methamphetamine use, monitored by twice weekly urine drug screens and self-reported use. Results The results of a linear mixed effects repeated measures model showed significantly reduced HAMD and BAI scores as early as week 2 when compared to baseline scores. This improvement was maintained through study completion. Brain phosphocreatine concentrations were higher at the second phosphorus magnetic resonance spectroscopy scan compared to the baseline scan; Mbaseline = 0.223 (SD = 0.013) vs. Mpost-treatment = 0.233 (SD = 0.009), t(9) = 2.905, p < .01, suggesting that creatine increased phosphocreatine levels. Also, a reduction in methamphetamine

  19. Isotopic effects in mechanistic studies of biotransformations of fluorine derivatives of L-alanine catalysed by L-alanine dehydrogenase.

    PubMed

    Szymańska-Majchrzak, Jolanta; Pałka, Katarzyna; Kańska, Marianna

    2017-05-01

    Synthesis of 3-fluoro-[2-(2)H]-L-alanine (3-F-[(2)H]-L-Ala) in reductive amination of 3-fluoropyruvic acid catalysed by L-alanine dehydrogenase (AlaDH) was described. Fluorine derivative was used to study oxidative deamination catalysed by AlaDH applied kinetic (for 3-F-L-Ala in H2O - KIE's on Vmax: 1.1; on Vmax/KM: 1.2; for 3-F-L-Ala in (2)H2O - on Vmax: 1.4; on Vmax/KM: 2.1) and solvent isotope effect methods (for 3-F-L-Ala - SIE's on Vmax: 1.0; on Vmax/KM: 0.87; for 3-F-[2-(2)H]-L-Ala - on Vmax: 1.4; on Vmax/KM: 1.5). Studies explain some details of reaction mechanism.

  20. Effects of two and five days of creatine loading on muscular strength and anaerobic power in trained athletes.

    PubMed

    Law, Yu Li Lydia; Ong, Wee Sian; GillianYap, Tsien Lin; Lim, Su Ching Joselin; Von Chia, Ee

    2009-05-01

    The purpose of this study was to establish the effects of 2 and 5 days of creatine loading, coupled with resistance training, on muscular strength and anaerobic performance in trained athletes. Seventeen trained men were randomly assigned to a creatine or a placebo group. The creatine supplementation group consumed 20 g of creatine per day (4 doses of 5 g per day), whereas the placebo group was given a placebo similar in appearance and taste over the 5-day supplementation duration. Anaerobic power and strength performance measures, in addition to blood and urine analysis, were conducted in the morning before the supplementation began and on the third and sixth day to establish the effect of 2 and 5 days of creatine loading, respectively. The study found that a 5-day creatine loading regime coupled with resistance training resulted in significant improvements in both average anaerobic power, as measured by the 30-second Wingate test and back squat strength compared with just training alone. However, 2 days of supplementation was not sufficient to produce similar performance gains as that observed at the end of 5 days of loading in trained men, despite increases in creatine uptake in the body. The standard 5-day loading regime should hence be prescribed to individuals supplementing with creatine for enhanced strength and power.

  1. 75 FR 17769 - In the Matter of Certain Products Advertised as Containing Creatine Ethyl Ester; Notice of...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-04-07

    ... COMMISSION In the Matter of Certain Products Advertised as Containing Creatine Ethyl Ester; Notice of Commission Issuance of a Limited Exclusion Order Against the Products Advertised as Containing Creatine Ethyl... sale within the United States after importation of certain products advertised as containing...

  2. VDAC electronics: 3. VDAC-Creatine kinase-dependent generation of the outer membrane potential in respiring mitochondria.

    PubMed

    Lemeshko, Victor V

    2016-07-01

    Mitochondrial energy in cardiac cells has been reported to be channeled into the cytosol through the intermembrane contact sites formed by the adenine nucleotide translocator, creatine kinase and VDAC. Computational analysis performed in this study showed a high probability of the outer membrane potential (OMP) generation coupled to such a mechanism of energy channeling in respiring mitochondria. OMPs, positive inside, calculated at elevated concentrations of creatine are high enough to restrict ATP release from mitochondria, to significantly decrease the apparent K(m,ADP) for state 3 respiration and to maintain low concentrations of Ca(2+) in the mitochondrial intermembrane space. An inhibition by creatine of Ca(2+)-induced swelling of isolated mitochondria and other protective effects of creatine reported in the literature might be explained by generated positive OMP. We suggest that VDAC-creatine kinase-dependent generation of OMP represents a novel physiological factor controlling metabolic state of mitochondria, cell energy channeling and resistance to death.

  3. A Protein That Binds Specifically to the M-Line of Skeletal Muscle Is Identified as the Muscle Form of Creatine Kinase

    PubMed Central

    Turner, David C.; Wallimann, Theo; Eppenberger, Hans M.

    1973-01-01

    Published information on the properties of two proteins from chicken muscle, creatine kinase (MM-creatine kinase) and an M-line protein, suggested that they might be identical molecules. Different published procedures were used to purify the two proteins to homogeneity, and the properties of the two preparations were compared. Creatine kinase specific activity increased during purification of M-line protein, reaching a value comparable to that of purified MM-creatine kinase. The two proteins migrated identically in two electrophoretic systems and, after electrophoresis, both could be stained for creatine kinase activity. Double immunodiffusion tests with antibody prepared against MM-creatine kinase established the serological identity of the two protein preparations. Immunofluorescent studies showed that antiserum against MM-creatine kinase was bound in a regular pattern at the centers of the A-band regions of isolated myofibrils. These data show conclusively that the M-line protein and MM-creatine kinase are identical. Images PMID:4197625

  4. Effect of creatine supplementation during the last week of gestation on birth intervals, stillbirth, and preweaning mortality in pigs.

    PubMed

    Vallet, J L; Miles, J R; Rempel, L A

    2013-05-01

    We hypothesized that creatine supplementation would reduce birth intervals, stillbirth rate, and preweaning survival in pigs because of its reported improvement of athletic performance in humans. In Exp. 1, gilts (n = 42) and first parity sows (n = 75) were mated at estrus. Beginning on d 110 of gestation, dams received either no treatment or 20 g creatine daily until farrowing. At farrowing in November 2008, pigs were monitored by video camera to determine individual piglet birth intervals. On d 1, piglets were weighed, euthanized, and the cerebellum, brain stem, and spinal cord were collected from the largest and smallest piglets in each litter to measure myelin basic proteins, myelin cholesterol, glucocerebrosides, phosphatidylethanolamine, phosphatidylcholine, and sphingomyelin. Preweaning mortality of the remaining piglets was recorded, including whether a piglet had been overlayed by the dam. A second experiment was performed using gilts (n = 90), farrowing in July 2010, to test differential effects of creatine supplementation during hot, humid weather when dams typically have more difficulty farrowing. Once again, gilts were provided either no supplementation or 20 g creatine daily from d 110 to the day of farrowing. Gilts were video recorded during farrowing, piglets were weighed on d 1, and preweaning mortality (including overlays) was recorded. In Exp. 1, creatine supplementation had no effect on birth intervals or stillbirth rate. Creatine supplementation improved the amount of myelin lipids in brain regions of piglets, particularly the brain stem. Creatine supplementation also reduced overlays of low birth weight piglets from gilts but not second parity sows. Data from Exp. 2 were combined with gilt data from Exp. 1 to examine the effect of creatine, season, and their interaction. There were no effects of treatment or season on birth intervals, stillbirth rates, or overall preweaning mortality. Creatine treatment reduced the incidence of overlays in low

  5. Thermal stability, pH dependence and inhibition of four murine kynurenine aminotransferases

    PubMed Central

    2010-01-01

    Background Kynurenine aminotransferase (KAT) catalyzes the transamination of kynunrenine to kynurenic acid (KYNA). KYNA is a neuroactive compound and functions as an antagonist of alpha7-nicotinic acetylcholine receptors and is the only known endogenous antagonist of N-methyl-D-aspartate receptors. Four KAT enzymes, KAT I/glutamine transaminase K/cysteine conjugate beta-lyase 1, KAT II/aminoadipate aminotransferase, KAT III/cysteine conjugate beta-lyase 2, and KAT IV/glutamic-oxaloacetic transaminase 2/mitochondrial aspartate aminotransferase, have been reported in mammalian brains. Because of the substrate overlap of the four KAT enzymes, it is difficult to assay the specific activity of each KAT in animal brains. Results This study concerns the functional expression and comparative characterization of KAT I, II, III, and IV from mice. At the applied test conditions, equimolar tryptophan with kynurenine significantly inhibited only mouse KAT I and IV, equimolar methionine inhibited only mouse KAT III and equimolar aspartate inhibited only mouse KAT IV. The activity of mouse KAT II was not significantly inhibited by any proteinogenic amino acids at equimolar concentrations. pH optima, temperature preferences of four KATs were also tested in this study. Midpoint temperatures of the protein melting, half life values at 65°C, and pKa values of mouse KAT I, II, III, and IV were 69.8, 65.9, 64.8 and 66.5°C; 69.7, 27.4, 3.9 and 6.5 min; pH 7.6, 5.7, 8.7 and 6.9, respectively. Conclusion The characteristics reported here could be used to develop specific assay methods for each of the four murine KATs. These specific assays could be used to identify which KAT is affected in mouse models for research and to develop small molecule drugs for prevention and treatment of KAT-involved human diseases. PMID:20482848

  6. Structures of aspartate aminotransferases from Trypanosoma brucei, Leishmania major and Giardia lamblia

    PubMed Central

    Abendroth, Jan; Choi, Ryan; Wall, Abigail; Clifton, Matthew C.; Lukacs, Christine M.; Staker, Bart L.; Van Voorhis, Wesley; Myler, Peter; Lorimer, Don D.; Edwards, Thomas E.

    2015-01-01

    The structures of three aspartate aminotransferases (AATs) from eukaryotic pathogens were solved within the Seattle Structural Genomics Center for Infectious Disease (SSGCID). Both the open and closed conformations of AAT were observed. Pyridoxal phosphate was bound to the active site via a Schiff base to a conserved lysine. An active-site mutant showed that Trypanosoma brucei AAT still binds pyridoxal phosphate even in the absence of the tethering lysine. The structures highlight the challenges for the structure-based design of inhibitors targeting the active site, while showing options for inhibitor design targeting the N-terminal arm. PMID:25945710

  7. Structures of aspartate aminotransferases from Trypanosoma brucei, Leishmania major and Giardia lamblia.

    PubMed

    Abendroth, Jan; Choi, Ryan; Wall, Abigail; Clifton, Matthew C; Lukacs, Christine M; Staker, Bart L; Van Voorhis, Wesley; Myler, Peter; Lorimer, Don D; Edwards, Thomas E

    2015-05-01

    The structures of three aspartate aminotransferases (AATs) from eukaryotic pathogens were solved within the Seattle Structural Genomics Center for Infectious Disease (SSGCID). Both the open and closed conformations of AAT were observed. Pyridoxal phosphate was bound to the active site via a Schiff base to a conserved lysine. An active-site mutant showed that Trypanosoma brucei AAT still binds pyridoxal phosphate even in the absence of the tethering lysine. The structures highlight the challenges for the structure-based design of inhibitors targeting the active site, while showing options for inhibitor design targeting the N-terminal arm.

  8. Creatine Kinase Activity Weakly Correlates to Volume Completed Following Upper Body Resistance Exercise

    ERIC Educational Resources Information Center

    Machado, Marco; Willardson, Jeffrey M.; Silva, Dailson P.; Frigulha, Italo C.; Koch, Alexander J.; Souza, Sergio C.

    2012-01-01

    In the current study, we examined the relationship between serum creatine kinase (CK) activity following upper body resistance exercise with a 1- or 3-min rest between sets. Twenty men performed two sessions, each consisting of four sets with a 10-repetition maximum load. The results demonstrated significantly greater volume for the 3-min…

  9. Automated urinalysis technique determines concentration of creatine and creatinine by colorimetry

    NASA Technical Reports Server (NTRS)

    Rho, J. H.

    1967-01-01

    Continuous urinalysis technique is useful in the study of muscle wastage in primates. Creatinine concentration in urine is determined in an aliquot mixture by a color reaction. Creatine is determined in a second aliquot by converting it to creatinine and measuring the difference in color intensity between the two aliquots.

  10. The Effects of Creatine Supplementation on Exercise-Induced Muscle Damage.

    ERIC Educational Resources Information Center

    Rawson, Eric S.; Gunn, Bridget; Clarkson, Priscilla M.

    2001-01-01

    Investigated the effects of oral creatine (Cr) supplementation on markers of exercise-induced muscle damage following high-force eccentric exercise in men randomly administered Cr or placebo. Results indicated that 5 days of Cr supplementation did not reduce indirect makers of muscle damage or enhance recovery from high-force eccentric exercise.…

  11. The effects of pre versus post workout supplementation of creatine monohydrate on body composition and strength

    PubMed Central

    2013-01-01

    Background Chronic supplementation with creatine monohydrate has been shown to promote increases in total intramuscular creatine, phosphocreatine, skeletal muscle mass, lean body mass and muscle fiber size. Furthermore, there is robust evidence that muscular strength and power will also increase after supplementing with creatine. However, it is not known if the timing of creatine supplementation will affect the adaptive response to exercise. Thus, the purpose of this investigation was to determine the difference between pre versus post exercise supplementation of creatine on measures of body composition and strength. Methods Nineteen healthy recreational male bodybuilders (mean ± SD; age: 23.1 ± 2.9; height: 166.0 ± 23.2 cm; weight: 80.18 ± 10.43 kg) participated in this study. Subjects were randomly assigned to one of the following groups: PRE-SUPP or POST-SUPP workout supplementation of creatine (5 grams). The PRE-SUPP group consumed 5 grams of creatine immediately before exercise. On the other hand, the POST-SUPP group consumed 5 grams immediately after exercise. Subjects trained on average five days per week for four weeks. Subjects consumed the supplement on the two non-training days at their convenience. Subjects performed a periodized, split-routine, bodybuilding workout five days per week (Chest-shoulders-triceps; Back-biceps, Legs, etc.). Body composition (Bod Pod®) and 1-RM bench press (BP) were determined. Diet logs were collected and analyzed (one random day per week; four total days analyzed). Results 2x2 ANOVA results - There was a significant time effect for fat-free mass (FFM) (F = 19.9; p = 0.001) and BP (F = 18.9; p < 0.001), however, fat mass (FM) and body weight did not reach significance. While there were trends, no significant interactions were found. However, using magnitude-based inference, supplementation with creatine post workout is possibly more beneficial in comparison to pre workout supplementation

  12. Changes of creatine kinase structure upon ligand binding as seen by small-angle scattering

    NASA Astrophysics Data System (ADS)

    Forstner, Michael; Kriechbaum, Manfred; Laggner, Peter; Wallimann, Theo

    1996-09-01

    Small-angle X-ray and neutron scattering have been used to investigate structural changes upon binding of individual substrates or a transition state analogue complex (TSAC), consisting of Mg-ADP, creatine and KNO 3 to creatine kinase isoenzymes (dimeric M-CK and octameric Mi-CK) and monomeric arginine kinase (AK). Considerable changes in the shape and the size of the molecules occurred upon binding of Mg-ATP and TSAC, whereas creatine alone had only a small effect. In Mi-CK, the radius of gyration was reduced from 55.6 Å (free enzyme) to 48.9 Å (enzyme + Mg-ATP) and to 48.2 Å (enzyme + TSAC). The experiments performed with M-CK showed similar changes from 28.0 Å (free enzyme) to 25.6 Å (enzyme + Mg-ATP) and to 25.5 Å (enzyme + TSAC). Creatine alone did not lead to significant changes in the radii of gyration, nor did free ATP or ADP. AK showed the same behaviour: a change of the radius of gyration from 21.5 Å (free enzyme) to 19.7 Å (enzyme + MG-ATP), whereas with arginine alone only a minor change could be observed. The primary change in structure as seen with monomeric AK seems to be a magnesium-nucleotide induced domain movement relative to each other, whereas the effect of substrate may be of local order only. In creatine kinase, however, further movements must be involved in the large conformational change.

  13. Creatine but not betaine supplementation increases muscle phosphorylcreatine content and strength performance.

    PubMed

    del Favero, Serena; Roschel, Hamilton; Artioli, Guilherme; Ugrinowitsch, Carlos; Tricoli, Valmor; Costa, André; Barroso, Renato; Negrelli, Ana Lua; Otaduy, Maria Concepción; da Costa Leite, Cláudia; Lancha-Junior, Antonio Herbert; Gualano, Bruno

    2012-06-01

    We aimed to investigate the role of betaine supplementation on muscle phosphorylcreatine (PCr) content and strength performance in untrained subjects. Additionally, we compared the ergogenic and physiological responses to betaine versus creatine supplementation. Finally, we also tested the possible additive effects of creatine and betaine supplementation. This was a double-blind, randomized, placebo-controlled study. Subjects were assigned to receive betaine (BET; 2 g/day), creatine (CR; 20 g/day), betaine plus creatine (BET+CR; 2+20 g/day, respectively) or placebo (PL). At baseline and after 10 days of supplementation, we assessed muscle strength and power, muscle PCr content, and body composition. The CR and BET+CR groups presented greater increase in muscle PCr content than PL (p=0.004 and p=0.006, respectively). PCr content was comparable between BET versus PL (p=0.78) and CR versus BET+CR (p=0.99). CR and BET+CR presented greater muscle power output than PL in the squat exercise following supplementation (p=0.003 and p=0.041, respectively). Similarly, bench press average power was significantly greater for the CR-supplemented groups. CR and BET+CR groups also showed significant pre- to post-test increase in 1-RM squat and bench press (CR: p=0.027 and p<0.0001; BET+CR: p=0.03 and p<0.0001 for upper- and lower-body assessments, respectively) No significant differences for 1-RM strength and power were observed between BET versus PL and CR versus BET+CR. Body composition did not differ between the groups. In conclusion, we reported that betaine supplementation does not augment muscle PCr content. Furthermore, we showed that betaine supplementation combined or not with creatine supplementation does not affect strength and power performance in untrained subjects.

  14. Programmed cell death genes are linked to elevated creatine kinase levels in unhealthy male nonagenarians

    PubMed Central

    Kim, Sangkyu; Simon, Eric; Myers, Leann; Hamm, L. Lee; Jazwinski, S. Michal

    2016-01-01

    Declining health in the oldest-old takes an energy toll for simple maintenance of body functions. The underlying mechanisms, however, differ in males and females. In females, the declines are explained by loss of muscle mass, but this is not the case in males in whom they are associated with increased levels of circulating creatine kinase. This relationship raises the possibility that muscle damage rather than muscle loss is the cause of the increased energy demands of unhealthy aging in males. We have now examined factors that contribute to the increase in creatine kinase. Much of it (60%) can be explained by a history of cardiac problems and lower kidney function, while being mitigated by moderate physical activity, reinforcing the notion that tissue damage is a likely source. In a search for genetic risk factors associated with elevated creatine kinase, the Ku70 gene XRCC6 and the ceramide synthase gene LASS1 were investigated because of their roles in telomere length and longevity and healthy aging, respectively. Single-nucleotide polymorphisms in these two genes were independently associated with creatine kinase levels. The XRCC6 variant was epistatic to one of the LASS1 variants but not to the other. These gene variants have potential regulatory activity. Ku70 is an inhibitor of the pro-apoptotic Bax, while the product of Lass1, ceramide, operates in both caspase-dependent and independent pathways of programmed cell death, providing a potential cellular mechanism for the effects of these genes on tissue damage and circulating creatine kinase. PMID:26913518

  15. Effect of free creatine therapy on cisplatin-induced renal damage.

    PubMed

    Genc, Gurkan; Okuyucu, Ali; Meydan, Bilge Can; Yavuz, Oguzhan; Nisbet, Ozlem; Hokelek, Murat; Bedir, Abdulkerim; Ozkaya, Ozan

    2014-08-01

    Abstract Cisplatin is one of the commonly used anticancer drugs and nephrotoxicity limits its use. The aim of this study is to investigate the possible protective effect of creatine supplementation on cisplatin-induced nephrotoxicity. Sixty male Sprague-Dawley rats were divided into three groups: Group I: Cisplatin (n=20) (7 mg/kg cisplatin intraperitoneal (i.p.) single dose), group II: Cisplatin+creatine monohydrate (n=20) (7 mg/kg cisplatin i.p. single dose and 300 mg/kg creatine p.o. daily for 30 days starting on first day of cisplatin injection), group III: Control group (n=20) (Serum physiologic, 2.5 mL/kg i.p.). Sacrifications were performed at first week and 30th day. Blood urea nitrogen (BUN) and serum creatinine levels, histopathological evaluation, mitochondrial deoxyribonucleic acid (mtDNA) common deletion rates, and body weights of rats were evaluated. A significant decrease in body weight, higher values of kidney function tests, histopathological scores, and mtDNA deletion ratios were observed in group I compared to control group at days 7 and 30 (p<0.05). In group II, there was a slight decrease in body weight at same days (p=0.931 and 0.084, respectively). Kidney function tests, histopathological scores, and mtDNA common deletion ratios were statistically better in group II than group I at 7th and 30th day (p<0.05). Although creatine significantly reversed kidney functions and pathological findings, this improvement was not sufficient to reach normal control group's results at days 7 and 30. In conclusion, the present study demonstrates that creatine administration is a promising adjuvant protective drug for reducing nephrotoxic effect of cisplatin.

  16. Evaluation of Intravascular Hemolysis With Erythrocyte Creatine in Patients With Aortic Stenosis.

    PubMed

    Sugiura, Tetsuro; Okumiya, Toshika; Kubo, Toru; Takeuchi, Hiroaki; Matsumura, Yoshihisa

    2016-07-27

    Chronic intravascular hemolysis has been identified in patients with cardiac valve prostheses, but only a few case reports have evaluated intravascular hemolysis in patients with native valvular heart disease. To detect intravascular hemolysis in patients with aortic stenosis, erythrocyte creatine was evaluated with hemodynamic indices obtained by echocardiography.Erythrocyte creatine, a marker of erythrocyte age, was assayed in 30 patients with aortic stenosis and 10 aged matched healthy volunteers. Peak flow velocity of the aortic valve was determined by continuous-wave Doppler echocardiography. Twenty of 30 patients with aortic stenosis had high erythrocyte creatine levels (> 1.8 µmol/g Hb) and erythrocyte creatine was significantly higher as compared with control subjects (1.98 ± 0.49 versus 1.52 ± 0.19 µmol/g Hb, P = 0.007). Peak transvalvular pressure gradient ranged from 46 to 142 mmHg and peak flow velocity ranged from 3.40 to 5.95 m/second. Patients with aortic stenosis had a significantly lower erythrocyte count (387 ± 40 versus 436 ± 42 × 10(4) µL, P = 0.002) and hemoglobin (119 ± 11 versus 135 ± 11 g/L, P < 0.001) as compared with control subjects. Erythrocyte creatine had a fair correlation with peak flow velocity (r = 0.55, P = 0.002).In conclusion, intravascular hemolysis due to destruction of erythrocytes was detected in patients with moderate to severe aortic stenosis and the severity of intravascular hemolysis was related to valvular flow velocity of the aortic valve.

  17. PRECREST: A phase II prevention and biomarker trial of creatine in at-risk Huntington disease

    PubMed Central

    Doros, Gheorghe; Gevorkian, Sona; Malarick, Keith; Reuter, Martin; Coutu, Jean-Philippe; Triggs, Tyler D.; Wilkens, Paul J.; Matson, Wayne; Salat, David H.; Hersch, Steven M.

    2014-01-01

    Objective: To assess the safety and tolerability of high-dose creatine, the feasibility of enrolling premanifest and 50% at-risk subjects in a prevention trial, and the potential of cognitive, imaging, and blood markers. Methods: Sixty-four eligible consenting participants were randomly allocated (1:1) to 15 g twice daily of creatine monohydrate or placebo for a 6-month double-blind phase followed by a 12-month open-label extension. Subjects included premanifest (tested) and at-risk (not tested) individuals without clinical symptoms or signs of Huntington disease (HD). Primary outcomes were safety and tolerability. Exploratory endpoints included fine motor, visuospatial, and memory performance; structural and diffusion MRI; and selected blood markers. Results: Forty-seven HD carriers and 17 non-HD controls were enrolled. Fifteen discontinued treatment (2 assigned to placebo); all were followed for the entire study period. Primary analysis was by intent to treat. The most common adverse events were gastrointestinal. Neuroimaging demonstrated treatment-related slowing of cortical and striatal atrophy at 6 and 18 months. Conclusion: We describe a design that preserves the autonomy of subjects not wanting genetic testing while including controls for assessing the specificity of treatment effects. Our results demonstrate the feasibility of prevention trials for HD and the safety of high-dose creatine, provide possible evidence of disease modification, support future studies of creatine, and illustrate the value of prodromal biomarkers. Classification of evidence: This study provides Class I evidence that high-dose creatine is safe and tolerable. PMID:24510496

  18. Role of creatine supplementation on exercise-induced cardiovascular function and oxidative stress

    PubMed Central

    Cunningham, Daniel; Mason, Laura; Kilduff, Liam P; McEneny, Jane

    2009-01-01

    Many degenerative diseases are associated with increased oxidative stress. Creatine has the potential to act as an indirect and direct antioxidant; however, limited data exist to evaluate the antioxidant capabilities of creatine supplementation within in vivo human systems. This study aimed to investigate the effects of oral creatine supplementation on markers of oxidative stress and antioxidant defenses following exhaustive cycling exercise. Following preliminary testing and two additional familiarization sessions, 18 active males repeated two exhaustive incremental cycling trials (T1 and T2) separated by exactly 7 days. The subjects were assigned, in a double-blind manner, to receive either 20 g of creatine (Cr) or a placebo (P) for the 5 days preceding T2. Breath-by-breath respiratory data and heart rate were continually recorded throughout the exercise protocol and blood samples were obtained at rest (preexercise), at the end of exercise (postexercise), and the day following exercise (post24 h). Serum hypdroperoxide concentrations were elevated at postexercise by 17 ± 5% above preexercise values (p = 0.030). However, supplementation did not influence lipid peroxidation (serum hypdroperoxide concentrations), resistance of low density lipoprotein to oxidative stress (t1/2max LDL oxidation) and plasma concentrations of non-enzymatic antioxidants (retinol, α-carotene, β-carotene, α-tocopherol, γ-tocopherol, lycopene and vitamin C). Heart rate and oxygen uptake responses to exercise were not affected by supplementation. These findings suggest that short-term creatine supplementation does not enhance non-enzymatic antioxidant defence or protect against lipid peroxidation induced by exhaustive cycling in healthy males. PMID:20716911

  19. Effects of combined creatine and sodium bicarbonate supplementation on repeated sprint performance in trained men.

    PubMed

    Barber, James J; McDermott, Ann Y; McGaughey, Karen J; Olmstead, Jennifer D; Hagobian, Todd A

    2013-01-01

    Creatine and sodium bicarbonate supplementation independently increase exercise performance, but it remains unclear whether combining these 2 supplements is more beneficial on exercise performance. The purpose of this study was to evaluate the impact of combining creatine monohydrate and sodium bicarbonate supplementation on exercise performance. Thirteen healthy, trained men (21.1 ± 0.6 years, 23.5 ± 0.5 kg·m(-2), 66.7 ± 5.7 ml·(kg·m)(-1) completed 3 conditions in a double-blinded, crossover fashion: (a) Placebo (Pl; 20 g maltodextrin + 0.5 g·kg(-1) maltodextrin), (b) Creatine (Cr; 20 g + 0.5 g·kg(-1) maltodextrin), and (c) Creatine plus sodium bicarbonate (Cr + Sb; 20 g + 0.5 g·kg(-1) sodium bicarbonate). Each condition consisted of supplementation for 2 days followed by a 3-week washout. Peak power, mean power, relative peak power, and bicarbonate concentrations were assessed during six 10-second repeated Wingate sprint tests on a cycle ergometer with a 60-second rest period between each sprint. Compared with Pl, relative peak power was significantly higher in Cr (4%) and Cr + Sb (7%). Relative peak power was significantly lower in sprints 4-6, compared with that in sprint 1, in both Pl and Cr. However, in Cr + Sb, sprint 6 was the only sprint significantly lower compared with sprint 1. Pre-Wingate bicarbonate concentrations were significantly higher in Cr + Sb (10%), compared with in Pl and Cr, and mean concentrations remained higher after sprint 6, although not significantly. Combining creatine and sodium bicarbonate supplementation increased peak and mean power and had the greatest attenuation of decline in relative peak power over the 6 repeated sprints. These data suggest that combining these 2 supplements may be advantageous for athletes participating in high-intensity, intermittent exercise.

  20. β-N-methylamino-l-alanine causes neurological and pathological phenotypes mimicking Amyotrophic Lateral Sclerosis (ALS): the first step towards an experimental model for sporadic ALS.

    PubMed

    de Munck, Estefanía; Muñoz-Sáez, Emma; Miguel, Begoña G; Solas, M Teresa; Ojeda, Irene; Martínez, Ana; Gil, Carmen; Arahuetes, Rosa Ma

    2013-09-01

    β-N-methylamino-l-alanine (L-BMAA) is a neurotoxic amino acid that has been related to various neurodegenerative diseases. The aim of this work was to analyze the biotoxicity produced by L-BMAA in vivo in rats, trying to elucidate its physiopathological mechanisms and to search for analogies between the found effects and pathologies like Amyotrophic Lateral Sclerosis (ALS). Our data demonstrated that the neurotoxic effects in vivo were dosage-dependent. For evaluating the state of the animals, a neurological evaluation scale was developed as well as a set of functional tests. Ultrastructural cell analysis of spinal motoneurons has revealed alterations both in endoplasmic reticulum and mitochondria. Since GSK3β could play a role in some neuropathological processes, we analyzed the alterations occurring in GSK3β levels in L-BMAA treated rats, we have observed an increase in the active form of GSK3β levels in lumbar spinal cord and motor cerebral cortex. On the other hand, (TAR)-DNA-binding protein 43 (TDP-43) increased in L-BMAA treated animals. Our results indicated that N-acetylaspartate (NAA) declined in animals treated with L-BMAA, and the ratio of N-acetylaspartate/choline (NAA/Cho), N-acetylaspartate/creatine (NAA/Cr) and N-acetylaspartate/choline+creatine (NAA/Cho+Cr) tended to decrease in lumbar spinal cord and motor cortex. This project offers some encouraging results that could help establishing the progress in the development of an animal model of sporadic ALS and L-BMAA could be a useful tool for this purpose.

  1. The anomalous kinetics of coupled aspartate aminotransferase and malate dehydrogenase. Evidence for compartmentation of oxaloacetate.

    PubMed Central

    Bryce, C F; Williams, D C; John, R A; Fasella, P

    1976-01-01

    Cytoplasmic aspartate aminotransferase and malate dehydrogenase were purified from pig heart. Kinetic parameters were determined for the separate reaction catalysed by each enzyme and used to predict the course of the coupled reaction: (see article). Although a lag phase should have been easily seen, none was detected. The same coupled reaction was also carried out by using radioactive aspartate in the presence of unlabelled oxaloacetate. The reaction was quenched with HClO4 after 70 ms and the specific radioactivity of the malate produced in this system was found to be essentially the same as that of the original aspartate. These results show that oxaloacetate produced by the aspartate aminotransferase is converted into malate by malate dehydrogenase before it equilibrates with the pool of unlabelled oxaloacetate and are consistent with a proposal that the enzymes are associated in a complex. However, no physical evidence of the existence of a complex could be found. An alternative means of compartmentation of the intermediate as an unstable isomer is considered. Images Fig. 2. Fig. 3. PMID:942372

  2. The amino acid sequence of the aspartate aminotransferase from baker's yeast (Saccharomyces cerevisiae).

    PubMed Central

    Cronin, V B; Maras, B; Barra, D; Doonan, S

    1991-01-01

    1. The single (cytosolic) aspartate aminotransferase was purified in high yield from baker's yeast (Saccharomyces cerevisiae). 2. Amino-acid-sequence analysis was carried out by digestion of the protein with trypsin and with CNBr; some of the peptides produced were further subdigested with Staphylococcus aureus V8 proteinase or with pepsin. Peptides were sequenced by the dansyl-Edman method and/or by automated gas-phase methods. The amino acid sequence obtained was complete except for a probable gap of two residues as indicated by comparison with the structures of counterpart proteins in other species. 3. The N-terminus of the enzyme is blocked. Fast-atom-bombardment m.s. was used to identify the blocking group as an acetyl one. 4. Alignment of the sequence of the enzyme with those of vertebrate cytosolic and mitochondrial aspartate aminotransferases and with the enzyme from Escherichia coli showed that about 25% of residues are conserved between these distantly related forms. 5. Experimental details and confirmatory data for the results presented here are given in a Supplementary Publication (SUP 50164, 25 pages) that has been deposited at the British Library Document Supply Centre, Boston Spa. Wetherby, West Yorkshire LS23 7 BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1991) 273, 5. PMID:1859361

  3. An aminotransferase from Lactococcus lactis initiates conversion of amino acids to cheese flavor compounds.

    PubMed Central

    Yvon, M; Thirouin, S; Rijnen, L; Fromentier, D; Gripon, J C

    1997-01-01

    The enzymatic degradation of amino acids in cheese is believed to generate aroma compounds and therefore to be involved in the complex process of cheese flavor development. In lactococci, transamination is the first step in the degradation of aromatic and branched-chain amino acids which are precursors of aroma compounds. Here, the major aromatic amino acid aminotransferase of a Lactococcus lactis subsp. cremoris strain was purified and characterized. The enzyme transaminates the aromatic amino acids, leucine, and methionine. It uses the ketoacids corresponding to these amino acids and alpha-ketoglutarate as amino group acceptors. In contrast to most bacterial aromatic aminotransferases, it does not act on aspartate and does not use oxaloacetate as second substrate. It is essential for the transformation of aromatic amino acids to flavor compounds. It is a pyridoxal 5'-phosphate-dependent enzyme and is composed of two identical subunits of 43.5 kDa. The activity of the enzyme is optimal between pH 6.5 and 8 and between 35 and 45 degrees C, but it is still active under cheese-ripening conditions. PMID:9023921

  4. Prodynorphine opioid peptides and aspartate aminotransferase studied in spinal cord and sensory neurons

    SciTech Connect

    Sweetnam, P.M.

    1985-01-01

    An objective of this research was to obtain evidence for the synthesis and release of newly discovered opioid peptides, such as dynorphin, in spinal cord and sensory neurons. Several specific antisera were used to visualize dynorphin and related peptides in spinal cord and dorsal root ganglion neurons in dissociated cell culture. Antisera specific for the midportion of the dynorphin molecule revealed a subpopulation of spinal cord neurons with dense immunoreactive dynorphin in cell perikarya, but none in their associated neurites. Antisera specific for either the amino or carboxy terminal sequences of the molecule produced intense immunoreactivity in both cell perikarya and neurites of spinal neurons. These data suggest the cleavage products of dynorphin and not the complete molecule are possible neurotransmitters in the spinal cord. Additional evidence in support of this hypothesis was derived from radioimmunoassays of these cells and their culture medium following depolarization induced by elevated extracellular potassium. Antisera against aspartate aminotransferase revealed no differentially elevated immunoreactive aspartate aminotransferase in tissue sections of spinal cord or dorsal root ganglia.

  5. NMR studies on /sup 15/N-labeled creatine (CR), creatinine (CRN), phosphocreatine (PCR), and phosphocreatinine (PCRN), and on barriers to rotation in creatine kinase-bound creatine in the enzymatic reaction

    SciTech Connect

    Kenyon, G.L.; Reddick, R.E.

    1986-05-01

    Recently, the authors have synthesized /sup 15/N-2-Cr, /sup 15/N-3-Crn, /sup 15/N-2-Crn, /sup 15/N-3-PCrn, /sup 15/N-3-PCr, and /sup 15/N-2-PCr. /sup 1/H, /sup 15/N, /sup 31/P NMR data show that Crn protonates exclusively at the non-methylated ring nitrogen, confirm that PCrn is phosphorylated at the exocyclic nitrogen, and demonstrate that the /sup 31/P-/sup 15/N one-bond coupling constant in /sup 15/N-3-PCr is 18 Hz, not 3 Hz as previously reported by Brindle, K.M., Porteous, R. and Radda, G.K.. The authors have found that creatine kinase is capable of catalyzing the /sup 14/N//sup 15/N positional isotope exchange of 3-/sup 15/N-PCr in the presence of MgADP, but not in its absence. Further, the exchange does not take place when labeled PCr is resynthesized exclusively from the ternary complex E X Cr X MgATP as opposed to either E X Cr or free Cr. This suggests that the enzyme both imparts an additional rotational barrier to creatine in the complex and catalyzes the transfer of phosphoryl group with essentially complete regiospecificity.

  6. Thermodynamics of Deca-alanine Folding in Water.

    PubMed

    Hazel, Anthony; Chipot, Christophe; Gumbart, James C

    2014-07-08

    The determination of the folding dynamics of polypeptides and proteins is critical in characterizing their functions in biological systems. Numerous computational models and methods have been developed for studying structure formation at the atomic level. Due to its small size and simple structure, deca-alanine is used as a model system in molecular dynamics (MD) simulations. The free energy of unfolding in vacuum has been studied extensively using the end-to-end distance of the peptide as the reaction coordinate. However, few studies have been conducted in the presence of explicit solvent. Previous results show a significant decrease in the free energy of extended conformations in water, but the α-helical state is still notably favored over the extended state. Although sufficient in vacuum, we show that end-to-end distance is incapable of capturing the full complexity of deca-alanine folding in water. Using α-helical content as a second reaction coordinate, we deduce a more descriptive free-energy landscape, one which reveals a second energy minimum in the extended conformations that is of comparable free energy to the α-helical state. Equilibrium simulations demonstrate the relative stability of the extended and α-helical states in water as well as the transition between the two states. This work reveals both the necessity and challenge of determining a proper reaction coordinate to fully characterize a given process.

  7. The effect of immunonutrition (glutamine, alanine) on fracture healing

    PubMed Central

    Küçükalp, Abdullah; Durak, Kemal; Bayyurt, Sarp; Sönmez, Gürsel; Bilgen, Muhammed S.

    2014-01-01

    Background There have been various studies related to fracture healing. Glutamine is an amino acid with an important role in many cell and organ functions. This study aimed to make a clinical, radiological, and histopathological evaluation of the effects of glutamine on fracture healing. Methods Twenty rabbits were randomly allocated into two groups of control and immunonutrition. A fracture of the fibula was made to the right hind leg. All rabbits received standard food and water. From post-operative first day for 30 days, the study group received an additional 2 ml/kg/day 20% L-alanine L-glutamine solution via a gastric catheter, and the control group received 2 ml/kg/day isotonic via gastric catheter. At the end of 30 days, the rabbits were sacrificed and the fractures were examined clinically, radiologically, and histopathologically in respect to the degree of union. Results Radiological evaluation of the control group determined a mean score of 2.5 according to the orthopaedists and 2.65 according to the radiologists. In the clinical evaluation, the mean score was 1.875 for the control group and 2.0 for the study group. Histopathological evaluation determined a mean score of 8.5 for the control group and 9.0 for the study group. Conclusion One month after orally administered glutamine–alanine, positive effects were observed on fracture healing radiologically, clinically, and histopathologically, although no statistically significant difference was determined.

  8. Formation of chloroform during chlorination of alanine in drinking water.

    PubMed

    Chu, Wen-Hai; Gao, Nai-Yun; Deng, Yang; Dong, Bing-Zhi

    2009-11-01

    Currently, dissolved nitrogenous organic matters in water, important precursors of disinfection by-products (DBPs), are of significant concern. This study was to explore the formation of chloroform (CF) during chlorination of alanine (Ala), an important nitrogenous organic compound commonly present in water sources. Our results indicated that the CF yield reached a maximum value of 0.143% at the molar ratio of chlorine atom to nitrogen atom (Cl/N)=1.0 over a Cl/N range of 0.2-5.0 (pH=7.0, reaction time=5d, and initial Ala=0.1mM). At an acidic-neutral condition (pH 4-7), the formation of CF was suppressed. However, the highest CF yield (0.227%) occurred at weakly alkaline condition (pH 8.0) (initial Ala=0.1mM, and Cl/N=1.0). The increase of Br(-) in water can increase total trihalomethanes (THMs) and bromo-THMs. However, the bromo-THMs level reached a plateau at Br(-)/Cl>0.04. Finally, based on the computation of frontier electron density and identification and measurement of key intermediates during Ala chlorination, we proposed a formation pathway of CF from Ala chlorination: Ala-->monochloro-N-alanine (MC-N-Ala)-->acetaldehyde (AAld)-->monochloroacetaldehyde acetaldehyde (MCAld)-->dichloroacetaldehyde (DCAld)-->trichloroacetaldehyde (TCAld)-->CF.

  9. Cardiac troponin T and creatine kinase MB isoenzyme as biochemical markers of ischemia after heart preservation and transplantation.

    PubMed

    Carrier, M; Solymoss, B C; Cartier, R; Leclerc, Y; Pelletier, L C

    1994-01-01

    An ischemic preservation period of less than 4 to 6 hours for the donor heart is considered safe in heart transplantation. To determine the severity of myocardial cell damage, we measured serum creatine kinase MB isoenzyme activity, creatine kinase MB isoenzyme mass concentration, and troponin T release in 14 patients during the first 48 hours after heart transplantation. All donors had normal cardiac function at echocardiographic evaluation. The heart was arrested with cold crystalloid cardioplegic solution and preserved in a hypothermic solution. All patients survived the first week after transplantation. Total ischemic time averaged 126 +/- 33 minutes (range 88 to 195 minutes). Maximal creatine kinase MB isoenzyme activity, creatine kinase MB isoenzyme mass concentration, and troponin T serum values after transplantation averaged 130 +/- 44 IU/L, 140 +/- 121 ng/ml, and 3.3 +/- 1.4 ng/ml, respectively. No significant correlation was found between ischemic time and peak levels of creatine kinase MB isoenzyme activity (r = 0.22), creatine kinase MB isoenzyme mass (r = 0.37) and troponin T (r = 0.12). A moderate correlation between ischemic time and the initial slope of time-activity curve of creatine kinase MB isoenzyme mass (r = 0.66, p = 0.01) and of troponin T release (r = 0.55, p = 0.03) was observed. Ischemic time and donor age were significantly related to creatine kinase MB isoenzyme mass (R2 = 0.61) and to troponin T (R2 = 0.47) initial release slopes. In conclusion, during a short period of ischemic preservation, myocardial cell damage appears to be mild and best reflected by the elevation and the time-activity curves of release of cardiac troponin T and creatine kinase MB isoenzyme mass.

  10. Simultaneous determination of adenine nucleotides, creatine phosphate and creatine in rat liver by high performance liquid chromatography-electrospray ionization-tandem mass spectrometry.

    PubMed

    Jiang, Yang; Sun, Chengjun; Ding, Xueqin; Yuan, Ding; Chen, Kefei; Gao, Bo; Chen, Yi; Sun, Aimin

    2012-07-01

    A high performance liquid chromatography-electrospray ionization-tandem mass spectrometric method (HPLC-ESI-MS/MS) was developed for simultaneous determination of adenosine 5'-triphosphate (ATP), adenosine 5'-diphosphate (ADP), adenosine 5'-monophosphate (AMP), creatine phosphate (CP), and creatine in rat liver. After extraction with pre-cooled (4 °C) methanol/water (1:1, v/v), the analytes were separated on a porous graphitic carbon (Hypercarb) column (2.1 mm × 150 mm, 5 μm) using a programmed gradient elution with a mobile phase consisting of 2 mmol/L ammonium acetate in water and 2 mmol/L ammonium acetate in acetonitrile (pH=10.0). The analytes were detected in a way of multiple reaction monitoring (MRM) under negative scan mode by a triple quadrupole mass spectrometer with electrospray ionization (ESI). An external calibration method with linear ranges from 10 to 5000 ng/mL for the five target compounds was used for quantification with a correlation coefficients≥0.9973. The limits of detection and limits of quantification for all analytes were in ranges from 0.50 to 1.5 ng/mL and 1.6 to 0.5 ng/mL, respectively. The average recoveries spiked in three levels were from 77.2% to 102% and precisions expressed in RSDs were from 0.2% to 4.8%. The established method was successfully applied to determination of ATP, ADP, AMP, CP and creatine in liver tissue.

  11. Involvement of alanine racemase in germination of Bacillus cereus spores lacking an intact exosporium.

    PubMed

    Venir, Elena; Del Torre, Manuela; Cunsolo, Vincenzo; Saletti, Rosaria; Musetti, Rita; Stecchini, Mara Lucia

    2014-02-01

    The L-alanine mediated germination of food isolated Bacillus cereus DSA 1 spores, which lacked an intact exosporium, increased in the presence of D-cycloserine (DCS), which is an alanine racemase (Alr) inhibitor, reflecting the activity of the Alr enzyme, capable of converting L-alanine to the germination inhibitor D-alanine. Proteomic analysis of the alkaline extracts of the spore proteins, which include exosporium and coat proteins, confirmed that Alr was present in the B. cereus DSA 1 spores and matched to that encoded by B. cereus ATCC 14579, whose spore germination was strongly affected by the block of conversion of L- to D-alanine. Unlike ATCC 14579 spores, L-alanine germination of B. cereus DSA 1 spores was not affected by the preincubation with DCS, suggesting a lack of restriction in the reactant accessibility.

  12. The Helical Alanine Controversy: An (Ala)6 Insertion Dramatically Increases Helicity

    PubMed Central

    Lin, Jasper C.; Barua, Bipasha

    2013-01-01

    Employing chemical shift melts and hydrogen/deuterium exchange NMR techniques, we have determined the stabilization of the Trp-cage miniprotein due to multiple alanine insertions within the N-terminal α-helix. Alanine is shown to be uniquely helix-stabilizing and this stabilization is reflected in the global fold stability of the Trp-cage. The associated free energy change per alanine can be utilized to calculate the alanine propagation value. From the Lifson–Roig formulation, the calculated value (wAla = 1.6) is comparable to those obtained for short, solubilized, alanine-rich helices and is much larger than the values obtained by prior host–guest techniques or in N-terminally templated helices and peptides bearing long contiguous strings of alanines with no capping or solubilizing units present. PMID:15493925

  13. Expression, crystallization and preliminary X-ray crystallographic analysis of Xoo0352, D-alanine-D-alanine ligase A, from Xanthomonas oryzae pv. oryzae.

    PubMed

    Doan, Thanh Thi Ngoc; Kim, Jin-Kwang; Kim, Hyesoon; Ahn, Yeh-Jin; Kim, Jeong-Gu; Lee, Byoung-Moo; Kang, Lin-Woo

    2008-12-01

    Xanthomonas oryzae pv. oryzae (Xoo) causes bacterial blight (BB), which is one of the most devastating diseases of rice in most rice-growing countries. D-Alanine-D-alanine ligase A (DdlA), coded by the Xoo0352 gene, was expressed, purified and crystallized. DdlA is an enzyme that is involved in D-alanine metabolism and the biosynthesis of an essential bacterial peptidoglycan precursor, in which it catalyzes the formation of D-alanyl-D-alanine from two D-alanines, and is thus an attractive antibacterial drug target against Xoo. The DdlA crystals diffracted to 2.3 A resolution and belonged to the primitive tetragonal space group P4(3)2(1)2, with unit-cell parameters a = b = 83.0, c = 97.6 A. There is one molecule in the asymmetric unit, with a corresponding V(M) of 1.88 A(3) Da(-1) and a solvent content of 34.6%. The initial structure was determined by molecular replacement using D-alanine-D-alanine ligase from Staphylococcus aureus (PDB code 2i87) as a template model.

  14. Creatine co-ingestion with carbohydrate or cinnamon extract provides no added benefit to anaerobic performance.

    PubMed

    Islam, Hashim; Yorgason, Nick J; Hazell, Tom J

    2016-09-01

    The insulin response following carbohydrate ingestion enhances creatine transport into muscle. Cinnamon extract is promoted to have insulin-like effects, therefore this study examined if creatine co-ingestion with carbohydrates or cinnamon extract improved anaerobic capacity, muscular strength, and muscular endurance. Active young males (n = 25; 23.7 ± 2.5 y) were stratified into 3 groups: (1) creatine only (CRE); (2) creatine+ 70 g carbohydrate (CHO); or (3) creatine+ 500 mg cinnamon extract (CIN), based on anaerobic capacity (peak power·kg(-1)) and muscular strength at baseline. Three weeks of supplementation consisted of a 5 d loading phase (20 g/d) and a 16 d maintenance phase (5 g/d). Pre- and post-supplementation measures included a 30-s Wingate and a 30-s maximal running test (on a self-propelled treadmill) for anaerobic capacity. Muscular strength was measured as the one-repetition maximum 1-RM for chest, back, quadriceps, hamstrings, and leg press. Additional sets of the number of repetitions performed at 60% 1-RM until fatigue measured muscular endurance. All three groups significantly improved Wingate relative peak power (CRE: 15.4% P = .004; CHO: 14.6% P = .004; CIN: 15.7%, P = .003), and muscular strength for chest (CRE: 6.6% P < .001; CHO: 6.7% P < .001; CIN: 6.4% P < .001), back (CRE: 5.8% P < .001; CHO: 6.4% P < .001; CIN: 8.1% P < .001), and leg press (CRE: 11.7% P = .013; CHO: 10.0% P = .007; CIN: 17.3% P < .001). Only the CRE (10.4%, P = .021) and CIN (15.5%, P < .001) group improved total muscular endurance. No differences existed between groups post-supplementation. These findings demonstrate that three different methods of creatine ingestion lead to similar changes in anaerobic power, strength, and endurance.

  15. Effect of creatine supplementation on metabolism and performance in humans during intermittent sprint cycling.

    PubMed

    Finn, J P; Ebert, T R; Withers, R T; Carey, M F; Mackay, M; Phillips, J W; Febbraio, M A

    2001-03-01

    This double blind study investigated the effect of oral creatine supplementation (CrS) on 4 x 20 s of maximal sprinting on an air-braked cycle ergometer. Each sprint was separated by 20 s of recovery. A group of 16 triathletes [mean age 26.6 (SD 5.1) years. mean body mass 77.0 (SD 5.8) kg, mean body fat 12.9 (SD 4.6)%, maximal oxygen uptake 4.86 (SD 0.7) l.min-1] performed an initial 4 x 20 s trial after a muscle biopsy sample had been taken at rest. The subjects were then matched on their total intramuscular creatine content (TCr) before being randomly assigned to groups to take by mouth either a creatine supplement (CRE) or a placebo (CON) before a second 4 x 20 s trial. A muscle biopsy sample was also taken immediately before this second trial. The CrS of 100 g comprised 4 x 5 g for 5 days. The initial mean TCr were 112.5 (SD 8.7) and 112.5 (SD 10.7) mmol.kg-1 dry mass for CRE and CON, respectively. After creatine loading and placebo ingestion respectively, CRE [128.7 (SD 11.8) mmol.kg-1 dry mass] had a greater (P = 0.01) TCr than CON [112.0 (SD 10.0) mmol.kg-1 dry mass]. While the increase in free creatine for CRE was statistically significant (P = 0.034), this was not so for the changes in phosphocreatine content [trial 1: 75.7 (SD 6.9), trial 2: 84.7 (SD 11.0) mmol.kg-1 dry mass, P = 0.091]. There were no significant differences between CRE and CON for citrate synthase activity (P = 0.163). There was a tendency towards improved performance in terms of 1 s peak power (in watts P = 0.07; in watts per kilogram P = 0.05), 5 s peak power (in watts P = 0.08) and fatigue index (P = 0.08) after CrS for sprint 1 of the second trial. However, there was no improvement for mean power (in watts P = 0.15; in watts per kilogram P = 0.1) in sprint 1 or for any performance values in subsequent sprints. Our results suggest that, while CrS elevates the intramuscular stores of free creatine, this does not have an ergogenic effect on 4 x 20 s all-out cycle sprints with

  16. Dissociated expression of mitochondrial and cytosolic creatine kinases in the human brain: a new perspective on the role of creatine in brain energy metabolism

    PubMed Central

    Lowe, Matthew TJ; Kim, Eric H; Faull, Richard LM; Christie, David L; Waldvogel, Henry J

    2013-01-01

    The phosphocreatine/creatine kinase (PCr/CK) system in the brain is defined by the expression of two CK isozymes: the cytosolic brain-type CK (BCK) and the ubiquitous mitochondrial CK (uMtCK). The system plays an important role in supporting cellular energy metabolism by buffering adenosine triphosphate (ATP) consumption and improving the flux of high-energy phosphoryls around the cell. This system is well defined in muscle tissue, but there have been few detailed studies of this system in the brain, especially in humans. Creatine is known to be important for neurologic function, and its loss from the brain during development can lead to mental retardation. This study provides the first detailed immunohistochemical study of the expression pattern of BCK and uMtCK in the human brain. A strikingly dissociated pattern of expression was found: uMtCK was found to be ubiquitously and exclusively expressed in neuronal populations, whereas BCK was dominantly expressed in astrocytes, with a low and selective expression in neurons. This pattern indicates that the two CK isozymes are not widely coexpressed in the human brain, but rather are selectively expressed depending on the cell type. These results suggest that the brain cells may use only certain properties of the PCr/CK system depending on their energetic requirements. PMID:23715059

  17. Pseudolinkage of the duplicate loci for supernatant aspartate aminotransferase in brook trout, Salvelinus fontinalis.

    PubMed

    Wright, J E; May, B; Stoneking, M; Lee, G M

    1980-01-01

    Electrophoretic variation involving three alleles is described for the duplicated loci for supernatant aspartate aminotransferase (AAT-1,2), from muscle extracts of brook trout. Both loci exhibit largely disomic inheritance. Exceptional progeny types are proposed to be the result of a form of tetrasomic inheritance. Nonrandom segregation was found among the progeny of males doubly heterozygous for AAT markers; where so-called linkage phase was known, this nonrandom assortment was shown to be pseudolinkage (78.9 percent recombination). Analyses of joint segregation of triply heterozygous males for the AAT-(1,2) loci and for the single alpha glycerophosphate dehydrogenase locus (AGP-1) revealed true linkage of AGP-1 with one AAT locus (mean r = 11 percent), but pseudolinkage with the other AAT locus (r = 74 percent). Intraindividual variation for homoeologous multivalent pairing of two acrocentric with two metacentric chromosomes in males, but with bivalent pairing in females, is proposed to account for pseudolinkage and for the tetrasomically inherited types.

  18. Hepatitis A virus genotype IA-infected patient with marked elevation of aspartate aminotransferase levels.

    PubMed

    Miura, Yoshifumi; Kanda, Tatsuo; Yasui, Shin; Takahashi, Koji; Haga, Yuki; Sasaki, Reina; Nakamura, Masato; Wu, Shuang; Nakamoto, Shingo; Arai, Makoto; Nishizawa, Tsutomu; Okamoto, Hiroaki; Yokosuka, Osamu

    2017-02-01

    We describe a case of acute liver failure (ALF) without hepatic encephalopathy with marked elevation of aminotransferase due to hepatitis A, according to the revised Japanese criteria of ALF. This liver biopsy of the patient showed compatible to acute viral hepatitis and she immediately recovered without intensive care. She had no comorbid disorders. Of interest, phylogenetic tree analysis using almost complete genomes of hepatitis A virus (HAV) demonstrated that the HAV isolate from her belonged to the HAV subgenotype IA strain and was similar to the HAJFF-Kan12 strain (99% nucleotide identity) or FH1 strain (98% nucleotide identity), which is associated with severe or fulminant hepatitis A. Careful interpretation of the association between HAV genome variations and severity of hepatitis A is needed and the mechanism of the severe hepatitis should be explored.

  19. Ornithine Aminotransferase, an Important Glutamate-Metabolizing Enzyme at the Crossroads of Multiple Metabolic Pathways

    PubMed Central

    Ginguay, Antonin; Cynober, Luc; Curis, Emmanuel; Nicolis, Ioannis

    2017-01-01

    Ornithine δ-aminotransferase (OAT, E.C. 2.6.1.13) catalyzes the transfer of the δ-amino group from ornithine (Orn) to α-ketoglutarate (aKG), yielding glutamate-5-semialdehyde and glutamate (Glu), and vice versa. In mammals, OAT is a mitochondrial enzyme, mainly located in the liver, intestine, brain, and kidney. In general, OAT serves to form glutamate from ornithine, with the notable exception of the intestine, where citrulline (Cit) or arginine (Arg) are end products. Its main function is to control the production of signaling molecules and mediators, such as Glu itself, Cit, GABA, and aliphatic polyamines. It is also involved in proline (Pro) synthesis. Deficiency in OAT causes gyrate atrophy, a rare but serious inherited disease, a further measure of the importance of this enzyme. PMID:28272331

  20. Antibacterial Activity of Alanine-Derived Gemini Quaternary Ammonium Compounds.

    PubMed

    Piecuch, Agata; Obłąk, Ewa; Guz-Regner, Katarzyna

    The antibacterial activity of alanine-derived gemini quaternary ammonium salts (chlorides and bromides) with various spacer and alkyl chain lengths was investigated. The studied compounds exhibited a strong bactericidal effect, especially bromides with 10 and 12 carbon alkyl chains and 3 carbon spacer groups (TMPAL-10 Br and TMPAL-12 Br), with a short contact time. Both salts dislodged biofilms of Pseudomonas aeruginosa and Staphylococcus epidermidis, and were lethal to adherent cells of S. epidermidis. Bromide with 2 carbon spacer groups and 12 carbon alkyl chains (TMEAL-12 Br) effectively reduced microbial adhesion by coating polystyrene and silicone surfaces. The results obtained suggest that, after further studies, gemini QAS might be considered as antimicrobial agents in medicine or industry.

  1. Charge dependent photodynamic activity of alanine based zinc phthalocyanines.

    PubMed

    Wang, Ao; Li, Yejing; Zhou, Lin; Yuan, Linxin; Lu, Shan; Lin, Yun; Zhou, Jiahong; Wei, Shaohua

    2014-12-01

    In this paper, to minimize the effects of different structure, three alanine-based zinc phthalocyanines (Pcs) of differing charges were engineered and synthesized with the same basic structure. On this premise, the relationship between nature of charge and photodynamic activity was studied. Besides, further verification and explanation of some inconsistent results were also carried out. The results showed that charge can influence the aggregation state, singlet oxygen generation ability and cellular uptake of Pcs, thereby affecting their photodynamic activity. In addition, the biomolecules inside cells may interact with Pcs of differing charges, which can also influence the aggregation state and singlet oxygen generation of the Pcs, and then influence the relationship between nature of charge and photodynamic activity.

  2. Radiolysis of alanine adsorbed in a clay mineral

    SciTech Connect

    Aguilar-Ovando, Ellen Y.; Negron-Mendoza, Alicia

    2013-07-03

    Optical activity in molecules is a chemical characteristic of living beings. In this work, we examine the hypothesis of the influence of different mineral surfaces on the development of a specific chirality in organic molecules when subjected to conditions simulating the primitive Earth during the period of chemical evolution. By using X-ray diffraction techniques and HPLC/ELSD to analyze aqueous suspensions of amino acids adsorbed on minerals irradiated in different doses with a cobalt-60 gamma source, the experiments attempt to prove the hypothesis that some solid surfaces (like clays and meteorite rocks) may have a concentration capacity and protective role against external sources of ionizing radiation (specifically {gamma}-ray) for some organic compounds (like some amino acids) adsorbed on them. Preliminary results show a slight difference in the adsorption and radiolysis of the D-and L-alanine.

  3. First-principles study of fluorination of L-Alanine

    NASA Astrophysics Data System (ADS)

    Sreepad, H. R.; Ravi, H. R.; Ahmed, Khaleel; Dayananda, H. M.; Umakanth, K.; Manohara, B. M.

    2013-02-01

    First-principles calculations based on Density Functional Theory have been done on effect of fluorination of an important amino acid - L-Alanine. Its structure has been simulated. The unit cell is orthorhombic with lattice parameters a=5.90Å, b=13.85Å and c=5.75Å with volume 470 (Å)3. Bond lengths and bond angles have been estimated. Electronic Density of States calculations show that the material has a band gap of 4.47eV. Electronic band structure indicates that the material can be effectively used for NLO applications. The electronic contribution to the dielectric constant has been calculated and its average value comes out to be 2.165.

  4. Rabbit muscle creatine phosphokinase. CDNA cloning, primary structure and detection of human homologues.

    PubMed

    Putney, S; Herlihy, W; Royal, N; Pang, H; Aposhian, H V; Pickering, L; Belagaje, R; Biemann, K; Page, D; Kuby, S

    1984-12-10

    A cDNA library was constructed from rabbit muscle poly(A) RNA. Limited amino acid sequence information was obtained on rabbit muscle creatine phosphokinase and this was the basis for design and synthesis of two oligonucleotide probes complementary to a creatine kinase cDNA sequence which encodes a pentapeptide. Colony hybridizations with the probes and subsequent steps led to isolation of two clones, whose cDNA segments partially overlap and which together encode the entire protein. The primary structure was established from the sequence of two cDNA clones and from independently determined sequences of scattered portions of the polypeptide. The reactive cysteine has been located to position 282 within the 380 amino acid polypeptide. The rabbit cDNA hybridizes to digests of human chromosomal DNA. This reveals a restriction fragment length polymorphism associated with the human homologue(s) which hybridizes to the rabbit cDNA.

  5. [Time peak of creatine kinase activity in various sites of myocardial infarct].

    PubMed

    Sochman, J; Fabián, J; Englis, M

    1989-05-12

    The authors studied the differences in the period of time spanning the onset of anginous pain and the peak of plasma creatine kinase activity in different areas of the left ventricle in patients with acute myocardial infarction. The patients were given either intravenous or intracoronary thrombolytic agents. The involvement of different areas of the cardiac muscle had a major effect on the lenght of time until the peak of creatine kinase activity was reached. This reduces the significance of this parameter which as so far been seen as a noninvasive indicator of the patency of the artery supplying the site of the infarction. The authors also refer to other familiar factors likely to alter the role of this parameter.

  6. DL-canaline and 5-fluoromethylornithine. Comparison of two inactivators of ornithine aminotransferase.

    PubMed Central

    Bolkenius, F N; Knödgen, B; Seiler, N

    1990-01-01

    5-Fluoromethylornithine (5FMOrn) is an enzyme-activated irreversible inhibitor or ornithine aminotransferase (L-ornithine:2-oxo-acid 5-aminotransferase, OAT). For purified rat liver OAT, Ki(app.) was found to be 30 microM. and tau 1/2 = 4 min. Of the four stereomers of 5FMOrn only one reacts with OAT. The formation of a chromophore with an absorption maximum at 458 nm after inactivation of OAT by 5FMOrn suggests the formation of an enamine intermediate, which is slowly hydrolysed to release an unsaturated ketone. L-Canaline [(S)-2-amino-4-amino-oxybutyric acid] is a well-known irreversible inhibitor of OAT. Not only the natural L-enantiomer but also the D-enantiomer reacts by oxime formation with pyridoxal 5'-phosphate in the active site of the enzyme, although considerably more slowly. This demonstrates that the stereochemistry at C-2 of ornithine is not absolutely stringent. In vitro, canaline reacted faster than 5FMOrn with OAT. In vivo, however, only incomplete OAT inhibition was observed with canaline. Whereas intraperitoneal administration of 10 mg of 5FMOrn/kg body wt. to mice was sufficient to inactivate OAT in brain and liver by 90% for 24 h, 500 mg of DL-canaline/kg body wt. only produced a transient inhibition of 65-70%. The accumulation of ornithine in these tissues was considerably slower and the maximum concentrations lower than were achieved with 5FMOrn. It appears that DL-canaline, in contrast with 5FMOrn, is not useful as a tool in studies of biological consequences of OAT inhibition. PMID:2363680

  7. Histidine degradation via an aminotransferase increases the nutritional flexibility of Candida glabrata.

    PubMed

    Brunke, Sascha; Seider, Katja; Richter, Martin Ernst; Bremer-Streck, Sibylle; Ramachandra, Shruthi; Kiehntopf, Michael; Brock, Matthias; Hube, Bernhard

    2014-06-01

    The ability to acquire nutrients during infections is an important attribute in microbial pathogenesis. Amino acids are a valuable source of nitrogen if they can be degraded by the infecting organism. In this work, we analyzed histidine utilization in the fungal pathogen of humans Candida glabrata. Hemiascomycete fungi, like C. glabrata or Saccharomyces cerevisiae, possess no gene coding for a histidine ammonia-lyase, which catalyzes the first step of a major histidine degradation pathway in most other organisms. We show that C. glabrata instead initializes histidine degradation via the aromatic amino acid aminotransferase Aro8. Although ARO8 is also present in S. cerevisiae and is induced by extracellular histidine, the yeast cannot use histidine as its sole nitrogen source, possibly due to growth inhibition by a downstream degradation product. Furthermore, C. glabrata relies only on Aro8 for phenylalanine and tryptophan utilization, since ARO8, but not its homologue ARO9, was transcriptionally activated in the presence of these amino acids. Accordingly, an ARO9 deletion had no effect on growth with aromatic amino acids. In contrast, in S. cerevisiae, ARO9 is strongly induced by tryptophan and is known to support growth on aromatic amino acids. Differences in the genomic structure of the ARO9 gene between C. glabrata and S. cerevisiae indicate a possible disruption in the regulatory upstream region. Thus, we show that, in contrast to S. cerevisiae, C. glabrata has adapted to use histidine as a sole source of nitrogen and that the aromatic amino acid aminotransferase Aro8, but not Aro9, is the enzyme required for this process.

  8. Phenylalanine-pyruvate aminotransferase activity in chicks subjected to phenylalanine imbalance or phenylalanine toxicity.

    PubMed

    Lu, J; Austic, R E

    2009-11-01

    Two experiments were done to determine the influence of Phe imbalance and excess on Phe-pyruvate aminotransferase (PAT) activity in the chick. Five replicates of 3 chicks (experiment 1) or 2 chicks (experiment 2) of a commercial brown egg layer strain were fed a semipurified diet for 1 wk and then received experimental diets for 10 d. Three diets were used in experiment 1: the basal diet contained 0.46% Phe; the imbalance diet was similar to the basal diet except that it contained a 10% mixture of indispensable amino acids lacking Phe (IAA - Phe) to create a Phe imbalance; the imbalance corrected diet was similar to the imbalance diet except that it was supplemented with 1.12% Phe to correct the imbalance. A 3 x 2 factorial arrangement of treatments in experiment 2 provided 3 dietary levels (0.46, 1.58, and 2.46%) of Phe and either no supplement or 10% supplement of IAA - Phe. Nonfasted chicks were killed and livers were sampled in experiment 1, and livers, kidneys, brains, and pectoralis major muscles were sampled in experiment 2. In experiment 1, liver PAT activity per gram of liver was 80 and 55% higher (P < 0.01) in chicks fed the imbalance and imbalance corrected diets than in chicks fed the basal diet. In experiment 2, the livers and kidneys, but not brains and muscles, of chicks that received the 10% supplement of IAA - Phe had higher activities of PAT per gram of tissue per minute and per milligram of tissue protein extract per minute than chicks that did not receive IAA - Phe (P < 0.001). No effect of dietary Phe on PAT activity was detected (P > 0.05). Phenylalanine-pyruvate aminotransferase activity appears to be regulated in response to dietary content of indispensable amino acids but not by the dietary level of Phe.

  9. An Evaluation of the Possible Association of Malignant Hyperpyrexia with the Noonan Syndrome Using Serum Creatine Phosphokinase Levels

    ERIC Educational Resources Information Center

    Hunter, Alasdair; Pinsky, Leonard

    1975-01-01

    Examined for malignant hyperpyrexia (extremely high fever) were serum creatine phosphokinase (enzyme) levels of 27 children from 1-to 17-years-old with Noonan syndrome which is characterized by webbed neck, short stature and low set ears. (CL)

  10. Long-term creatine supplementation improves muscular performance during resistance training in older women.

    PubMed

    Aguiar, Andreo Fernando; Januário, Renata Selvatici Borges; Junior, Raymundo Pires; Gerage, Aline Mendes; Pina, Fábio Luiz Cheche; do Nascimento, Matheus Amarante; Padovani, Carlos Roberto; Cyrino, Edilson Serpeloni

    2013-04-01

    This study examined the effects of long-term creatine supplementation combined with resistance training (RT) on the one-repetition maximum (1RM) strength, motor functional performance (e.g., 30-s chair stand, arm curl, and getting up from lying on the floor tests) and body composition (e.g., fat-free mass, muscle mass, and % body fat using DEXA scans) in older women. Eighteen healthy women (64.9 ± 5.0 years) were randomly assigned in a double-blind fashion to either a creatine (CR, N = 9) or placebo (PL, N = 9) group. Both groups underwent a 12-week RT program (3 days week(-1)), consuming an equivalent amount of either creatine (5.0 g day(-1)) or placebo (maltodextrin). After 12 week, the CR group experienced a greater (P < 0.05) increase (Δ%) in training volume (+164.2), and 1RM bench press (+5.1), knee extension (+3.9) and biceps curl (+8.8) performance than the PL group. Furthermore, CR group gained significantly more fat-free mass (+3.2) and muscle mass (+2.8) and were more efficient in performing submaximal-strength functional tests than the PL group. No changes (P > 0.05) in body mass or % body fat were observed from pre- to post-test in either group. These results indicate that long-term creatine supplementation combined with RT improves the ability to perform submaximal-strength functional tasks and promotes a greater increase in maximal strength, fat-free mass and muscle mass in older women.

  11. Effect of caffeine ingestion after creatine supplementation on intermittent high-intensity sprint performance.

    PubMed

    Lee, Chia-Lun; Lin, Jung-Charng; Cheng, Ching-Feng

    2011-08-01

    The aim of this study was to investigate the effects of acute caffeine ingestion on intermittent high-intensity sprint performance after 5 days of creatine loading. After completing a control trial (no ergogenic aids, CON), twelve physically active men were administered in a double-blind, randomized crossover protocol to receive CRE + PLA (0.3 g kg(-1) day(-1) of creatine for 5 days then followed by 6 mg kg(-1) of placebo) and CRE + CAF (0.3 g kg(-1) day(-1) of creatine for 5 days and followed by 6 mg kg(-1) of caffeine), after which they performed a repeated sprint test. Each test consisted of six 10-s intermittent high-intensity sprints on a cycling ergometer, with 60-s rest intervals between sprints. Mean power, peak power, rating of perceived exertion (RPE), and heart rates were measured during the test. Blood samples for lactate, glucose, and catecholamine concentrations were drawn at specified intervals. The mean and peak power observed in the CRE + CAF were significantly higher than those found in the CON during Sprints 1 and 3; and the CRE + CAF showed significantly higher mean and peak power than that in the CRE + PLA during Sprints 1 and 2. The mean and peak power during Sprint 3 in the CRE + PLA was significantly greater than that in the CON. Heart rates, plasma lactate, and glucose increased significantly with CRE + CAF during most sprints. No significant differences were observed in the RPE among the three trials. The present study determined that caffeine ingestion after creatine supplements augmented intermittent high-intensity sprint performance.

  12. Creatine prevents the imbalance of redox homeostasis caused by homocysteine in skeletal muscle of rats.

    PubMed

    Kolling, Janaína; Scherer, Emilene B S; Siebert, Cassiana; Marques, Eduardo Peil; Dos Santos, Tiago Marcom; Wyse, Angela T S

    2014-07-15

    Homocystinuria is a neurometabolic disease caused by severe deficiency of cystathionine beta-synthase activity, resulting in severe hyperhomocysteinemia. Affected patients present several symptoms including a variable degree of motor dysfunction, being that the pathomechanism is not fully understood. In the present study we investigated the effect of chronic hyperhomocysteinemia on some parameters of oxidative stress, namely 2'7'dichlorofluorescein (DCFH) oxidation, levels of thiobarbituric acid-reactive substances (TBARS), antioxidant enzyme activities (SOD, CAT and GPx), reduced glutathione (GSH), total sulfhydryl and carbonyl content, as well as nitrite levels in soleus skeletal muscle of young rats subjected to model of severe hyperhomocysteinemia. We also evaluated the effect of creatine on biochemical alterations elicited by hyperhomocysteinemia. Wistar rats received daily subcutaneous injection of homocysteine (0.3-0.6 μmol/g body weight), and/or creatine (50mg/kg body weight) from their 6th to the 28th days age. Controls and treated rats were decapitated at 12h after the last injection. Chronic homocysteine administration increased 2'7'dichlorofluorescein (DCFH) oxidation, an index of production of reactive species and TBARS levels, an index of lipoperoxidation. Antioxidant enzyme activities, such as SOD and CAT were also increased, but GPx activity was not altered. The content of GSH, sulfhydril and carbonyl were decreased, as well as levels of nitrite. Creatine concurrent administration prevented some homocysteine effects probably by its antioxidant properties. Our data suggest that the oxidative insult elicited by chronic hyperhomocystenemia may provide insights into the mechanisms by which homocysteine exerts its effects on skeletal muscle function. Creatine prevents some alterations caused by homocysteine.

  13. Is dilution important: Factitious Total Creatine Kinase in case of Rhabdomyolysis?

    PubMed Central

    Dinakaran, Asha; Ray, Lopamudra

    2016-01-01

    Factitious test reports may result in incorrect diagnosis and incorrect management. Such incorrect diagnosis can be prevented by a vigilant biochemist. We report a case of Rhabdomyolysis presenting with extremely low total Creatine Kinase (CK) levels which was factitious. Running the sample in dilution resulted in a very high value of total CK which could have been missed if the sample was not run in dilution and the diagnosis of Rhabdomyolysis could have been missed. PMID:27891332

  14. Is dilution important: Factitious Total Creatine Kinase in case of Rhabdomyolysis?

    PubMed

    Nanda, Sunil Kumar; Dinakaran, Asha; Ray, Lopamudra

    2016-10-01

    Factitious test reports may result in incorrect diagnosis and incorrect management. Such incorrect diagnosis can be prevented by a vigilant biochemist. We report a case of Rhabdomyolysis presenting with extremely low total Creatine Kinase (CK) levels which was factitious. Running the sample in dilution resulted in a very high value of total CK which could have been missed if the sample was not run in dilution and the diagnosis of Rhabdomyolysis could have been missed.

  15. Efficient L-Alanine Production by a Thermo-Regulated Switch in Escherichia coli.

    PubMed

    Zhou, Li; Deng, Can; Cui, Wen-Jing; Liu, Zhong-Mei; Zhou, Zhe-Min

    2016-01-01

    L-Alanine has important applications in food, pharmaceutical and veterinary and is used as a substrate for production of engineered thermoplastics. Microbial fermentation could reduce the production cost and promote the application of L-alanine. However, the presence of L-alanine significantly inhibit cell growth rate and cause a decrease in the ultimate L-alanine productivity. For efficient L-alanine production, a thermo-regulated genetic switch was designed to dynamically control the expression of L-alanine dehydrogenase (alaD) from Geobacillus stearothermophilus on the Escherichia coli B0016-060BC chromosome. The optimal cultivation conditions for the genetically switched alanine production using B0016-060BC were the following: an aerobic growth phase at 33 °C with a 1-h thermo-induction at 42 °C followed by an oxygen-limited phase at 42 °C. In a bioreactor experiment using the scaled-up conditions optimized in a shake flask, B0016-060BC accumulated 50.3 g biomass/100 g glucose during the aerobic growth phase and 96 g alanine/100 g glucose during the oxygen-limited phase, respectively. The L-alanine titer reached 120.8 g/l with higher overall and oxygen-limited volumetric productivities of 3.09 and 4.18 g/l h, respectively, using glucose as the sole carbon source. Efficient cell growth and L-alanine production were reached separately, by switching cultivation temperature. The results revealed the application of a thermo-regulated strategy for heterologous metabolic production and pointed to strategies for improving L-alanine production.

  16. Effects of acute creatine loading with or without carbohydrate on repeated bouts of maximal swimming in high-performance swimmers.

    PubMed

    Theodorou, Apostolos S; Havenetidis, Konstantinos; Zanker, Cathy L; O'Hara, John P; King, Roderick F G J; Hood, Colin; Paradisis, Giorgios; Cooke, Carlton B

    2005-05-01

    The addition of carbohydrate (CHO) to an acute creatine (Cr) loading regimen has been shown to increase muscle total creatine content significantly beyond that achieved through creatine loading alone. However, the potential ergogenic effects of combined Cr and CHO loading have not been assessed. The purpose of this study was to compare swimming performance, assessed as mean swimming velocity over repeated maximal intervals, in high-performance swimmers before and after an acute loading regimen of either creatine alone (Cr) or combined creatine and carbohydrate (Cr + CHO). Ten swimmers (mean +/- SD of age and body mass: 17.8 +/- 1.8 years and 72.3 +/- 6.8 kg, respectively) of international caliber were recruited and were randomized to 1 of 2 groups. Each swimmer ingested five 5 g doses of creatine for 4 days, with the Cr + CHO group also ingesting approximately 100 g of simple CHO 30 minutes after each dose of creatine. Performance was measured on 5 separate occasions: twice at "baseline" (prior to intervention, to assess the repeatability of the performance test), within 48 hours after intervention, and then 2 and 4 weeks later. All subjects swam faster after either dietary loading regimen (p < 0.01, both regimens); however, there was no difference in the extent of improvement of performance between groups. In addition, all swimmers continued to produce faster swim times for up to 4 weeks after intervention. Our findings suggest that no performance advantage was gained from the addition of carbohydrate to a creatine-loading regimen in these high-caliber swimmers.

  17. Effects of creatine monohydrate supplementation and exercise on depression-like behaviors and raphe 5-HT neurons in mice

    PubMed Central

    Ahn, Nari; Leem, Yea Hyun; Kato, Morimasa; Chang, Hyukki

    2016-01-01

    [Purpose] The effects of creatine and exercise on chronic stress-induced depression are unclear. In the present study, we identified the effects of 4-week supplementation of creatine monohydrate and/or exercise on antidepressant behavior and raphe 5-HT expression in a chronic mild stress-induced depressed mouse model. [Methods] Seven-week-old male C57BL/6 mice (n=48) were divided randomly into 5 groups: (1) non-stress control (CON, n=10), (2) stress control (ST-CON, n=10), (3) stress and creatine intake (ST-Cr, n=10), (4) stress and exercise (ST-Ex, n=9), and (5) combined stress, exercise, and creatine intake (ST-Cr+Ex, n=9). After five weeks’ treatment, we investigated using both anti-behavior tests (the Tail Suspension Test (TST) and the Forced Swimming Test (FST)), and 5-HT expression in the raphe nuclei (the dorsal raphe (DR) and median raphe (MnR)). [Results] Stress for 4 weeks significantly increased depressive behaviors in the mice. Treatment with creatine supplementation combined with exercise significantly decreased depressive behaviors as compared with the CON-ST group in both the TST and FST tests. With stress, 5-HT expression in the raphe nuclei decreased significantly. With combined creatine and exercise, 5-HT positive cells increased significantly and had a synergic effect on both DR and MnR. [Conclusion] The present study found that even a single treatment of creatine or exercise has partial effects as an antidepressant in mice with chronic mild stress-induced depression. Furthermore, combined creatine and exercise has synergic effects and is a more effective prescription than a single treatment. PMID:27757384

  18. An Fe3O4-nanoparticles-based amperometric biosensor for creatine determination.

    PubMed

    Kaçar, Ceren; Erden, Pinar Esra; Pekyardimci, Sule; Kiliç, Esma

    2013-02-01

    An amperometric biosensor for the detection of creatine was designed, based on carbon paste electrode modified with Fe(3)O(4) nanoparticles. Electron transfer properties of unmodified and Fe(3)O(4)-nanoparticles-modified carbon paste electrodes were investigated by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) methods. Fe(3)O(4) nanoparticles increased the surface area and electric conductivity of the electrode, thus enhancing the sensitivity of the electrode. Optimum pH, buffer concentration, working potential and enzyme loading were selected as 7.0; 0.05 mol L(-1); +0.30 V and 2.0 Unit creatinase (CI), 1.0 Unit sarcosine oxidase (SO), respectively. The purposed biosensor exhibited linear response from 2.0 × 10(-7) mol L(-1) to 3.8 × 10(-6) mol L(-1) and from 9.0 × 10(-6) mol L(-1) to 1.2 × 10(-4) mol L(-1) with a detection limit of 2.0 × 10(-7) mol L(-1). Biosensor was used for determination of creatine in commercial creatine powder samples and showed a good sensing performance.

  19. Impact of creatine supplementation in combination with resistance training on lean mass in the elderly

    PubMed Central

    Pinto, Camila Lemos; Botelho, Patrícia Borges; Carneiro, Juliana Alves

    2016-01-01

    Abstract Background Human ageing is a process characterized by loss of muscle mass, strength, and bone mass. We aimed to examine the efficacy of low‐dose creatine supplementation associated with resistance training on lean mass, strength, and bone mass in the elderly. Methods This was a 12‐week, parallel‐group, double‐blind, randomized, placebo‐controlled trial. The individuals were randomly allocated into one of the following groups: placebo plus resistance training (PL + RT) and creatine supplementation plus resistance training (CR + RT) . The participants were assessed at baseline and after 12 weeks. The primary outcomes were lean mass and strength, assessed by dual energy X‐ray absorptiometry (DXA) and ten‐repetition maximal tests (10 RM), respectively. Secondary outcomes included the lumbar spine, right and left femoral neck, both femur and whole body bone mineral density (BMD), and whole body bone mineral content (BMC), assessed by DXA. Results The CR + RT group had superior gains in lean mass when compared with the PL + RT group (P = 0.02). Changes in the 10 RM tests in bench press and leg press exercises, body composition, BMD, and BMC of all assessed sites did not significantly differ between the groups (P > 0.05). Conclusions Twelve weeks of low‐dose creatine supplementation associated with resistance training resulted in increases in lean mass in the elderly. PMID:27239423

  20. Administration of memantine and imipramine alters mitochondrial respiratory chain and creatine kinase activities in rat brain.

    PubMed

    Réus, Gislaine Z; Stringari, Roberto B; Rezin, Gislaine T; Fraga, Daiane B; Daufenbach, Juliana F; Scaini, Giselli; Benedet, Joana; Rochi, Natália; Streck, Emílio L; Quevedo, João

    2012-04-01

    Several studies have appointed for a role of glutamatergic system and/or mitochondrial function in major depression. In the present study, we evaluated the creatine kinase and mitochondrial respiratory chain activities after acute and chronic treatments with memantine (N-methyl-D: -aspartate receptor antagonist) and imipramine (tricyclic antidepressant) in rats. To this aim, rats were acutely or chronically treated for 14 days once a day with saline, memantine (5, 10 and 20 mg/kg) and imipramine (10, 20 and 30 mg/kg). After acute or chronic treatments, we evaluated mitochondrial respiratory chain complexes (I, II, II-III and IV) and creatine kinase activities in prefrontal cortex, hippocampus and striatum. Our results showed that both acute and chronic treatments with memantine or imipramine altered respiratory chain complexes and creatine kinase activities in rat brain; however, these alterations were different with relation to protocols (acute or chronic), complex, dose and brain area. Finally, these findings further support the hypothesis that the effects of imipramine and memantine could be involve mitochondrial function modulation.

  1. Sarcopenia: current theories and the potential beneficial effect of creatine application strategies.

    PubMed

    Candow, Darren G

    2011-08-01

    Sarcopenia, defined as the age-related loss of muscle mass, subsequently has a negative effect on strength, metabolic rate and functionality leading to a reduced quality of life. With the projected increase in life expectancy, the incidence of muscle loss may rise and further drain the health care system, with greater need for hospitalization, treatment, and rehabilitation. Without effective strategies to counteract aging muscle loss, a global health care crisis may be inevitable. Resistance training is well established to increase aging muscle mass and strength. However, muscle and strength loss is still evident in older adults who have maintained resistance training for most of their life, suggesting that other factors such as nutrition may affect aging muscle biology. Supplementing with creatine, a high-energy compound found in red meat and seafood, during resistance training has a beneficial effect on aging muscle. Emerging evidence now suggests that the timing and dosage of creatine supplementation may be important factors for aging muscle accretion. Unfortunately, the long-term effects of different creatine application strategies on aging muscle are relatively unknown.

  2. Effect of oral creatine supplementation on single-effort sprint performance in elite swimmers.

    PubMed

    Burke, L M; Pyne, D B; Telford, R D

    1996-09-01

    Oral supplementation with creatine monohydrate (Cr.H2O) has been reported to increase muscle creatine phosphate levels. The aim of the present study was to determine the effect of such supplementation on performance of a single-effort sprint by elite swimmers. Thirty-two elite swimmers (M = 18, F = 14; age = 17-25 years) from the Australian Institute of Sport were tested on two occasions, 1 week apart. Tests performed were 25-m, 50-m, and 100-m maximal effort sprints (electronically timed with dive start, swimmers performing their best stroke), each with approximately 10 min active recovery. A 10-s maximal leg ergometry test was also undertaken. Swimmers were divided into two groups matched for sex, stroke/event, and sprint time over 50 m, and groups were randomly assigned to 5 days of Cr.H2O supplementation (4 . day-1 x 5 g Cr.H2O + 2 g sucrose, n = 16) or placebo (4 . day-1 x 5 g Polycose + 2 g sucrose, n = 16) prior to the second trial. Results revealed no significant differences between the group means for sprint times or between 10-s maximal leg ergometry power and work. This study does not support the hypothesis that creatine supplementation enhances single-effort sprint ability of elite swimmers.

  3. Creatine supplementation improves the anaerobic performance of elite junior fin swimmers.

    PubMed

    Juhász, Imre; Györe, I; Csende, Zs; Rácz, L; Tihanyi, J

    2009-09-01

    The objective of this study was to determine whether creatine supplementation (CrS) could improve mechanical power output, and swimming performance in highly trained junior competitive fin swimmers. Sixteen male fin swimmers (age:15.9+/-1.6 years) were randomly and evenly assigned to either a creatine (CR, 4x5 g/day creatine monohydrate for 5 days) or placebo group (P, same dose of a dextrose-ascorbic acid placebo) in a double-blind research. Before and after CrS the average power output was determined by a Bosco-test and the swimming time was measured in two maximal 100 m fin swims. After five days of CrS the average power of one minute continuous rebound jumps increased by 20.2%. The lactate concentration was significantly less after 5 minutes restitution at the second measurement in both groups. The swimming time was significantly reduced in both first (pre: 50.69+/-1.41 s; post: 48.86+/-1.34 s) and second (pre: 50.39+/-1.38 s; post: 48.53+/-1.35 s) sessions of swimming in CR group, but remained almost unchanged in the P group.The results of this study indicate that five day Cr supplementation enhances the dynamic strength and may increase anaerobic metabolism in the lower extremity muscles, and improves performance in consecutive maximal swims in highly trained adolescent fin swimmers.

  4. Creatine kinase kinetics and myocardial infarction in different regions of the left ventricle.

    PubMed

    Sochman, J; Fabián, J; Málek, I; Belán, A; Englis, M

    1989-01-01

    The interval from the onset of infarction pain to culmination of plasma creatine kinase activity (t-peak) was measured in 68 patients with their first myocardial infarction. There is a major difference in this parameter in patients with infarction in the area of the right coronary artery and that supplied by the left anterior descending coronary artery (LAD). While, in 29 patients with infarction in the right coronary artery area, t-peak was 17.7 +/- 4.7 hours, in 39 subjects with infarction in area supplied by the LAD, t-peak was 13.2 +/- 4.6 hours (p less than 0.001). The type of thrombolytic treatment (intravenous or intracoronary, used no difference, just as the time from onset of pain to start of therapy, infarct size and presence or absence of collaterals. A detailed analysis of creatine kinase culmination in relation to the type of artery recanalization is given. The authors conclude that, besides the known factors, creatine kinase culmination is influenced also by the necrosis site, a fact somewhat modifying the informative value of this parameter. However, explanations of this phenomenon are only hypothetical at the present time.

  5. Detection of a novel intragenic rearrangement in the creatine transporter gene by next generation sequencing.

    PubMed

    Yu, Hui; van Karnebeek, Clara; Sinclair, Graham; Hill, Alan; Cui, Hong; Zhang, Victor Wei; Wong, Lee-Jun

    2013-12-01

    Deficiency caused by mutations in the creatine transporter gene (SLC6A8/CT1) is an X-linked form of intellectual disability. The presence of highly homologous pseudogenes and high GC content of SLC6A8 genomic sequence complicates the molecular diagnosis of this disorder. To minimize the pseudogene interference, exons 2 to 13 of SLC6A8 were amplified as a single PCR product using gene-specific long-range PCR (LR-PCR) primers. The GC-rich exon 1 and its flanking intronic sequences were amplified separately in a short fragment under GC-rich conditions and a touchdown PCR program. Traditional Sanger sequence analysis of all coding exons of SLC6A8 from a 3-year-old boy with creatine transporter deficiency did not detect deleterious mutations. The long-range PCR product was used as template followed by massively parallel sequencing (MPS) on HiSeq2000. We were able to detect a tandem duplication involving part of exons 11 and 12 in the SLC6A8 gene. The deduced c.1592_1639dup133 mutation was confirmed to be a hemizygous insertion by targeted genomic DNA and cDNA Sanger sequencing. Combination of deep sequencing technology with long-range PCR revealed a novel intragenic duplication in the SLC6A8 gene, providing a definitive molecular diagnosis of creatine transporter deficiency in a male patient.

  6. Reduced transglutaminase-catalyzed protein aggregation is observed in the presence of creatine using sedimentation velocity.

    PubMed

    Burguera, Elena F; Love, Brian J

    2006-03-01

    Transglutaminases (TGases) are enzymes that catalyze covalent isopeptide crosslinks between reactive lysine and glutamine residues in proteins. Higher than normal local concentrations of TGase have been correlated with increased protein aggregation in vivo. These insoluble protein aggregates are the hallmark of several neurodegenerative diseases, including Alzheimer's, Parkinson's, and Huntington's diseases, although each aggregating protein involved is disease specific. Because TGase is implicated in protein aggregation, there is evidence that its regulation may retard disease progression. Here we report on a laser light transmission technique as an in vitro tool to gauge the efficacy of creatine, a candidate inhibitor, to regulate aggregation. Sedimentation velocities of protein-coated particles in TGase-containing water-glycerol solutions were tracked with different levels of creatine. Sedimentation velocities were converted to apparent aggregate sizes using Stoke's law of sedimentation. The results indicated that creatine promoted up to a 20% reduction in protein aggregation in vitro. This technique may prove to be useful in identifying other functional TGase inhibitors.

  7. Is long term creatine and glutamine supplementation effective in enhancing physical performance of military police officers?

    PubMed

    da Silveira, Celismar Lázaro; de Souza, Thiago Siqueira Paiva; Batista, Gilmário Ricarte; de Araújo, Adenilson Targino; da Silva, Júlio César Gomes; de Sousa, Maria do Socorro Cirilo; Marta, Carlos; Garrido, Nuno Domingo

    2014-09-29

    The objective of this study was to analyze the effect of supplementation with creatine and glutamine on physical fitness of military police officers. Therefore, an experimental double blind study was developed, with the final sample composed by 32 men randomly distributed into three groups: a group supplemented with creatine (n=10), glutamine (n=10) and a placebo group (n=12) and evaluated in three distinct moments, in an interval of three months (T1, T2 and T3). The physical training had a weekly frequency of 5 sessions × 90 min, including strength exercises, local muscular resistance, flexibility and both aerobic and anaerobic capacity. After analyzing the effect of time, group and interaction (group × time) for measures that indicated the physical capabilities of the subjects, a significant effect of time for the entire variable was identified (p<0,05). However, these differences were not observed when the univaried intragroups and intergroups analysis was performed (p>0,05). In face of the results it was concluded that supplementation with creatine and glutamine showed no ergogenic effect on physical performance in military police officers.

  8. Upregulation of the creatine synthetic pathway in skeletal muscles of mature mdx mice

    PubMed Central

    McClure, Warrren C.; Rabon, Rick; Ogawa, Hirofumi; Tseng, Brian S.

    2009-01-01

    Duchenne muscular dystrophy (DMD) is a fatal neuromuscular human disease caused by dystrophin deficiency. The mdx mouse lacks dystrophin protein, yet does not exhibit the debilitating DMD phenotype. Investigating compensatory mechanisms in the mdx mouse is important. This study targets two metabolic genes, guanidinoacetate methyltransferase (GAMT) and arginine:glycine amidinotransferase (AGAT) which are required for creatine synthesis. We show that GAMT and AGAT mRNA are up-regulated 5.4 and 1.9-fold respectively in adult mdx muscle compared to C57. In addition, GAMT protein expression is up-regulated at least 2.5-fold in five different muscles of mdx vs. control. Furthermore, we find GAMT immunoreactivity in 80% of mature mdx muscle fibers in addition to small regenerating fibers and rare revertants; while GAMT immunoreactivity is equal to background levels in all muscle fibers of mature C57 mice. The up-regulation of the creatine synthetic pathway may help maintain muscle creatine levels and limit cellular energy failure in leaky mdx skeletal muscles. These results may help better understand the mild phenotype of the mdx mouse and may offer new treatment horizons for DMD. PMID:17588756

  9. Upregulation of the creatine synthetic pathway in skeletal muscles of mature mdx mice.

    PubMed

    McClure, Warren C; Rabon, Rick E; Ogawa, Hirofumi; Tseng, Brian S

    2007-08-01

    Duchenne muscular dystrophy (DMD) is a fatal neuromuscular human disease caused by dystrophin deficiency. The mdx mouse lacks dystrophin protein, yet does not exhibit the debilitating DMD phenotype. Investigating compensatory mechanisms in the mdx mouse may shed new insights into modifying DMD pathogenesis. This study targets two metabolic genes, guanidinoacetate methyltransferase (GAMT) and arginine:glycine amidinotransferase (AGAT) which are required for creatine synthesis. We show that GAMT and AGAT mRNA are up-regulated 5.4- and 1.9-fold respectively in adult mdx muscle compared to C57. In addition, GAMT protein expression is up-regulated at least 2.5-fold in five different muscles of mdx vs. control. Furthermore, we find GAMT immunoreactivity in up to 80% of mature mdx muscle fibers in addition to small regenerating fibers and rare revertants; while GAMT immunoreactivity is equal to background levels in all muscle fibers of mature C57 mice. The up-regulation of the creatine synthetic pathway may help maintain muscle creatine levels and limit cellular energy failure in leaky mdx skeletal muscles. These results may help better understand the mild phenotype of the mdx mouse and may offer new treatment horizons for DMD.

  10. Both Creatine and Its Product Phosphocreatine Reduce Oxidative Stress and Afford Neuroprotection in an In Vitro Parkinson’s Model

    PubMed Central

    Martín-de-Saavedra, Maria D.; Romero, Alejandro; Egea, Javier; Ludka, Fabiana K.; Tasca, Carla I.; Farina, Marcelo; Rodrigues, Ana Lúcia S.; López, Manuela G.

    2014-01-01

    Creatine is the substrate for creatine kinase in the synthesis of phosphocreatine (PCr). This energetic system is endowed of antioxidant and neuroprotective properties and plays a pivotal role in brain energy homeostasis. The purpose of this study was to investigate the neuroprotective effect of creatine and PCr against 6-hydroxydopamine (6-OHDA)-induced mitochondrial dysfunction and cell death in rat striatal slices, used as an in vitro Parkinson’s model. The possible involvement of the signaling pathway mediated by phosphatidylinositol-3 kinase (PI3K), protein kinase B (Akt), and glycogen synthase kinase-3β (GSK3β) was also evaluated. Exposure of striatal slices to 6-OHDA caused a significant disruption of the cellular homeostasis measured as 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide reduction, lactate dehydrogenase release, and tyrosine hydroxylase levels. 6-OHDA exposure increased the levels of reactive oxygen species and thiobarbituric acid reactive substances production and decreased mitochondrial membrane potential in rat striatal slices. Furthermore, 6-OHDA decreased the phosphorylation of Akt (Serine473) and GSK3β (Serine9). Coincubation with 6-OHDA and creatine or PCr reduced the effects of 6-OHDA toxicity. The protective effect afforded by creatine or PCr against 6-OHDA-induced toxicity was reversed by the PI3K inhibitor LY294002. In conclusion, creatine and PCr minimize oxidative stress in striatum to afford neuroprotection of dopaminergic neurons. PMID:25424428

  11. Both creatine and its product phosphocreatine reduce oxidative stress and afford neuroprotection in an in vitro Parkinson's model.

    PubMed

    Cunha, Mauricio Peña; Martín-de-Saavedra, Maria D; Romero, Alejandro; Egea, Javier; Ludka, Fabiana K; Tasca, Carla I; Farina, Marcelo; Rodrigues, Ana Lúcia S; López, Manuela G

    2014-01-01

    Creatine is the substrate for creatine kinase in the synthesis of phosphocreatine (PCr). This energetic system is endowed of antioxidant and neuroprotective properties and plays a pivotal role in brain energy homeostasis. The purpose of this study was to investigate the neuroprotective effect of creatine and PCr against 6-hydroxydopamine (6-OHDA)-induced mitochondrial dysfunction and cell death in rat striatal slices, used as an in vitro Parkinson's model. The possible involvement of the signaling pathway mediated by phosphatidylinositol-3 kinase (PI3K), protein kinase B (Akt), and glycogen synthase kinase-3β (GSK3β) was also evaluated. Exposure of striatal slices to 6-OHDA caused a significant disruption of the cellular homeostasis measured as 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide reduction, lactate dehydrogenase release, and tyrosine hydroxylase levels. 6-OHDA exposure increased the levels of reactive oxygen species and thiobarbituric acid reactive substances production and decreased mitochondrial membrane potential in rat striatal slices. Furthermore, 6-OHDA decreased the phosphorylation of Akt (Serine(473)) and GSK3β (Serine(9)). Coincubation with 6-OHDA and creatine or PCr reduced the effects of 6-OHDA toxicity. The protective effect afforded by creatine or PCr against 6-OHDA-induced toxicity was reversed by the PI3K inhibitor LY294002. In conclusion, creatine and PCr minimize oxidative stress in striatum to afford neuroprotection of dopaminergic neurons.

  12. 40 CFR 721.520 - Alanine, N-(2-carboxyethyl)-N-alkyl-, salt.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ...-, salt. 721.520 Section 721.520 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.520 Alanine, N-(2-carboxyethyl)-N-alkyl-, salt. (a) Chemical substance... alanine, N-(2-carboxyethyl)-N- alkyl-, salt (P-89-336) is subject to reporting under this section for...

  13. 40 CFR 721.520 - Alanine, N-(2-carboxyethyl)-N-alkyl-, salt.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ...-, salt. 721.520 Section 721.520 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.520 Alanine, N-(2-carboxyethyl)-N-alkyl-, salt. (a) Chemical substance... alanine, N-(2-carboxyethyl)-N- alkyl-, salt (P-89-336) is subject to reporting under this section for...

  14. Polymerization of alanine in the presence of a non-swelling montmorillonite

    NASA Technical Reports Server (NTRS)

    Paecht-Horowitz, M.; Lahav, N.

    1977-01-01

    Alanine, starting from alanine-adenylate, has been polymerized in the presence of non-swelling Al-montmorillonite. The yield of polymerization is much lower than that obtained in the presence of swelling Na-montmorillonite. The possibility that the changing interlayer spacing in Na-montmorillonite might be responsible for its catalytic properties, is discussed.

  15. Regulation of the ald gene encoding alanine dehydrogenase by AldR in Mycobacterium smegmatis.

    PubMed

    Jeong, Ji-A; Baek, Eun-Young; Kim, Si Wouk; Choi, Jong-Soon; Oh, Jeong-Il

    2013-08-01

    The regulatory gene aldR was identified 9