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1

The enzymatic reaction catalyzed by lactate dehydrogenase exhibits one dominant reaction path  

NASA Astrophysics Data System (ADS)

Enzymes are the most efficient chemical catalysts known, but the exact nature of chemical barrier crossing in enzymes is not fully understood. Application of transition state theory to enzymatic reactions indicates that the rates of all possible reaction paths, weighted by their relative probabilities, must be considered in order to achieve an accurate calculation of the overall rate. Previous studies in our group have shown a single mechanism for enzymatic barrier passage in human heart lactate dehydrogenase (LDH). To ensure that this result was not due to our methodology insufficiently sampling reactive phase space, we implement high-perturbation transition path sampling in both microcanonical and canonical regimes for the reaction catalyzed by human heart LDH. We find that, although multiple, distinct paths through reactive phase space are possible for this enzymatic reaction, one specific reaction path is dominant. Since the frequency of these paths in a canonical ensemble is inversely proportional to the free energy barriers separating them from other regions of phase space, we conclude that the rarer reaction paths are likely to have a negligible contribution. Furthermore, the non-dominate reaction paths correspond to altered reactive conformations and only occur after multiple steps of high perturbation, suggesting that these paths may be the result of non-biologically significant changes to the structure of the enzymatic active site.

Masterson, Jean E.; Schwartz, Steven D.

2014-10-01

2

Genetics Home Reference: Lactate dehydrogenase deficiency  

MedlinePLUS

... PubMed Recent literature OMIM Genetic disorder catalog Conditions > Lactate dehydrogenase deficiency On this page: Description Genetic changes ... names Glossary definitions Reviewed February 2012 What is lactate dehydrogenase deficiency? Lactate dehydrogenase deficiency is a condition ...

3

Lactate Dehydrogenases in Human Testes  

Microsoft Academic Search

A unique form of lactate dehydrogenase was observed in the starch-gel electrophoretic patterns of adult human testes. It was present in sperm, but absent in prepubertal testes. Its electrophoretic mobility, heat stability, kinetic behavior with pyridine nucleotide analogs, and chromatographic characteristics on diethylaminoethyl cellulose were intermediate to those observed for lactate dehydrogenase isozymes 3 and 4.

Antonio Blanco; William H. Zinkham

1963-01-01

4

Acrosome reaction in bovine spermatozoa: role of reactive oxygen species and lactate dehydrogenase C4.  

PubMed

After capacitation, mammalian spermatozoa accomplish the acrosome reaction (AR), a well-controlled exocytosis process crucial to fertilize mature oocytes that involves several protein kinases such as protein kinase A (PKA), C (PKC), and tyrosine kinase (PTK). Reactive oxygen species (ROS) are involved in both bovine sperm capacitation and AR. Lactate dehydrogenase C4 (LDH-C4) was associated with bovine and mouse sperm capacitation. Our aims were to study the participation of LDH-C4 to contribute with the status redox required for AR and the role of ROS in the regulation of PKA, PKC, and PTK involved in the exocytotic event. Sodium oxamate, an inhibitor of LDH-C4, prevented the AR induced by lysophosphatidylcholine (LPC) or NADH. Hydrogen peroxide promoted and superoxide dismutase (scavenger of superoxide), catalase (scavenger of hydrogen peroxide), diphenyleneiodinum, diphenyliodonium, cibacron blue, and lapachol (inhibitors of NADPH oxidase) prevented the AR, suggesting that ROS and a sperm oxidase are involved in the AR induced by these compounds. Inhibitors of PKA, PKC, and PTK also prevented the AR induced by LPC or NADH, suggesting the involvement of these kinases in the process. These results suggest that LDH-C4 may participate in the regulation of the redox status required to achieve the AR in bovine spermatozoa and that ROS are key elements in the regulation of protein kinases associated with the AR process. PMID:16112812

O'Flaherty, C; Breininger, E; Beorlegui, N; Beconi, M T

2005-10-30

5

21 CFR 862.1445 - Lactate dehydrogenase isoenzymes test system.  

Code of Federal Regulations, 2011 CFR

...2011-04-01 2011-04-01 false Lactate dehydrogenase isoenzymes test system...Chemistry Test Systems § 862.1445 Lactate dehydrogenase isoenzymes test system. (a) Identification. A lactate dehydrogenase isoenzymes test...

2011-04-01

6

21 CFR 862.1445 - Lactate dehydrogenase isoenzymes test system.  

Code of Federal Regulations, 2010 CFR

...2010-04-01 2010-04-01 false Lactate dehydrogenase isoenzymes test system...Chemistry Test Systems § 862.1445 Lactate dehydrogenase isoenzymes test system. (a) Identification. A lactate dehydrogenase isoenzymes test...

2010-04-01

7

21 CFR 862.1440 - Lactate dehydrogenase test system.  

Code of Federal Regulations, 2012 CFR

...2012-04-01 2012-04-01 false Lactate dehydrogenase test system. 862.1440...Chemistry Test Systems § 862.1440 Lactate dehydrogenase test system. (a) Identification. A lactate dehydrogenase test system is a...

2012-04-01

8

21 CFR 862.1445 - Lactate dehydrogenase isoenzymes test system.  

Code of Federal Regulations, 2014 CFR

...2014-04-01 2014-04-01 false Lactate dehydrogenase isoenzymes test system...Chemistry Test Systems § 862.1445 Lactate dehydrogenase isoenzymes test system. (a) Identification. A lactate dehydrogenase isoenzymes test...

2014-04-01

9

21 CFR 862.1440 - Lactate dehydrogenase test system.  

Code of Federal Regulations, 2011 CFR

...2011-04-01 2011-04-01 false Lactate dehydrogenase test system. 862.1440...Chemistry Test Systems § 862.1440 Lactate dehydrogenase test system. (a) Identification. A lactate dehydrogenase test system is a...

2011-04-01

10

21 CFR 862.1440 - Lactate dehydrogenase test system.  

Code of Federal Regulations, 2010 CFR

...2010-04-01 2010-04-01 false Lactate dehydrogenase test system. 862.1440...Chemistry Test Systems § 862.1440 Lactate dehydrogenase test system. (a) Identification. A lactate dehydrogenase test system is a...

2010-04-01

11

21 CFR 862.1445 - Lactate dehydrogenase isoenzymes test system.  

Code of Federal Regulations, 2013 CFR

...2013-04-01 2013-04-01 false Lactate dehydrogenase isoenzymes test system...Chemistry Test Systems § 862.1445 Lactate dehydrogenase isoenzymes test system. (a) Identification. A lactate dehydrogenase isoenzymes test...

2013-04-01

12

21 CFR 862.1445 - Lactate dehydrogenase isoenzymes test system.  

Code of Federal Regulations, 2012 CFR

...2012-04-01 2012-04-01 false Lactate dehydrogenase isoenzymes test system...Chemistry Test Systems § 862.1445 Lactate dehydrogenase isoenzymes test system. (a) Identification. A lactate dehydrogenase isoenzymes test...

2012-04-01

13

21 CFR 862.1440 - Lactate dehydrogenase test system.  

Code of Federal Regulations, 2014 CFR

...2014-04-01 2014-04-01 false Lactate dehydrogenase test system. 862.1440...Chemistry Test Systems § 862.1440 Lactate dehydrogenase test system. (a) Identification. A lactate dehydrogenase test system is a...

2014-04-01

14

21 CFR 862.1440 - Lactate dehydrogenase test system.  

Code of Federal Regulations, 2013 CFR

...2013-04-01 2013-04-01 false Lactate dehydrogenase test system. 862.1440...Chemistry Test Systems § 862.1440 Lactate dehydrogenase test system. (a) Identification. A lactate dehydrogenase test system is a...

2013-04-01

15

Activity, Stability and Structural Studies of Lactate Dehydrogenases Adapted to  

E-print Network

Activity, Stability and Structural Studies of Lactate Dehydrogenases Adapted to Extreme Thermal; accepted 18 September 2007 Available online 22 September 2007 Lactate dehydrogenase (LDH) catalyzes the conversion of pyruvate to lactate with concomitant oxidation of NADH during the last step in anaerobic

Glover, Mark

16

Biochemical and structural characterization of Cryptosporidium parvum Lactate dehydrogenase.  

PubMed

The protozoan parasite Cryptosporidium parvum causes waterborne diseases worldwide. There is no effective therapy for C. parvum infection. The parasite depends mainly on glycolysis for energy production. Lactate dehydrogenase is a major regulator of glycolysis. This paper describes the biochemical characterization of C. parvum lactate dehydrogenase and high resolution crystal structures of the apo-enzyme and four ternary complexes. The ternary complexes capture the enzyme bound to NAD/NADH or its 3-acetylpyridine analog in the cofactor binding pocket, while the substrate binding site is occupied by one of the following ligands: lactate, pyruvate or oxamate. The results reveal distinctive features of the parasitic enzyme. For example, C. parvum lactate dehydrogenase prefers the acetylpyridine analog of NADH as a cofactor. Moreover, it is slightly less sensitive to gossypol inhibition compared with mammalian lactate dehydrogenases and not inhibited by excess pyruvate. The active site loop and the antigenic loop in C. parvum lactate dehydrogenase are considerably different from those in the human counterpart. Structural features and enzymatic properties of C. parvum lactate dehydrogenase are similar to enzymes from related parasites. Structural comparison with malate dehydrogenase supports a common ancestry for the two genes. PMID:25542170

Cook, William J; Senkovich, Olga; Hernandez, Agustin; Speed, Haley; Chattopadhyay, Debasish

2015-03-01

17

Pleiotropic effects of lactate dehydrogenase inactivation in Lactobacillus casei  

Microsoft Academic Search

In lactic acid bacteria, conversion of pyruvic to lactic acid through the activity of lactate dehydrogenase (Ldh) constitutes the final step of the homofermentative pathway. Lactobacillus casei has two characterized genes encoding Ldh activities. The ldhL gene codes for an L-Ldh, which specifically catalyzes the formation of l-lactate, whereas the hicD gene codes for a d-hydroxyisocaproate dehydrogenase (HicDH), which catalyzes

Rosa Viana; María Jesús Yebra; José Luis Galán; Vicente Monedero; Gaspar Pérez-Martínez

2005-01-01

18

A Specific, Highly Active Malate Dehydrogenase by Redesign of a Lactate Dehydrogenase Framework  

Microsoft Academic Search

Three variations to the structure of the nicotinamide adenine dinucleotide (NAD)-dependent L-lactate dehydrogenase from Bacillus stearothermophilus were made to try to change the substrate specificity from lactate to malate: Asp197--> Asn, Thr246--> Gly, and Gln102--> Arg). Each modification shifts the specificity from lactate to malate, although only the last (Gln102--> Arg) provides an effective and highly specific catalyst for the

Helen M. Wilks; Keith W. Hart; Raymond Feeney; Cameron R. Dunn; Hilary Muirhead; William N. Chia; David A. Barstow; Tony Atkinson; Anthony R. Clarke; J. John Holbrook

1988-01-01

19

Lactate Dehydrogenase Undergoes a Substantial Structural Change to Bind its Substrate  

E-print Network

Lactate Dehydrogenase Undergoes a Substantial Structural Change to Bind its Substrate Linlin Qiu and thermodynamics of the formation of a very early ternary binding intermediate formed when lactate dehydrogenase is formed in lactate dehydrogenase (LDH). We focus on how the enzymeÁligand encounter complex is formed

Callender, Robert

20

Acta Cryst. (2000). D56, 8183 Lee et al. Lactate dehydrogenase 81 crystallization papers  

E-print Network

Acta Cryst. (2000). D56, 81±83 Lee et al. Lactate dehydrogenase 81 crystallization papers Acta Crystallographica Section D Biological Crystallography ISSN 0907-4449 Lactate dehydrogenase from of Crystallography Printed in Denmark ± all rights reserved l(+)-Lactate dehydrogenase (LDH) is a key enzyme

Suh, Se Won

21

Mutants of Escherichia coli deficient in the fermentative lactate dehydrogenase  

SciTech Connect

Mutants of Escherichia coli deficient in the fermentative NAD-linked lactate dehydrogenase (ldh) have been isolated. These mutants showed no growth defects under anaerobic conditions unless present together with a defect in pyruvate formate lyase (pfl). Double mutants (pfl ldh) were unable to grow anaerobically on glucose or other sugars even when supplemented with acetate, whereas pfl mutants can do so. The ldh mutation was found to map at 30.5 min on the E. coli chromosome. The ldh mutant FMJ39 showed no detectable lactate dehydrogenase activity and produced no lactic acid from glucose under anaerobic conditions as estimated by in vivo nuclear magnetic resonance measurements. We also found that in wild-type strains the fermentative lactate dehydrogenase was conjointly induced by anaerobic conditions and an acidic pH. Despite previous findings that phosphate concentrations affect the proportion of lactic acid produced during fermentation, we were unable to find any intrinsic effect of phosphate on lactate dehydrogenase activity, apart from the buffering effect of this ion.

Mat-Jan, F.; Alam, K.Y.; Clark, D.P. (Southern Illinois Univ., Carbondale (USA))

1989-01-01

22

Lactate dehydrogenase regulation of the metmyoglobin reducing system to improve color stability of bovine muscles through lactate enhancement  

E-print Network

The primary objectives of this research were to characterize the involvement of lactate dehydrogenase (LDH) in color stability of physiologically different bovine muscles, and to investigate the influence of lactate enhancement on the myoglobin...

Kim, Yuan Hwan

2009-05-15

23

Stability of lactate dehydrogenases. I. Chemical modification of lysines.  

PubMed

The lysine residues of lactate dehydrogenase (L-lactate: NAD+ oxidoreductase, EC 1.1.1.27) can be amidinated by methyl-4-hydroxy-3-nitrobenzimidate to introduce nitrophenolate anions. This modification results in lowered thermal stability, as does acetylation. The conversion of these groups into uncharged aminophenol groups without further modification of the enzyme itself stabilizes the enzymes from pig heart and muscle and from chicken muscle, as does acetamidination, but the unusually stable enzyme from chicken heart reverts only to the stability of the native form. The results allow for the following conclusions. Destabilization is brought about at many points at the surface of lactate dehydrogenases by neutralization of positive charges. Stabilization, in contrast, is concluded to be due to modification of one lysine at position 241 of the sequence. This lysine must have been changed to arginine during the evolution of heart-type lactate dehydrogenases in going from lower to higher reptiles. This exchange has been conserved in the enzymes from the hearts of birds and therefore the enzyme from chicken heart is very stable and cannot further be stabilized by modification of lysines. From X-ray structure analysis, the stabilization by exchange of Arg for Lys at position 241 or by amidination is explained by the formation of additional ion pairs with aspartic acid57 of the Q-related subunits. PMID:6793082

Müller, J

1981-07-28

24

Lactate dehydrogenase mutants of Streptococcus mutans: isolation and preliminary characterization.  

PubMed Central

Mutants of Streptococcus mutans were isolated which lack the enzyme activity L (+)-lactate dehydrogenase. Reversion studies indicate that the genetic defects are in the structural gene for the enzyme. The mutants produce less titratable acid from glucose, adhere better to hydroxyapatite, and accumulate more plaque when grown in the presence of sucrose than does the parent strain. These findings suggest a possible use for the mutants as effector strains in the replacement therapy of dental caries. Images PMID:30695

Hillman, J D

1978-01-01

25

Anti-peptide antibodies differentiate between plasmodial lactate dehydrogenases  

Microsoft Academic Search

Malaria lactate dehydrogenase, a glycolytic enzyme, is a malaria diagnostic target in lateral flow immunochromatographic rapid diagnostic tests. Recombinant Plasmodium yoelii LDH was cloned into the pET-28a vector, expressed and the expressed protein purified from a Ni-NTA affinity matrix. A pan-malarial LDH antibody directed against a common malaria LDH peptide (APGKSDKEWNRDDLL) and two anti-peptide antibodies, each targeting a unique Plasmodium

Ramona Hurdayal; Ikechukwu Achilonu; David Choveaux; Theresa H. T. Coetzer; J. P. Dean Goldring

2010-01-01

26

Probing the Role of Dynamics in Hydride Transfer Catalyzed by Lactate Dehydrogenase  

E-print Network

Probing the Role of Dynamics in Hydride Transfer Catalyzed by Lactate Dehydrogenase Nickolay Zhadin, Bronx, New York 10461 ABSTRACT The dynamic nature of the interconversion of pyruvate to lactate as catalyzed by lactate dehydrogenase (LDH) is characterized by laser-induced temperature jump relaxation

Callender, Robert

27

The Cost of Capturing Prey: Measuring Largemouth Bass Foraging Activity using Glycolytic Enzymes (Lactate Dehydrogenase)  

E-print Network

(Lactate Dehydrogenase) By Trevor M. Selch A thesis submitted in partial fulfillment of the requirements Enzymes (Lactate Dehydrogenase) This thesis is approved as a creditable and independent investigation THE COST OF CAPTURING PREY: MEASURING LARGEMOUTH BASS FORAGING ACTIVITY USING GLYCOLYTIC ENZYMES (LACTATE

28

Nuclear Lactate Dehydrogenase: Bridging Central Metabolism to Epigenetic Modifications in Mammalian Cellular Systems  

E-print Network

1 Nuclear Lactate Dehydrogenase: Bridging Central Metabolism to Epigenetic Modifications isoform of lactate dehydrogenase (LDH) and its role in modifying histone acetylation states due to its to serve as substrate for nuclear LDH. This enzyme catalyzes the interconversion of pyruvate to lactate

Appanna, Vasu

29

NAD-dependent lactate dehydrogenase catalyses the first step in respiratory utilization of lactate by Lactococcus lactis?  

PubMed Central

Lactococcus lactis can undergo respiration when hemin is added to an aerobic culture. The most distinctive feature of lactococcal respiration is that lactate could be consumed in the stationary phase concomitantly with the rapid accumulation of diacetyl and acetoin. However, the enzyme responsible for lactate utilization in this process has not yet been identified. As genes for fermentative NAD-dependent l-lactate dehydrogenase (l-nLDH) and potential electron transport chain (ETC)-related NAD-independent l-LDH (l-iLDH) exist in L. lactis, the activities of these enzymes were measured in this study using crude cell extracts prepared from respiratory and fermentation cultures. Further studies were conducted with purified preparations of recombinant LDH homologous proteins. The results showed that l-iLDH activity was hardly detected in both crude cell extracts and purified l-iLDH homologous protein while l-nLDH activity was very significant. This suggested that l-iLDHs were inactive in lactate utilization. The results of kinetic analyses and the effects of activator, inhibitor, substrate and product concentrations on the reaction equilibrium showed that l-nLDH was much more prone to catalyze the pyruvate reduction reaction but could reverse its role provided that the concentrations of NADH and pyruvate were extremely low while NAD and lactate were abundant. Metabolite analysis in respiratory culture revealed that the cellular status in the stationary phase was beneficial for l-nLDH to catalyze lactate oxidation. The factors accounting for the respiration- and stationary phase-dependent lactate utilization in L. lactis are discussed here. PMID:24251099

Zhao, Rui; Zheng, Sui; Duan, Cuicui; Liu, Fei; Yang, Lijie; Huo, Guicheng

2013-01-01

30

Detection of Necrosis by Release of Lactate Dehydrogenase (LDH) Activity  

PubMed Central

Summary Apoptosis and necrosis are two major forms of cell death observed in normal and disease pathologies. Although there are many assays for detection of apoptosis, relatively few assays are available for measuring necrosis. A key signature for necrotic cells is the permeabilization of plasma membrane. This event can be quantified in tissue culture settings by measuring the release of the enzyme lactate dehydrogenase (LDH). When combined with other methods, measuring LDH release is a useful method for detection of necrosis. In this chapter, we describe the step-by-step procedure for detection of LDH release from necrotic cells using a microtiter plate based colorimetric absorbance assay. PMID:23397389

Chan, Francis Ka-Ming; Moriwaki, Kenta; De Rosa, María José

2013-01-01

31

Activity patterns of phosphofructokinase, glyceraldehydephosphate dehydrogenase, lactate dehydrogenase and malate dehydrogenase in microdissected fast and slow fibres from rabbit psoas and soleus muscle  

Microsoft Academic Search

Methods for standardized determination of phosphofructokinase (PFK), glyceraldehydephosphate dehydrogenase (GAPDH), lactate dehydrogenase (LDH) and malate dehydrogenase (MDH) activities in nanogram samples of microdissected single fibres of rabbit psoas and soleus muscle are described. Fast and slow fibres in soleus muscle show lower absolute activities of these enzymes than the respective fibre types in psoas muscle. Slow fibres represent a more

Cornelia Spamer; Dirk Pette

1977-01-01

32

Measurement of the enzymes lactate dehydrogenase and creatine kinase using reflectance spectroscopy and reagent strips.  

PubMed Central

Two new methods for the assay of total activities of lactate dehydrogenase and creatine kinase are described, in which the enzyme activities are measured from a solid-state reagent strip during a kinetic reaction, the reaction being monitored in the ultra-violet region of the spectrum by reflectance spectroscopy. The performances of these methods are evaluated, and compared to conventional "wet" chemistry methods. The solid-phase reagent methods demonstrated precision and accuracy acceptable for diagnostic purposes, and were easy to use by trained operators. PMID:6655069

Stevens, J F; Tsang, W; Newall, R G

1983-01-01

33

Peroxisomal lactate dehydrogenase is generated by translational readthrough in mammals  

PubMed Central

Translational readthrough gives rise to low abundance proteins with C-terminal extensions beyond the stop codon. To identify functional translational readthrough, we estimated the readthrough propensity (RTP) of all stop codon contexts of the human genome by a new regression model in silico, identified a nucleotide consensus motif for high RTP by using this model, and analyzed all readthrough extensions in silico with a new predictor for peroxisomal targeting signal type 1 (PTS1). Lactate dehydrogenase B (LDHB) showed the highest combined RTP and PTS1 probability. Experimentally we show that at least 1.6% of the total cellular LDHB is targeted to the peroxisome by a conserved hidden PTS1. The readthrough-extended lactate dehydrogenase subunit LDHBx can also co-import LDHA, the other LDH subunit, into peroxisomes. Peroxisomal LDH is conserved in mammals and likely contributes to redox equivalent regeneration in peroxisomes. DOI: http://dx.doi.org/10.7554/eLife.03640.001 PMID:25247702

Schueren, Fabian; Lingner, Thomas; George, Rosemol; Hofhuis, Julia; Dickel, Corinna; Gärtner, Jutta; Thoms, Sven

2014-01-01

34

Mitochondrial Lactate Dehydrogenase Is Involved in Oxidative-Energy Metabolism in Human Astrocytoma  

E-print Network

Mitochondrial Lactate Dehydrogenase Is Involved in Oxidative-Energy Metabolism in Human Astrocytoma and Biochemistry, Laurentian University, Sudbury, Ontario, Canada Abstract Lactate has long been regarded as an end (CCF-STTG1) to consume lactate and to generate ATP via oxidative phosphorylation. 13 C-NMR and HPLC

Appanna, Vasu

35

Not only osmoprotectant: betaine increased lactate dehydrogenase activity and L-lactate production in lactobacilli.  

PubMed

Lactobacilli are commonly used for industrial production of polymer-grade L-lactic acid. The present study tested the Tween 80 alternative betaine in L-lactate production by several industrial lactobacilli. In flask fermentation of Lactobacillus casei, Lactobacillus buchneri, Lactobacillus lactis and Lactobacillus rhamnosus, the betaine addition (2g/l) had similar osmoprotectant effect with Tween 80 but had increased the lactate dehydrogenase activities and L-lactate production than Tween 80 control. In fed-batch fermentation of L. casei, betaine supplementation improved the L-lactic acid titer to 190 g/l, the yield to 95.5% (g L-lactic acid/g glucose), the productivity to 2.6g/lh, and the optical purity to 97.0%. The results demonstrated that supplementation of Tween 80 alternative - betaine in the fermentation medium is feasible for industrial l-lactic acid fermentation by lactobacilli, which will improve the lactate production but will not increase the process costs and modify any process conditions. PMID:24035452

Zou, Huibin; Wu, Zaiqiang; Xian, Mo; Liu, Hui; Cheng, Tao; Cao, Yujin

2013-11-01

36

The expression of lactate dehydrogenase in Zea mays seedlings under hypoxic and anoxic conditions  

E-print Network

in anoxic maize root tips, catalyzed by lactate dehydrogenase (LDH), prior to sustained ethanolic fermentation. The initial lactic acid production may contribute to cytoplasmic acidosis. However, barley roots, when deprived of oxygen, exhibit a long term...

MacAlpine, David Michael

1995-01-01

37

Acetate Utilization in Lactococcus lactis Deficient in Lactate Dehydrogenase: a Rescue Pathway for Maintaining Redox Balance  

PubMed Central

Acetate was shown to improve glucose fermentation in Lactococcus lactis deficient in lactate dehydrogenase. 13C and 1H nuclear magnetic resonance studies using [2-13C]glucose and [2-13C]acetate as substrates demonstrated that acetate was exclusively converted to ethanol. This novel pathway provides an alternative route for NAD+ regeneration in the absence of lactate dehydrogenase. PMID:10464231

Hols, Pascal; Ramos, Ana; Hugenholtz, Jeroen; Delcour, Jean; de Vos, Willem M.; Santos, Helena; Kleerebezem, Michiel

1999-01-01

38

Characterization of Hypoxically Inducible Lactate Dehydrogenase in Maize.  

PubMed Central

Oxygen deprivation induces a wide variety of genes, but the most extensively studied are those encoding enzymes of the glycolytic pathway. Lactate dehydrogenase (LDH, EC 1.1.1.27) activity increases up to 3.5-fold in maize (Zea mays L.) roots during several days of hypoxic induction. This increase in activity is accompanied by a decrease in in vitro enzyme stability. LDH activity in aerobic root extracts has an in vitro half-life of 240 min, decreasing to 100 min in 72-h hypoxically induced plant root extracts. The increase in enzyme activity during hypoxic induction is the result of increased protein levels, which correlate with increased transcript levels. Two ldh transcripts of 1.3 and 1.7 kb are induced, with maximum levels reached by 8 and 24 h, respectively. This suggests that the two ldh genes are differentially regulated. Treatment with the protein synthesis inhibitor cycloheximide does not preclude ldh induction during the first few hours of hypoxic stress, suggesting that new protein synthesis may not be essential for elevation of ldh transcript levels under hypoxic conditions. The rapid and substantial increase in ldh mRNA levels under hypoxic conditions and in the presence of cycloheximide suggests that the ldh gene may be valuable in analyzing the hypoxic signal transduction pathway. PMID:12226430

Christopher, M. E.; Good, A. G.

1996-01-01

39

Mitochondrial and plasma membrane lactate transporter and lactate dehydrogenase isoform expression in breast cancer cell lines  

PubMed Central

We hypothesized that dysregulation of lactate/pyruvate (monocarboxylate) transporters (MCT) and lactate dehydrogenase (LDH) isoforms contribute to the Warburg effect in cancer. Therefore, we assayed for the expression levels and the localizations of MCT (1, 2, and 4), and LDH (A and B) isoforms in breast cancer cell lines MCF-7 and MDA-MB-231 and compared results with those from a control, untransformed primary breast cell line, HMEC 184. Remarkably, MCT1 is not expressed in MDA-MB-231, but MCT1 is expressed in MCF-7 cells, where its abundance is less than in control HMEC 184 cells. When present in HMEC 184 and MCF-7 cells, MCT1 is localized to the plasma membrane. MCT2 and MCT4 were expressed in all the cell lines studied. MCT4 expression was higher in MDA-MB-231 compared with MCF-7 and HMEC 184 cells, whereas MCT2 abundance was higher in MCF-7 compared with MDA-MB-231 and HMEC 184 cells. Unlike MCT1, MCT2 and MCT4 were localized in mitochondria in addition to the plasma membrane. LDHA and LDHB were expressed in all the cell-lines, but abundances were higher in the two cancer cell lines than in the control cells. MCF-7 cells expressed mainly LDHB, while MDA-MB-231 and control cells expressed mainly LDHA. LDH isoforms were localized in mitochondria in addition to the cytosol. These localization patterns were the same in cancerous and control cell lines. In conclusion, MCT and LDH isoforms have distinct expression patterns in two breast cancer cell lines. These differences may contribute to divergent lactate dynamics and oxidative capacities in these cells, and offer possibilities for targeting cancer cells. PMID:21177384

Hussien, Rajaa

2011-01-01

40

Sequence analysis of teleost retina-specific lactate dehydrogenase C: evolutionary implications for the vertebrate lactate dehydrogenase gene family.  

PubMed Central

At least two gene duplication events have led to the three lactate dehydrogenase (LDH; EC 1.1.1.27) isozymes (LDH-A, LDH-B, and LDH-C) of chordates. The prevailing model for the evolution of the LDH loci involves duplication of a primordial LDH locus near the origin of vertebrates, giving rise to Ldh-A and Ldh-B. A third locus, designated Ldh-C, is expressed in the spermatocytes of mammals and a single family of birds and in the eye or liver tissues of teleost fishes. Ldh-C might have arisen independently in these taxa as duplications of either Ldh-A or Ldh-B. Several authors have challenged this traditional hypothesis on the basis of amino acid sequence and immunological similarity of the three LDH isozymes. They suggest that the primordial LDH gene was duplicated to form Ldh-C and a locus that later gave rise to Ldh-A and Ldh-B. We have differentiated between these hypotheses by determining the cDNA sequence of the retina-specific LDH-C from a teleost, Fundulus heteroclitus. On the basis of amino acid sequence similarity, we conclude that the LDH-C isozymes in fish and mammals are not orthologous but derive from independent gene duplications. Furthermore, our phylogenetic analyses support previous hypotheses that teleost Ldh-C is derived from a duplication of the Ldh-B locus. PMID:8419929

Quattro, J M; Woods, H A; Powers, D A

1993-01-01

41

Energy landscape of the Michaelis complex of lactate dehydrogenase: relationship to catalytic mechanism.  

PubMed

Lactate dehydrogenase (LDH) catalyzes the interconversion between pyruvate and lactate with nicotinamide adenine dinucleotide (NAD) as a cofactor. Using isotope-edited difference Fourier transform infrared spectroscopy on the "live" reaction mixture (LDH·NADH·pyruvate ? LDH·NAD(+)·lactate) for the wild-type protein and a mutant with an impaired catalytic efficiency, a set of interconverting conformational substates within the pyruvate side of the Michaelis complex tied to chemical activity is revealed. The important structural features of these substates include (1) electronic orbital overlap between pyruvate's C2?O bond and the nicotinamide ring of NADH, as shown from the observation of a delocalized vibrational mode involving motions from both moieties, and (2) a characteristic hydrogen bond distance between the pyruvate C2?O group and active site residues, as shown by the observation of at least four C2?O stretch bands indicating varying degrees of C2?O bond polarization. These structural features form a critical part of the expected reaction coordinate along the reaction path, and the ability to quantitatively determine them as well as the substate population ratios in the Michaelis complex provides a unique opportunity to probe the structure-activity relationship in LDH catalysis. The various substates have a strong variance in their propensity toward on enzyme chemistry. Our results suggest a physical mechanism for understanding the LDH-catalyzed chemistry in which the bulk of the rate enhancement can be viewed as arising from a stochastic search through an available phase space that, in the enzyme system, involves a restricted ensemble of more reactive conformational substates as compared to the same chemistry in solution. PMID:24576110

Peng, Huo-Lei; Deng, Hua; Dyer, R Brian; Callender, Robert

2014-03-25

42

Energy Landscape of the Michaelis Complex of Lactate Dehydrogenase: Relationship to Catalytic Mechanism  

PubMed Central

Lactate dehydrogenase (LDH) catalyzes the interconversion between pyruvate and lactate with nicotinamide adenine dinucleotide (NAD) as a cofactor. Using isotope-edited difference Fourier transform infrared spectroscopy on the “live” reaction mixture (LDH·NADH·pyruvate ? LDH·NAD+·lactate) for the wild-type protein and a mutant with an impaired catalytic efficiency, a set of interconverting conformational substates within the pyruvate side of the Michaelis complex tied to chemical activity is revealed. The important structural features of these substates include (1) electronic orbital overlap between pyruvate’s C2=O bond and the nicotinamide ring of NADH, as shown from the observation of a delocalized vibrational mode involving motions from both moieties, and (2) a characteristic hydrogen bond distance between the pyruvate C2=O group and active site residues, as shown by the observation of at least four C2=O stretch bands indicating varying degrees of C2=O bond polarization. These structural features form a critical part of the expected reaction coordinate along the reaction path, and the ability to quantitatively determine them as well as the substate population ratios in the Michaelis complex provides a unique opportunity to probe the structure–activity relationship in LDH catalysis. The various substates have a strong variance in their propensity toward on enzyme chemistry. Our results suggest a physical mechanism for understanding the LDH-catalyzed chemistry in which the bulk of the rate enhancement can be viewed as arising from a stochastic search through an available phase space that, in the enzyme system, involves a restricted ensemble of more reactive conformational substates as compared to the same chemistry in solution. PMID:24576110

2015-01-01

43

Direct Evidence of Catalytic Heterogeneity in Lactate Dehydrogenase by Temperature Jump Infrared Spectroscopy  

PubMed Central

Protein conformational heterogeneity and dynamics are known to play an important role in enzyme catalysis, but their influence has been difficult to observe directly. We have studied the effects of heterogeneity in the catalytic reaction of pig heart lactate dehydrogenase using isotope edited infrared spectroscopy, laser-induced temperature jump relaxation, and kinetic modeling. The isotope edited infrared spectrum reveals the presence of multiple reactive conformations of pyruvate bound to the enzyme, with three major reactive populations having substrate C2 carbonyl stretches at 1686, 1679, and 1674 cm?1, respectively. The temperature jump relaxation measurements and kinetic modeling indicate that these substates form a heterogeneous branched reaction pathway, and each substate catalyzes the conversion of pyruvate to lactate with a different rate. Furthermore, the rate of hydride transfer is inversely correlated with the frequency of the C2 carbonyl stretch (the rate increases as the frequency decreases), consistent with the relationship between the frequency of this mode and the polarization of the bond, which determines its reactivity toward hydride transfer. The enzyme does not appear to be optimized to use the fastest pathway preferentially but rather accesses multiple pathways in a search process that often selects slower ones. These results provide further support for a dynamic view of enzyme catalysis where the role of the enzyme is not just to bring reactants together but also to guide the conformational search for chemically competent interactions. PMID:25149276

Reddish, Michael; Peng, Huo-Lei; Deng, Hua; Panwar, Kunal S.; Callender, Robert; Dyer, R. Brian

2014-01-01

44

Incorporating expression data in metabolic modeling: a case study of lactate dehydrogenase  

E-print Network

Integrating biological information from different sources to understand cellular processes is an important problem in systems biology. We use data from mRNA expression arrays and chemical kinetics to formulate a metabolic model relevant to K562 erythroleukemia cells. MAP kinase pathway activation alters the expression of metabolic enzymes in K562 cells. Our array data show changes in expression of lactate dehydrogenase (LDH) isoforms after treatment with phorbol 12-myristate 13-acetate (PMA), which activates MAP kinase signaling. We model the change in lactate production which occurs when the MAP kinase pathway is activated, using a non-equilibrium, chemical-kinetic model of homolactic fermentation. In particular, we examine the role of LDH isoforms, which catalyze the conversion of pyruvate to lactate. Changes in the isoform ratio are not the primary determinant of the production of lactate. Rather, the total concentration of LDH controls the lactate concentration.

Joshua Downer; Joel R. Sevinsky; Natalie G. Ahn; Katheryn A. Resing; M. D. Betterton

2005-11-12

45

Ligand Binding and Protein Dynamics in Lactate Dehydrogenase J. R. Exequiel T. Pineda,* Robert Callender,y  

E-print Network

Ligand Binding and Protein Dynamics in Lactate Dehydrogenase J. R. Exequiel T. Pineda,* Robert Einstein College of Medicine, Bronx, New York ABSTRACT Recent experimental studies suggest that lactate an enzyme binds its substrate at the earliest stages of binding. We specifically ex- amine the lactate

Callender, Robert

46

LACTIC ACID PRODUCTION BY SACCHAROMYCES CEREVISIAE EXPRESSING A RHIZOPUS ORYZAE LACTATE DEHYDROGENASE GENE  

Technology Transfer Automated Retrieval System (TEKTRAN)

This work demonstrates the first example of a fungal LDH expressed in yeast. A L(+)-lactate dehydrogenase gene, ldhA, from the filamentous fungus Rhizopus oryzae was modified to be expressed under control of the Saccharomyces cerevisiae adhl promoter and terminator, then placed in a 2 micron contai...

47

Lactate dehydrogenase isoenzymes in prostate and testis of wild animals and some histological remarks  

Microsoft Academic Search

The lactate dehydrogenase (LDH) isoenzyme pattern of prostate and of testis of 27 wild animals showed that in several species more than 5 isoenzymes are detected, with electrophoretic mobilities different to those found in humans. The LDH-X band, found in testis from mature humans is also observed in the testis of wild animals. However, in several species more than one

K. Van Camp; M. Van Sande

1988-01-01

48

Increasing the heme-dependent respiratory efficiency of Lactococcus lactis by inhibition of lactate dehydrogenase.  

PubMed

The discovery of heme-induced respiration in Lactococcus lactis has radically improved the industrial processes used for the biomass production of this species. Here, we show that inhibition of the lactate dehydrogenase activity of L. lactis during growth under respiration-permissive conditions can stimulate aerobic respiration, thereby increasing not only growth efficiency but also the robustness of this organism. PMID:23064338

Arioli, Stefania; Zambelli, Daniele; Guglielmetti, Simone; De Noni, Ivano; Pedersen, Martin B; Pedersen, Per Dedenroth; Dal Bello, Fabio; Mora, Diego

2013-01-01

49

THE ROLE OF LIPID PHASE STRUCTURE IN THE INTERACTION OF LACTATE DEHYDROGENASE WITH PHOSPHATIDYLSERINE. ACTIVITY STUDIES  

Microsoft Academic Search

Lactate dehydrogenase is one of the enzymes of the glycolytic path. It has been shown to be able to bind in vitro to cellular membranes. The presence of anionic phospholipids induces changes in the catalytic properties of the enzyme similar to those found when the enzyme is bound to natural membranes. In this study, a nonionic detergent (Tween 20), at

GRZEGORZ TERLECKI; JAN GUTOWICZ

2002-01-01

50

Resting Oxygen Consumption Varies among Lactate Dehydrogenase Genotypes in the Sow Bug, Porcellio scaber  

Microsoft Academic Search

Laboratory studies of respiration in the sow bug, Porcellio scaber, reveal that respiration rates are related to genetic variation at the lactate dehydrogenase (Ldh) locus. In population samples taken from Burlington, North Carolina and Pacific Grove, California, respiration rates differed among Ldh genotypes, but not among genotypes at the other enzyme polymorphisms. In both population samples, the respiration rate of

Jeffry B. Mitton; Patrick A. Carter; Adam Digiacomo

1997-01-01

51

Intracellular lactate dehydrogenase concentration as an index of cytotoxicity in rat hepatocyte primary culture  

Microsoft Academic Search

In searching for a reliable index for cytotoxicity testing in rat hepatocyte primary culture, lactate dehydrogenase (LDH) concentrations in lysates of attached hepatocytes and LDH released into the culture medium were compared under conditions of exposure to various dosages of sodium chloride, sodium salicylate, R-warfarin, acetaminophen, phenylbutazone, and furosemide (frusemide). The amount of intracellular LDH was assessed by inducing the

Eddie S. Chao; Deborah Dunbar; Laurence S. Kaminsky

1988-01-01

52

Modification of Rhizopus lactate dehydrogenase for improved resistance to fructose 1,6-bisphosphate  

Technology Transfer Automated Retrieval System (TEKTRAN)

Rhizopus oryzae is frequently used for fermentative production of lactic acid. We determined that one of the key enzymes, lactate dehydrogenase (LDH), involved in synthesis of lactic acid by R. oryzae was significantly inhibited by fructose 1,6-bisphosphate (FBP) at physiological concentrations. Thi...

53

Relationship of lactate dehydrogenase activity to body measurements of Angus x Charolais cows and calves  

Technology Transfer Automated Retrieval System (TEKTRAN)

Objectives were to examine 1) relationships between lactate dehydrogenase (LDH) activity and body measurements of grazing beef cows, and 2) the association between maternal LDH activity in late gestation and subsequent calf birth weight (BRW), hip height (HH) at weaning, and adjusted weaning weight ...

54

Relationship of lactate dehydrogenase activity with body measeurements of Angus x Charolais cows and calves  

Technology Transfer Automated Retrieval System (TEKTRAN)

Angus x Charolais cows (n = 87) and their Angus-sired, spring-born calves (n = 86) were utilized to examine relationships between lactate dehydrogenase (LDH) activity and body measurements of beef cows; and the relationship between maternal LDH activity in late gestation and subsequent calf birth we...

55

Acta Cryst. (1970). B26, 998 Crystallographic Studies on Lactate Dehydrogenase at -75C  

E-print Network

, Indiana 47907, U.S.A. (Received14 July 1969) Crystals of lactate dehydrogenase (LDH) were frozen of radiation damage for frozen crystals was tenfold less than for crystals at room temperature. The physical properties of frozen crystals are dis- cussed. Analysis of 3"5A data collected at -75°C for native LDH

Rossmann, Michael G.

1970-01-01

56

Interdemic variation in haematocrit and lactate dehydrogenase in the African cyprinid Barbus  

E-print Network

National Park, Uganda differed in traits related to aerobic and anaerobic metabolic potential. Haematocrit was measured as an index of blood oxygen-carrying capacity, and tissue activities and isozyme composition of lactate dehydrogenase (LDH) were measured as indices of tissue anaerobic capacity. To address whether site

Chapman, Lauren J.

57

Metabolic Imaging: A link between Lactate Dehydrogenase A, Lactate and Tumor Phenotype  

PubMed Central

Purpose We compared the metabolic profiles and the association between LDH-A expression and lactate production in two isogenic murine breast cancer cell lines and tumors (67NR and 4T1). These cell lines were derived from a single mammary tumor and have different growth and metabolic phenotypes. Experimental Design LDH-A expression, lactate concentration, glucose utilization and oxygen consumption were measured in cells, and the potential relationship between tumor lactate levels (measured by magnetic resonance spectroscopic imaging (MRSI)) and tumor glucose utilization (measured by [18F] 2-deoxy-2-fluoro-D-glucose positron emission tomography ([18F]FDG-PET)) was assessed in orthotopic breast tumors derived from these cell lines. Results We show a substantial difference in LDH-A expression between 67NR and 4T1 cells under normoxia and hypoxia. We also show that small orthotopic 4T1 tumors generate tenfold more lactate than corresponding 67NR tumors. The high lactate levels in small primary 4T1 tumors are associated with intense pimonidazole staining (a hypoxia indicator). Less intense hypoxia staining was observed in the larger 67NR tumors, and is consistent with the gradual increase and plateau of lactate concentration in enlarging 67NR tumors. Conclusions Lactate-MRSI has a greater dynamic range than [18F]FDG-PET and may be a more sensitive measure with which to evaluate the aggressive and metastatic potential of primary breast tumors. PMID:21844011

Thakur, Sunitha B.; Vider, Jelena; Russell, James; Blasberg, Ronald; Koutcher, Jason A.

2014-01-01

58

The core of allosteric motion in Thermus caldophilus L-lactate dehydrogenase.  

PubMed

For Thermus caldophilus L-lactate dehydrogenase (TcLDH), fructose 1,6-bisphosphate (FBP) reduced the pyruvate S(0.5) value 10(3)-fold and increased the V(max) value 4-fold at 30 °C and pH 7.0, indicating that TcLDH has a much more T state-sided allosteric equilibrium than Thermus thermophilus L-lactate dehydrogenase, which has only two amino acid replacements, A154G and H179Y. The inactive (T) and active (R) state structures of TcLDH were determined at 1.8 and 2.0 Å resolution, respectively. The structures indicated that two mobile regions, MR1 (positions 172-185) and MR2 (positions 211-221), form a compact core for allosteric motion, and His(179) of MR1 forms constitutive hydrogen bonds with MR2. The Q4(R) mutation, which comprises the L67E, H68D, E178K, and A235R replacements, increased V(max) 4-fold but reduced pyruvate S(0.5) only 5-fold in the reaction without FBP. In contrast, the P2 mutation, comprising the R173Q and R216L replacements, did not markedly increase V(max), but 10(2)-reduced pyruvate S(0.5), and additively increased the FBP-independent activity of the Q4(R) enzyme. The two types of mutation consistently increased the thermal stability of the enzyme. The MR1-MR2 area is a positively charged cluster, and its center approaches another positively charged cluster (N domain cluster) across the Q-axis subunit interface by 5 Å, when the enzyme undergoes the T to R transition. Structural and kinetic analyses thus revealed the simple and unique allosteric machinery of TcLDH, where the MR1-MR2 area pivotally moves during the allosteric motion and mediates the allosteric equilibrium through electrostatic repulsion within the protein molecule. PMID:25258319

Ikehara, Yoko; Arai, Kazuhito; Furukawa, Nayuta; Ohno, Tadashi; Miyake, Tatsuya; Fushinobu, Shinya; Nakajima, Masahiro; Miyanaga, Akimasa; Taguchi, Hayao

2014-11-01

59

Understanding interactions of functionalized nanoparticles with proteins: a case study on lactate dehydrogenase.  

PubMed

Nanomaterials in biological solutions are known to interact with proteins and have been documented to affect protein function, such as enzyme activity. Understanding the interactions of nanoparticles with biological components at the molecular level will allow for rational designs of nanomaterials for use in medical technologies. Here we present the first detailed molecular mechanics model of functionalized gold nanoparticle (NP) interacting with an enzyme (L-lactate dehydrogenase (LDH) enzyme). Molecular dynamics (MD) simulations of the response of LDH to the NP binding demonstrate that although atomic motions (dynamics) of the main chain exhibit only a minor response to the binding, the dynamics of side chains are significantly constrained in all four active sites that predict alteration in kinetic properties of the enzyme. It is also demonstrated that the 5 nm gold NPs cause a decrease in the maximal velocity of the enzyme reaction (V(max)) and a trend towards a reduced affinity (increased K(m)) for the ?-NAD binding site, while pyruvate enzyme kinetics (K(m) and V(max)) are not significantly altered in the presence of the gold NPs. These results demonstrate that modeling of NP:protein interactions can be used to understand alterations in protein function. PMID:24591162

Stueker, Oliver; Ortega, Van A; Goss, Greg G; Stepanova, Maria

2014-05-28

60

Lactate dehydrogenase activity in bovine and porcine muscle as influenced by electrical stimulation, aging, freezing, thawing and heating  

E-print Network

LACTATE DEHYDROGENASE ACTIVITY IN BOVINF. AND PORCINE MUSCLE AS INFLUENCED BY ELECTRICAL STIMULATION, AGING, FREEZING, THA&v'ING AiVD HEATING A Thesis by SHAREN SUE COLLINS Submitted to the Graduate College of Texas ARM University in partial... fulfillment of the requirements for the degree of MASTER OF SCIENCE May 1987 Major Subject: Animal Science LACTATE DEHYDROGENASE ACTIVITY IN BOVINE AND PORCINE MUSCLE AS INFLUENCED BY ELECTRICAL STIMULATION, AGING, FREEZING, THAWING AND HEATING A Thesis...

Collins, Sharen Sue

1987-01-01

61

Characteristics of Monoclonal Antibodies to the Lactate Dehydrogenase-Elevating Virus  

Microsoft Academic Search

Summary Spleen cells, from BALB\\/c mice that had been infected with lactate dehydrogenase-elevating virus (LDV) for 6 days and that had or had not been previously immunized with glutaraldehyde-inactivated LDV, were fused with NS-1 myeloma cells. The fusion frequency was at least 10 times higher than with spleen cells of normal or chronically infected mice. Only 1 of 297 wells

John T. Harty; Stephen P. K. Chan; Peter G. W. Plagemann

1987-01-01

62

Lactate Dehydrogenase Undergoes a Substantial Structural Change to Bind its Substrate  

Microsoft Academic Search

Employing temperature-jump relaxation spectroscopy, we investigate the kinetics and thermodynamics of the formation of a very early ternary binding intermediate formed when lactate dehydrogenase (LDH) binds a substrate mimic on its way to forming the productive LDH\\/NADH·substrate Michaelis complex. Temperature-jump scans show two distinct submillisecond processes are involved in the formation of this ternary binding intermediate, called the encounter complex

Linlin Qiu; Miriam Gulotta; Robert Callender

2007-01-01

63

Effect of water activity on inactivation of Listeria monocytogenes and lactate dehydrogenase during high pressure processing  

Microsoft Academic Search

The aim of this study was to investigate the effect of water activity (aw) on the inactivation of Listeria monocytogenes and lactate dehydrogenase (LDH) during high pressure processing (HPP). For microbial inactivation lyophilized cells of L. monocytogenes 19,115 were left dry or were suspended in 10 ml of 0.1% peptone water, 10 ml of glycerol, or mixtures of glycerol and peptone water.

Melinda M. Hayman; Gilles K. Kouassi; Ramaswamy C. Anantheswaran; John D. Floros; Stephen J. Knabel

2008-01-01

64

The use of molecular modelling in the understanding of configurational specificity (R or S) in asymmetric reactions catalyzed by Saccharomyces cerevisiae or isolated dehydrogenases  

Microsoft Academic Search

This method gives a general ideal how to use crystallographic information of enzymes to understand reactions catalyzed by these biocatalysts, commonly used by biochemists to produce chiral products. The interactions of three acetoacetic esters with the enzymes L-lactate dehydrogenase and alcohol dehydrogenase were studied through molecular modelling computer program. These artificial substrates have been widely used to produce chiral synthons.

Ricardo de Souza Pereira; Fernando Pavão; Glaucius Oliva

1998-01-01

65

Cloning, nucleotide sequence, and transcriptional analysis of the Pediococcus acidilactici L-(+)-lactate dehydrogenase gene.  

PubMed Central

Recombinant plasmids containing the Pediococcus acidilactici L-(+)-lactate dehydrogenase gene (ldhL) were isolated by complementing for growth under anaerobiosis of an Escherichia coli lactate dehydrogenase-pyruvate formate lyase double mutant. The nucleotide sequence of the ldhL gene predicted a protein of 323 amino acids showing significant similarity with other bacterial L-(+)-lactate dehydrogenases and especially with that of Lactobacillus plantarum. The ldhL transcription start points in P. acidilactici were defined by primer extension, and the promoter sequence was identified as TCAAT-(17 bp)-TATAAT. This sequence is closely related to the consensus sequence of vegetative promoters from gram-positive bacteria as well as from E. coli. Northern analysis of P. acidilactici RNA showed a 1.1-kb ldhL transcript whose abundance is growth rate regulated. These data, together with the presence of a putative rho-independent transcriptional terminator, suggest that ldhL is expressed as a monocistronic transcript in P. acidilactici. PMID:7887607

Garmyn, D; Ferain, T; Bernard, N; Hols, P; Delcour, J

1995-01-01

66

Molecular and Kinetic Characterization of Babesia microti Gray Strain Lactate Dehydrogenase as a Potential Drug Target  

PubMed Central

Babesia microti is an emerging zoonotic protozoan organism that causes “malaria-like” symptoms that can be fatal in immunocompromised people. Owing to lack of specific therapeutic regiment against the disease, we cloned and characterized B. microti lactate dehydrogenase (BmLDH) as a potential molecular drug receptor. The in vitro kinetic properties of BmLDH enzyme was evaluated using nicotinamide adenine dinucleotide (NAD+) as a co-factor and lactate as a substrate. Inhibitory assay was also done using gossypol as BmLDH inhibitor to determine the inhibitory concentration 50 (IC50). The result showed that the 0.99 kbp BmLDH gene codes for a barely soluble 36 kDa protein (332 amino acids) localized in both the cytoplasm and nucleus of the parasite. In vitro enzyme kinetic studies further revealed that BmLDH is an active enzyme with a high catalytic efficiency at optimal pH of 10.2. The Km values of NAD+ and lactate were 8.7 ± 0.57 mM and 99.9 ± 22.33 mM, respectively. The IC50 value for gossypol was 0.345 ?M, while at 2.5 ?M, gossypol caused 100% inhibition of BmLDH catalytic activity. These findings, therefore, provide initial evidence that BmLDH could be a potential drug target, although further in vivo studies are needed to validate the practical application of lactate dehydrogenase inhibitors against B. microti infection. PMID:25125971

Vudriko, Patrick; Masatani, Tatsunori; Cao, Shinuo; Terkawi, Mohamad Alla; Kamyingkird, Ketsarin; Mousa, Ahmed A; Adjou Moumouni, Paul F; Nishikawa, Yoshifumi; Xuan, Xuenan

2014-01-01

67

Clinical research on neuroblastoma based on serum lactate dehydrogenase.  

PubMed

In recent years, more and more scholars tend to study neuroblastoma (NB) since it possesses increasing morbidity, but lack of effective treatment. This paper aims to investigate variation and clinical significance of the neuron-specific enolase (NSE) and lactic dehydrogenase (LDH) level in serum of children with NB before and after Auto Peripheral Blood Stem Cell Transplantation (APBSCT). A total of 90 children with NB from various hospitals were included in this research, and we analyzed the relationship between levels of NSE and LDH and the change of disease by comparing the two levels before and after APBSCT treatment. The results indicated that the positive rate of NSE in serum was high before treatment, and the levels of NSE and LDH were remarkably higher than those when the treatment was valid; after comprehensive treatment of chemotherapy, excision and radiotherapy, there was a significant difference of NSE and LDH levels in serum between children with complete remission (CR) and those with partial remission (PR); however, no significant differences of NSE and LDH levels were found among children in progressive stage compared to before treatment. It is believed that NSE and LDH levels are associated to the recurrence and treatment effect of NB, proving that both can reflect tumor load, therefore they can be taken as the auxiliary indicators for monitoring curative effects of NB treatment. PMID:25864749

Pang, Q M; Li, K; Ma, L J; Sun, R P

2015-01-01

68

Modulation of DNA polymerases ?, ? and ? by lactate dehydrogenase and 3-phosphoglycerate kinase 1 Enzymes: DNA polymerases ?, ?, and ?, deoxynucleoside-triphosphate:DNA deoxynucleotidyltransferase (DNA-directed) (EC 2.7.7.7); lactate dehydrogenase, l-lactate:NAD + oxidoreductase (EC 1.1.1.27); 3-phosphoglycerate kinase, ATP:3-phospho- d-glycerate 1-phosphotransferase (EC 2.7.2.3.). 1  

Microsoft Academic Search

Literature documents that glycolytic enzymes (among them lactate dehydrogenase and 3-phosphoglycerate kinase) can reside in nuclei of mammalian cells and exert functions in DNA replication, transcription and DNA repair, in addition to their role as catalysts in the cytoplasm. Transfer of glycolytic enzymes to cell nuclei requires modification, for example phosphorylation. We studied the effects of phosphorylated lactate dehydrogenase and

Odilia Popanda; Gabriele Fox; Heinz Walter Thielmann

1998-01-01

69

Spinal Fluid Lactate Dehydrogenase Level Differentiates between Structural and Metabolic Etiologies of Altered Mental Status in Children  

PubMed Central

Objective Altered mental status is a common cause of intensive care unit admission in children. Differentiating structural causes of altered mental status from metabolic etiologies is of utmost importance in diagnostic approach and management of the patients. Among many biomarkers proposed to help stratifying patients with altered mental status, spinal fluid lactate dehydrogenase appears to be the most promising biomarker to predict cellular necrosis. Materials & Methods In this cross sectional study we measured spinal fluid level of lactate dehydrogenase in children 2 months to 12 years of age admitted to a single center intensive care unit over one year. Spinal fluid level of lactate dehydrogenase in 40 pediatric cases of febrile seizure was also determined as the control group. Results The study group included 35 boys (58.3%) and 25 girls (41.7%). Their mean age was 2.7+/-3 years and their mean spinal fluid lactate dehydrogenase level was 613.8+/-190.4 units/liter. The control group included 24 boys (55.8%) and 19 girls (44.2%). Their mean age was 1.3+/-1.2 years and their mean spinal fluid lactate dehydrogenase level was 18.9+/-7.5 units/liter. The mean spinal fluid lactate dehydrogenase level in children with abnormal head CT scan was 246.3+/-351.5 units/liter compared to 164.5+/-705.7 in those with normal CT scan of the head (p=0.001). Conclusion Spinal fluid lactate dehydrogenase level is useful in differentiating structural and metabolic causes of altered mental status in children. PMID:25767536

KHOSROSHAHI, Nahid; ALIZADEH, Parastoo; KHOSRAVI, Mehdi; SALAMATI, Peyman; KAMRANI, Kamyar

2015-01-01

70

Multichannel Simultaneous Determination of Activities of Lactate Dehydrogenase  

SciTech Connect

It is very important to find the best conditions for some enzymes to do the best catalysis in current pharmaceutical industries. Based on the results above, we could say that this set-up could be widely used in finding the optimal condition for best enzyme activity of a certain enzyme. Instead of looking for the best condition for enzyme activity by doing many similar reactions repeatedly, we can complete this assignment with just one run if we could apply enough conditions.

Ma, L.

2000-09-12

71

Lactate dehydrogenase is the key enzyme for pneumococcal pyruvate metabolism and pneumococcal survival in blood.  

PubMed

Streptococcus pneumoniae is a fermentative microorganism and causes serious diseases in humans, including otitis media, bacteremia, meningitis, and pneumonia. However, the mechanisms enabling pneumococcal survival in the host and causing disease in different tissues are incompletely understood. The available evidence indicates a strong link between the central metabolism and pneumococcal virulence. To further our knowledge on pneumococcal virulence, we investigated the role of lactate dehydrogenase (LDH), which converts pyruvate to lactate and is an essential enzyme for redox balance, in the pneumococcal central metabolism and virulence using an isogenic ldh mutant. Loss of LDH led to a dramatic reduction of the growth rate, pinpointing the key role of this enzyme in fermentative metabolism. The pattern of end products was altered, and lactate production was totally blocked. The fermentation profile was confirmed by in vivo nuclear magnetic resonance (NMR) measurements of glucose metabolism in nongrowing cell suspensions of the ldh mutant. In this strain, a bottleneck in the fermentative steps is evident from the accumulation of pyruvate, revealing LDH as the most efficient enzyme in pyruvate conversion. An increase in ethanol production was also observed, indicating that in the absence of LDH the redox balance is maintained through alcohol dehydrogenase activity. We also found that the absence of LDH renders the pneumococci avirulent after intravenous infection and leads to a significant reduction in virulence in a model of pneumonia that develops after intranasal infection, likely due to a decrease in energy generation and virulence gene expression. PMID:25245810

Gaspar, Paula; Al-Bayati, Firas A Y; Andrew, Peter W; Neves, Ana Rute; Yesilkaya, Hasan

2014-12-01

72

Cationic Surfactant-Based Colorimetric Detection of Plasmodium Lactate Dehydrogenase, a Biomarker for Malaria, Using the Specific DNA Aptamer  

PubMed Central

A simple, sensitive, and selective colorimetric biosensor for the detection of the malarial biomarkers Plasmodium vivax lactate dehydrogenase (PvLDH) and Plasmodium falciparum LDH (PfLDH) was demonstrated using the pL1 aptamer as the recognition element and gold nanoparticles (AuNPs) as probes. The proposed method is based on the aggregation of AuNPs using hexadecyltrimethylammonium bromide (CTAB). The AuNPs exhibited a sensitive color change from red to blue, which could be seen directly with the naked eye and was monitored using UV-visible absorption spectroscopy and transmission electron microscopy (TEM). The reaction conditions were optimized to obtain the maximum color intensity. PvLDH and PfLDH were discernible with a detection limit of 1.25 pM and 2.94 pM, respectively. The applicability of the proposed biosensor was also examined in commercially available human serum. PMID:24992632

Lee, Seonghwan; Manjunatha, D H; Jeon, Weejeong; Ban, Changill

2014-01-01

73

An electrochemical biosensor with nanointerface for lactate detection based on lactate dehydrogenase immobilized on zinc oxide nanorods.  

PubMed

Hepatic immaturity is observed particularly in children whose age is under three, when the lactate concentration is greater than the normal level in blood. An electrochemical lactate biosensor was developed by immobilizing lactate dehydrogenase (LDH) on to ZnO nanorods at pH 7.4 via chitosan. Growth of polycrystalline ZnO nanorods towards (101) plane was confirmed using XRD. The FE-SEM study revealed the formation of ZnO nanorods with an aspect ratio of 3.24. Immobilization of LDH on ZnO nanorods was confirmed using FTIR spectra and surface coverage. Electrochemical studies were carried out through cyclic voltammetry and amperometry using three electrode system with Au/NanoZnO/LDH as working electrode, Ag/AgCl in 0.1 M KCl as reference electrode and Pt wire as counter electrode. The sensitivity of the biosensor was found to be 1.832 ?A ?mol(-1) L exhibiting linearity 0.2-0.8 ?mol L(-1) with the detection and quantification limits of 4.73 and 15.75 nmol L(-1) respectively. The response time of Au/NanoZnO/LDH bioelectrode was found to be <1 s. Prediction band for net current was framed to enhance specificity. Michaelis-Menten constant (KM(app)) and maximum rate (Imax) values for immobilized LDH were found to be 0.38 ?mol L(-1) and 2.798 ?A respectively. Repeatability and reproducibility of LDH biosensor were also reported. PMID:24231089

Nesakumar, Noel; Thandavan, Kavitha; Sethuraman, Swaminathan; Krishnan, Uma Maheswari; Rayappan, John Bosco Balaguru

2014-01-15

74

Resting oxygen consumption varies among lactate dehydrogenase genotypes in the sow bug, Porcellio scaber  

PubMed Central

Laboratory studies of respiration in the sow bug, Porcellio scaber, reveal that respiration rates are related to genetic variation at the lactate dehydrogenase (Ldh) locus. In population samples taken from Burlington, North Carolina and Pacific Grove, California, respiration rates differed among Ldh genotypes, but not among genotypes at the other enzyme polymorphisms. In both population samples, the respiration rate of the common Ldh homozygote exceeded the respiration rate of the heterozygote by more than 50 per cent. The differences in respiration rates are consistent with previously reported viability differentials at the Ldh polymorphism.

Mitton, J. B.; Carter, P. A.; DiGiacomo, A.

1997-01-01

75

Identification of 3,6-disubstituted dihydropyrones as inhibitors of human lactate dehydrogenase.  

PubMed

A series of 3,6-disubstituted dihydropyrones were identified as inhibitors of human lactate dehydrogenase (LDH)-A. Structure activity relationships were explored and a series of 6,6-spiro analogs led to improvements in LDHA potency (IC50 <350 nM). An X-ray crystal structure of an improved compound bound to human LDHA was obtained and it illustrated additional opportunities to enhance the potency of these compounds, resulting in the identification of 51 (IC50=30 nM). PMID:25467161

Fauber, Benjamin P; Dragovich, Peter S; Chen, Jinhua; Corson, Laura B; Ding, Charles Z; Eigenbrot, Charles; Labadie, Sharada; Malek, Shiva; Peterson, David; Purkey, Hans E; Robarge, Kirk; Sideris, Steve; Ultsch, Mark; Wei, BinQing; Yen, Ivana; Yue, Qin; Zhou, Aihe

2014-12-15

76

Interaction Between Nanoparticulate Anatase TiO 2 and Lactate Dehydrogenase  

Microsoft Academic Search

In order to study the mechanisms underlying the effects of TiO2 nanoparticles on lactate dehydrogenase (LDH, EC1.1.1.27), Institute of Cancer Research region mice were injected with nanoparticulate\\u000a anatase TiO2 (5 nm) of various doses into the abdominal cavity daily for 14 days. We then examined LDH activity in vivo and in vitro and\\u000a direct evident for interaction between nanoparticulate anatase TiO2 and

Yanmei Duan; Na Li; Chao Liu; Huiting Liu; Yaling Cui; Han Wang; Fashui Hong

2010-01-01

77

The maximum activities of hexokinase, phosphorylase, phosphofructokinase, glycerol phosphate dehydrogenases, lactate dehydrogenase, octopine dehydrogenase, phosphoenolpyruvate carboxykinase, nucleoside diphosphatekinase, glutamate-oxaloacetate transaminase and arginine kinase in relation to carbohydrate utilization in muscles from marine invertebrates.  

PubMed Central

Comparison of the activities of hexokinase, phosphorylase and phosphofructokinase in muscles from marine invertebrates indicates that they can be divided into three groups. First, the activities of the three enzymes are low in coelenterate muscles, catch muscles of molluscs and muscles of echinoderms; this indicates a low rate of carbohydrate (and energy) utilization by these muscles. Secondly, high activities of phosphorylase and phosphofructokinase relative to those of hexokinase are found in, for example, lobster abdominal and scallop snap muscles; this indicates that these muscles depend largely on anaerobic degradation of glycogen for energy production. Thirdly, high activities of hexokinase are found in the radular muscles of prosobranch molluscs and the fin muscles of squids; this indicates a high capacity for glucose utilization, which is consistent with the high activities of enzymes of the tricarboxylic acid cycle in these muscles [Alp, Newsholme & Zammit (1976) Biochem. J. 154, 689-700]. 2. The activities of lactate dehydrogenase, octopine dehydrogenase, phosphoenolpyruvate carboxykinase, cytosolic and mitochondrial glycerol 3-phosphate dehydrogenase and glutamate-oxaloacetate transaminase were measured in order to provide a qualitative indication of the importance of different processes for oxidation of glycolytically formed NADH. The muscles are divided into four groups: those that have a high activity of lactate dehydrogenase relative to the activities of phosphofructokinase (e.g. crustacean muscles); those that have high activities of octopine dehydrogenase but low activities of lactate dehydrogenase (e.g. scallop snap muscle); those that have moderate activities of both lactate dehydrogenase and octopine dehydrogenase (radular muscles of prosobranchs), and those that have low activities of both lactate dehydrogenase and octopine dehydrogenase, but which possess activities of phosphoenolpyruvate carboxykinase (oyster adductor muscles). It is suggested that, under anaerobic conditions, muscles of marine invertebrates form lactate and/or octopine or succinate (or similar end product) according to the activities of the enzymes present in the muscles (see above). The muscles investigated possess low activities of cytosolic glycerol 3-phosphate dehydrogenase, which indicates that glycerol phosphate formation is quantitatively unimportant under anaerobic conditions, and low activities of mitochondrial glycerol phosphate dehydrogenase, which indicates that the glycerol phosphate cycle is unimportant in the re-oxidation of glycolytically produced NADH in these muscles under aerobic conditions. Conversely, high activities of glutamate-oxaloacetate transaminase are present in some muscles, which indicates that the malate-aspartate cycle may be important in oxidation of glycolytically produced NADH under aerobic conditions. 3. High activities of nucleoside diphosphate kinase were found in muscles that function for prolonged periods under anaerobic conditions (e.g... PMID:13783

Zammit, V A; Newsholme, E A

1976-01-01

78

Placental 17 beta-hydroxysteroid oxidoreductase, lactate dehydrogenase and malate dehydrogenase during the latter half of pregnancy in the mouse.  

PubMed

The specific activity of 17 beta-hydroxysteroid oxidoreductase (17-HOR) with estradiol-17 beta (E2), estrone (E1) and testosterone (T), as well as that of lactate dehydrogenase (LDH) and malate dehydrogenase (MDH) were measured in homogenates of CF-1 mouse placenta during the latter half of pregnancy. 17-HOR activity with E2 and T increased over 100-fold between days 9 and 12, and 3- to 4-fold between days 15 and 19, with no further change to day 21. In contrast, activity with E1 increased 39-fold between days 9 and 12, 3.8-fold between days 15 and 19 but then decreased between days 19 and 21. The E2/T activity ratio was constant while the E2/E1 ratio increased between days 9 and 21. LDH increased 2-fold between days 9 and 12 with no further increase to day 19. MDH was constant from day 9 to 19. Activity with E2 was inhibited by T, 5 alpha-dihydrotestosterone (5 alpha-DHT) and DHA but not by E1, androstenedione (A) or 20 alpha-dihydroprogesterone. Activity with T was inhibited by E2, 5 alpha-DHT and DHA, but not by A. In contrast, activity with E1 was inhibited by A and DHA but not by E2, T or 5 alpha-DHT. The results suggest placental 17-HOR is developmentally regulated. Although the results are also suggestive of multiple forms of 17-HOR, a single enzyme with an ordered kinetic mechanism cannot be ruled out. PMID:8338791

Blomquist, C H; Hensleigh, H C; Block, D L; Feeney, L A

1993-07-01

79

Novel control of lactate dehydrogenase from the freeze tolerant wood frog: role of posttranslational modifications  

PubMed Central

Lactate dehydrogenase (LDH), the terminal enzyme of anaerobic glycolysis, plays a crucial role both in sustaining glycolytic ATP production under oxygen-limiting conditions and in facilitating the catabolism of accumulated lactate when stress conditions are relieved. In this study, the effects on LDH of in vivo freezing and dehydration stresses (both of which impose hypoxia/anoxia stress on tissues) were examined in skeletal muscle of the freeze-tolerant wood frog, Rana sylvatica. LDH from muscle of control, frozen and dehydrated wood frogs was purified to homogeneity in a two-step process. The kinetic properties and stability of purified LDH were analyzed, revealing no significant differences in Vmax, Km and I50 values between control and frozen LDH. However, control and dehydrated LDH differed significantly in Km values for pyruvate, lactate, and NAD, I50 urea, and in temperature, glucose, and urea effects on these parameters. The possibility that posttranslational modification of LDH was responsible for the stable differences in enzyme behavior between control and dehydrated states was assessed using ProQ diamond staining to detect phosphorylation and immunoblotting to detect acetylation, methylation, ubiquitination, SUMOylation and nitrosylation of the enzyme. LDH from muscle of dehydrated wood frogs showed significantly lower levels of acetylation, providing one of the first demonstrations of a potential role for protein acetylation in the stress-responsive control of a metabolic enzyme. PMID:23638346

Abboud, Jean

2013-01-01

80

Regulation of transplacental virus infection by developmental and immunological factors: studies with lactate dehydrogenase-elevating virus  

Microsoft Academic Search

Placental and fetal infections with lactate dehydrogenase-elevating virus (LDV) were determined by virus titration, indirect fluorescence antibody (IFA), and in situ hybridization with cDNA probes. Experiments were designed to determine the effects of gestational age, timing of maternal LDV infection, and immunological (antibody and cytokine) factors on mouse placental and fetal LDV infection. Virus infection of the placenta was detected

Thomas R Haven; Raymond R. R Rowland; Peter G. W Plagemann; Grace H. W Wong; Sarahann E Bradley; William A Cafruny

1996-01-01

81

THE EFFECT OF EXERCISE ON PLASMA ACTIVITIES OF LACTATE DEHYDROGENASE AND CREATINE KINASE IN RED-TAILED HAWKS (Buteo jamaicensis)  

Microsoft Academic Search

Plasma activities of lactate dehydrogenase (LD) and creatine kinase (CK) have been used as diagnostic indicators of muscle fitness and damage, respectively, in mammals. Activities of these enzymes were measured in three groups of red-tailed hawks (Buteojamaicensis) differing in flight capability (trained, untrained, and disabled) to determine whether their plasma enzyme activities were indicative of muscle fitness and flight training

SHANNON T. KNUTH; SUSAN B. CHAPLIN

82

Renal lactate dehydrogenase (LDH) isoenzyme pattern in short-term experimental obstructive nephropathy.  

PubMed

The lactate dehydrogenase isoenzyme pattern of different zones of rat kidney has been investigated at different stages after unilateral ureteral obstruction. In the obstructed kidney, the cortical tissue showed an early (3 days) progressive shift toward a cathodic zymogram; at 7 days LDH-5 isoenzyme was the predominant fraction (44.7 +/- 2.5). In the outer medullary tissue a similar change was observed even though middle fractions were always quite evident. No significant changes were found in the inner medullary tissue. The observed enzymatic abnormalities are related to renal hypoxia and to the occurrence of a less differentiated nephronic cell population. They represent a metabolic marker of the morphologic and functional alterations which occur in obstructive nephropathy. PMID:447486

Cestonaro, G; Emanuelli, G; Calcamuggi, G; Anfossi, G; Gatti, G

1979-07-01

83

Genomic structure and promoter activity of the human testis lactate dehydrogenase gene.  

PubMed

The structure of the gene encoding the human testis-specific isozyme of lactate dehydrogenase (LDH) has been characterized and a regulatory region identified by promoter activity. The single-copy ldh-c gene has two alternative 5' noncoding exons and seven coding exons comprising an approximately 40-kb locus. The gene does not contain the canonical TATA or CAAT promoter sequences, and ribonuclease protection experiments suggest multiple transcription start sites. In the present study an immortalized murine germ cell line was used to detect promoter activity driven by 5' sequence of human ldh-c with lacZ as the reporter gene. Reporter gene activity was nondetectable when promoter constructs were transfected into nongerminal cells. PMID:8318584

Cooker, L A; Brooke, C D; Kumari, M; Hofmann, M C; Millán, J L; Goldberg, E

1993-06-01

84

Fever and high lactate dehydrogenase in HIV-positive patients from the Antilles and Surinam: histoplasmosis?  

PubMed

We describe four cases of HIV-positive patients, two from Surinam, one from the Dutch Antilles and one from Nigeria, who presented with a febrile illness and a high lactate dehydrogenase plasma level. In all four, the diagnosis of disseminated histoplasmosis was made, in three of them by liver biopsy. Two patients had retinal abnormalities compatible with a systemic fungal infection. Three patients were treated successfully with antifungal agents. One patient died. Between 2000 and 2006, only 14 patients with HIV have been found to have histoplasmosis in the Netherlands. Although histoplasmosis is not endemic in the Netherlands, physicians are more likely to see cases because of a growing number of HIV -positive immigrants from endemic regions. PMID:16990694

Peters, E J G; Kauffmann, R H; Blok, P

2006-09-01

85

Identification of substituted 3-hydroxy-2-mercaptocyclohex-2-enones as potent inhibitors of human lactate dehydrogenase.  

PubMed

A novel class of 3-hydroxy-2-mercaptocyclohex-2-enone-containing inhibitors of human lactate dehydrogenase (LDH) was identified through a high-throughput screening approach. Biochemical and surface plasmon resonance experiments performed with a screening hit (LDHA IC50=1.7 ?M) indicated that the compound specifically associated with human LDHA in a manner that required simultaneous binding of the NADH co-factor. Structural variation of this screening hit resulted in significant improvements in LDHA biochemical inhibition activity (best IC50=0.18 ?M). Two crystal structures of optimized compounds bound to human LDHA were obtained and explained many of the observed structure-activity relationships. In addition, an optimized inhibitor exhibited good pharmacokinetic properties after oral administration to rats (F=45%). PMID:25037916

Dragovich, Peter S; Fauber, Benjamin P; Boggs, Jason; Chen, Jinhua; Corson, Laura B; Ding, Charles Z; Eigenbrot, Charles; Ge, HongXiu; Giannetti, Anthony M; Hunsaker, Thomas; Labadie, Sharada; Li, Chiho; Liu, Yichin; Liu, Yingchun; Ma, Shuguang; Malek, Shiva; Peterson, David; Pitts, Keith E; Purkey, Hans E; Robarge, Kirk; Salphati, Laurent; Sideris, Steve; Ultsch, Mark; VanderPorten, Erica; Wang, Jing; Wei, BinQing; Xu, Qing; Yen, Ivana; Yue, Qin; Zhang, Huihui; Zhang, Xuying; Zhou, Aihe

2014-08-15

86

In situ Regeneration of NADH via Lipoamide Dehydrogenase-catalyzed Electron Transfer Reaction Evidenced by Spectroelectrochemistry  

SciTech Connect

NAD/NADH is a coenzyme found in all living cells, carrying electrons from one reaction to another. We report on characterizations of in situ regeneration of NADH via lipoamide dehydrogenase (LD)-catalyzed electron transfer reaction to regenerate NADH using UV-vis spectroelectrochemistry. The Michaelis-Menten constant (Km) and maximum velocity (Vmax) of NADH regeneration were measured as 0.80 {+-} 0.15 mM and 1.91 {+-} 0.09 {micro}M s-1 in a 1-mm thin-layer spectroelectrochemical cell using gold gauze as the working electrode at the applied potential -0.75 V (vs. Ag/AgCl). The electrocatalytic reduction of the NAD system was further coupled with the enzymatic conversion of pyruvate to lactate by lactate dehydrogenase to examine the coenzymatic activity of the regenerated NADH. Although the reproducible electrocatalytic reduction of NAD into NADH is known to be difficult compared to the electrocatalytic oxidation of NADH, our spectroelectrochemical results indicate that the in situ regeneration of NADH via LD-catalyzed electron transfer reaction is fast and sustainable and can be potentially applied to many NAD/NADH-dependent enzyme systems.

Tam, Tsz Kin; Chen, Baowei; Lei, Chenghong; Liu, Jun

2012-08-01

87

Addressable self-immobilization of lactate dehydrogenase across multiple length scales.  

PubMed

Successful nanobiotechnology implementation largely depends on control over the interfaces between inorganic materials and biological molecules. Controlling the orientations of biomolecules and their spatial arrangements on the surface may transform many technologies including sensors, to energy. Here, we demonstrate the self-organization of L-lactate dehydrogenase (LDH), which exhibits enhanced enzymatic activity and stability on a variety of gold surfaces ranging from nanoparticles to electrodes, by incorporating a gold-binding peptide tag (AuBP2) as the fusion partner for Bacillus stearothermophilus LDH (bsLDH). Binding kinetics and enzymatic assays verified orientation control of the enzyme on the gold surface through the genetically incorporated peptide tag. Finally, redox catalysis efficiency of the immobilized enzyme was detected using cyclic voltammetry analysis in enzyme-based biosensors for lactate detection as well as in biofuel cell energy systems as the anodic counterpart. Our results demonstrate that the LDH enzyme can be self-immobilized onto different gold substrates using the short peptide tag under a biologically friendly environment. Depending on the desired inorganic surface, the proposed peptide-mediated path could be extended to any surface to achieve single-step oriented enzyme immobilization for a wide range of applications. PMID:23386458

Cetinel, Sibel; Caliskan, H Burak; Yucesoy, Deniz T; Donatan, A Senem; Yuca, Esra; Urgen, Mustafa; Karaguler, Nevin G; Tamerler, Candan

2013-02-01

88

The importance of arginine 171 in substrate binding by Bacillus stearothermophilus lactate dehydrogenase.  

PubMed

A variant of lactate dehydrogenase from Bacillus stearothermophilus has been engineered by site-directed mutagenesis in which an active-site arginine residue at position 171 in the protein sequence is replaced by lysine. Replacement of this arginine by lysine has no effect on co-enzyme binding, a relatively small effect on the rate of turnover of the enzyme, but causes a 2000-fold increase in the Michaelis constant for pyruvate, a 6000-fold increase in the dissociation constant for oxamate and results in a Michaelis constant for lactate which is too high to measure. The decrease in binding energy for these carboxylate-containing substrates caused by this mutation is very large, around 5.5 kcal.mol-1 and in part, is explained by the small increase in the distance of a lysine-substrate carboxylate interaction at this site and the absence of the additional hydrogen bond from a two-point arginine-carboxylate interaction. Consistent with this last observation, the ability of this mutant enzyme to stabilize an NAD+-sulphite compound in its active site (an alternative enzyme-substrate complex which does not involve bifurcated bonding to arginine) is only reduced 14-fold. PMID:3606622

Hart, K W; Clarke, A R; Wigley, D B; Chia, W N; Barstow, D A; Atkinson, T; Holbrook, J J

1987-07-15

89

Molecular cloning and characterization of lactate dehydrogenase gene 1 in the silkworm, Bombyx mori.  

PubMed

Lactate dehydrogenase (LDH) catalyzes the reduction of pyruvate into lactate and constitutes a major checkpoint of anaerobic glycolysis. Recently, LDH draws a great deal of attention for its potential to be used as a novel diagnostic and therapeutic target for various diseases, including cancer and malaria. Insect LDHs have been mainly identified from fruit fly and mosquitoes, but not from silkworm. In this study, a novel LDH homologue, designated as BmLDH1, was firstly identified and characterized from the silkworm, Bombyx mori. The BmLDH1 cDNA contains an open reading frame of 996 bp, and encodes a protein of 331 amino acid residues with calculated molecular mass of 36 kDa. Sequence comparison showed BmLDH1 is a highly conserved protein. RT-PCR revealed BmLDH1 is transcripted in all tissues and in all developmental stages tested, indicating its essential roles for silkworm physiology and development. The BmLDH1 gene was subcloned and expressed in E. coli, and was further characterized by Western blot and Mass Spectrometry. The expressed protein contained the LDH activity, and could be inhibited by reduced glutathione in vitro. Immunofluoresence showed that the BmLDH1 was located in the cytoplasm. The cloned BmLDH1 sequence was deposited in the GenBank (accession number EU334850). PMID:20852941

Xia, Hengchuan; Wu, Chao; Xu, Qinggang; Shi, Jing; Feng, Fan; Chen, Keping; Yao, Qin; Wang, Yong; Wang, Lin

2011-03-01

90

Surface modification of silicon dioxide, silicon nitride and titanium oxynitride for lactate dehydrogenase immobilization.  

PubMed

Three different types of surface, silicon dioxide (SiO2), silicon nitride (Si3N4), and titanium oxynitride (TiON) were modified for lactate dehydrogenase (LDH) immobilization using (3-aminopropyl)triethoxysilane (APTES) to obtain an amino layer on each surface. The APTES modified surfaces can directly react with LDH via physical attachment. LDH can be chemically immobilized on those surfaces after incorporation with glutaraldehyde (GA) to obtain aldehyde layers of APTES-GA modified surfaces. The wetting properties, chemical bonding composition, and morphology of the modified surface were determined by contact angle (CA) measurement, Fourier transform infrared (FTIR) spectroscopy, and scanning electron microscopy (SEM), respectively. In this experiment, the immobilized protein content and LDH activity on each modified surface was used as an indicator of surface modification achievement. The results revealed that both the APTES and APTES-GA treatments successfully link the LDH molecule to those surfaces while retaining its activity. All types of tested surfaces modified with APTES-GA gave better LDH immobilizing efficiency than APTES, especially the SiO2 surface. In addition, the SiO2 surface offered the highest LDH immobilization among tested surfaces, with both APTES and APTES-GA modification. However, TiON and Si3N4 surfaces could be used as alternative candidate materials in the preparation of ion-sensitive field-effect transistor (ISFET) based biosensors, including lactate sensors using immobilized LDH on the ISFET surface. PMID:25108848

Saengdee, Pawasuth; Chaisriratanakul, Woraphan; Bunjongpru, Win; Sripumkhai, Witsaroot; Srisuwan, Awirut; Jeamsaksiri, Wutthinan; Hruanun, Charndet; Poyai, Amporn; Promptmas, Chamras

2015-05-15

91

Structural characterization of the apo form and NADH binary complex of human lactate dehydrogenase.  

PubMed

Lactate dehydrogenase A (LDH-A) is a key enzyme in anaerobic respiration that is predominantly found in skeletal muscle and catalyses the reversible conversion of pyruvate to lactate in the presence of NADH. LDH-A is overexpressed in many tumours and has therefore emerged as an attractive target for anticancer drug discovery. Crystal structures of human LDH-A in the presence of inhibitors have been described, but currently no structures of the apo or binary NADH-bound forms are available for any mammalian LDH-A. Here, the apo structure of human LDH-A was solved at a resolution of 2.1 Å in space group P4122. The active-site loop adopts an open conformation and the packing and crystallization conditions suggest that the crystal form is suitable for soaking experiments. The soaking potential was assessed with the cofactor NADH, which yielded a ligand-bound crystal structure in the absence of any inhibitors. The structures show that NADH binding induces small conformational changes in the active-site loop and an adjacent helix. A comparison with other eukaryotic apo LDH structures reveals the conservation of intra-loop interactions. The structures provide novel insight into cofactor binding and provide the foundation for soaking experiments with fragments and inhibitors. PMID:24816116

Dempster, Sally; Harper, Stephen; Moses, John E; Dreveny, Ingrid

2014-05-01

92

Evolutionary implications of the cDNA sequence of the single lactate dehydrogenase of a lamprey.  

PubMed Central

All vertebrates other than lampreys exhibit multiple loci encoding lactate dehydrogenase +ADL-LDH; (S)-lactate:NAD+ oxidoreductase, EC 1.1.1.27+BD. Of these loci, Ldh-A is expressed predominantly in muscle, Ldh-B is expressed predominantly in heart, and Ldh-C (where present) exhibits different tissue-restricted patterns of expression depending on the taxon. To examine the relationship of the single LDH of lampreys to other vertebrate LDHs, we have determined the cDNA sequence of the LDH of the sea lamprey Petromyzon marinus and compared it to previously published sequences from bacteria, plants, and vertebrates. The lamprey sequence exhibits a mixture of features of both LDH-A and LDH-B at the amino acid level that may account for its intermediate kinetic properties. Both distance and maximum parsimony analyses strongly reject a relationship of lamprey LDH with mammalian LDH-C but do not significantly distinguish among remaining alternative phylogenetic hypotheses. Evolutionary parsimony analyses suggest that the lamprey LDH is related to Ldh-A and that the single locus condition has arisen as a result of the loss of Ldh-B (prior to the appearance of Ldh-C). The collection of LDH sequences for further studies of the evolution of the vertebrate LDH gene family will be facilitated by the PCR approach that we have used to obtain the lamprey sequence. PMID:1542673

Stock, D W; Whitt, G S

1992-01-01

93

Physiological and fermentation properties of Bacillus coagulans and a mutant lacking fermentative lactate dehydrogenase activity.  

PubMed

Bacillus coagulans, a sporogenic lactic acid bacterium, grows optimally at 50-55 °C and produces lactic acid as the primary fermentation product from both hexoses and pentoses. The amount of fungal cellulases required for simultaneous saccharification and fermentation (SSF) at 55 °C was previously reported to be three to four times lower than for SSF at the optimum growth temperature for Saccharomyces cerevisiae of 35 °C. An ethanologenic B. coagulans is expected to lower the cellulase loading and production cost of cellulosic ethanol due to SSF at 55 °C. As a first step towards developing B. coagulans as an ethanologenic microbial biocatalyst, activity of the primary fermentation enzyme L-lactate dehydrogenase was removed by mutation (strain Suy27). Strain Suy27 produced ethanol as the main fermentation product from glucose during growth at pH 7.0 (0.33 g ethanol per g glucose fermented). Pyruvate dehydrogenase (PDH) and alcohol dehydrogenase (ADH) acting in series contributed to about 55% of the ethanol produced by this mutant while pyruvate formate lyase and ADH were responsible for the remainder. Due to the absence of PDH activity in B. coagulans during fermentative growth at pH 5.0, the l-ldh mutant failed to grow anaerobically at pH 5.0. Strain Suy27-13, a derivative of the l-ldh mutant strain Suy27, that produced PDH activity during anaerobic growth at pH 5.0 grew at this pH and also produced ethanol as the fermentation product (0.39 g per g glucose). These results show that construction of an ethanologenic B. coagulans requires optimal expression of PDH activity in addition to the removal of the LDH activity to support growth and ethanol production. PMID:20677017

Su, Yue; Rhee, Mun Su; Ingram, Lonnie O; Shanmugam, K T

2011-03-01

94

Development of an enzymatic chromatography strip with nicotinamide adenine dinucleotide-tetrazolium coupling reactions for quantitative l-lactate analysis.  

PubMed

In this study, a dry assay of l-lactate via the enzymatic chromatographic test (ECT) was developed. An l-lactate dehydrogenase plus a nicotinamide adenine dinucleotide (NADH) regeneration reaction were applied simultaneously. Various tetrazolium salts were screened to reveal visible color intensities capable of determining the lactate concentrations in the sample. The optimal analysis conditions were as follows. The diaphorase (0.5?l, 2(-6)U/?l) was immobilized in the test line of the ECT strip. Nitrotetrazolium blue chloride (5?l, 12mM), l-lactate dehydrogenase (1?l, 0.25U/?l), and NAD(+) (2?l, 1.5×10(-5)M) were added into the mobile phase (100?l) composed of 0.1% (w/w) Tween 20 in 10mM phosphate buffer (pH 9.0), and the process was left to run for 10min. This detection had a linear range of 0.039 to 5mM with a detection limit of 0.047mM. This quantitative analysis process for l-lactate was easy to operate with good stability and was proper for the point-of-care testing applications. PMID:25454507

Kan, Shu-Chen; Chang, Wei-Feng; Lan, Min-Chi; Lin, Chia-Chi; Lai, Wei-Shiang; Shieh, Chwen-Jen; Hsiung, Kuang-Pin; Liu, Yung-Chuan

2015-02-15

95

Rapid Detection of Lactate Dehydrogenase and Genotyping of Plasmodium falciparum in Saliva of Children with Acute Uncomplicated Malaria  

PubMed Central

The diagnosis of malaria in biological fluids other than blood using non-invasive, rapid diagnostic techniques provides a valuable approach in case management and epidemiological studies of malaria. Rapid detection of Plasmodium falciparum lactate dehydrogenase (pLDH) in saliva samples from 130 of 144 children with microscopically confirmed P. falciparum infection was evaluated using Optimal-IT dipsticks. Genotyping of parasites was also performed in saliva and blood samples from a cohort of patients by polymerase chain reaction (PCR). The sensitivity of the dipstick in whole-blood, whole-saliva, or supernatant of spun saliva samples was 97.2%, 77.9%, and 48.4%, respectively. The sensitivity of the dipstick in whole-saliva samples was significantly higher than in supernatant of spun saliva samples (P < 0.0005). Mutant T76 allele was detectable in 60% and 57% of blood and saliva samples, respectively. This finding shows rapid detection of pLDH in patient saliva. PMID:20810809

Gbotosho, Grace O.; Happi, Christian T.; Folarin, Onikepe; Keyamo, Ochuko; Sowunmi, Akintunde; Oduola, Ayoade M. J.

2010-01-01

96

Automated High Throughput Protein Crystallization Screening at Nanoliter Scale and Protein Structural Study on Lactate Dehydrogenase  

SciTech Connect

The purposes of our research were: (1) To develop an economical, easy to use, automated, high throughput system for large scale protein crystallization screening. (2) To develop a new protein crystallization method with high screening efficiency, low protein consumption and complete compatibility with high throughput screening system. (3) To determine the structure of lactate dehydrogenase complexed with NADH by x-ray protein crystallography to study its inherent structural properties. Firstly, we demonstrated large scale protein crystallization screening can be performed in a high throughput manner with low cost, easy operation. The overall system integrates liquid dispensing, crystallization and detection and serves as a whole solution to protein crystallization screening. The system can dispense protein and multiple different precipitants in nanoliter scale and in parallel. A new detection scheme, native fluorescence, has been developed in this system to form a two-detector system with a visible light detector for detecting protein crystallization screening results. This detection scheme has capability of eliminating common false positives by distinguishing protein crystals from inorganic crystals in a high throughput and non-destructive manner. The entire system from liquid dispensing, crystallization to crystal detection is essentially parallel, high throughput and compatible with automation. The system was successfully demonstrated by lysozyme crystallization screening. Secondly, we developed a new crystallization method with high screening efficiency, low protein consumption and compatibility with automation and high throughput. In this crystallization method, a gas permeable membrane is employed to achieve the gentle evaporation required by protein crystallization. Protein consumption is significantly reduced to nanoliter scale for each condition and thus permits exploring more conditions in a phase diagram for given amount of protein. In addition, evaporation rate can be controlled or adjusted in this method during the crystallization process to favor either nucleation or growing processes for optimizing crystallization process. The protein crystals gotten by this method were experimentally proven to possess high x-ray diffraction qualities. Finally, we crystallized human lactate dehydrogenase 1 (H4) complexed with NADH and determined its structure by x-ray crystallography. The structure of LDH/NADH displays a significantly different structural feature, compared with LDH/NADH/inhibitor ternary complex structure, that subunits in LDH/NADH complex show open conformation or two conformations on the active site while the subunits in LDH/NADH/inhibitor are all in close conformation. Multiple LDH/NADH crystals were obtained and used for x-ray diffraction experiments. Difference in subunit conformation was observed among the structures independently solved from multiple individual LDH/NADH crystals. Structural differences observed among crystals suggest the existence of multiple conformers in solution.

Fenglei Li

2006-08-09

97

Calpain-like and lactate dehydrogenase activities in the cerebrospinal fluid of patients with amyotrophic lateral sclerosis  

Microsoft Academic Search

Amyotrophic lateral sclerosis (ALS) is neurodegenerative disease accompanied by the death of motor neurons. To clarify the\\u000a role of calpain in the ALS-induced death of neurons, we measured the calpainlike and lactate dehydrogenase (LDH) activities\\u000a in the cerebrospinal fluid (CSF) of ALS patients and patients of a control group. The LDH activity measured in the CSF of\\u000a patients with ALS

L. V. Brylev; A. A. Yakovlev; M. V. Onufriev; M. N. Zakharova; I. A. Zavalishin; N. V. Gulyaeva

2007-01-01

98

A double mutant of highly purified Geobacillus stearothermophilus lactate dehydrogenase recognises l-mandelic acid as a substrate.  

PubMed

Lactate dehydrogenase from the thermophilic organism Geobacillus stearothermophilus (formerly Bacillus stearothermophilus) (bsLDH) has a crucial role in producing chirally pure hydroxyl compounds. ?-Hydroxy acids are used in many industrial situations, ranging from pharmaceutical to cosmetic dermatology products. One drawback of this enzyme is its limited substrate specificity. For instance, l-lactate dehydrogenase exhibits no detectable activity towards the large side chain of 2-hydroxy acid l-mandelic acid, an ?-hydroxy acid with anti-bacterial activity. Despite many attempts to engineer bsLDH to accept ?-hydroxy acid substrates, there have been no attempts to introduce the industrially important l-mandelic acid to bsLDH. Herein, we describe attempts to change the reactivity of bsLDH towards l-mandelic acid. Using the Insight II molecular modelling programme (except 'program' in computers) and protein engineering techniques, we have successfully introduced substantial mandelate dehydrogenase activity to the enzyme. Energy minimisation modelling studies suggested that two mutations, T246G and I240A, would allow the enzyme to utilise l-mandelic acid as a substrate. Genes encoding for the wild-type and mutant enzymes were constructed, and the resulting bsLDH proteins were overexpressed in Escherichia coli and purified using the TAGZyme system. Enzyme assays showed that insertion of this double mutation into highly purified bsLDH switched the substrate specificity from lactate to l-mandelic acid. PMID:23608509

Binay, Bar??; Sessions, Richard B; Karagüler, Nevin Gül

2013-05-10

99

Lactate Dehydrogenase Activity in Gingival Crevicular Fluid as a Marker in Orthodontic Tooth Movement  

PubMed Central

Objectives: This study aims at analyzing the changes in gingival crevicular fluid (GCF) lactate dehydrogenase (LDH) activity during orthodontic movement. Methods: Twenty patients all requiring first premolar extractions were selected and treated with conventional straight wire mechanotherapy. Canine retraction was done using 125 g Nitinol closed coil springs. The maxillary canine on one side served as the experimental site while the contralateral canine served as the control. GCF was collected from the canines before initiation of retraction, then 1 hour after initiating canine retraction, followed by 1 day, 7 days, 14 days and 21 days. GCF LDH levels were estimated and compared with the control site. Results The results revealed significantly higher LDH levels on the 7th, 14th and 21st day at the sites where orthodontic force had been applied. The levels also showed a significant increase from 0 hour to the 21st day. Peak levels were seen on 14th and 21st day following initiation of retraction. Conclusions: The study showed that LDH could be successfully estimated in the GCF and its increased levels could indicate active tooth movement, which could aid the clinician in monitoring active orthodontic tooth movement. PMID:21760863

Alfaqeeh, Sarah A; Anil, Sukumaran

2011-01-01

100

The effects of lanthanoid on the structure–function of lactate dehydrogenase from mice heart.  

PubMed

The activity of lactate dehydrogenase (LDH, EC1.1.1.27) is often changed upon inflammatory responses in animals. Lanthanoid (Ln) was shown to provoke various inflammatory responses both in rats and mice; however, the molecular mechanism by which Ln3+ exert its toxicity has not been completely understood, especially that we know little about the mechanism of the interaction between Ln with 4f electron shell and alternation valence and LDH. In this report, we investigated the mechanisms of LaCl3, CeCl3, and NdCl3 on LDH activity in vivo and in vitro. Our results showed that La3+, Ce3+, and Nd3+ could significantly activate LDH in vivo and in vitro; the order of activation was Ce3+>Nd3+> La3+>control. The affinity of LDH for Ce3+ was higher than Nd3+ and La3+; the saturated binding sites for Ce3+ on the LDH protein were 1.2 and for La3+ and Nd3+ 1.55. Ln3+ caused the reduction of exposure degree of cysteine or tryptophan/tyrosine of LDH, the increase of space resistance, and the enhancement of ?-helix in secondary structure of LDH, which was greatest in Ce3+ treatment, medium in Nd3+ treatment, and least in La3+ treatment. It implied that the changes of structure-function on LDH caused by Ln3+ were closely related to the characteristics of 4f electron shell and alternation valence in Ln. PMID:19396407

Li, Na; Duan, Yanmei; Zhou, Min; Liu, Chao; Hong, Fashui

2009-12-01

101

Glycoconjugates Influence Caspase Release and Minimize Production of Lactate Dehydrogenase upon Pathogen Exposure  

NASA Astrophysics Data System (ADS)

Many pathogens stimulate cell death of immune cells while promoting survival of pathogens. Early cell death is characterized by the release of mediators, namely Caspases (Cas). Infections caused by pathogens can be eradicated if immune cells could resist cell death and kill pathogens upon exposure. In this research, we studied whether glycoconjugates (GCs) influence Cas release and cytotoxicity upon pathogen damage. GC1 and GC3 constituted samples studied principally. Bacterial spores were used as a pathogen model. GC effects were determined "prior to," "during," and "following" pathogen exposure throughout phagocytosis. Cytotoxic damage was assessed by measuring lactate dehydrogenase (LDH) production. Our data show that GC3 was more effective than GC1 during phagocytosis. GC3 controls Cas release under all three exposure conditions. Minimum production of LDH was noticed in the "following" exposure condition compared to the "prior to" and "during" exposure conditions for GC1 and GC3. The present study provided the selection method of GC ligands bearing anti-cytotoxic and anti-apoptotic properties.

Eassa, Souzan; Tarasenko, Olga

2010-04-01

102

Detection of an intermediate late in the unfolding pathway of bacillus stearothermophilus lactate dehydrogenase  

NASA Astrophysics Data System (ADS)

In vivo proteins fold to form one active structure in minutes or seconds, ruling out the possibility that a polypeptide samples all possible conformational space during folding. We have used site directed mutagenesis to produce 15 single tryptophan containing mutants of Bacillus stearothermophilus lactate dehydrogenase (BS LDH) thus enabling the equilibria of unfolding to be seen from 15 defined positions. These mutant versions of BS LDH have the same X-ray structure as the wild type protein8. Previously Smith et al.11 had detected and assigned structures to 4 folding states. The first intermediate, a monomer with secondary and super secondary structure largely intact, is formed after the dimer dissociates at 0.55 M guanidinium hydrochloride (GuHCl). The second intermediate on the unfolding pathway is stable at 2.2 M GuHCl. It had been assumed previously that the transition from this molten-globule structure to the fully denatured form occurred as a single process. We have now identified a core folding motif. In this, helix (alpha) -1F forms a helix-sheet interaction with (beta) -K and (beta) -K has interactions with both (alpha) -2G and (alpha) -3G. This super secondary interaction forms the most stable folding motif in BS LDH and is lost at 2.8 M GuHCl leaving helix (alpha) -1F, (alpha) -2G, and (alpha) -3G which are stable until 3 M GuHCl.

Sleigh, Roger N.; Halsall, David J.; Clarke, Anthony R.; Behan-Martin, Moira; Holbrook, J. J.

1994-08-01

103

A single amino acid substitution deregulates a bacterial lactate dehydrogenase and stabilizes its tetrameric structure.  

PubMed

We have engineered a variant of the lactate dehydrogenase enzyme from Bacillus stearothermophilus in which arginine-173 at the proposed regulatory site has been replaced by glutamine. Like the wild-type enzyme, this mutant undergoes a reversible, protein-concentration-dependent subunit assembly, from dimer to tetramer. However, the mutant tetramer is much more stable (by a factor of 400) than the wild type and is destabilized rather than stabilized by binding the allosteric regulator, fructose 1,6-biphosphate (Fru-1,6-P2). The mutation has not significantly changed the catalytic properties of the dimer (Kd NADH, Km pyruvate, Ki oxamate and kcat), but has weakened the binding of Fru-1,6-P2 to both the dimeric and tetrameric forms of the enzyme and has almost abolished any stimulatory effect. We conclude that the Arg-173 residue in the wild-type enzyme is directly involved in the binding of Fru-1,6-P2, is important for allosteric communication with the active site, and, in part, regulates the state of quaternary structure through a charge-repulsion mechanism. PMID:3580377

Clarke, A R; Wigley, D B; Barstow, D A; Chia, W N; Atkinson, T; Holbrook, J J

1987-05-27

104

An atomic-resolution view of neofunctionalization in the evolution of apicomplexan lactate dehydrogenases  

PubMed Central

Malate and lactate dehydrogenases (MDH and LDH) are homologous, core metabolic enzymes that share a fold and catalytic mechanism yet possess strict specificity for their substrates. In the Apicomplexa, convergent evolution of an unusual LDH from MDH produced a difference in specificity exceeding 12 orders of magnitude. The mechanisms responsible for this extraordinary functional shift are currently unknown. Using ancestral protein resurrection, we find that specificity evolved in apicomplexan LDHs by classic neofunctionalization characterized by long-range epistasis, a promiscuous intermediate, and few gain-of-function mutations of large effect. In canonical MDHs and LDHs, a single residue in the active-site loop governs substrate specificity: Arg102 in MDHs and Gln102 in LDHs. During the evolution of the apicomplexan LDH, however, specificity switched via an insertion that shifted the position and identity of this ‘specificity residue’ to Trp107f. Residues far from the active site also determine specificity, as shown by the crystal structures of three ancestral proteins bracketing the key duplication event. This work provides an unprecedented atomic-resolution view of evolutionary trajectories creating a nascent enzymatic function. DOI: http://dx.doi.org/10.7554/eLife.02304.001 PMID:24966208

Boucher, Jeffrey I; Jacobowitz, Joseph R; Beckett, Brian C; Classen, Scott; Theobald, Douglas L

2014-01-01

105

Establishment of permanent chimerism in a lactate dehydrogenase-deficient mouse mutant with hemolytic anemia  

SciTech Connect

Pluripotent hemopoietic stem cell function was investigated in the homozygous muscle type lactate dehydrogenase (LDH-A) mutant mouse using bone marrow transplantation experiments. Hemopoietic tissues of LDH-A mutants showed a marked decreased in enzyme activity that was associated with severe hemolytic anemia. This condition proved to be transplantable into wild type mice (+/+) through total body irradiation (TBI) at a lethal dose of 8.0 Gy followed by engraftment of mutant bone marrow cells. Since the mutants are extremely radiosensitive (lethal dose50/30 4.4 Gy vs 7.3 Gy in +/+ mice), 8.0-Gy TBI followed by injection of even high numbers of normal bone marrow cells did not prevent death within 5-6 days. After a nonlethal dose of 4.0 Gy and grafting of normal bone marrow cells, a transient chimerism showing peripheral blood characteristics of the wild type was produced that returned to the mutant condition within 12 weeks. The transfusion of wild type red blood cells prior to and following 8.0-Gy TBI and reconstitution with wild type bone marrow cells prevented the early death of the mutants and permanent chimerism was achieved. The chimeras showed all hematological parameters of wild type mice, and radiosensitivity returned to normal. It is concluded that the mutant pluripotent stem cells are functionally comparable to normal stem cells, emphasizing the significance of this mouse model for studies of stem cell regulation.

Datta, T.; Doermer, P.

1987-12-01

106

Muscular cholinesterase and lactate dehydrogenase activities in deep-sea fish from the NW Mediterranean.  

PubMed

Organisms inhabiting submarine canyons can be potentially exposed to higher inputs of anthropogenic chemicals than their counterparts from the adjacent areas. To find out to what extend this observation applies to a NW Mediterranean canyon (i.e. Blanes canyon) off the Catalan coast, four deep-sea fish species were collected from inside the canyon (BC) and the adjacent open slope (OS). The selected species were: Alepocephalus rostratus, Lepidion lepidion, Coelorinchus mediterraneus and Bathypterois mediterraneus. Prior to the choice of an adequate sentinel species, the natural variation of the selected parameters (biomarkers) in relation to factors such as size, sex, sampling depth and seasonality need to be characterised. In this study, the activities of cholinesterases (ChEs) and lactate dehydrogenase (LDH) enzymes were determined in the muscle of the four deep-sea fish. Of all ChEs, acetylcholinesterase (AChE) activity was dominant and selected for further monitoring. Overall, AChE activity exhibited a significant relationship with fish size whereas LDH activity was mostly dependent on the sex and gonadal development status, although in a species-dependent manner. The seasonal variability of LDH activity was more marked than for AChE activity, and inside-outside canyon (BC-OS) differences were not consistent in all contrasted fish species, and in fact they were more dependent on biological traits. Thus, they did not suggest a differential stress condition between sites inside and outside the canyon. PMID:24296242

Koenig, Samuel; Solé, Montserrat

2014-03-01

107

Regulation of the synthesis of lactate dehydrogenase-X during spermatogenesis in the mouse  

PubMed Central

Total mouse testis RNA directs the synthesis of the sperm-specific C subunit of lactate dehydrogenase-X (LDH-X) when translated in a cell- free system derived from rabbit reticulocytes. The newly synthesized C subunits were isolated by immunoprecipitation with antibody specific for this isozyme, and quantitated by electrophoresis on SDS polyacrylamide gels. The amount of radioactivity incorporated into the enzyme subunit was directly proportional to the amount of testis RNA added to the translational system, thereby providing a sensitive and reliable method for assessing relative LDH-X mRNA activity. A combination of sucrose gradient centrifugation and oligo(dT)-cellulose chromatography resulted in a 23-fold purification of LDH-X mRNA over total cytoplasmic testis RNA. Analysis of LDH-X mRNA activity in the developing testis indicated that the appearance of functional LDH-X mRNA activity coincides with the appearance of LDH-X catalytic activity at 14 d postpartum. Measurement of LDH-X mRNA levels in separated testis cell populations prepared by centrifugal elutriation demonstrated that LDH-X mRNA represents 0.17-0.18% of the total functional mRNA activity in fractions enriched in pachytene spermatocytes and round spermatids, but only 0.09-0.10% of the translation products of elongated spermatids. PMID:7217199

1981-01-01

108

Source of catalysis in the lactate dehydrogenase system. Ground-state interactions in the enzyme-substrate complex.  

PubMed

The Raman spectra of both the NAD-pyruvate and the pyridine aldehyde adenine dinucleotide (PAAD)-pyruvate bound to pig heart, pig muscle, and Bacillus stearothermophilus lactate dehydrogenases were measured and are nearly the same, which is consistent with the conserved shell of residues surrounding the active-site cavity in these enzymes. The symmetrical stretching mode of the pyruvate carboxylate group, found at 1398 cm-1, is shifted only slightly when complexed to these enzymes, which shows that the group remains ionized in the ion pair complex with Arg-171 on the enzyme. The vibrational mode for the carbonyl stretch of the bound pyruvate moiety is shifted about 35 cm-1 to a lower frequency than observed for the carbonyl of unliganded pyruvate in the bacterial enzyme because of polarization of the carbonyl bond. Thus, the bacterial enzyme shows the same substrate activation because of the C(+)-O- charge separation that was seen previously with the mammalian enzymes. On the basis of an empirical Badger-Bauer relationship between frequency shift and interaction enthalpy, this shift in frequency is equivalent to an approximately -14 to -17 kcal/mol interaction between the enzyme and the adduct C = O coordinate, a substantial part of which is an electrostatic interaction (hydrogen bond) between the C V O and the protonated His-195. Thus, while the C = O bond is polarized on the enzyme (which requires energy), the overall ground-state enthalpy of the carbonyl imidazolium part of the reaction coordinate is stability substantially relative to its value in solution, and this is the dominant enthalpic effect on the entire reaction coordinate since the other internal coordinates for the hydride transfer are not much affected during formation of the ternary complex.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8117687

Deng, H; Zheng, J; Clarke, A; Holbrook, J J; Callender, R; Burgner, J W

1994-03-01

109

Decreasing lactate level and increasing antibody production in Chinese Hamster Ovary cells (CHO) by reducing the expression of lactate dehydrogenase and pyruvate dehydrogenase kinases  

Microsoft Academic Search

Large-scale fed-batch cell culture processes of CHO cells are the standard platform for the clinical and commercial production of monoclonal antibodies. Lactate is one of the major by-products of CHO fed-batch culture. In pH-controlled bioreactors, accumulation of high levels of lactate is accompanied by high osmolality due to the addition of base to control pH of the cell culture medium,

Meixia Zhou; Yongping Crawford; Domingos Ng; Jack Tung; Abigail F. J. Pynn; Angela Meier; Inn H. Yuk; Natarajan Vijayasankaran; Kimberly Leach; John Joly; Bradley Snedecor; Amy Shen

2011-01-01

110

Epidermal growth factor or serum stimulation of rat fibroblasts induces an elevation in mRNA levels for lactate dehydrogenase and other glycolytic enzymes.  

PubMed Central

We have isolated cloned cDNAs corresponding to five mRNAs whose level is increased following stimulation of quiescent rat fibroblasts by either epidermal growth factor or serum. Partial sequencing followed by a computer search of data banks has shown that the cloned cDNAs correspond to mRNAs encoding proteins with extensive homology to lactate dehydrogenase, glyceraldehyde 3-phosphate dehydrogenase, enolase, triose phosphate isomerase, and actin. The complete nucleotide sequence of a rat fibroblast lactate dehydrogenase is presented. Images PMID:3873645

Matrisian, L M; Rautmann, G; Magun, B E; Breathnach, R

1985-01-01

111

The lactate dehydrogenase–reduced nicotinamide–adenine dinucleotide–pyruvate complex. Kinetics of pyruvate binding and quenching of coenzyme fluorescence  

PubMed Central

The stopped-flow kinetic studies described in this and the following paper (Südi, 1974) demonstrate that a Haldane-type description of the reversible lactate dehydrogenase reaction presents an experimentally feasible task. Combined results of these two papers yield numerical values for the six rate constants defined by the following equilibrium scheme, where E represents lactate dehydrogenase: [Formula: see text] The experiments were carried out at pH8.4 at a relatively low temperature (6.3°C) with the pig heart enzyme. Identification of the above two intermediates and determination of the corresponding rate constants actually involve four series of independent observations in these studies, since (a) the reaction can be followed in both directions, and (b) both the u.v. absorption and the fluorescence of the coenzymes are altered in the reaction, and it is shown that these two spectral changes do not occur simultaneously. Kinetic observations made in the reverse direction are reported in this paper. It is demonstrated that the fluorescence of NADH can no longer be observed in the ternary complex ENADHPyr. Even though the oxidation–reduction reaction rapidly follows the formation of this complex, the numerical values of k?4 (8.33×105m?1·s?1) and k+4 (222s?1) are easily obtained from a directly observed second-order reaction step in which fluorescent but not u.v.-absorbing material is disappearing. U.v.-absorption measurements do not clearly resolve the subsequent oxidation–reduction step from the dissociation of lactate. It is shown that this must be due partly to the instrumental dead time, and partly to a low transient concentration of ENAD+Lac in the two-step sequential reaction in which the detectable disappearance of u.v.-absorbing material takes place. It is estimated that about one-tenth of the total change in u.v. absorption is due to a `burst reaction' in which ENAD+Lac is produced, and this estimation yields, from kobs.=120s?1, k?2=1200s?1. PMID:4377095

Südi, J.

1974-01-01

112

Lactate dehydrogenase regulation in aged skeletal muscle: Regulation by anabolic steroids and functional overload.  

PubMed

Aging alters the skeletal muscle response to overload-induced growth. The onset of functional overload is characterized by increased myoblast proliferation and an altered muscle metabolic profile. The onset of functional overload is associated with increased energy demands that are met through the interconversion of lactate and pyruvate via the activity of lactate dehydrogenase (LDH). Testosterone targets many of the processes activated at the onset of functional overload. However, the effect of aging on this metabolic plasticity at the onset of functional overload and how anabolic steroid administration modulates this response is not well understood. The purpose of this study was to determine if aging would alter overload-induced LDH activity and expression at the onset of functional overload and whether anabolic steroid administration would modulate this response. Five-month and 25-month male Fischer 344xF1 BRN were given nandrolone decanoate (ND) or sham injections for 14days and then the plantaris was functionally overloaded (OV) for 3days by synergist ablation. Aging reduced muscle LDH-A & LDH-B activity 70% (p<0.05). Aging also reduced LDH-A mRNA abundance, however there was no age effect on LDH-B mRNA abundance. In 5-month muscle, both ND and OV decreased LDH-A and LDH-B activity. However, there was no synergistic or additive effect. In 5-month muscle, ND and OV decreased LDH-A mRNA expression with no change in LDH-B expression. In 25-month muscle, ND and OV increased LDH-A and LDH-B activity. LDH-A mRNA expression was not altered by ND or OV in aged muscle. However, there was a main effect of OV to decrease LDH-B mRNA expression. There was also an age-induced LDH isoform shift. ND and OV treatment increased the "fast" LDH isoforms in aged muscle, whereas ND and OV increased the "slow" isoforms in young muscle. Our study provides evidence that aging alters aspects of skeletal muscle metabolic plasticity normally induced by overload and anabolic steroid administration. PMID:24835193

Washington, Tyrone A; Healey, Julie M; Thompson, Raymond W; Lowe, Larry L; Carson, James A

2014-09-01

113

Temporal changes in the involvement of pyruvate dehydrogenase complex in muscle lactate accumulation during lipopolysaccharide infusion in rats  

PubMed Central

A characteristic manifestation of sepsis is muscle lactate accumulation. This study examined any putative (causative) association between pyruvate dehydrogenase complex (PDC) inhibition and lactate accumulation in the extensor digitorum longus (EDL) muscle of rats infused with lipopolysaccharide (LPS), and explored the involvement of increased transcription of muscle-specific pyruvate dehydrogenase kinase (PDK) isoenzymes. Conscious, male Sprague–Dawley rats were infused i.v. with saline (0.4 ml h?1, control) or LPS (150 ?g kg?1 h?1) for 2 h, 6 h or 24 h (n = 6–8). Muscle lactate concentration was elevated after 2, 6 and 24 h LPS infusion. Muscle PDC activity was the same at 2 h and 6 h, but was 65% lower after 24 h of LPS infusion (P < 0.01), when there was a 47% decrease in acetylcarnitine concentration (P < 0.05), and a 24-fold increase in PDK4 mRNA expression (P < 0.001). These changes were preceded by marked increases in tumour necrosis factor-? and interleukin-6 mRNA expression at 2 h. The findings indicate that the early (2 and 6 h) elevation in muscle lactate concentration during LPS infusion was not attributable to limited muscle oxygen availability or ATP production (evidenced by unchanged ATP and phosphocreatine (PCr) concentrations) or to PDC inhibition, whereas after 24 h, muscle lactate accumulation appears to have resulted from PDC activation status limiting pyruvate flux, most probably due to cytokine-mediated up-regulation of PDK4 transcription. PMID:18218678

Alamdari, N; Constantin-Teodosiu, D; Murton, A J; Gardiner, S M; Bennett, T; Layfield, R; Greenhaff, P L

2008-01-01

114

Regulation of the Activity of Lactate Dehydrogenases from Four Lactic Acid Bacteria*  

PubMed Central

Despite high similarity in sequence and catalytic properties, the l-lactate dehydrogenases (LDHs) in lactic acid bacteria (LAB) display differences in their regulation that may arise from their adaptation to different habitats. We combined experimental and computational approaches to investigate the effects of fructose 1,6-bisphosphate (FBP), phosphate (Pi), and ionic strength (NaCl concentration) on six LDHs from four LABs studied at pH 6 and pH 7. We found that 1) the extent of activation by FBP (Kact) differs. Lactobacillus plantarum LDH is not regulated by FBP, but the other LDHs are activated with increasing sensitivity in the following order: Enterococcus faecalis LDH2 ? Lactococcus lactis LDH2 < E. faecalis LDH1 < L. lactis LDH1 ? Streptococcus pyogenes LDH. This trend reflects the electrostatic properties in the allosteric binding site of the LDH enzymes. 2) For L. plantarum, S. pyogenes, and E. faecalis, the effects of Pi are distinguishable from the effect of changing ionic strength by adding NaCl. 3) Addition of Pi inhibits E. faecalis LDH2, whereas in the absence of FBP, Pi is an activator of S. pyogenes LDH, E. faecalis LDH1, and L. lactis LDH1 and LDH2 at pH 6. These effects can be interpreted by considering the computed binding affinities of Pi to the catalytic and allosteric binding sites of the enzymes modeled in protonation states corresponding to pH 6 and pH 7. Overall, the results show a subtle interplay among the effects of Pi, FBP, and pH that results in different regulatory effects on the LDHs of different LABs. PMID:23720742

Feldman-Salit, Anna; Hering, Silvio; Messiha, Hanan L.; Veith, Nadine; Cojocaru, Vlad; Sieg, Antje; Westerhoff, Hans V.; Kreikemeyer, Bernd; Wade, Rebecca C.; Fiedler, Tomas

2013-01-01

115

Evolutionary factors affecting Lactate dehydrogenase A and B variation in the Daphnia pulex species complex  

PubMed Central

Background Evidence for historical, demographic and selective factors affecting enzyme evolution can be obtained by examining nucleotide sequence variation in candidate genes such as Lactate dehydrogenase (Ldh). Two closely related Daphnia species can be distinguished by their electrophoretic Ldh genotype and habitat. Daphnia pulex populations are fixed for the S allele and inhabit temporary ponds, while D. pulicaria populations are fixed for the F allele and inhabit large stratified lakes. One locus is detected in most allozyme surveys, but genome sequencing has revealed two genes, LdhA and LdhB. Results We sequenced both Ldh genes from 70 isolates of these two species from North America to determine if the association between Ldh genotype and habitat shows evidence for selection, and to elucidate the evolutionary history of the two genes. We found that alleles in the pond-dwelling D. pulex and in the lake-dwelling D. pulicaria form distinct groups at both loci, and the substitution of Glutamine (S) for Glutamic acid (F) at amino acid 229 likely causes the electrophoretic mobility shift in the LDHA protein. Nucleotide diversity in both Ldh genes is much lower in D. pulicaria than in D. pulex. Moreover, the lack of spatial structuring of the variation in both genes over a wide geographic area is consistent with a recent demographic expansion of lake populations. Neutrality tests indicate that both genes are under purifying selection, but the intensity is much stronger on LdhA. Conclusions Although lake-dwelling D. pulicaria hybridizes with the other lineages in the pulex species complex, it remains distinct ecologically and genetically. This ecological divergence, coupled with the intensity of purifying selection on LdhA and the strong association between its genotype and habitat, suggests that experimental studies would be useful to determine if variation in molecular function provides evidence that LDHA variants are adaptive. PMID:21767386

2011-01-01

116

Regulation of liver lactate dehydrogenase by reversible phosphorylation in response to anoxia in a freshwater turtle.  

PubMed

Lactate dehydrogenase (LDH) is the terminal enzyme of anaerobic glycolysis and key to hypoxia/anoxia survival by most animals. In this study, the effects of anoxic submergence (20 h at 7°C in nitrogen-bubbled water) were assessed on LDH from liver of an anoxia-tolerant freshwater turtle, the red-eared slider (Trachemys scripta elegans). Liver LDH from aerobic and anoxic turtles was purified to homogeneity in two steps. The kinetic properties and thermal stability of purified LDH were analyzed, revealing significant differences between the two enzyme forms in V(max), K(m) pyruvate, and I(50) pyruvate as well as melting temperature determined by differential scanning fluorimetry. The phosphorylation state of aerobic and anoxic forms of LDH was visualized by ProQ Diamond phosphoprotein staining, the results indicating that the anoxic form had a higher phosphorylation state. Incubation studies that promoted protein kinase versus protein phosphatase actions showed that changes in the phosphorylation state of aerobic and anoxic forms mimicked the anoxia-responsive changes in K(m) pyruvate and I(50) pyruvate. The high phosphate form of liver LDH that occurs in anoxic turtles appears to be a less active form. Turtle liver LDH was also subject to another form of posttranslational modification, protein acetylation, with a 70% higher content of acetylated lysine residues on anoxic versus aerobic LDH. This is the first study to show that LDH function in an anoxia-tolerant animal can be differentially modified between aerobic and anoxic states via the mechanism of posttranslational modification. PMID:22735190

Xiong, Zi Jian; Storey, Kenneth B

2012-10-01

117

Direct electrochemistry of lactate dehydrogenase immobilized on silica sol-gel modified gold electrode and its application.  

PubMed

The direct electrochemistry of lactate dehydrogenase (LDH) immobilized in silica sol-gel film on gold electrode was investigated, and an obvious cathodic peak at about -200 mV (versus SCE) was found for the first time. The LDH-modified electrode showed a surface controlled irreversible electrode process involving a one electron transfer reaction with the charge-transfer coefficient (alpha) of 0.79 and the apparent heterogeneous electron transfer rate constant (K(s)) of 3.2 s(-1). The activated voltammetric response and decreased charge-transfer resistance of Ru(NH(3))(6)(2+/3+) on the LDH-modified electrode provided further evidence. The surface morphologies of silica sol-gel and the LDH embedded in silica sol-gel film were characterized by SEM. A potential application of the LDH-modified electrode as a biosensor for determination of lactic acid was also investigated. The calibration range of lactic acid was from 2.0 x 10(-6) to 3.0 x 10(-5) mol L(-1) and the detection limit was 8.0 x 10(-7) mol L(-1) at a signal-to-noise ratio of 3. Finally, the effect of environmental pollutant resorcinol on the direct electrochemical behavior of LDH was studied. The experimental results of voltammetry indicated that the conformation of LDH molecule was altered by the interaction between LDH and resorcinol. The modified electrode can be applied as a biomarker to study the pollution effect in the environment. PMID:17869089

Di, Junwei; Cheng, Jiongjia; Xu, Quan; Zheng, Huie; Zhuang, Jingyue; Sun, Yongbo; Wang, Keyu; Mo, Xiangyin; Bi, Shuping

2007-12-15

118

Assessment of lactate dehydrogenase, alkaline phosphatase and aspartate aminotransferase activities in cow's milk as an indicator of subclinical mastitis.  

PubMed

This study examined the activities of lactate dehydrogenase (LDH), alkaline phosphatase (ALP) and aspartate aminotransferase (AST) in the milk of lactating Holstein cows in association with subclinical mastitis (SCM). A total of 94 milk samples were collected from 58 lactating dairy cows representing stages of lactation from the second to the tenth week after calving. Those which were classified as positive by California mastitis test (CMT) were deemed to have subclinical mastitis. All the milk samples were skimmed by centrifugation at 10 000g at 0 degrees C and were used for enzyme activities estimations. The mean activities of LDH and ALP were higher in the milk from udders with SCM than in the milk from healthy udders (p < 0.05). There were no significant differences in AST values. The maximum agreement rates between the CMT results and LDH and ALP values were seen at thresholds of > 180 IU/L and > 40 IU/L respectively (kappa values 0.65 and 0.79, respectively). However, the sensitivity of the tests for identifying SCM at these thresholds was higher for ALP (96.4%) than for LDH (68.5%). In this study, LDH and ALP tests were standardized for cow's milk and results showed that only the ALP test was reliable in the early diagnosis of subclinical mastitis. PMID:17268916

Babaei, H; Mansouri-Najand, L; Molaei, M M; Kheradmand, A; Sharifan, M

2007-05-01

119

The reaction of choline dehydrogenase with some electron acceptors.  

PubMed Central

1. The choline dehydrogenase (EC 1.1.99.1) WAS SOLUBILIZED FROM ACETONE-DRIED POWDERS OF RAT LIVER MITOCHONDRIA BY TREATMENT WITH Naja naja venom. 2. The kinetics of the reaction of enzyme with phenazine methosulphate and ubiquinone-2 as electron acceptors were investigated. 3. With both electron acceptors the reaction mechanism appears to involve a free, modified-enzyme intermediate. 4. With some electron acceptors the maximum velocity of the reaction is independent of the nature of the acceptor. With phenazine methosulphate and ubiquinone-2 as acceptors the Km value for choline is also independent of the nature of the acceptor molecule. 5. The mechanism of the Triton X-100-solubilized enzyme is apparently the smae as that for the snake venom solubilized enzyme. PMID:1218095

Barrett, M C; Dawson, A P

1975-01-01

120

Effect of water activity on inactivation of Listeria monocytogenes and lactate dehydrogenase during high pressure processing.  

PubMed

The aim of this study was to investigate the effect of water activity (aw) on the inactivation of Listeria monocytogenes and lactate dehydrogenase (LDH) during high pressure processing (HPP). For microbial inactivation lyophilized cells of L. monocytogenes 19,115 were left dry or were suspended in 10 ml of 0.1% peptone water, 10 ml of glycerol, or mixtures of glycerol and peptone water. All samples of various aws were high pressure (HP) processed at ambient temperature at 600 MPa for 300 s. Following HPP, samples were serially diluted in 0.1% peptone and spread-plated on Tryptic Soy agar supplemented with Yeast Extract. For enzyme inactivation, 4.2 mg of lyophilized LDH was suspended in 2 ml of 100 mM phosphate buffer (pH 7.4), 2 ml of peptone water or glycerol, or in 2 ml mixtures of glycerol and peptone water. A lyophilized sample with no added liquid was also included. All enzyme samples were subjected to HPP as described above. After HPP, LDH was diluted to 0.28 microg/ml in 100 mM phosphate buffer (pH 7.4). LDH activity was assessed by measuring the change in concentration of beta-NADH as a function of time. Dynamic light scattering analysis (DLS) was performed to examine the size distribution, polydispersity, and hydrodynamic radius of LDH before and after HPP. No significant difference in CFU/g was observed between lyophilized cells not subjected to HPP and lyophilized cells subjected to 600 MPa for 300 s (P<0.05). However, lyophilized cells that were suspended in 100% to 60% peptone water showed a approximately 7.5-log(10) reduction when subjected to HPP. Survival of L. monocytogenes following HPP significantly increased (P<0.05) when the peptone water concentration was decreased below 60% (aw approximately 0.8). DLS results revealed that LDH suspended in buffer underwent aggregation following HPP (600 MPa, 300 s). Inactivation rate constants obtained using a first-order kinetic model indicated that untreated and HP processed lyophilized LDH had similar activities. When LDH was subject to HPP in solutions containing glycerol, enzyme activity decreased as the water content increased (r2=0.95). Lyophilization completely protected L. monocytogenes and LDH from inactivation by high pressure. Furthermore, enzyme activity and cell survival increased as water activity was decreased. We postulate low aw results in protein stabilization, which prevents protein denaturation and cell death during HPP. PMID:18403036

Hayman, Melinda M; Kouassi, Gilles K; Anantheswaran, Ramaswamy C; Floros, John D; Knabel, Stephen J

2008-05-10

121

Effect of the inactivation of lactate dehydrogenase, ethanol dehydrogenase, and phosphotransacetylase on 2,3-butanediol production in Klebsiella pneumoniae strain  

PubMed Central

Background 2,3-Butanediol (2,3-BD) is a high-value chemical usually produced petrochemically but which can also be synthesized by some bacteria. To date, Klebsiella pneumoniae is the most powerful 2,3-BD producer which can utilize a wide range of substrates. However, many by-products are also produced by K. pneumoniae, such as ethanol, lactate, and acetate, which negatively regulate the 2,3-BD yield and increase the costs of downstream separation and purification. Results In this study, we constructed K. pneumoniae mutants with lactate dehydrogenase (LDH), acetaldehyde dehydrogenase (ADH), and phosphotransacetylase (PTA) deletion individually by suicide vector conjugation. These mutants showed different behavior of production formation. Knock out of ldhA had little influence on the yield of 2,3-BD, whereas knock out of adhE or pta significantly improved the formation of 2,3-BD. The accumulation of the intermediate of 2,3-BD biosynthesis, acetoin, was decreased in all the mutants. The mutants were then tested in five different carbon sources and increased 2,3-BD was observed. Also a double mutant strain with deletion of adhE and ldhA was constructed which resulted in accelerated fermentation and higher 2,3-BD production. In fed-batch culture this strain achieved more than 100 g/L 2,3-BD from glucose with a relatively high yield of 0.49 g/g. Conclusion 2,3-BD production was dramatically improved with the inactivation of adhE and pta. The inactivation of ldhA could advance faster cell growth and shorter fermentation time. The double mutant strain with deletion of adhE and ldhA resulted in accelerated fermentation and higher 2,3-BD production. These results provide new insights for industrial production of 2,3-BD by K. pneumoniae. PMID:24669952

2014-01-01

122

Efficient Production of (R)-2-Hydroxy-4-Phenylbutyric Acid by Using a Coupled Reconstructed d-Lactate Dehydrogenase and Formate Dehydrogenase System  

PubMed Central

Background (R)-2-Hydroxy-4-phenylbutyric acid [(R)-HPBA] is a key precursor for the production of angiotensin-converting enzyme inhibitors. However, the product yield and concentration of reported (R)-HPBA synthetic processes remain unsatisfactory. Methodology/Principal Findings The Y52L/F299Y mutant of NAD-dependent d-lactate dehydrogenase (d-nLDH) in Lactobacillus bulgaricus ATCC 11842 was found to have high bio-reduction activity toward 2-oxo-4-phenylbutyric acid (OPBA). The mutant d-nLDHY52L/F299Y was then coexpressed with formate dehydrogenase in Escherichia coli BL21 (DE3) to construct a novel biocatalyst E. coli DF. Thus, a novel bio-reduction process utilizing whole cells of E. coli DF as the biocatalyst and formate as the co-substrate for cofactor regeneration was developed for the production of (R)-HPBA from OPBA. The biocatalysis conditions were then optimized. Conclusions/Significance Under the optimum conditions, 73.4 mM OPBA was reduced to 71.8 mM (R)-HPBA in 90 min. Given its high product enantiomeric excess (>99%) and productivity (47.9 mM h?1), the constructed coupling biocatalysis system is a promising alternative for (R)-HPBA production. PMID:25089519

Sheng, Binbin; Zheng, Zhaojuan; Lv, Min; Zhang, Haiwei; Qin, Tong; Gao, Chao; Ma, Cuiqing; Xu, Ping

2014-01-01

123

Reduced diagnostic value of lactate dehydrogenase (LDH) in the presence of radiographic contrast media.  

PubMed

Isoforms of the enzyme lactate dehydrogenase (LDH) were found in almost all cells of the organism and an elevated activity of LDH in the circulation is thought to be a clear indicator of elevated cell destruction coinciding with an increased release of components from the cellular cytoplasm, e.g. LDH. Here, we report on an in-vitro examination to test whether radiographic contrast media (RCM) could induce cell destruction followed by an increase in LDH release. The RCM were tested in non-flow cultures of human umbilical venous endothelial cells (HUVEC) of the fourth passage seeded on extracellular matrix and the results were compared to those from control cultures not exposed to contrast media. The examination revealed that the addition of contrast media to the cell culture media supplemented with pooled human serum (HSP) as source of exogenous LDH was followed by a strong decrease in LDH activity both in the absence and presence of HUVEC. Within 1.5 min after the addition of contrast media to the culture medium supplemented with HSP (30% vol of the culture medium were replaced by either of two contrast media, Iodixanol or Iopromide) the LDH activity decreased about 80% compared to the initial values. In contrast, the LDH activity did not change in cell culture media not supplemented with RCM. The partial replacement of HSP supplemented cell culture medium by RCM will cause a dilution of cell culture medium constituents. The decrease of LDH activity, however, was much stronger than the decrease thought to be attributable to the effects of dilution of cell culture medium, so that the role of dilution seems to be a minor one in this case. It has to be assumed that the RCM could interact with the LDH available in the culture medium as well as with the substrates delivered with the measurement system for the assessment of LDH activity, so that both, the amount of LDH and the activities of enzymes involved might be influenced. In the presence of HUVEC a similar effect was observed. Here, a little less strong decrease of LDH activity occurred compared to the decrease in cell culture medium without HUVEC. This was unexpected because a considerable amount of HUVEC were detached after the addition of contrast media and many of these cells were damaged seriously so that a significant amount of endogenous LDH should have been released. These unexpected results make it necessary to re-evaluate those past time examinations focussed on cell damage/destruction in the presence of contrast media, where the measurement of LDH activity was used as indicator or cell vitality and where cell decease rates were correlated to questionable toxic influences. According to the results of the examination reported here it is difficult to uphold the interpretation of recently published findings that contrast media almost exclusively induce cellular apoptosis and not necrosis. PMID:20675892

Franke, R-P; Fuhrmann, R; Mrowietz, C; Rickert, D; Hiebl, B; Jung, F

2010-01-01

124

Changes in milk L-lactate, lactate dehydrogenase, serum albumin, and IgG during milk ejection and their association with somatic cell count.  

PubMed

In both conventional and automatic milking systems (AMS), sensitive and reliable mastitis detection is important for profitable milk production. Mastitis detection parameters must be able to detect mastitis when the somatic cell count (SCC) is only slightly elevated. Owing to the pre-milking teat cleaning process in AMS, sampling cannot take place before the occurrence of alveolar milk ejection and importantly, this can affect the ability of parameters to detect mastitis. The aim of the present study was to examine the effect of alveolar milk ejection on l-lactate, lactate dehydrogenase (LDH), serum albumin (SA) and immunoglobulin G (IgG) compared with SCC, a commonly used indicator of mastitis. In this experiment, milk samples were collected every 20 s from one quarter during a 120-s manual teat stimulation in ten cows. Samples were analysed for SCC, l-lactate, LDH, SA and IgG. Quarters were grouped by low (<5·0 log10 cells/ml), mid (5·0-5·7 log10 cells/ml), and high (>5·7 log10 cells/ml) SCC using the sample at t=0 s. Neither l-lactate nor LDH could statistically differentiate between low and mid-SCC quarters, but there were a significant difference in levels between the high-SCC quarters and low and mid-SCC quarters. SA could not differentiate between the low and mid-SCC quarters, but the SA levels for the high SCC quarters remained statistically different compared with low and mid-SCC quarters throughout the experiment. IgG could statistically differentiate between low and mid-SCC, although the high-SCC quarters were not statistically different from the mid-SCC quarters after 60 s. In the high-SCC quarters, a decrease was shown in all parameters during milk ejection, after t=60 s. In conclusion, alveolar milk ejection reduces the effectiveness of detection parameters when compared with SCC. With the exception of IgG, the ability of other tested parameters was not satisfactory to differentiate between quarters with low to mid-SCC levels. PMID:25467384

Lehmann, Mirjam; Wall, Samantha K; Wellnitz, Olga; Bruckmaier, Rupert M

2015-05-01

125

Combined inactivation of the Clostridium cellulolyticum lactate and malate dehydrogenase genes substantially increases ethanol yield from cellulose and switchgrass fermentations  

SciTech Connect

Background: The model bacterium Clostridium cellulolyticum efficiently hydrolyzes crystalline cellulose and hemicellulose, using cellulosomes to degrade lignocellulosic biomass. Although it imports and ferments both pentose and hexose sugars to produce a mixture of ethanol, acetate, lactate, H2 and CO2, the proportion of ethanol is low, which impedes its use in consolidated bioprocessing for biofuels. Therefore genetic engineering will likely be required to improve the ethanol yield. Random mutagenesis, plasmid transformation, and heterologous expression systems have previously been developed for C. cellulolyticum, but targeted mutagenesis has not been reported for this organism. Results: The first targeted gene inactivation system was developed for C. cellulolyticum, based on a mobile group II intron originating from the Lactococcus lactis L1.LtrB intron. This markerless mutagenesis system was used to disrupt both the paralogous L-lactate dehydrogenase (Ccel_2485; ldh) and L-malate dehydrogenase (Ccel_0137; mdh) genes, distinguishing the overlapping substrate specificities of these enzymes. Both mutations were then combined in a single strain. This double mutant produced 8.5-times more ethanol than wild-type cells growing on crystalline cellulose. Ethanol constituted 93% of the major fermentation products (by molarity), corresponding to a molar ratio of ethanol to organic acids of 15, versus 0.18 in wild-type cells. During growth on acid-pretreated switchgrass, the double mutant also produced four-times as much ethanol as wild-type cells. Detailed metabolomic analyses identified increased flux through the oxidative branch of the mutant s TCA pathway. Conclusions: The efficient intron-based gene inactivation system produced the first gene-targeted mutations in C. cellulolyticum. As a key component of the genetic toolbox for this bacterium, markerless targeted mutagenesis enables functional genomic research in C. cellulolyticum and rapid genetic engineering to significantly alter the mixture of fermentation products. The initial application of this system successfully engineered a strain with high ethanol productivity from complex biomass substrates.

Li, Yongchao [ORNL; Tschaplinski, Timothy J [ORNL; Engle, Nancy L [ORNL; Hamilton, Choo Yieng [ORNL; Rodriguez, Jr., Miguel [ORNL; Liao, James C [ORNL; Schadt, Christopher Warren [ORNL; Guss, Adam M [ORNL; Yang, Yunfeng [ORNL; Graham, David E [ORNL

2012-01-01

126

Decreased Hematocrit-To-Viscosity Ratio and Increased Lactate Dehydrogenase Level in Patients with Sickle Cell  

E-print Network

with Sickle Cell Anemia and Recurrent Leg Ulcers Philippe Connes1,2,3* , Yann Lamarre1,2 , Marie-à-Pitre, Guadeloupe Abstract Leg ulcer is a disabling complication in patients with sickle cell anemia (SCA Dehydrogenase Level in Patients with Sickle Cell Anemia and Recurrent Leg Ulcers. PLoS ONE 8(11): e79680. doi:10

Paris-Sud XI, Université de

127

Production of l-lactic acid by the yeast Candida sonorensis expressing heterologous bacterial and fungal lactate dehydrogenases  

PubMed Central

Background Polylactic acid is a renewable raw material that is increasingly used in the manufacture of bioplastics, which offers a more sustainable alternative to materials derived from fossil resources. Both lactic acid bacteria and genetically engineered yeast have been implemented in commercial scale in biotechnological production of lactic acid. In the present work, genes encoding l-lactate dehydrogenase (LDH) of Lactobacillus helveticus, Bacillus megaterium and Rhizopus oryzae were expressed in a new host organism, the non-conventional yeast Candida sonorensis, with or without the competing ethanol fermentation pathway. Results Each LDH strain produced substantial amounts of lactate, but the properties of the heterologous LDH affected the distribution of carbon between lactate and by-products significantly, which was reflected in extra-and intracellular metabolite concentrations. Under neutralizing conditions C. sonorensis expressing L. helveticus LDH accumulated lactate up to 92 g/l at a yield of 0.94 g/g glucose, free of ethanol, in minimal medium containing 5 g/l dry cell weight. In rich medium with a final pH of 3.8, 49 g/l lactate was produced. The fermentation pathway was modified in some of the strains studied by deleting either one or both of the pyruvate decarboxylase encoding genes, PDC1 and PDC2. The deletion of both PDC genes together abolished ethanol production and did not result in significantly reduced growth characteristic to Saccharomyces cerevisiae deleted of PDC1 and PDC5. Conclusions We developed an organism without previous record of genetic engineering to produce L-lactic acid to a high concentration, introducing a novel host for the production of an industrially important metabolite, and opening the way for exploiting C. sonorensis in additional biotechnological applications. Comparison of metabolite production, growth, and enzyme activities in a representative set of transformed strains expressing different LDH genes in the presence and absence of a functional ethanol pathway, at neutral and low pH, generated a comprehensive picture of lactic acid production in this yeast. The findings are applicable in generation other lactic acid producing yeast, thus providing a significant contribution to the field of biotechnical production of lactic acid. PMID:23706009

2013-01-01

128

Enzymes Related to Lactate Metabolism in Green Algae and Lower Land Plants 1  

PubMed Central

Cell-free extracts of Chlorella pyrenoidosa contained two enzymes capable of oxidizing d-lactate; these were glycolate dehydrogenase and NAD+-dependent d-lactate dehydrogenase. The two enzymes could be distinguished by differential centrifugation, glycolate dehydrogenase being largely particulate and NAD+-d-lactate dehydrogenase being soluble. The reduction of pyruvate by NADH proceeded more rapidly than the reverse reaction, and the apparent Michaelis constants for pyruvate and NADH were lower than for d-lactate and NAD+. These data indicated that under physiological conditions, the NAD+-linked d-lactate dehydrogenase probably functions to produce d-lactate from pyruvate. Lactate dehydrogenase activity dependent on NAD+ was found in a number of other green algae and in the green tissues of a few lower land plants. When present in species which contain glycolate oxidase rather than glycolate dehydrogenase, the enzyme was specific for l-lactate rather than d-lactate. A cyclic system revolving around the production and utilization of d-lactate in some species and l-lactate in certain others is proposed. PMID:16658670

Gruber, Peter J.; Frederick, Sue Ellen; Tolbert, N. E.

1974-01-01

129

Lactation  

PubMed Central

Lactation is the most energy-efficient way to provide for the dietary needs of young mammals, their mother's milk being actively protective, immunomodulatory, and ideal for their needs. Intrauterine mammary gland development in the human female is already apparent by the end of the sixth week of gestation. During puberty and adolescence secretions of the anterior pituitary stimulate the maturation of the graafian follicles in the ovaries and stimulate the secretion of follicular estrogens, which stimulate development of the mammary ducts. Pregnancy has the most dramatic effect on the breast, but development of the glandular breast tissue and deposition of fat and connective tissue continue under the influence of cyclic sex-hormone stimulation. Many changes occur in the nipple and breast during pregnancy and at delivery as a prelude to lactation. Preparation of the breasts is so effective that lactation could commence even if pregnancy were discontinued at 16 weeks. Following birth, placental inhibition of milk synthesis is removed, and a woman's progesterone blood levels decline rapidly. The breasts fill with milk, which is a high-density, low-volume feed called colostrum until about 30 hours after birth. Because it is not the level of maternal hormones, but the efficiency of infant suckling and/or milk removal that governs the volume of milk produced in each breast, mothers who permit their infants to feed ad libitum commonly observe that they have large volumes of milk 24-48 hours after birth. The two maternal reflexes involved in lactation are the milk-production and milk-ejection reflex. A number of complementary reflexes are involved when the infant feeds: the rooting reflex (which programmes the infant to search for the nipple), the sucking reflex (rhythmic jaw action creating negative pressure and a peristaltic action of the tongue), and the swallowing reflex. The infant's instinctive actions need to be consolidated into learned behaviour in the postpartum period when the use of artificial teats and dummies (pacifiers) may condition the infant to different oral actions that are inappropriate for breast-feeding. Comparisons of breast milk and cow's milk fail to describe the many important differences between them, e.g., the structural and qualitative differences in proteins and fats, and the bioavailability of minerals. The protection against infection and allergies conferred on the infant, which is impossible to attain through any other feeding regimen, is one of breast milk's most outstanding qualities. The maximum birth-spacing effect of lactation is achieved when an infant is fully, or nearly fully, breast-fed and the mother consequently remains amenorrhoeic. PMID:20604468

1989-01-01

130

Combined inactivation of the Clostridium cellulolyticum lactate and malate dehydrogenase genes substantially increases ethanol yield from cellulose and switchgrass fermentations  

PubMed Central

Background The model bacterium Clostridium cellulolyticum efficiently degrades crystalline cellulose and hemicellulose, using cellulosomes to degrade lignocellulosic biomass. Although it imports and ferments both pentose and hexose sugars to produce a mixture of ethanol, acetate, lactate, H2 and CO2, the proportion of ethanol is low, which impedes its use in consolidated bioprocessing for biofuels production. Therefore genetic engineering will likely be required to improve the ethanol yield. Plasmid transformation, random mutagenesis and heterologous expression systems have previously been developed for C. cellulolyticum, but targeted mutagenesis has not been reported for this organism, hindering genetic engineering. Results The first targeted gene inactivation system was developed for C. cellulolyticum, based on a mobile group II intron originating from the Lactococcus lactis L1.LtrB intron. This markerless mutagenesis system was used to disrupt both the paralogous L-lactate dehydrogenase (Ccel_2485; ldh) and L-malate dehydrogenase (Ccel_0137; mdh) genes, distinguishing the overlapping substrate specificities of these enzymes. Both mutations were then combined in a single strain, resulting in a substantial shift in fermentation toward ethanol production. This double mutant produced 8.5-times more ethanol than wild-type cells growing on crystalline cellulose. Ethanol constituted 93% of the major fermentation products, corresponding to a molar ratio of ethanol to organic acids of 15, versus 0.18 in wild-type cells. During growth on acid-pretreated switchgrass, the double mutant also produced four times as much ethanol as wild-type cells. Detailed metabolomic analyses identified increased flux through the oxidative branch of the mutant's tricarboxylic acid pathway. Conclusions The efficient intron-based gene inactivation system produced the first non-random, targeted mutations in C. cellulolyticum. As a key component of the genetic toolbox for this bacterium, markerless targeted mutagenesis enables functional genomic research in C. cellulolyticum and rapid genetic engineering to significantly alter the mixture of fermentation products. The initial application of this system successfully engineered a strain with high ethanol productivity from cellobiose, cellulose and switchgrass. PMID:22214220

2012-01-01

131

Warburg effect in chemosensitivity: Targeting lactate dehydrogenase-A re-sensitizes Taxol-resistant cancer cells to Taxol  

PubMed Central

Background Taxol is one of the most effective chemotherapeutic agents for the treatment of patients with breast cancer. Despite impressive clinical responses initially, the majority of patients eventually develop resistance to Taxol. Lactate dehydrogenase-A (LDH-A) is one of the predominant isoforms of LDH expressed in breast tissue, which controls the conversion of pyruvate to lactate and plays an important role in glucose metabolism. In this study we investigated the role of LDH-A in mediating Taxol resistance in human breast cancer cells. Results Taxol-resistant subclones, derived from the cancer cell line MDA-MB-435, sustained continuous growth in high concentrations of Taxol while the Taxol-sensitive cells could not. The increased expression and activity of LDH-A were detected in Taxol-resistant cells when compared with their parental cells. The downregulation of LDH-A by siRNA significantly increased the sensitivity of Taxol-resistant cells to Taxol. A higher sensitivity to the specific LDH inhibitor, oxamate, was found in the Taxol-resistant cells. Furthermore, treating cells with the combination of Taxol and oxamate showed a synergistical inhibitory effect on Taxol-resistant breast cancer cells by promoting apoptosis in these cells. Conclusion LDH-A plays an important role in Taxol resistance and inhibition of LDH-A re-sensitizes Taxol-resistant cells to Taxol. This supports that Warburg effect is a property of Taxol resistant cancer cells and may play an important role in the development of Taxol resistance. To our knowledge, this is the first report showing that the increased expression of LDH-A plays an important role in Taxol resistance of human breast cancer cells. This study provides valuable information for the future development and use of targeted therapies, such as oxamate, for the treatment of patients with Taxol-resistant breast cancer. PMID:20144215

2010-01-01

132

Lactate Dehydrogenase-B Is Silenced by Promoter Methylation in a High Frequency of Human Breast Cancers  

PubMed Central

Objective Under normoxia, non-malignant cells rely on oxidative phosphorylation for their ATP production, whereas cancer cells rely on Glycolysis; a phenomenon known as the Warburg effect. We aimed to elucidate the mechanisms contributing to the Warburg effect in human breast cancer. Experimental design Lactate Dehydrogenase (LDH) isoenzymes were profiled using zymography. LDH-B subunit expression was assessed by reverse transcription PCR in cells, and by Immunohistochemistry in breast tissues. LDH-B promoter methylation was assessed by sequencing bisulfite modified DNA. Results Absent or decreased expression of LDH isoenzymes 1-4, were seen in T-47D and MCF7 cells. Absence of LDH-B mRNA was seen in T-47D cells, and its expression was restored following treatment with the demethylating agent 5'Azacytadine. LDH-B promoter methylation was identified in T-47D and MCF7 cells, and in 25/ 25 cases of breast cancer tissues, but not in 5/ 5 cases of normal breast tissues. Absent immuno-expression of LDH-B protein (<10% cells stained), was seen in 23/ 26 (88%) breast cancer cases, and in 4/8 cases of adjacent ductal carcinoma in situ lesions. Exposure of breast cancer cells to hypoxia (1% O2), for 48 hours resulted in significant increases in lactate levels in both MCF7 (14.0 fold, p?=?0.002), and T-47D cells (2.9 fold, p?=?0.009), but not in MDA-MB-436 (-0.9 fold, p?=?0.229), or MCF10AT (1.2 fold, p?=?0.09) cells. Conclusions Loss of LDH-B expression is an early and frequent event in human breast cancer occurring due to promoter methylation, and is likely to contribute to an enhanced glycolysis of cancer cells under hypoxia. PMID:23437403

Brown, Nicola J.; Higham, Sue E.; Perunovic, Branko; Arafa, Mohammad; Balasubramanian, Sabapathy; Rehman, Ishtiaq

2013-01-01

133

Membrane association of the C-terminal half of the open reading frame 1a protein of lactate dehydrogenase-elevating virus  

Microsoft Academic Search

Summary ORF 1a of lactate dehydrogenase-elevating virus, strain P (LDV-P), encodes a protein of 2206 amino acids. Eisenberg hydrophobic moment analysis of the protein predicted the presence of eleven transmembrane segments in the C-terminal half of the molecule (amino acids 980–1852) that flank the serine protease domain. cDNAs encoding ORF 1a protein segments encompassing transmembrane segments 5 to 11 and

K. S. Faaberg; P. G. W. Plagemann

1996-01-01

134

The interaction of solutes and temperature on A4-lactate dehydrogenase orthologs from warm-adapted and cold-adapted marine fishes  

Microsoft Academic Search

We examined the effects of temperature and stabilizing solutes on A4-lactate dehydrogenase (A4-LDH) from warm- and cold-adapted fishes, to determine how extrinsic stabilizers affect orthologs with different intrinsic stabilities. Conformational changes during substrate binding are rate- limiting for A4-LDH, thus stabilization due to intrinsic or extrinsic factors leads to decreased activity. A4-LDH from a warm-temperate goby (Gillichthys mirabilis ), which

Peter A. Fields; Benjamin D. Wahlstrand; George N. Somero

135

Diacetyl and ?-Acetolactate Overproduction by Lactococcus lactis subsp. lactis Biovar Diacetylactis Mutants That Are Deficient in ?-Acetolactate Decarboxylase and Have a Low Lactate Dehydrogenase Activity  

PubMed Central

Lactococcus lactis subsp. lactis biovar diacetylactis strains are utilized in several industrial processes for producing the flavoring compound diacetyl or its precursor ?-acetolactate. Using random mutagenesis with nitrosoguanidine, we selected mutants that were deficient in ?-acetolactate decarboxylase and had low lactate dehydrogenase activity. The mutants produced large amounts of ?-acetolactate in anaerobic milk cultures but not in aerobic cultures, except when the medium was supplemented with catalase, yeast extract, or hemoglobin. PMID:11097941

Monnet, Christophe; Aymes, Frédéric; Corrieu, Georges

2000-01-01

136

Clinicopathological Significance and Prognostic Value of Lactate Dehydrogenase A Expression in Gastric Cancer Patients  

PubMed Central

Introduction LDH-A, the enzyme responsible for transforming pyruvate into lactate, has been demonstrated to be up-regulated in many types of cancer and to give rise to more aggressive behavior by regulating proliferation and anti-apoptosis. However, its expression in gastric cancer (GC) has not been characterized thoroughly. The purpose of this study was to clarify the expression and potential impact of LDH-A in GC. Methods We examined LDH-A expression by immunohistochemistry on GC tissue microarray (TMA) and using Western blot on fresh GC tissues and cell lines. Prognostic value and correlation with other clinicopathologic factors were evaluated. We transfected siRNA into GC cells against LDH-A. LDH-A was analyzed by Western blotting and real-time RT-PCR. Cell growth was evaluated in vitro and in vivo. Lactate and ATP production by cells were determined. Results There was significantly higher LDH-A expression in carcinoma than in non-neoplastic mucosa (NNM). There was a positive correlation of LDH-A expression with age, histological type and Lymph node metastases. Survival analysis demonstrated that high expression of LDH-A in GC was associated with lower overall survival (OS). When stratified by Lauren grade and histological classification, significance appeared in diffuse type and undifferentiated type GC. In multivariate analysis, the LDH-A expression in GC was an independent prognostic risk factor for OS (hazard ratio?=?1.829, 95%CI 1.375–2.433,P<0.0001). Specific siRNA against LDH-A in GC cell line retarded cell growth both in vitro and in mouse models. LDH-A knockdown also reduced lactate and ATP production in GC cells. Conclusions Our study indicated the oncogenic role of LDH-A in GC. LDH-A expression is an independent prognostic risk factor in GC patients and up-regulated expression of LDH-A could be predictive of poor outcomes in diffuse type and undifferentiated type GC. Our results suggested that LDH-A might be a potential therapeutic target in gastric cancer. PMID:24608789

Sun, Xuren; Sun, Zhe; Zhu, Zhi; Guan, Haixia; Zhang, Junyan; Zhang, Yining; Xu, Huimian; Sun, Mingjun

2014-01-01

137

Escherichia coli derivatives lacking both alcohol dehydrogenase and phosphotransacetylase grow anaerobically by lactate fermentation  

SciTech Connect

Escherichia coli mutants lacking alcohol dehydrogenase (adh mutants) cannot synthesize the fermentation product ethanol and are unable to grow anaerobically on glucose and other hexoses. Similarly, phosphotransacetylase-negative mutants (pta mutants) neither excrete acetate nor grow anaerobically. However, when a strain carrying an adh deletion was selected for anaerobic growth on glucose, spontaneous pta mutants were isolated. Strains carrying both adh and pta mutations were observed by in vivo nuclear magnetic resonance and shown to produce lactic acid as the major fermentation product. Various combinations of adh pta double mutants regained the ability to grow anaerobically on hexoses, by what amounts to a homolactic fermentation. Unlike wild-type strains, such adh pta double mutants were unable to grow anaerobically on sorbitol or on glucuronic acid. The growth properties of strains carrying various mutations affecting the enzymes of fermentation are discussed terms of redox balance.

Gupta, S.; Clark, D.P. (Southern Illinois Univ., Carbondale (USA))

1989-07-01

138

Large Scale Dynamics of the Michaelis Complex in B. stearothermophilus Lactate Dehydrogenase Revealed by Single Tryptophan Mutants Study  

PubMed Central

Large scale dynamics within the Michaelis complex mimic of Bacillus stearothermophilus thermophylic lactate dehydrogenase, bsLDH•NADH•oxamate, were studied with site specific resolution by laser induced temperature jump relaxation spectroscopy having a time resolution of 20 ns. NADH emission and Trp emission from the wild type and a series of single-tryptophan bsLDH mutants, with the tryptophan positions at different distances from the active site, were used as reporters of evolving structure in response to the rapid change in temperature. Several distinct dynamical events were observed on the ms - ?s time-scale involving motion of atoms spread over the protein, some occurring concomitantly or nearly concomitantly with structural changes at the active site. This suggests that a large portion of the protein-substrate complex moves in a rather concerted fashion to bring about catalysis. The catalytically important surface loop undergoes two distinct movements, both needed for a competent enzyme. Our results also suggest that what is called `loop motion' is not just localized to the loop and active site residues. Rather, it involves the motion of atoms spread over the protein, even some quite distal from the active site. How these results bear on catalytic mechanism of bsLDH is discussed. PMID:23428201

Nie, Beining; Deng, Hua; Desamero, Ruel; Callender, Robert

2013-01-01

139

Large scale dynamics of the Michaelis complex in Bacillus stearothermophilus lactate dehydrogenase revealed by a single-tryptophan mutant study.  

PubMed

Large scale dynamics within the Michaelis complex mimic of Bacillus stearothermophilus thermophilic lactate dehydrogenase, bsLDH·NADH·oxamate, were studied with site specific resolution by laser-induced temperature jump relaxation spectroscopy with a time resolution of 20 ns. NADH emission and Trp emission from the wild type and a series of single-tryptophan bsLDH mutants, with the tryptophan positions different distances from the active site, were used as reporters of evolving structure in response to the rapid change in temperature. Several distinct dynamical events were observed on the millisecond to microsecond time scale involving motion of atoms spread over the protein, some occurring concomitantly or nearly concomitantly with structural changes at the active site. This suggests that a large portion of the protein-substrate complex moves in a rather concerted fashion to bring about catalysis. The catalytically important surface loop undergoes two distinct movements, both needed for a competent enzyme. Our results also suggest that what is called "loop motion" is not just localized to the loop and active site residues. Rather, it involves the motion of atoms spread over the protein, even some quite distal from the active site. How these results bear on the catalytic mechanism of bsLDH is discussed. PMID:23428201

Nie, Beining; Deng, Hua; Desamero, Ruel; Callender, Robert

2013-03-19

140

Introduction of a (poly)histidine tag in L-lactate dehydrogenase produces a mixture of active and inactive molecules.  

PubMed

A (poly)histidine tag was fused to either the N- or the C-terminus of L-lactate dehydrogenase (LDH) of Bacillus stearothermophilus to facilitate purification and immobilization of these enzymes. The C-terminally tagged enzyme displayed lower activity compared both to the wild-type and to the N-terminally tagged variant. The reason for this loss of activity was investigated by affinity chromatography of the enzymes on a 5'-AMP-Sepharose resin and by size-exclusion chromatography. The C-terminally tagged enzyme could be separated into an inactive, unbound fraction and an active, bound fraction. Further differences between the C-terminally tagged enzyme and the N-terminally tagged and wild-type LDH were observed on size-exclusion chromatography of the three enzymes. These data suggest that the introduction of a "his-tag" at the C-terminus may induce misfolding of the LDH and serve as a warning that the introduction of a (poly)histidine tag can produce unforseen changes in a protein. PMID:11488630

Halliwell, C M; Morgan, G; Ou, C P; Cass, A E

2001-08-15

141

Adsorption of lactate dehydrogenase enzyme on carbon nanotubes: how to get accurate results for the cytotoxicity of these nanomaterials.  

PubMed

Carbon nanotube (CNT) cytotoxicity is frequently investigated using in vitro classical toxicology assays. However, these cellular tests, usually based on the use of colorimetric or fluorimetric dyes, were designed for chemicals and may not be suitable for nanosized materials. Indeed, because of their unique physicochemical properties CNT can interfere with the assays and bias the results. To get accurate data and draw reliable conclusions, these artifacts should be carefully taken into account. The aim of this study was to evaluate qualitatively and quantitatively the interferences occurring between CNT and the commonly used lactate dehydrogenase (LDH) assay. Experiments under cell-free conditions were performed, and it was clearly demonstrated that artifacts occurred. They were due to the intrinsic absorbance of CNT on one hand and the adsorption of LDH at the CNT surface on the other hand. The adsorption of LDH on CNT was modeled and was found to fit the Langmuir model. The Kads and neq constants were defined, allowing the correction of results obtained from cellular experiments to get more accurate data and lead to proper conclusions on the cytotoxicity of CNT. PMID:25768724

Forest, Valérie; Figarol, Agathe; Boudard, Delphine; Cottier, Michèle; Grosseau, Philippe; Pourchez, Jérémie

2015-03-31

142

Purification and Properties of White Muscle Lactate Dehydrogenase from the Anoxia-Tolerant Turtle, the Red-Eared Slider, Trachemys scripta elegans  

PubMed Central

Lactate dehydrogenase (LDH; E.C. 1.1.1.27) is a crucial enzyme involved in energy metabolism in muscle, facilitating the production of ATP via glycolysis during oxygen deprivation by recycling NAD+. The present study investigated purified LDH from the muscle of 20?h anoxic and normoxic T. s. elegans, and LDH from anoxic muscle showed a significantly lower (47%) Km for L-lactate and a higher Vmax value than the normoxic form. Several lines of evidence indicated that LDH was converted to a low phosphate form under anoxia: (a) stimulation of endogenously present protein phosphatases decreased the Km of L-lactate of control LDH to anoxic levels, whereas (b) stimulation of kinases increased the Km of L-lactate of anoxic LDH to normoxic levels, and (c) dot blot analysis shows significantly less serine (78%) and threonine (58%) phosphorylation in anoxic muscle LDH as compared to normoxic LDH. The physiological consequence of anoxia-induced LDH dephosphorylation appears to be an increase in LDH activity to promote the reduction of pyruvate in muscle tissue, converting the glycolytic end product to lactate to maintain a prolonged glycolytic flux under energy-stressed anoxic conditions. PMID:23533717

Dawson, Neal J.; Bell, Ryan A. V.; Storey, Kenneth B.

2013-01-01

143

An alternative allosteric regulation mechanism of an acidophilic l-lactate dehydrogenase from Enterococcus mundtii 15-1A  

PubMed Central

A plant-derived Enterococcus mundtii 15-1A, that has been previously isolated from Brassica rapa L. subsp. nipposinica (L.H. Bailey) Hanelt var. linearifolia by our group, possesses two kinds of l-lactate dehydrogenase (l-LDH): LDH-1 and LDH-2. LDH-1 was activated under low concentration of fluctose-1,6-bisphosphate (FBP) at both pH 5.5 and 7.5. Although LDH-2 was also activated under the low concentration of FBP at pH 5.5, a high concentration of FBP is necessary to activate it at pH 7.5. The present study shows the crystal structures of the acidophilic LDH-2 in a complex with and without FBP and NADH. Although the tertiary structure of the ligands-bound LDH-2 is similar to that of the active form of other bacterial l-LDHs, the structure without the ligands is different from that of any other previously determined l-LDHs. Major structural alterations between the two structures of LDH-2 were observed at two regions in one subunit. At the N-terminal parts of the two regions, the ligands-bound form takes an ?-helical structure, while the form without ligands displays more disordered and extended structures. A vacuum-ultraviolet circular dichroism analysis showed that the ?-helix content of LDH-2 in solution is approximately 30% at pH 7.5, which is close to that in the crystal structure of the form without ligands. A D241N mutant of LDH-2, which was created by us to easily form an ?-helix at one of the two parts, exhibited catalytic activity even in the absence of FBP at both pH 5.5 and 7.5. PMID:25379380

Matoba, Yasuyuki; Miyasako, Masashi; Matsuo, Koichi; Oda, Kosuke; Noda, Masafumi; Higashikawa, Fumiko; Kumagai, Takanori; Sugiyama, Masanori

2014-01-01

144

Elevated lactate dehydrogenase activity and increased cardiovascular mortality in the arsenic-endemic areas of southwestern Taiwan  

SciTech Connect

Arsenic ingestion has been linked to increasing global prevalence of and mortality from cardiovascular disease (CVD); arsenic can be removed from drinking water to reduce related health effects. Lactate dehydrogenase (LDH) is used for the evaluation of acute arsenic toxicity in vivo and in vitro, but it is not validated for the evaluation of long-term, chronic arsenic exposure. The present study examined the long-term effect of chronic arsenic exposure on CVD and serum LDH levels, after consideration of arsenic metabolism capacity. A total of 380 subjects from an arseniasis-endemic area and 303 from a non-endemic area of southwestern Taiwan were recruited in 2002. Various urinary arsenic species were analyzed using high-performance liquid chromatography (HPLC) and hydride generation systems. Fasting serum was used for quantitative determination of the total LDH activity. A significant dose–response relationship was observed between arsenic exposure and LDH elevation, independent of urinary arsenic profiles (P < 0.001). Furthermore, abnormal LDH elevation was associated with CVD mortality after adjustment for Framingham risk scores for 10-year CVD and arsenic exposure (hazard ratio, 3.98; 95% confidence interval, 1.07–14.81). LDH was elevated in subjects with arsenic exposure in a dose-dependent manner. LDH is a marker of arsenic toxicity associated with CVD mortality. Results of this study have important implications for use in ascertaining long-term arsenic exposure risk of CVD. -- Highlights: ? We showed that arsenic exposure was correlated with LDH elevation. ? LDH elevation was related to arsenic methylation capacity. ? Abnormal LDH elevation can be a marker of susceptibility to CVD mortality.

Liao, Ya-Tang [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China) [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China); Graduate Institute of Epidemiology and Preventive Medicine, College of Public Health, National Taiwan University, Taiwan (China); Genomics Research Center, Academia Sinica, Taiwan (China); Chen, Chien-Jen [Graduate Institute of Epidemiology and Preventive Medicine, College of Public Health, National Taiwan University, Taiwan (China) [Graduate Institute of Epidemiology and Preventive Medicine, College of Public Health, National Taiwan University, Taiwan (China); Genomics Research Center, Academia Sinica, Taiwan (China); Li, Wan-Fen [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China)] [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China); Hsu, Ling-I [Genomics Research Center, Academia Sinica, Taiwan (China)] [Genomics Research Center, Academia Sinica, Taiwan (China); Tsai, Li-Yu; Huang, Yeou-Lih [Department of Medical Laboratory Science and Biotechnology, Kaohsiung Medical University, Taiwan (China)] [Department of Medical Laboratory Science and Biotechnology, Kaohsiung Medical University, Taiwan (China); Sun, Chien-Wen [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China)] [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China); Chen, Wei J., E-mail: wjchen@ntu.edu.tw [Graduate Institute of Epidemiology and Preventive Medicine, College of Public Health, National Taiwan University, Taiwan (China); Genetic Epidemiology Core Laboratory, National Taiwan University Center for Genomic Medicine, Taiwan (China); Wang, Shu-Li, E-mail: slwang@nhri.org.tw [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China) [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China); Department of Public Health, College of Public Health, China Medical University, Taichung, Taiwan (China)

2012-08-01

145

Effect of high-pressure processing on activity and structure of alkaline phosphatase and lactate dehydrogenase in buffer and milk.  

PubMed

Changes in the activity and structure of alkaline phosphatase (ALP) and L-lactate dehydrogenase (LDH) were investigated after high pressure processing (HPP). HPP treatments (206-620 MPa for 6 and 12 min) were applied to ALP and LDH prepared in buffer, fat-free milk, and 2% fat milk. Enzyme activities were measured using enzymatic assays, and changes in structure were investigated using far-ultraviolet circular dichroism (CD) spectroscopy and dynamic light scattetering (DLS). Kinetic data indicated that the activity of ALP was not affected after 6 min of pressure treatments (206-620 MPa), regardless of the medium in which the enzyme was prepared. Increasing the processing time to 12 min did significantly reduce the activity of ALP at 620 MPa (P < 0.001). However, even the lowest HPP treatment of 206 MPa induced a reduction in LDH activity, and the course of reduction increased with HPP treatment until complete inactivation at 482, 515, and 620 MPa. CD data demonstrated a partial change in the secondary structure of ALP at 620 MPa, whereas the structure of LDH showed gradual denaturation after exposure at 206 MPa for 6 min, leading to a random coil structure at both 515 and 620 MPa. DLS results indicated aggregation of ALP only at HPP treatment of 206 MPa and not above and enzyme precipitation as well as aggregation at 345, 415, 482, and 515 MPa. The loss of LDH activity with increasing pressure and time treatment was due to the combined effects of denaturation and aggregation. PMID:17944537

Kouassi, Gilles K; Anantheswaran, Ramaswamy C; Knabel, Stephen J; Floros, John D

2007-11-14

146

Serum lactate dehydrogenase profile as a retrospective indicator of uterine preparedness for labor: a prospective, observational study  

PubMed Central

Background Lactate dehydrogenase (LDH) isoenzymes are required for adenosine triphosphate production, with each of five different isoenzymes having varying proficiencies in anaerobic versus aerobic environments. With advancing pregnancy, the isoenzyme profile in uterine muscle shifts toward a more anaerobic profile, speculatively to facilitate uterine efficiency during periods of low oxygen that accompany labor contractions. Profile shifting may even occur throughout labor. Maternal serum LDH levels between 24–48 hours following delivery predominantly originate from uterine muscle, reflecting the enzymatic state of the myometrium during labor. Our purpose was to describe serum LDH isoenzymes 24–30 hours post-delivery to determine if cervical dilation rates following labor admission were associated with a particular LDH profile. We also compared differences in post-delivery LDH isoenzyme profiles between women admitted in pre-active versus established active labor. Methods Low-risk, nulliparous women with spontaneous labor onset were sampled (n?=?91). Maternal serum LDH was measured at labor admission and 24–30 hours post-vaginal delivery. Rates of cervical dilation during the first four hours after admission were also measured. Spearman’s rho coefficients were used for association testing and t tests evaluated for group and paired-sample differences. Results More efficient dilation following admission was associated with decreased LDH1 (p?=?0.029) and increased LDH3 and LDH4 (p?=?0.017 and p?=?0.017, respectively) in the post-delivery period. Women admitted in established active labor had higher relative serum levels of LDH3 (t?=?2.373; p?=?0.023) and LDH4 (t?=?2.268; p?=?0.029) and lower levels of LDH1 (t?=?2.073; p?=?0.045) and LDH5 (t?=?2.041; p?=?0.048) when compared to women admitted in pre-active labor. Despite having similar dilatations at admission (3.4?±?0.5 and 3.7?±?0.6 cm, respectively), women admitted in pre-active labor had longer in-hospital labor durations (12.1?±?4.3 vs. 5.3?±?1.4 hours; p?

2013-01-01

147

Lactate dehydrogenase-5 (LDH-5) overexpression in non-small-cell lung cancer tissues is linked to tumour hypoxia, angiogenic factor production and poor prognosis.  

PubMed

Lactate dehydrogenase-5 (LDH-5) catalyses the reversible transformation of pyruvate to lactate, having a principal position in the anaerobic cellular metabolism. Induction of LDH-5 occurs during hypoxia and LDH-5 transcription is directly regulated by the hypoxia-inducible factor 1 (HIF1). Serum LDH levels have been correlated with poor prognosis and resistance to chemotherapy and radiotherapy in various neoplastic diseases. The expression, however, of LDH in tumours has never been investigated in the past. In the present study, we established an immunohistochemical method to evaluate the LDH-5 overexpression in tumours, using two novel antibodies raised against the rat muscle LDH-5 and the human LDH-5 (Abcam, UK). The subcellular patterns of expression in cancer cells were mixed nuclear and cytoplasmic. In direct contrast to cancer cells, stromal fibroblasts were reactive for LDH-5 only in a minority of cases. Serum LDH, although positively correlated with, does not reliably reflect the intratumoral LDH-5 status. Lactate dehydrogenase-5 overexpression was directly related to HIF1alpha and 2alpha, but not with the carbonic anhydrase 9 expression. Patients with tumours bearing high LDH-5 expression had a poor prognosis. Tumours with simultaneous LDH-5 and HIF1alpha (or HIF2alpha) overexpression, indicative of a functional HIF pathway, had a particularly aggressive behaviour. It is concluded that overexpression of LDH-5 is a common event in non-small-cell lung cancer, can be easily assessed in paraffin-embedded material and provides important prognostic information, particularly when combined with other endogenous markers of hypoxia and acidity. PMID:12942121

Koukourakis, M I; Giatromanolaki, A; Sivridis, E; Bougioukas, G; Didilis, V; Gatter, K C; Harris, A L

2003-09-01

148

The Effects of Heart and Skeletal Muscle Inflammation and Cardiomyopathy Syndrome on Creatine Kinase and Lactate Dehydrogenase Levels in Atlantic Salmon (Salmo salar L.)  

PubMed Central

Heart and skeletal muscle inflammation (HSMI) and cardiomyopathy syndrome (CMS) are putative viral cardiac diseases of Atlantic salmon. This study examined the levels and correlated the serum enzymes creatine kinase (CK) and lactate dehydrogenase (LDH) to the histopathology of clinical outbreaks of HSMI and chronic CMS in farmed Atlantic salmon. A total of 75 fish from 3 different HSMI outbreaks, 30 chronic CMS fish, and 68 fish from 3 nondiseased fish groups were used as the study population (N = 173). Serum CK and LDH levels correlated significantly with the total inflammation and total necrosis scores for HSMI fish (P = 0.001). However, no correlation was identified for enzyme levels and histopathology scores for chronic CMS fish. The significantly increased CK and LDH levels and their positive correlations to histopathology differentiate HSMI from CMS clinically suggesting the potential use of enzymes for screening for HSMI is promising. PMID:22701371

Yousaf, Muhammad Naveed; Powell, Mark D.

2012-01-01

149

Lactate dehydrogenase genes of caiman and Chinese soft-shelled turtle, with emphasis on the molecular phylogenetics and evolution of reptiles.  

PubMed

L-Lactate dehydrogenase (LDH) cDNAs encoding for LDH-A(4) (muscle) and LDH-B(4) (heart) isozymes from caiman (Caiman crocodilus apaporiensis) belonging to the order Crocodilia and Chinese soft-shelled turtle (Pelodiscus sinensis) belonging to the order Chelonia were sequenced. The phylogenetic relationships of the newly determined cDNA and their deduced protein sequences, as well as the previously published sequences of vertebrate LDH isozymes, were analyzed by various phylogenetic tree construction methods. These results indicated that Chelonia is indeed more closely related to Crocodilia. The divergent times between caiman and alligator, turtle and soft-shelled turtle, and Chelonia and Crocodilia were estimated to be approximately 36, 100 and 177 million years, respectively. PMID:11722846

Liao, C H; Ho, W Z; Huang, H W; Kuo, C H; Lee, S C; Li, S S

2001-11-14

150

The effects of heart and skeletal muscle inflammation and cardiomyopathy syndrome on creatine kinase and lactate dehydrogenase levels in Atlantic salmon (Salmo salar L.).  

PubMed

Heart and skeletal muscle inflammation (HSMI) and cardiomyopathy syndrome (CMS) are putative viral cardiac diseases of Atlantic salmon. This study examined the levels and correlated the serum enzymes creatine kinase (CK) and lactate dehydrogenase (LDH) to the histopathology of clinical outbreaks of HSMI and chronic CMS in farmed Atlantic salmon. A total of 75 fish from 3 different HSMI outbreaks, 30 chronic CMS fish, and 68 fish from 3 nondiseased fish groups were used as the study population (N = 173). Serum CK and LDH levels correlated significantly with the total inflammation and total necrosis scores for HSMI fish (P = 0.001). However, no correlation was identified for enzyme levels and histopathology scores for chronic CMS fish. The significantly increased CK and LDH levels and their positive correlations to histopathology differentiate HSMI from CMS clinically suggesting the potential use of enzymes for screening for HSMI is promising. PMID:22701371

Yousaf, Muhammad Naveed; Powell, Mark D

2012-01-01

151

Oxamate-mediated inhibition of lactate dehydrogenase induces protective autophagy in gastric cancer cells: involvement of the Akt-mTOR signaling pathway.  

PubMed

Cancer cells produce a substantial amount of energy through aerobic glycolysis even in the presence of adequate oxygen. Lactate dehydrogenase (LDH), a key regulator of glycolysis, reversibly catalyzes the conversion of pyruvate to lactate. Recently, oxamate, an inhibitor of LDH, has been shown to be a promising anticancer agent. However, the detailed mechanism remains largely unclear. In this study, we demonstrate that oxamate inhibits the viability of human gastric cancer cells in a dose- and time-dependent manner. In addition, treatment with oxamate induces protective autophagy in gastric cancer cells. Moreover, autophagy inhibited by chloroquine or Beclin 1 small interfering RNA (siRNA) enhances oxamate-induced apoptosis and proliferation inhibition. Further study has shown that oxamate treatment significantly augments reactive oxygen species (ROS) production. Furthermore, cells pretreated with N-acetyl cysteine (NAC), a ROS inhibitor, display significantly reduced ROS production and attenuated oxamate-induced autophagy. Finally, functional studies reveal that the Akt-mTOR signaling pathway, a major negative regulator of autophagy, is inhibited by oxamate. Together, our results provide new insights regarding the biological and anti-proliferative activities of oxamate against gastric cancer, and may offer a promising therapeutic strategy for gastric cancer. PMID:25524555

Zhao, Zhi; Han, Fanghai; Yang, Shibin; Wu, Jianhai; Zhan, Wenhua

2015-03-01

152

Stable Suppression of Lactate Dehydrogenase Activity during Anoxia in the Foot Muscle of Littorina littorea and the Potential Role of Acetylation as a Novel Posttranslational Regulatory Mechanism  

PubMed Central

The intertidal marine snail, Littorina littorea, has evolved to withstand extended bouts of oxygen deprivation brought about by changing tides or other potentially harmful environmental conditions. Survival is dependent on a strong suppression of its metabolic rate and a drastic reorganization of its cellular biochemistry in order to maintain energy balance under fixed fuel reserves. Lactate dehydrogenase (LDH) is a crucial enzyme of anaerobic metabolism as it is typically responsible for the regeneration of NAD+, which allows for the continued functioning of glycolysis in the absence of oxygen. This study compared the kinetic and structural characteristics of the D-lactate specific LDH (E.C. 1.1.1.28) from foot muscle of aerobic control versus 24?h anoxia-exposed L. littorea. Anoxic LDH displayed a near 50% decrease in Vmax (pyruvate-reducing direction) as compared to control LDH. These kinetic differences suggest that there may be a stable modification and regulation of LDH during anoxia, and indeed, subsequent dot-blot analyses identified anoxic LDH as being significantly less acetylated than the corresponding control enzyme. Therefore, acetylation may be the regulatory mechanism that is responsible for the suppression of LDH activity during anoxia, which could allow for the production of alternative glycolytic end products that in turn would increase the ATP yield under fixed fuel reserves. PMID:24233354

Shahriari, Ali; Dawson, Neal J.; Bell, Ryan A. V.; Storey, Kenneth B.

2013-01-01

153

Expression, purification, crystallization and preliminary X-ray crystallographic analysis of L-lactate dehydrogenase and its H171C mutant from Bacillus subtilis  

SciTech Connect

L-Lactate dehydrogenase (LDH) is an important enzyme involved in the last step of glycolysis that catalyzes the reversible conversion of pyruvate to L-lactate with the simultaneous oxidation of NADH to NAD{sup +}. In this study, wild-type LDH from Bacillus subtilis (BsLDH-WT) and the H171C mutant (BsLDH-H171C) were expressed in Escherichia coli and purified to near-homogeneity. BsLDH-WT was crystallized in the presence of fructose 1,6-bisphosphate (FBP) and NAD{sup +} and the crystal diffracted to 2.38 {angstrom} resolution. The crystal belonged to space group P3, with unit-cell parameters a = b = 171.04, c = 96.27 {angstrom}. BsLDH-H171C was also crystallized as the apoenzyme and in complex with NAD{sup +}, and data sets were collected to 2.20 and 2.49 {angstrom} resolution, respectively. Both BsLDH-H171C crystals belonged to space group P3, with unit-cell parameters a = b = 133.41, c = 99.34 {angstrom} and a = b = 133.43, c = 99.09 {angstrom}, respectively. Tetramers were observed in the asymmetric units of all three crystals.

Zhang, Yanfeng; Gao, Xiaoli (MSU)

2012-08-31

154

Reaction rate studies of glucose-6-phosphate dehydrogenase activity in sections of rat liver using four tetrazolium salts  

Microsoft Academic Search

Summary  The reaction rate of glucose-6-phosphate dehydrogenase activity in liver sections from fed and starved rats has been monitored by the continuous measurement at 37 C of the reaction product as it is formed using scanning and integrating microdensitometry. Control media lacked either substrate or both substrate and coenzyme. All reactions were nonlinear; however, subtraction of either of the controls from

R. G. Butcher; C. J. F. Van Noorden

1985-01-01

155

Reaction rate studies of glucose-6-phosphate dehydrogenase in rat tracheal epithelium; the effect of section thickness  

Microsoft Academic Search

The reaction velocity of glucose-6-phosphate dehydrogenase has been quantified by continuous monitoring on a Vickers microdensitometer of the reaction product as it formed in sections of different thickness of rat tracheal epithelium. Reaction velocity was directly proportional to section thickness when either tetranitro BT or neotetrazolium was used as the final acceptor; the rate was the same with each tetrazolium

R. G. Butcher

1984-01-01

156

Dependence of Substrate Irradiation Reaction Rate Stimulation on Lactic Dehydrogenase Source  

E-print Network

Stimulation of LDH initial reaction rates by timed pre-irradiation of crystalline sodium pyruvate and lithium lactate is reported for enzymes isolated from rabbit muscle, pig heart, human erythrocytes and chicken heart. The phenomenon investigated is referred to as the Comorosan effect. For the mammalian source enzymes, the pyruvate irradiation stimulations occurred at irradiation times of 5 and 35 sec. and the lactate irradiation times at 15 and 45 seconds. In contrast, for the chicken heart enzyme, the pyruvate irradiation stimulations occurred at 15 and 35 sec., while those for lactate occurred at 5 and 20 sec. Thus, a shift in stimulatory irradiation times is found on going from the mammalian enzymes to the avian enzyme. A similar shift between mammalian and yeast enzymes has been established by Comorosan and co-workers. For the chicken heart LDH, the separation between successive irradiation times is different for the forward and reverse reactions. This is the first reported incidence of the separation not being the same.

George Bass; James Chenevey

2013-04-16

157

The enzymatic reaction-induced configuration change of the prosthetic group PQQ of methanol dehydrogenase.  

PubMed

Methanol dehydrogenase is a heterotetrameric enzyme containing the prosthetic group pyrroloquinoline quinone (PQQ), which catalyzes the oxidation of methanol to formaldehyde. The crystal structure of methanol dehydrogenase from Methylophilus W3A1, previously determined at high resolution, exhibits a non-planar configuration of the PQQ ring system and lends support for a hydride transfer mechanism of the enzymatic reaction catalyzed by the enzyme. To investigate why PQQ is in the C5-reduced form and to better understand the catalytic mechanism of the enzyme, three structures of this enzyme in a new crystal form have been determined at higher resolution. Two of the three crystals were grown in the presence of 1 and 50 mM methanol, respectively, both structures of which show non-planar configurations of the PQQ ring system, confirming the previous conclusion; the other was crystallized in the presence of 50 mM ethanol, the structure of which displays a planar ring system for PQQ. Comparison of these structures reveals that the configuration change of PQQ is induced by the enzymatic reaction. The reaction takes place and the C5-reduced PQQ intermediate is produced when the enzyme co-crystallizes with methanol, but the enzymatic reaction does not take place and the PQQ ring retains a planar configuration of the oxidized orthoquinone form when ethanol instead of methanol is present in the crystallization solution. PMID:21356200

Li, Jie; Gan, Jian-Hua; Mathews, F Scott; Xia, Zong-Xiang

2011-03-25

158

Use of tritiated prostaglandins in metabolism studies. I: Evaluation of the kinetic isotope effect in the prostaglandin dehydrogenase reactions  

SciTech Connect

Although numerous data exist concerning tritium kinetic isotope effect in enzymic reactions, little is related to the metabolism of tritiated prostaglandins. The present study reports an evaluation of the kinetic isotope effect which occurs during the oxidation of 15-hydroxyl group of tritium-labeled prostaglandins E2 and F2 alpha by the 15-hydroxyprostaglandin dehydrogenase and during the oxidation of 9-hydroxyl group of tritium-labeled prostaglandin F2 alpha by the 9-hydroxyprostaglandin dehydrogenase. The large kinetic isotope effect tends to limit the validity of the dehydrogenase assay using tritium-labeled prostaglandins as substrate. However these assays can be considered to be an indication of relative enzyme activity.

Moussard, C.; Alber, D.; Perruche, C.; Henry, J.C.

1986-03-01

159

Catalytic reaction of cytokinin dehydrogenase: preference for quinones as electron acceptors.  

PubMed Central

The catalytic reaction of cytokinin oxidase/dehydrogenase (EC 1.5.99.12) was studied in detail using the recombinant flavoenzyme from maize. Determination of the redox potential of the covalently linked flavin cofactor revealed a relatively high potential dictating the type of electron acceptor that can be used by the enzyme. Using 2,6-dichlorophenol indophenol, 2,3-dimethoxy-5-methyl-1,4-benzoquinone or 1,4-naphthoquinone as electron acceptor, turnover rates with N6-(2-isopentenyl)adenine of approx. 150 s(-1) could be obtained. This suggests that the natural electron acceptor of the enzyme is quite probably a p-quinone or similar compound. By using the stopped-flow technique, it was found that the enzyme is rapidly reduced by N6-(2-isopentenyl)adenine (k(red)=950 s(-1)). Re-oxidation of the reduced enzyme by molecular oxygen is too slow to be of physiological relevance, confirming its classification as a dehydrogenase. Furthermore, it was established for the first time that the enzyme is capable of degrading aromatic cytokinins, although at low reaction rates. As a result, the enzyme displays a dual catalytic mode for oxidative degradation of cytokinins: a low-rate and low-substrate specificity reaction with oxygen as the electron acceptor, and high activity and strict specificity for isopentenyladenine and analogous cytokinins with some specific electron acceptors. PMID:14965342

Frébortová, Jitka; Fraaije, Marco W; Galuszka, Petr; Sebela, Marek; Pec, Pavel; Hrbác, Jan; Novák, Ondrej; Bilyeu, Kristin D; English, James T; Frébort, Ivo

2004-01-01

160

Catalytic reaction of cytokinin dehydrogenase: preference for quinones as electron acceptors  

Microsoft Academic Search

The catalytic reaction of cytokinin oxidase\\/dehydrogenase (EC 1.5.99.12) was studied in detail using the recombinant flavoenzyme from maize. Determination of the redox potential of the covalently linked flavin cofactor revealed a relatively high potential dictating the type of electron acceptor that can be used by the enzyme. Using 2,6-dichlorophenol indophenol, 2,3-dimethoxy-5-methyl-1,4-benzoquinone or 1,4-naphthoquinone as electron acceptor, turnover rates with N6-(2-isopentenyl)adenine

James T. English; Kristin D. Bilyeu; Ond?ej Novák; Pavel Pe?; Marek Šebela; Petr Galuszka; Marco W. Fraaije; Ivo Frébort; Jan Hrbá?; Jitka Frébortová

2004-01-01

161

Insulin, CCAAT/Enhancer-Binding Proteins and Lactate Regulate the Human 11?-Hydroxysteroid Dehydrogenase Type 2 Gene Expression in Colon Cancer Cell Lines  

PubMed Central

11?-Hydroxysteroid dehydrogenases (11beta-HSD) modulate mineralocorticoid receptor transactivation by glucocorticoids and regulate access to the glucocorticoid receptor. The isozyme 11beta-HSD2 is selectively expressed in mineralocorticoid target tissues and its activity is reduced in various disease states with abnormal sodium retention and hypertension, including the apparent mineralocorticoid excess. As 50% of patients with essential hypertension are insulin resistant and hyperinsulinemic, we hypothesized that insulin downregulates the 11beta-HSD2 activity. In the present study we show that insulin reduced the 11beta-HSD2 activity in cancer colon cell lines (HCT116, SW620 and HT-29) at the transcriptional level, in a time and dose dependent manner. The downregulation was reversible and required new protein synthesis. Pathway analysis using mRNA profiling revealed that insulin treatment modified the expression of the transcription factor family C/EBPs (CCAAT/enhancer-binding proteins) but also of glycolysis related enzymes. Western blot and real time PCR confirmed an upregulation of C/EBP beta isoforms (LAP and LIP) with a more pronounced increase in the inhibitory isoform LIP. EMSA and reporter gene assays demonstrated the role of C/EBP beta isoforms in HSD11B2 gene expression regulation. In addition, secretion of lactate, a byproduct of glycolysis, was shown to mediate insulin-dependent HSD11B2 downregulation. In summary, we demonstrate that insulin downregulates HSD11B2 through increased LIP expression and augmented lactate secretion. Such mechanisms are of interest and potential significance for sodium reabsorption in the colon. PMID:25133511

Alikhani-Koupaei, Rasoul; Ignatova, Irena D.; Guettinger, Andreas; Frey, Felix J.; Frey, Brigitte M.

2014-01-01

162

21 CFR 184.1311 - Ferrous lactate.  

Code of Federal Regulations, 2011 CFR

...reacting calcium lactate or sodium lactate with ferrous sulfate, direct reaction of lactic acid with iron filings...chloride with sodium lactate, or reaction of ferrous sulfate with ammonium lactate. (b) The ingredient meets...

2011-04-01

163

21 CFR 184.1311 - Ferrous lactate.  

Code of Federal Regulations, 2014 CFR

...reacting calcium lactate or sodium lactate with ferrous sulfate, direct reaction of lactic acid with iron filings...chloride with sodium lactate, or reaction of ferrous sulfate with ammonium lactate. (b) The ingredient meets...

2014-04-01

164

21 CFR 184.1311 - Ferrous lactate.  

Code of Federal Regulations, 2013 CFR

...reacting calcium lactate or sodium lactate with ferrous sulfate, direct reaction of lactic acid with iron filings...chloride with sodium lactate, or reaction of ferrous sulfate with ammonium lactate. (b) The ingredient meets...

2013-04-01

165

21 CFR 184.1311 - Ferrous lactate.  

Code of Federal Regulations, 2012 CFR

...reacting calcium lactate or sodium lactate with ferrous sulfate, direct reaction of lactic acid with iron filings...chloride with sodium lactate, or reaction of ferrous sulfate with ammonium lactate. (b) The ingredient meets...

2012-04-01

166

21 CFR 184.1311 - Ferrous lactate.  

Code of Federal Regulations, 2010 CFR

...reacting calcium lactate or sodium lactate with ferrous sulfate, direct reaction of lactic acid with iron filings...chloride with sodium lactate, or reaction of ferrous sulfate with ammonium lactate. (b) The ingredient meets...

2010-04-01

167

Lactate oxidation at the mitochondria: a lactate-malate-aspartate shuttle at work  

PubMed Central

Lactate, the conjugate base of lactic acid occurring in aqueous biological fluids, has been derided as a “dead-end” waste product of anaerobic metabolism. Catalyzed by the near-equilibrium enzyme lactate dehydrogenase (LDH), the reduction of pyruvate to lactate is thought to serve to regenerate the NAD+ necessary for continued glycolytic flux. Reaction kinetics for LDH imply that lactate oxidation is rarely favored in the tissues of its own production. However, a substantial body of research directly contradicts any notion that LDH invariably operates unidirectionally in vivo. In the current Perspective, a model is forwarded in which the continuous formation and oxidation of lactate serves as a mitochondrial electron shuttle, whereby lactate generated in the cytosol of the cell is oxidized at the mitochondria of the same cell. From this perspective, an intracellular lactate shuttle operates much like the malate-aspartate shuttle (MAS); it is also proposed that the two shuttles are necessarily interconnected in a lactate-MAS. Among the requisite features of such a model, significant compartmentalization of LDH, much like the creatine kinase of the phosphocreatine shuttle, would facilitate net cellular lactate oxidation in a variety of cell types. PMID:25505376

Kane, Daniel A.

2014-01-01

168

Lactate and succinate oxidoreductases in marine invertebrates  

Microsoft Academic Search

Nineteen species of littoral marine invertebrates, representing Porifera, Cnidaria, Ctenophora, Brachiopoda, Mollusca, and Arthropoda, were studied with respect to the ability of tissue extracts to catalyze the lactate and succinate dehydrogenase reactions in both directions. Pyruvate reductase (PR) activity varied tremendously with species, from 0.014 µmole\\/min\\/g of tissue in the etenophore Mnemiopsis leidyi to 145 µ-moles\\/min in leg muscle of

C. S. Hammen

1969-01-01

169

Depressive symptoms of female nursing staff working in stressful environments and their association with serum creatine kinase and lactate dehydrogenase – a preliminary study  

PubMed Central

Background The activity of creatine kinase (CK) in serum has recently been reported to be potentially associated with several types of depression. The aim of this study is to evaluate whether serum enzymes, including CK, vary even in a healthy population with depressive symptoms caused by work-related stress. We gave questionnaires and blood examinations to 93 healthy female nursing home workers and did an enzyme-linked immunosorbent assay for the quantitative detection of CK isozyme muscle-type M chain (CK-MM) in serum. Findings Depressive symptoms were determined using the Center for Epidemiologic Studies Depression (CES-D) scale and compared with the results of the blood examination and serum CK-MM levels. The CES-D results showed significant negative correlations with total CK and lactate dehydrogenase (LDH) activities and CK-MM level (r?=?-0.29, p?=?0.0062; r?=?-0.29, p?=?0.0065; r?=?-0.33, p?=?0.0016, respectively). Conclusions Total CK and LDH activities and serum CK-MM level appear to be associated with the depressive symptoms of healthy nurses working in stressful environments, although the significance level was relatively low. The simultaneous detection of serum CK and LDH activities or serum CK-MM level and LDH activity may be useful as an indicator of depressive symptoms, at least for female nursing staff with work-related stress. PMID:25243019

2014-01-01

170

A population pharmacodynamic model for lactate dehydrogenase and neuron specific enolase to predict tumor progression in small cell lung cancer patients.  

PubMed

The development of individualized therapies poses a major challenge in oncology. Significant hurdles to overcome include better disease monitoring and early prediction of clinical outcome. Current clinical practice consists of using Response Evaluation Criteria in Solid Tumors (RECIST) to categorize response to treatment. However, the utility of RECIST is restricted due to limitations on the frequency of measurement and its categorical rather than continuous nature. We propose a population modeling framework that relates circulating biomarkers in plasma, easily obtained from patients, to tumor progression levels assessed by imaging scans (i.e., RECIST categories). We successfully applied this framework to data regarding lactate dehydrogenase (LDH) and neuron specific enolase (NSE) concentrations in patients diagnosed with small cell lung cancer (SCLC). LDH and NSE have been proposed as independent prognostic factors for SCLC. However, their prognostic and predictive value has not been demonstrated in the context of standard clinical practice. Our model incorporates an underlying latent variable ("disease level") representing (unobserved) tumor size dynamics, which is assumed to drive biomarker production and to be influenced by exposure to treatment; these assumptions are in agreement with the known physiology of SCLC and these biomarkers. Our model predictions of unobserved disease level are strongly correlated with disease progression measured by RECIST criteria. In conclusion, the proposed framework enables prediction of treatment outcome based on circulating biomarkers and therefore can be a powerful tool to help clinicians monitor disease in SCLC. PMID:24740245

Buil-Bruna, Núria; López-Picazo, José-María; Moreno-Jiménez, Marta; Martín-Algarra, Salvador; Ribba, Benjamin; Trocóniz, Iñaki F

2014-05-01

171

Incompatibility of silver nanoparticles with lactate dehydrogenase leakage assay for cellular viability test is attributed to protein binding and reactive oxygen species generation.  

PubMed

A growing number of studies report that conventional cytotoxicity assays are incompatible with certain nanoparticles (NPs) due to artifacts caused by the distinctive characteristics of NPs. Lactate dehydrogenase (LDH) leakage assays have inadequately detected cytotoxicity of silver nanoparticles (AgNPs), leading to research into the underlying mechanism. When ECV304 endothelial-like umbilical cells were treated with citrate-capped AgNPs (cAgNPs) or bare AgNPs (bAgNPs), the plasma membrane was disrupted, but the LDH leakage assay failed to detect cytotoxicity, indicating interference with the assay by AgNPs. Both cAgNPs and bAgNPs inactivated LDH directly when treated to cell lysate as expected. AgNPs adsorbed LDH and thus LDH, together with AgNPs, was removed from assay reactants during sample preparation, with a resultant underestimation of LDH leakage from cells. cAgNPs, but not bAgNPs, generated reactive oxygen species (ROS), which were successfully scavenged by N-acetylcysteine or ascorbic acid. LDH inhibition by cAgNPs could be restored partially by simultaneous treatment with those antioxidants, suggesting the contribution of ROS to LDH inactivation. Additionally, the composition of the protein corona surrounding AgNPs was identified employing liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. In sum, the LDH leakage assay, a conventional cell viability test method, should be employed with caution when assessing cytotoxicity of AgNPs. PMID:24463055

Oh, Seok-Jeong; Kim, Hwa; Liu, Yingqiu; Han, Hyo-Kyung; Kwon, Kyenghee; Chang, Kyung-Hwa; Park, Kwangsik; Kim, Younghun; Shim, Kyuhwan; An, Seong Soo A; Lee, Moo-Yeol

2014-03-21

172

Baseline Serum Lactate Dehydrogenase Levels for Patients Treated With Intensity-Modulated Radiotherapy for Nasopharyngeal Carcinoma: A Predictor of Poor Prognosis and Subsequent Liver Metastasis  

SciTech Connect

Purpose: To evaluate the prognostic value of baseline serum lactate dehydrogenase (LDH) levels in patients with nasopharyngeal carcinoma (NPC) treated with intensity-modulated radiotherapy (IMRT). Methods and Materials: Cases of NPC (n = 465) that involved treatment with IMRT with or without chemotherapy were retrospectively analyzed. Results: The mean ({+-}SD) and median baseline serum LDH levels for this cohort were 172.77 {+-} 2.28 and 164.00 IU/L, respectively. Levels of LDH were significantly elevated in patients with locoregionally advanced disease (p = 0.016). Elevated LDH levels were identified as a prognostic factor for rates of overall survival (OS), disease-free survival (DFS), and distant metastasis-free survival (DMFS), with p values <0.001 in the univariate analysis and p < 0.001, p = 0.004, and p = 0.003, respectively, in the multivariate analysis. Correspondingly, the prognostic impact of patient LDH levels was found to be statistically significant for rates of OS, DFS, and DMFS (p = 0.028, 0.024, and 0.020, respectively). For patients who experienced subsequent liver failure after treatment, markedly higher pretreatment serum LDH levels were detected compared with patients experiencing distant metastasis events at other sites (p = 0.032). Conclusions: Elevated baseline LDH levels are associated with clinically advanced disease and are a poor prognosticator for OS, DFS, and DMFS for NPC patients. These results suggest that elevated serum levels of LDH should be considered when evaluating treatment options.

Zhou Guanqun; Tang Linglong; Mao Yanping; Chen Lei; Li Wenfei; Sun Ying [Department of Radiation Oncology, Cancer Center, State Key Laboratory of Oncology in South China, Sun Yat-sen University, Guangzhou (China); Liu Lizhi; Li Li [Imaging Diagnosis and Interventional Center, Cancer Center, State Key Laboratory of Oncology in South China, Sun Yat-sen University, Guangzhou (China); Lin Aihua [Department of Medical Statistics and Epidemiology, School of Public Health, Sun Yat-sen University, Guangzhou (China); Ma Jun, E-mail: drjunma@hotmail.com [Department of Radiation Oncology, Cancer Center, State Key Laboratory of Oncology in South China, Sun Yat-sen University, Guangzhou (China)

2012-03-01

173

Duck lens epsilon-crystallin and lactate dehydrogenase B4 are identical: a single-copy gene product with two distinct functions.  

PubMed Central

To investigate whether or not duck lens epsilon-crystallin and duck heart lactate dehydrogenase (LDH) B4 are the product of the same gene, we have isolated and sequenced cDNA clones of duck epsilon-crystallin. By using these clones we demonstrate that there is a single-copy Ldh-B gene in duck and in chicken. In the duck lens this gene is overexpressed, and its product is subject to posttranslational modification. Reconstruction of the evolutionary history of the LDH protein family reveals that the mammalian Ldh-C gene most probably originated from an ancestral Ldh-A gene and that the amino acid replacement rate in LDH-C is approximately 4 times the rate in LDH-A. Molecular modeling of LDH-B sequences shows that the increased thermostability of the avian tetramer might be explained by mutations that increase the number of ion pairs. Furthermore, the replacement of bulky side chains by glycines on the corners of the duck protein suggests an adaptation to facilitate close packing in the lens. Images PMID:3174623

Hendriks, W; Mulders, J W; Bibby, M A; Slingsby, C; Bloemendal, H; de Jong, W W

1988-01-01

174

Immune evasion mediated by tumor-derived lactate dehydrogenase induction of NKG2D ligands on myeloid cells in glioblastoma patients  

PubMed Central

Myeloid cells are key regulators of the tumor microenvironment, governing local immune responses. Here we report that tumor-infiltrating myeloid cells and circulating monocytes in patients with glioblastoma multiforme (GBM) express ligands for activating the Natural killer group 2, member D (NKG2D) receptor, which cause down-regulation of NKG2D on natural killer (NK) cells. Tumor-infiltrating NK cells isolated from GBM patients fail to lyse NKG2D ligand-expressing tumor cells. We demonstrate that lactate dehydrogenase (LDH) isoform 5 secreted by glioblastoma cells induces NKG2D ligands on monocytes isolated from healthy individuals. Furthermore, sera from GBM patients contain elevated amounts of LDH, which correlate with expression of NKG2D ligands on their autologous circulating monocytes. NKG2D ligands also are present on circulating monocytes isolated from patients with breast, prostate, and hepatitis C virus-induced hepatocellular carcinomas. Together, these findings reveal a previously unidentified immune evasion strategy whereby tumors produce soluble factors that induce NKG2D ligands on myeloid cells, subverting antitumor immune responses. PMID:25136121

Crane, Courtney A.; Austgen, Kathryn; Haberthur, Kristen; Hofmann, Carly; Moyes, Kara White; Avanesyan, Lia; Fong, Lawrence; Campbell, Michael J.; Cooper, Stewart; Oakes, Scott A.; Parsa, Andrew T.; Lanier, Lewis L.

2014-01-01

175

Serum lactate dehydrogenase with a systemic inflammation score is useful for predicting response and survival in patients with newly diagnosed diffuse large B-cell lymphoma.  

PubMed

We evaluated the relationship between serum lactate dehydrogenase (LDH) level with systemic inflammation score and survival in 213 patients with diffuse large B-cell lymphoma (DLBCL) receiving R-CHOP chemotherapy. The patients were classified into 3 groups based on LDH with the Glasgow Prognostic Score (L-GPS). A score of 2 was assigned to patients with elevated C-reactive protein, hypoalbuminemia and elevated LDH, a score of 1 to those with one or two abnormalities and a score of 0 to those with no abnormality. In multivariate analysis, independent poor prognostic factors for progression-free survival were L-GPS 2 [hazard ratio (HR) 5.415, p = 0.001], Eastern Cooperative Oncology Group performance status (ECOG PS) ?2 (HR 3.504, p = 0.001) and bulky lesion (HR 2.030, p = 0.039). Independent poor prognostic factors for overall survival were L-GPS 2 (HR 5.898, p = 0.001) and ECOG PS ?2 (HR 3.525, p = 0.001). The overall response rate for the R-CHOP chemotherapy decreased according to the L-GPS; it was 96.7% at L-GPS 0, 87% at L-GPS 1 and 75% at L-GPS 2 (p = 0.009). L-GPS based on systemic inflammatory indicators may be a useful clinical prognostic indicator for survival, and predicts the response for R-CHOP chemotherapy in patients with newly diagnosed DLBCL. PMID:24969101

Jung, Sung-Hoon; Yang, Deok-Hwan; Ahn, Jae-Sook; Kim, Yeo-Kyeoung; Kim, Hyeoung-Joon; Lee, Je-Jung

2015-01-01

176

A comparison of the primary structures of lactate dehydrogenase isozymes M4 from giant panda, red panda, black bear and dog.  

PubMed

Lactate dehydrogenase isozymes M4 have been isolated and purified from red panda (Ailurus fulgens), black bear (Selenarctos thibetanus) and dog (Canis familiars) by affinity chromatography and compared with that from giant panda (Ailuropoda melanoleuca). Experimental results have shown that the N-termini, C-termini and the molecular weights of LDH-M subunits of red panda, black bear and dog are the same as those of the LDH-M subunit of giant panda. Analysis and comparison of HPLC peptide maps from the tryptic digests of the isozymes of red panda, black bear and dog have shown that most of their peptide fragments had the same retention time and amino acid composition as the corresponding peptide fragments from giant panda. Fragments with different retention times and/or amino acid compositions were sequenced. Careful examination of those variant amino acid residues demonstrated clearly that the primary structure of giant panda LDH-M subunit is unique and it appears that the giant panda might be classified as an independent family. PMID:3629217

Liang, S P; Zhang, L X

1987-03-01

177

Structural characterization of tartrate dehydrogenase: a versatile enzyme catalyzing multiple reactions  

SciTech Connect

The first structure of an NAD-dependent tartrate dehydrogenase (TDH) has been solved to 2 {angstrom} resolution by single anomalous diffraction (SAD) phasing as a complex with the intermediate analog oxalate, Mg{sup 2+} and NADH. This TDH structure from Pseudomonas putida has a similar overall fold and domain organization to other structurally characterized members of the hydroxy-acid dehydrogenase family. However, there are considerable differences between TDH and these functionally related enzymes in the regions connecting the core secondary structure and in the relative positioning of important loops and helices. The active site in these complexes is highly ordered, allowing the identification of the substrate-binding and cofactor-binding groups and the ligands to the metal ions. Residues from the adjacent subunit are involved in both the substrate and divalent metal ion binding sites, establishing a dimer as the functional unit and providing structural support for an alternating-site reaction mechanism. The divalent metal ion plays a prominent role in substrate binding and orientation, together with several active-site arginines. Functional groups from both subunits form the cofactor-binding site and the ammonium ion aids in the orientation of the nicotinamide ring of the cofactor. A lysyl amino group (Lys192) is the base responsible for the water-mediated proton abstraction from the C2 hydroxyl group of the substrate that begins the catalytic reaction, followed by hydride transfer to NAD. A tyrosyl hydroxyl group (Tyr141) functions as a general acid to protonate the enolate intermediate. Each substrate undergoes the initial hydride transfer, but differences in substrate orientation are proposed to account for the different reactions catalyzed by TDH.

Malik, Radhika; Viola, Ronald E. (Toledo)

2010-10-28

178

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

SciTech Connect

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.

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

2008-02-01

179

Biocatalytic carbon capture via reversible reaction cycle catalyzed by isocitrate dehydrogenase.  

PubMed

The practice of carbon capture and storage (CCS) requires efficient capture and separation of carbon dioxide from its gaseous mixtures such as flue gas, followed by releasing it as a pure gas which can be subsequently compressed and injected into underground storage sites. This has been mostly achieved via reversible thermochemical reactions which are generally energy-intensive. The current work examines a biocatalytic approach for carbon capture using an NADP(H)-dependent isocitrate dehydrogenase (ICDH) which catalyzes reversibly carboxylation and decarboxylation reactions. Different from chemical carbon capture processes that rely on thermal energy to realize purification of carbon dioxide, the biocatalytic strategy utilizes pH to leverage the reaction equilibrium, thereby realizing energy-efficient carbon capture under ambient conditions. Results showed that over 25 mol of carbon dioxide could be captured and purified from its gas mixture for each gram of ICDH applied for each carboxylation/decarboxylation reaction cycle by varying pH between 6 and 9. This work demonstrates the promising potentials of pH-sensitive biocatalysis as a green-chemistry route for carbon capture. PMID:25152403

Xia, Shunxiang; Frigo-Vaz, Benjamin; Zhao, Xueyan; Kim, Jungbae; Wang, Ping

2014-09-12

180

Comparative characterization of a temperature responsive gene (lactate dehydrogenase-B, ldh-b) in two congeneric tropical fish, Lates calcarifer and Lates niloticus  

PubMed Central

The characterization of candidate loci is a critical step in obtaining insight into adaptation and acclimation of organisms. In this study of two non-model tropical (to sub-tropical) congeneric perciformes (Lates calcarifer and Lates niloticus) we characterized both coding and non-coding regions of lactate dehydrogenase-B (ldh-b), a locus which exhibits temperature-adaptive differences among temperate and sub-tropical populations of the North American killifish Fundulus heteroclitus. Ldh-b was 5,004 and 3,527 bp in length in L. calcarifer and L. niloticus, respectively, with coding regions comprising 1,005 bp in both species. A high level of sequence homology existed between species for both coding and non-coding regions of ldh-b (> 97% homology), corresponding to a 98.5% amino acid sequence homology. All six known functional sites within the encoded protein sequence (LDH-B) were conserved between the two Lates species. Ten simple sequence repeat (SSR) motifs (mono-, di-, tri- and tetranucleotide) and thirty putative microRNA elements (miRNAs) were identified within introns 1, 2, 5 and 6 of both Lates species. Five single nucleotide polymorphisms (SNPs) were also identified within miRNA containing intron regions. Such SNPs are implicated in several complex human conditions and/or diseases (as demonstrated by extensive genome-wide association studies). This novel characterization serves as a platform to further examine how non-model species may respond to changes in their native temperatures, which are expected to increase by up to 6°C over the next century. PMID:19787021

Edmunds, Richard C.; van Herwerden, Lynne; Smith-Keune, Carolyn; Jerry, Dean R.

2009-01-01

181

Long term intensive exercise training leads to a higher plasma malate/lactate dehydrogenase (M/L) ratio and increased level of lipid mobilization in horses.  

PubMed

Continuous high intensity training may induce alterations to enzyme activities related to glucose and lipid metabolism in horses. In our study, five Thoroughbred race horses (3 male and 2 female, avg age=5 yrs old) were compared against five riding horses (1 male, 1 female, 3 gelding, avg age=13 yrs old) in terms of energy metabolism, by examining plasma malate (MDH) and lactate (LDH) dehydrogenase activities and M/L ratio. MDH is involved in NADH and ATP generation, whereas LDH can convert NADH back into NAD(+) for ATP generation. An increase in plasma M/L ratio can reflect heightened energy metabolism in the liver and skeletal muscle of horses adapted to continuous intensive exercise. Moreover, plasma lipid metabolism analytes (adiponectin, NEFA, total cholesterol (T-Cho), and triglycerides (TG)) can reflect changes to lipolysis rate, which can also indicate a change in energy metabolism. Overall, race horses demonstrated increased MDH and LDH activity in plasma (4x and 2x greater, respectively), in addition to a plasma M/L ratio twice as high as that of riding horses (2.0 vs 1.0). In addition, race horses also demonstrated significantly higher levels of plasma NEFA (50% greater), TG (2x greater), and T-Cho (20% greater) as compared to riding horses. Therefore, race horse muscles may have adapted to prolonged high intensity endurance exercise by gaining a higher oxidative capacity and an increased capacity for fat utilization as an energy source, resulting in heightened energy metabolism and increased rate of lipid mobilization. PMID:22297553

Li, Gebin; Lee, Peter; Mori, Nobuko; Yamamoto, Ichiro; Arai, Toshiro

2012-06-01

182

Karnofsky Performance Status and Lactate Dehydrogenase Predict the Benefit of Palliative Whole-Brain Irradiation in Patients With Advanced Intra- and Extracranial Metastases From Malignant Melanoma  

SciTech Connect

Purpose: To determine prognostic factors that allow the selection of melanoma patients with advanced intra- and extracerebral metastatic disease for palliative whole-brain radiation therapy (WBRT) or best supportive care. Methods and Materials: This was a retrospective study of 87 patients who underwent palliative WBRT between 1988 and 2009 for progressive or multiple cerebral metastases at presentation. Uni- and multivariate analysis took into account the following patient- and tumor-associated factors: gender and age, Karnofsky performance status (KPS), neurologic symptoms, serum lactate dehydrogenase (LDH) level, number of intracranial metastases, previous resection or stereotactic radiosurgery of brain metastases, number of extracranial metastasis sites, and local recurrences as well as regional lymph node metastases at the time of WBRT. Results: In univariate analysis, KPS, LDH, number of intracranial metastases, and neurologic symptoms had a significant influence on overall survival. In multivariate survival analysis, KPS and LDH remained as significant prognostic factors, with hazard ratios of 3.3 (95% confidence interval [CI] 1.6-6.5) and 2.8 (95% CI 1.6-4.9), respectively. Patients with KPS ?70 and LDH ?240 U/L had a median survival of 191 days; patients with KPS ?70 and LDH >240 U/L, 96 days; patients with KPS <70 and LDH ?240 U/L, 47 days; and patients with KPS <70 and LDH >240 U/L, only 34 days. Conclusions: Karnofsky performance status and serum LDH values indicate whether patients with advanced intra- and extracranial tumor manifestations are candidates for palliative WBRT or best supportive care.

Partl, Richard, E-mail: richard.partl@medunigraz.at [Department of Therapeutic Radiology and Oncology, Medical University of Graz, Graz (Austria)] [Department of Therapeutic Radiology and Oncology, Medical University of Graz, Graz (Austria); Richtig, Erika [Department of Dermatology, Medical University of Graz, Graz (Austria)] [Department of Dermatology, Medical University of Graz, Graz (Austria); Avian, Alexander; Berghold, Andrea [Institute for Medical Informatics, Statistics and Documentation, Medical University of Graz, Graz (Austria)] [Institute for Medical Informatics, Statistics and Documentation, Medical University of Graz, Graz (Austria); Kapp, Karin S. [Department of Therapeutic Radiology and Oncology, Medical University of Graz, Graz (Austria)] [Department of Therapeutic Radiology and Oncology, Medical University of Graz, Graz (Austria)

2013-03-01

183

Evaluation of Milk Trace Elements, Lactate Dehydrogenase, Alkaline Phosphatase and Aspartate Aminotransferase Activity of Subclinical Mastitis as and Indicator of Subclinical Mastitis in Riverine Buffalo (Bubalus bubalis).  

PubMed

Mastitis is a highly morbid disease that requires detection at the subclinical stage. Tropical countries like India mainly depend on milch buffaloes for milk. The present study was conducted to investigate whether the trace minerals viz. copper (Cu), iron (Fe), zinc (Zn), cobalt (Co) and manganese (Mn) and enzyme activity of lactate dehydrogenase (LDH), alkaline phosphatase (ALP) and aspartate aminotransferase (AST) in riverine buffalo milk can be used as an indicator of subclinical mastitis (SCM) with the aim of developing suitable diagnostic kit for SCM. Trace elements and enzyme activity in milk were estimated with Atomic absorption Spectrophotometer, GBC 932 plus and biochemical methods, respectively. Somatic cell count (SCC) was done microscopically. The cultural examination revealed Gram positive bacteria as the most prevalent etiological agent. A statistically significant (p<0.01) increase in SCC, Fe, Zn, Co and LDH occurred in SCM milk containing gram positive bacterial agents only. ALP was found to be elevated in milk infected by both gram positive and negative bacteria. The percent sensitivity, specificity and accuracy, predictive values and likelihood ratios were calculated taking bacterial culture examination and SCC?2×10(5) cells/ml of milk as the benchmark. Only ALP and Zn, the former being superior, were found to be suitable for diagnosis of SCM irrespective of etiological agents. LDH, Co and Fe can be introduced in the screening programs where Gram positive bacteria are omnipresent. It is recommended that both ALP and Zn be measured together in milk to diagnose buffalo SCM, irrespective of etiology. PMID:25049573

Guha, Anirban; Gera, Sandeep; Sharma, Anshu

2012-03-01

184

Metabolic engineering of Lactococcus lactis: influence of the overproduction of alpha-acetolactate synthase in strains deficient in lactate dehydrogenase as a function of culture conditions.  

PubMed Central

The als gene for alpha-acetolactate synthase of Lactococcus lactis MG1363 was cloned on a multicopy plasmid under the control of the inducible L. lactis lacA promoter. More than a hundredfold overproduction of alpha-acetolactate synthase was obtained in L. lactis under inducing conditions as compared with that of the host strain, which contained a single chromosomal copy of the als gene. The effect of alpha-acetolactate synthase overproduction on the formation of end products in various L. lactis strains was studied under different fermentation conditions. Under aerobic conditions and with an initial pH of 6.0, overexpression of the als gene resulted in significant acetoin production that amounted to more than one-third of the pyruvate converted. However, the effect of the alpha-acetolactate synthase overproduction was even more pronounced in the lactate dehydrogenase-deficient strain L. lactis NZ2700. Anaerobic cultivation of this strain resulted in a doubling of the butanediol formation of up to 40% of the converted pyruvate. When cultivated aerobically at an initial pH of 6.8, overexpression of the als gene in L. lactis NZ2700 resulted in the conversion of more than 60% of the pyruvate into acetoin, while no butanediol was formed. Moreover, at an initial pH of 6.0, similar amounts of acetoin were obtained, but in addition approximately 20% of the pyruvate was converted into butanediol. These metabolic engineering studies indicate that more than 80% of the lactose can be converted via the activity of the overproduced alpha-acetolactate synthase in L. lactis. PMID:8526510

Platteeuw, C; Hugenholtz, J; Starrenburg, M; van Alen-Boerrigter, I; de Vos, W M

1995-01-01

185

Evidence for Hysteretic Substrate Channeling in the Proline Dehydrogenase and ?1-Pyrroline-5-carboxylate Dehydrogenase Coupled Reaction of Proline Utilization A (PutA)*  

PubMed Central

PutA (proline utilization A) is a large bifunctional flavoenzyme with proline dehydrogenase (PRODH) and ?1-pyrroline-5-carboxylate dehydrogenase (P5CDH) domains that catalyze the oxidation of l-proline to l-glutamate in two successive reactions. In the PRODH active site, proline undergoes a two-electron oxidation to ?1-pyrroline-5-carboxlylate, and the FAD cofactor is reduced. In the P5CDH active site, l-glutamate-?-semialdehyde (the hydrolyzed form of ?1-pyrroline-5-carboxylate) undergoes a two-electron oxidation in which a hydride is transferred to NAD+-producing NADH and glutamate. Here we report the first kinetic model for the overall PRODH-P5CDH reaction of a PutA enzyme. Global analysis of steady-state and transient kinetic data for the PRODH, P5CDH, and coupled PRODH-P5CDH reactions was used to test various models describing the conversion of proline to glutamate by Escherichia coli PutA. The coupled PRODH-P5CDH activity of PutA is best described by a mechanism in which the intermediate is not released into the bulk medium, i.e., substrate channeling. Unexpectedly, single-turnover kinetic experiments of the coupled PRODH-P5CDH reaction revealed that the rate of NADH formation is 20-fold slower than the steady-state turnover number for the overall reaction, implying that catalytic cycling speeds up throughput. We show that the limiting rate constant observed for NADH formation in the first turnover increases by almost 40-fold after multiple turnovers, achieving half of the steady-state value after 15 turnovers. These results suggest that EcPutA achieves an activated channeling state during the approach to steady state and is thus a new example of a hysteretic enzyme. Potential underlying causes of activation of channeling are discussed. PMID:24352662

Moxley, Michael A.; Sanyal, Nikhilesh; Krishnan, Navasona; Tanner, John J.; Becker, Donald F.

2014-01-01

186

Studies on the phenazine methosulphate-tetrazolium salt capture reaction in NAD(P) + -dependent dehydrogenase cytochemistry. I. Localization artefacts caused by the escape of reduced co-enzyme during cytochemical reactions for NAD(P) + -dependent dehydrogenases  

Microsoft Academic Search

Summary  The correct localization of oxidative enzymes using cytochemical tetrazolium methods, in which low molecular weight electron carriers such as NAD(P)H and reduced phenazine methosulphate (PMSH) are used, can be endangered by the escape of the reduced intermediates before they react to form the insoluble formazan at the true enzyme-containing sites. To investigate this phenomenon, the glucose-6-phosphate dehydrogenase reaction was studied

A. K. Raap; G. R. M. Van Hoof; P. Van Duijn

1983-01-01

187

Highly efficient L-lactate production using engineered Escherichia coli with dissimilar temperature optima for L-lactate formation and cell growth  

PubMed Central

L-Lactic acid, one of the most important chiral molecules and organic acids, is produced via pyruvate from carbohydrates in diverse microorganisms catalyzed by an NAD+-dependent L-lactate dehydrogenase. Naturally, Escherichia coli does not produce L-lactate in noticeable amounts, but can catabolize it via a dehydrogenation reaction mediated by an FMN-dependent L-lactate dehydrogenase. In aims to make the E. coli strain to produce L-lactate, three L-lactate dehydrogenase genes from different bacteria were cloned and expressed. The L-lactate producing strains, 090B1 (B0013-070, ?ldhA::diflldD::Pldh-ldhLca), 090B2 (B0013-070, ?ldhA::diflldD::Pldh-ldhStrb) and 090B3 (B0013-070, ?ldhA::diflldD::Pldh-ldhBcoa) were developed from a previously developed D-lactate over-producing strain, E. coli strain B0013-070 (ack-ptappspflBdldpoxBadhEfrdA) by: (1) deleting ldhA to block D-lactate formation, (2) deleting lldD to block the conversion of L-lactate to pyruvate, and (3) expressing an L-lactate dehydrogenase (L-LDH) to convert pyruvate to L-lactate under the control of the ldhA promoter. Fermentation tests were carried out in a shaking flask and in a 25-l bioreactor. Strains 090B1, 090B2 or 090B3 were shown to metabolize glucose to L-lactate instead of D-lactate. However, L-lactate yield and cell growth rates were significantly different among the metabolically engineered strains which can be attributed to a variation between temperature optimum for cell growth and temperature optimum for enzymatic activity of individual L-LDH. In a temperature-shifting fermentation process (cells grown at 37°C and L-lactate formed at 42°C), E. coli 090B3 was able to produce 142.2 g/l of L-lactate with no more than 1.2 g/l of by-products (mainly acetate, pyruvate and succinate) accumulated. In conclusion, the production of lactate by E. coli is limited by the competition relationship between cell growth and lactate synthesis. Enzymatic properties, especially the thermodynamics of an L-LDH can be effectively used as a factor to regulate a metabolic pathway and its metabolic flux for efficient L-lactate production. Highlights The enzymatic thermodynamics was used as a tool for metabolic regulation. ? minimizing the activity of L-lactate dehydrogenase in growth phase improved biomass accumulation. ? maximizing the activity of L-lactate dehydrogenase improved lactate productivity in production phase. PMID:24884499

2014-01-01

188

(1-3)-beta-D-glucan in association with lactate dehydrogenase as biomarkers of Pneumocystis pneumonia (PcP) in HIV-infected patients.  

PubMed

Pneumocystis pneumonia (PcP) is a major HIV-related illness caused by Pneumocystis jirovecii. Definitive diagnosis of PcP requires microscopic detection of P. jirovecii in pulmonary specimens. The objective of this study was to evaluate the usefulness of two serum markers in the diagnosis of PcP. Serum levels of (1-3)-beta-d-glucan (BG) and lactate dehydrogenase (LDH) were investigated in 100 HIV-positive adult patients and 50 healthy blood donors. PcP cases were confirmed using indirect immunofluorescence with monoclonal anti-Pneumocystis antibodies and nested-PCR to amplify the large subunit mitochondrial rRNA gene of P. jirovecii in pulmonary specimens. BG and LDH levels in serum were measured using quantitative microplate-based assays. BG and LDH positive sera were statistically associated with PcP cases (P???0.001). Sensitivity, specificity, positive/negative predictive values (PPV/NPV), and positive/negative likelihood ratios (PLR/NLR) were 91.3 %, 61.3 %, 85.1 %, 79.2 %, 2.359, and 0.142, respectively, for the BG kit assay, and 91.3 %, 35.5 %, 75.9 %, 64.7 %, 1.415 and 0.245, respectively, for the LDH test. Serologic markers levels combined with the clinical diagnostic criteria for PcP were evaluated for their usefulness in diagnosis of PcP. The most promising cutoff levels for diagnosis of PcP were determined to be 400 pg/ml of BG and 350 U/l of LDH, which combined with clinical data presented 92.8 % sensitivity, 83.9 % specificity, 92.8 % PPV, 83.9 % NPV, 5.764 PLR and 0.086 NLR (P?

Esteves, F; Lee, C-H; de Sousa, B; Badura, R; Seringa, M; Fernandes, C; Gaspar, J F; Antunes, F; Matos, O

2014-07-01

189

The risk of thrombo-embolic events is increased in patients with germ-cell tumours and can be predicted by serum lactate dehydrogenase and body surface area  

PubMed Central

The aim of this study was to evaluate the risk of thrombo-embolic events (TEE) in patients with germ-cell tumours (GCT) who receive cisplatin-based chemotherapy, to compare this risk to that of a matched control group of non-GCT cancer patients, and to identify risk factors of TEE. The rate of TEE during the 6 months following the initiation of chemotherapy was assessed in 100 consecutive patients with GCT and in 100 controls with various neoplasms who were matched on sex and age, and who received first-line cisplatin-based chemotherapy during the same period of time at Institut Gustave Roussy, Villejuif, France. Data were subsequently tested on a validation group of 77 GCT patients treated in Lyon, France. A total of 19 patients (19%) (95% confidence interval (CI): 13–28) and six patients (6%) (95% CI: 3–13) had a TEE in the GCT group and the non-GCT control group, respectively (relative risk (RR): 3.4; P<0.01). Three patients from the GCT group died of pulmonary embolism. In multivariate analysis, two factors had independent predictive value for TEE: a high body surface area (>1.9?m2) (RR: 5 (1.8–13.9)) and an elevated serum lactate dehydrogenase (LDH) (RR: 6.4 (2.3–18.2)). Patients with no risk factor (n=26) and those with at least one risk factor (n=71) had a probability of having a TEE of 4% (95% CI: 1–19) and 26% (95% CI: 17–37), respectively. In the GCT validation set, 10 (13%) patients had a TEE; patients with no risk factor and those with at least one risk factor had a probability of having a TEE of 0 and 17% (95% CI: 10–29), respectively. Patients with GCT are at a higher risk for TEE than patients with non-GCT cancer while on cisplatin-based chemotherapy. This risk can be accurately predicted by serum LDH and body surface area. This predictive index may help to study prospectively the impact of thromboprophylaxis in GCT patients. PMID:16205699

Piketty, A-C; Fléchon, A; Laplanche, A; Nouyrigat, E; Droz, J-P; Théodore, C; Fizazi, K

2005-01-01

190

Sugar-glycerol cofermentations in lactobacilli: the fate of lactate.  

PubMed Central

The simultaneous fermentation of glycerol and sugar by lactobacillus brevis B22 and Lactobacillus buchneri B190 increases both the growth rate and total growth. The reduction of glycerol to 1,3-propanediol by the lactobacilli was found to influence the metabolism of the sugar cofermented by channelling some of the intermediate metabolites (e.g., pyruvate) towards NADH-producing (rather than NADH-consuming) reactions. Ultimately, the absolute requirement for NADH to prevent the accumulation of 3-hydroxypropionaldehyde leads to a novel lactate-glycerol cofermentation. As a result, additional ATP can be made not only by (i) converting pyruvate to acetate via acetyl phosphate rather than to the ethanol usually found and (ii) oxidizing part of the intermediate pyruvate to acetate instead of the usual reduction to lactate but also by (iii) reoxidation of accumulated lactate to acetate via pyruvate. The conversion of lactate to pyruvate is probably catalyzed by NAD-independent lactate dehydrogenases that are found only in the cultures oxidizing lactate and producing 1,3-propanediol, suggesting a correlation between the expression of these enzymes and a raised intracellular NAD/NADH ratio. The enzymes metabolizing glycerol (glycerol dehydratase and 1,3-propanediol dehydrogenase) were expressed in concert without necessary induction by added glycerol, although their expression may also be influenced by the intracellular NAD/NADH ratio set by the different carbohydrates fermented. PMID:1732191

Veiga da Cunha, M; Foster, M A

1992-01-01

191

Application of artificial neural networks and DFT-based parameters for prediction of reaction kinetics of ethylbenzene dehydrogenase.  

PubMed

Artificial neural networks (ANNs) are used for classification and prediction of enzymatic activity of ethylbenzene dehydrogenase from EbN1 Azoarcus sp. bacterium. Ethylbenzene dehydrogenase (EBDH) catalyzes stereo-specific oxidation of ethylbenzene and its derivates to alcohols, which find its application as building blocks in pharmaceutical industry. ANN systems are trained based on theoretical variables derived from Density Functional Theory (DFT) modeling, topological descriptors, and kinetic parameters measured with developed spectrophotometric assay. Obtained models exhibit high degree of accuracy (100% of correct classifications, correlation between predicted and experimental values of reaction rates on the 0.97 level). The applicability of ANNs is demonstrated as useful tool for the prediction of biochemical enzyme activity of new substrates basing only on quantum chemical calculations and simple structural characteristics. Multi Linear Regression and Molecular Field Analysis (MFA) are used in order to compare robustness of ANN and both classical and 3D-quantitative structure-activity relationship (QSAR) approaches. PMID:16779618

Szaleniec, Maciej; Witko, Ma?gorzata; Tadeusiewicz, Ryszard; Goclon, Jakub

2006-03-01

192

Lactic acid conversion to 2,3-pentanedione and acrylic acid over silica-supported sodium nitrate: Reaction optimization and identification of sodium lactate as the active catalyst  

SciTech Connect

Lactic acid is converted to 2,3-pentanedione, acrylic acid, and other products in vapor-phase reactions over silica-supported sodium lactate formed from sodium nitrate. Multiparameter optimization of reaction conditions using a Box-Benkhen experimental design shows that the highest yield and selectivity to 2,3-pentanedione are achieved at low temperature, elevated pressure, and long contact time, while yield and selectivity to acrylic acid are most favorable at high temperature, low pressure, and short contact time. Post-reaction Fourier transform infrared spectroscopic analyses of the catalyst indicate that sodium nitrate as the initial catalyst material is transformed to sodium lactate at the onset of reaction via proton transfer from lactic acid to nitrate. The resultant nitric acid vaporizes as it is formed, leaving sodium lactate as the sole sodium-bearing species on the catalyst during reaction. 19 refs., 8 figs., 5 tabs.

Wadley, D.C.; Tam, M.S.; Miller, D.J. [Michigan State Univ., East Lansing, MI (United States)] [and others] [Michigan State Univ., East Lansing, MI (United States); and others

1997-01-15

193

Fiber optic biosensors for hydrogen peroxide and L-lactate  

NASA Astrophysics Data System (ADS)

An optical fiber biosensor for the selective determination of hydrogen peroxide has been developed as the base sensor for the construction of multienzyme optodes involving lactate converting enzymes for the analysis of lactic acid. The optode uses the H2O2 dependent oxidation of homovanillic acid by horseradish peroxidase (HRP) as the sensing reaction. The fluorescence of the dimeric product formed is used as the measuring signal related to the concentration of H2O2. HRP was immobilized on a membrane and combined with a bifurcated fiber optic probe. Under optimized conditions the sensor responds linearly to hydrogen peroxide between 1 micrometers ol/l and 0.12 mmol/l and exhibits a half life of 90 days. Using a lactate oxidase-HRP membrane, the sensor is suitable for lactate measurement with a linear range of 3 micrometers ol/l-0.2 mmol/l. To increase the sensitivity for lactate, lactate dehydrogenase was coimmobilized on the sensor membrane. In the presence of NADH the signal for lactate is amplified fourfold through the internal analyte recycling accomplished by the lactate-converting enzymes.

Schubert, Florian; Rinneberg, Herbert H.; Wang, Fang

1995-02-01

194

Reversibility of the mitochondrial isocitrate dehydrogenase reaction in the perfused rat liver. Evidence from isotopomer analysis of citric acid cycle intermediates.  

PubMed

The reversal of the mitochondrial isocitrate dehydrogenase reaction was investigated in rat livers perfused with [U-13C5]glutamate or [U-13C5]glutamine. The mass isotopomer distribution of citric acid cycle intermediates extracted from the livers was determined by gas chromatography-mass spectrometry. Citrate was enriched in an isotopomer containing five 13C. The formation of this isotopomer can only be explained by the reversal of the isocitrate dehydrogenase reaction. Calculation of kinetic parameters from the mass isotopomer data reveals a rapid interconversion of isocitrate and alpha-ketoglutarate. This interconversion results in an isotopic exchange between carbon 6 of citrate and mitochondrial CO2 that can affect the calculation of citric acid cycle kinetic parameters. Thus, the reversal of the isocitrate dehydrogenase reaction should be included in isotope labeling models of the citric acid cycle. PMID:7961626

Des Rosiers, C; Fernandez, C A; David, F; Brunengraber, H

1994-11-01

195

Pyruvate dehydrogenase and pyruvate dehydrogenase kinase expression in non small cell lung cancer and tumor-associated stroma.  

PubMed

Pyruvate dehydrogenase (PDH) catalyzes the conversion of pyruvate to acetyl-coenzyme A, which enters into the Krebs cycle, providing adenosine triphosphate (ATP) to the cell. PDH activity is under the control of pyruvate dehydrogenase kinases (PDKs). Under hypoxic conditions, conversion of pyruvate to lactate occurs, a reaction catalyzed by lactate dehydrogenase 5 (LDH5). In cancer cells, however pyruvate is transformed to lactate occurs, regardless of the presence of oxygen (aerobic glycolysis/Warburg effect). Although, hypoxic intratumoral conditions account for HIF1alpha stabilization and induction of anaerobic metabolism, recent data suggest that high pyruvate concentrations also result in HIF1alpha stabilization independently of hypoxia. In the present immunohistochemical study, we provide evidence that the PDH/PDK pathway is repressed in 73% of non small cell lung carcinomas, which may be a key reason for HIF1alpha stabilization and "aerobic glycolysis." However, about half of PDH-HIF pathway, and patients harboring these tumors have an excellent postoperative outcome. A small subgroup of clinically aggressive tumors maintains a coherent PDH and HIF/LDH5 expression. In contrast to cancer cells, fibroblasts in the tumor supporting stroma exhibit an intense PDH but reduced PDK1 expression favoring maximum PDH activity. This means that stroma may use lactic acid produced by tumor cells, preventing the creation of an intolerable intratumoral acidic environment at the same time. PMID:15736311

Koukourakis, Michael I; Giatromanolaki, Alexandra; Sivridis, Efthimios; Gatter, Kevin C; Harris, Adrian L

2005-01-01

196

Lactate Test  

MedlinePLUS

... of this website will be limited. Search Help? Lactate Share this page: Was this page helpful? Also ... there anything I can do to decrease my lactate level? Generally, no. However, if your elevated lactate ...

197

Potentiometric CO titrations of carbon monoxide dehydrogenase and CO-inhibition of the NI-removing reaction with 1,10--phenanthroline  

E-print Network

Carbon monoxide dehydrogenase from Clostridium thermoaceticum catalyzes two reactions involving redox chemistry (the reversible oxidation Of CO to C02, and the synthesis of acetyl-CoA) using three types of Ni and Fe-S structures called the A-, B...

Russell, William Kent

1996-01-01

198

Lactation Consultant  

MedlinePLUS

... specialized knowledge and clinical expertise in breastfeeding and human lactation. Job description Lactation consultants educate women, families, health professionals, and the community about breast feeding ...

199

Temperature optima of enzyme-catalysed reactions in microemulsion systems  

Microsoft Academic Search

Ternary phase systems (water\\/surfactant\\/organic solvent) were utilised to increase and broaden the temperature optima of enzyme-catalysed reactions. Alcohol dehydrogenases from yeast and Thermoanaerobium brockii (EC 1.1.1.1 and EC 1.1.1.2), lactate dehydrogenase from Lactobacillus delbrueckii (EC 1.1.1.28) and the particulate hydrogenase from Ralstonia eutropha (EC 1.18.99.1) were used as model enzymes in microemulsions, consisting of the surfactant Aerosol OT, and various

K. Mlejnek; B. Seiffert; T. Demberg; M. Kämper; M. Hoppert

2004-01-01

200

Measurement of lactate formation from glucose using (6- sup 3 H)- and (6- sup 14 C)glucose in humans  

SciTech Connect

To assess the validity of determining the origin of plasma lactate from the ratio of lactate and glucose specific activities (SA) during infusion of labeled glucose, normal subjects received infusions of (6-3H)- and (6-14C)glucose for 4 h after a 12 h fast, and, on another day, cold glucose labeled with both tracers during 4-6 h of hyperinsulinemia (approximately 650 microU/ml). Basally, less lactate was derived from plasma glucose when measured with (6-3H)glucose (27 +/- 2%) than with (6-14C)glucose (40 +/- 2%, P less than 0.001). Insulin did not increase the percent of lactate derived from plasma glucose when measured with (6-3H)glucose (29 +/- 2%) but did increase when measured with (6-14C)glucose (60 +/- 4%). The arterialized blood (A) (3H)lactate SA was 30-40% higher (P less than 0.01) than deep venous blood (V) (3H)lactate SA, whereas A and V (14C)lactate SA were similar. During conversion of alanine to lactate with glutamic-pyruvic transaminase (GPT) and lactate dehydrogenase (LDH) in vitro, 32 +/- 2% of 3H in (3-3H)alanine was found in water and 68 +/- 2% in lactate. During infusion of (6-3H)- and (6-14C)glucose, the ratio of (14C)alanine to lactate SA (0.88 +/- 0.05) was less than the ratio of (3H)alanine to lactate SA (0.31 +/- 0.03, P less than 0.001). In conclusion (1) loss of 3H relative to 14C from position 6 in glucose occurs during lactate formation in extrahepatic tissues possibly due to the GPT reaction (alanine conversion to pyruvate), and (2) even under supraphysiologic hyperinsulinemic conditions not all of plasma lactate originates from plasma glucose.

Virkamaeki, A.P.; Puhakainen, I.; Nurjhan, N.; Gerich, J.E.; Yki-Jaervinen, H. (Helsinki Univ. (Finland))

1990-09-01

201

13C NMR Characterization of an Exchange Reaction between CO and CO2 Catalyzed by Carbon Monoxide Dehydrogenase†  

PubMed Central

Carbon monoxide dehydrogenase (CODH) catalyzes the reversible oxidation of CO to CO2 at a nickel?iron?sulfur cluster (the C-cluster). CO oxidation follows a ping-pong mechanism involving two-electron reduction of the C-cluster followed by electron transfer through an internal electron transfer chain to external electron acceptors. We describe 13C NMR studies demonstrating a CODH-catalyzed steady-state exchange reaction between CO and CO2 in the absence of external electron acceptors. This reaction is characterized by a CODH-dependent broadening of the 13CO NMR resonance; however, the chemical shift of the 13CO resonance is unchanged, indicating that the broadening is in the slow exchange limit of the NMR experiment. The 13CO line broadening occurs with a rate constant (1080 s?1 at 20 °C) that is approximately equal to that of CO oxidation. It is concluded that the observed exchange reaction is between 13CO and CODH-bound 13CO2 because 13CO line broadening is pH-independent (unlike steady-state CO oxidation), because it requires a functional C-cluster (but not a functional B-cluster) and because the 13CO2 line width does not broaden. Furthermore, a steady-state isotopic exchange reaction between 12CO and 13CO2 in solution was shown to occur at the same rate as that of CO2 reduction, which is approximately 750-fold slower than the rate of 13CO exchange broadening. The interaction between CODH and the inhibitor cyanide (CN?) was also probed by 13C NMR. A functional C-cluster is not required for 13CN? broadening (unlike for 13CO), and its exchange rate constant is 30-fold faster than that for 13CO. The combined results indicate that the 13CO exchange includes migration of CO to the C-cluster, and CO oxidation to CO2, but not release of CO2 or protons into the solvent. They also provide strong evidence of a CO2 binding site and of an internal proton transfer network in CODH. 13CN? exchange appears to monitor only movement of CN? between solution and its binding to and release from CODH. PMID:18589895

2008-01-01

202

Assessment of the drug susceptibility of Plasmodium falciparum clinical isolates from africa by using a Plasmodium lactate dehydrogenase immunodetection assay and an inhibitory maximum effect model for precise measurement of the 50-percent inhibitory concentration.  

PubMed

The extension of drug resistance among malaria-causing Plasmodium falciparum parasites in Africa necessitates implementation of new combined therapeutic strategies. Drug susceptibility phenotyping requires precise measurements. Until recently, schizont maturation and isotopic in vitro assays were the only methods available, but their use was limited by technical constraints. This explains the revived interest in the development of replacement methods, such as the Plasmodium lactate dehydrogenase (pLDH) immunodetection assay. We evaluated a commercially controlled pLDH enzyme-linked immunosorbent assay (ELISA; the ELISA-Malaria antigen test; DiaMed AG, Cressier s/Morat, Switzerland) to assess drug susceptibility in a standard in vitro assay using fairly basic laboratory equipment to study the in vitro resistance of malaria parasites to major antimalarials. Five Plasmodium falciparum clones and 121 clinical African isolates collected during 2003 and 2004 were studied by the pLDH ELISA and the [8-(3)H]hypoxanthine isotopic assay as a reference with four antimalarials. Nonlinear regression with a maximum effect model was used to estimate the 50% inhibitory concentration (IC(50)) and its confidence intervals. The two methods were observed to have similar reproducibilities, but the pLDH ELISA demonstrated a higher sensitivity. The high correlation (r = 0.98) and the high phenotypic agreement (kappa = 0.88) between the two methods allowed comparison by determination of the IC(50)s. Recently collected Plasmodium falciparum African isolates were tested by pLDH ELISA and showed drug resistance or decreased susceptibilities of 62% to chloroquine and 11.5% to the active metabolite of amodiaquine. No decreased susceptibility to lumefantrine or the active metabolite of artemisinin was detected. The availability of this simple and highly sensitive pLDH immunodetection assay will provide an easier method for drug susceptibility testing of malaria parasites. PMID:17005815

Kaddouri, Halima; Nakache, Serge; Houzé, Sandrine; Mentré, France; Le Bras, Jacques

2006-10-01

203

Assessment of the Drug Susceptibility of Plasmodium falciparum Clinical Isolates from Africa by Using a Plasmodium Lactate Dehydrogenase Immunodetection Assay and an Inhibitory Maximum Effect Model for Precise Measurement of the 50-Percent Inhibitory Concentration  

PubMed Central

The extension of drug resistance among malaria-causing Plasmodium falciparum parasites in Africa necessitates implementation of new combined therapeutic strategies. Drug susceptibility phenotyping requires precise measurements. Until recently, schizont maturation and isotopic in vitro assays were the only methods available, but their use was limited by technical constraints. This explains the revived interest in the development of replacement methods, such as the Plasmodium lactate dehydrogenase (pLDH) immunodetection assay. We evaluated a commercially controlled pLDH enzyme-linked immunosorbent assay (ELISA; the ELISA-Malaria antigen test; DiaMed AG, Cressier s/Morat, Switzerland) to assess drug susceptibility in a standard in vitro assay using fairly basic laboratory equipment to study the in vitro resistance of malaria parasites to major antimalarials. Five Plasmodium falciparum clones and 121 clinical African isolates collected during 2003 and 2004 were studied by the pLDH ELISA and the [8-3H]hypoxanthine isotopic assay as a reference with four antimalarials. Nonlinear regression with a maximum effect model was used to estimate the 50% inhibitory concentration (IC50) and its confidence intervals. The two methods were observed to have similar reproducibilities, but the pLDH ELISA demonstrated a higher sensitivity. The high correlation (r = 0.98) and the high phenotypic agreement (? = 0.88) between the two methods allowed comparison by determination of the IC50s. Recently collected Plasmodium falciparum African isolates were tested by pLDH ELISA and showed drug resistance or decreased susceptibilities of 62% to chloroquine and 11.5% to the active metabolite of amodiaquine. No decreased susceptibility to lumefantrine or the active metabolite of artemisinin was detected. The availability of this simple and highly sensitive pLDH immunodetection assay will provide an easier method for drug susceptibility testing of malaria parasites. PMID:17005815

Kaddouri, Halima; Nakache, Serge; Houzé, Sandrine; Mentré, France; Le Bras, Jacques

2006-01-01

204

Reconstruction of lactate utilization system in Pseudomonas putida KT2440: a novel biocatalyst for l-2-hydroxy-carboxylate production  

PubMed Central

As an important method for building blocks synthesis, whole cell biocatalysis is hindered by some shortcomings such as unpredictability of reactions, utilization of opportunistic pathogen, and side reactions. Due to its biological and extensively studied genetic background, Pseudomonas putida KT2440 is viewed as a promising host for construction of efficient biocatalysts. After analysis and reconstruction of the lactate utilization system in the P. putida strain, a novel biocatalyst that only exhibited NAD-independent d-lactate dehydrogenase activity was prepared and used in l-2-hydroxy-carboxylates production. Since the side reaction catalyzed by the NAD-independent l-lactate dehydrogenase was eliminated in whole cells of recombinant P. putida KT2440, two important l-2-hydroxy-carboxylates (l-lactate and l-2-hydroxybutyrate) were produced in high yield and high optical purity by kinetic resolution of racemic 2-hydroxy carboxylic acids. The results highlight the promise in biocatalysis by the biotechnologically important organism P. putida KT2440 through genomic analysis and recombination. PMID:25373400

Wang, Yujiao; Lv, Min; Zhang, Yingxin; Xiao, Xieyue; Jiang, Tianyi; Zhang, Wen; Hu, Chunhui; Gao, Chao; Ma, Cuiqing; Xu, Ping

2014-01-01

205

A novel D-mandelate dehydrogenase used in three-enzyme cascade reaction for highly efficient synthesis of non-natural chiral amino acids.  

PubMed

A novel NAD(+)-dependent D-mandelate dehydrogenase was identified from Lactobacillus brevis (LbDMDH). After purified to homogeneity, the optimum pH and temperature for oxidation of D-mandelate were pH 10.0 and 40 °C, and the Km and kcat were 1.1 mM and 355 s(-1) respectively. Employing the LbDMDH together with a mandelate racemase from Pseudomonas putida and a leucine dehydrogenase (EsLeuDH) from Exiguobacterium sibiricum, we established a three-step one-pot domino reaction system for preparing chiral L-phenylglycine from racemic mandelic acid with internal cofactor recycling. Under the optimum conditions, 30.4 g rac-mandelic acid (0.2 M) at 1L scale had been converted into chiral L-phenylglycine, with 96.4% conversion, 86.5% isolation yield, >99% eep and 50.4 gL(-1)d(-1) space-time yield. PMID:25449542

Fan, Chang-Wei; Xu, Guo-Chao; Ma, Bao-Di; Bai, Yun-Peng; Zhang, Jie; Xu, Jian-He

2015-02-10

206

Lactate metabolism in the fetal rabbit lung  

SciTech Connect

Lactate is frequently overlooked as a potential substrate for the fetal lung, even though it is present in the fetal circulation in concentrations as high as 8 mM. These high concentrations, coupled with the relatively low levels of glucose in the fetal blood, may indicate that lactate can substitute for glucose in pulmonary energy generation and phospholipid synthesis. A series of experiments was therefore undertaken in order to investigate the role of lactate in perinatal pulmonary development. Explants from 30 day gestation fetal rabbit lungs were incubated in Krebs-Ringer bicarbonate buffer supplemented with 3 mM (U-/sup 14/C)-glucose and varying levels of lactate. In the absence of medium lactate, fetal rabbit lung explants were capable of producing lactate at a rate of approximately 200 etamoles/mg protein/hour. The addition of lactate to the bathing medium immediately reduced net lactate production and above 4 mM, fetal rabbit lung explants became net utilizers of lactate. Media lactate concentrations of 2.5 mM, 5 mM and 10 mM also decreased glucose incorporation into total tissue disaturated phosphatidylcholine by approximately 20%, 35%, and 45%, respectively. Glucose incorporation into surfactant phosphatidylcholine was also reduced by approximately 50%, when lactate was present in the incubation medium at a concentration of 5 mM. Additional experiments also revealed that fetal lung lactate dehydrogenase activity was almost twice that found in the adult rabbit lung. These data indicate that lactate may be an important carbon source for the developing lung and could be a significant component in the manufacture of surfactant phosphatidylcholine during late gestation.

Engle, M.J.; Brown, D.J.; Dooley, M.

1986-05-01

207

Lactate is always the end product of glycolysis  

PubMed Central

Through much of the history of metabolism, lactate (La?) has been considered merely a dead-end waste product during periods of dysoxia. Congruently, the end product of glycolysis has been viewed dichotomously: pyruvate in the presence of adequate oxygenation, La? in the absence of adequate oxygenation. In contrast, given the near-equilibrium nature of the lactate dehydrogenase (LDH) reaction and that LDH has a much higher activity than the putative regulatory enzymes of the glycolytic and oxidative pathways, we contend that La? is always the end product of glycolysis. Cellular La? accumulation, as opposed to flux, is dependent on (1) the rate of glycolysis, (2) oxidative enzyme activity, (3) cellular O2 level, and (4) the net rate of La? transport into (influx) or out of (efflux) the cell. For intracellular metabolism, we reintroduce the Cytosol-to-Mitochondria Lactate Shuttle. Our proposition, analogous to the phosphocreatine shuttle, purports that pyruvate, NAD+, NADH, and La? are held uniformly near equilibrium throughout the cell cytosol due to the high activity of LDH. La? is always the end product of glycolysis and represents the primary diffusing species capable of spatially linking glycolysis to oxidative phosphorylation. PMID:25774123

Rogatzki, Matthew J.; Ferguson, Brian S.; Goodwin, Matthew L.; Gladden, L. Bruce

2015-01-01

208

Glycolysis and the significance of lactate in traumatic brain injury  

PubMed Central

In traumatic brain injury (TBI) patients, elevation of the brain extracellular lactate concentration and the lactate/pyruvate ratio are well-recognized, and are associated statistically with unfavorable clinical outcome. Brain extracellular lactate was conventionally regarded as a waste product of glucose, when glucose is metabolized via glycolysis (Embden-Meyerhof-Parnas pathway) to pyruvate, followed by conversion to lactate by the action of lactate dehydrogenase, and export of lactate into the extracellular fluid. In TBI, glycolytic lactate is ascribed to hypoxia or mitochondrial dysfunction, although the precise nature of the latter is incompletely understood. Seemingly in contrast to lactate's association with unfavorable outcome is a growing body of evidence that lactate can be beneficial. The idea that the brain can utilize lactate by feeding into the tricarboxylic acid (TCA) cycle of neurons, first published two decades ago, has become known as the astrocyte-neuron lactate shuttle hypothesis. Direct evidence of brain utilization of lactate was first obtained 5 years ago in a cerebral microdialysis study in TBI patients, where administration of 13C-labeled lactate via the microdialysis catheter and simultaneous collection of the emerging microdialysates, with 13C NMR analysis, revealed 13C labeling in glutamine consistent with lactate utilization via the TCA cycle. This suggests that where neurons are too damaged to utilize the lactate produced from glucose by astrocytes, i.e., uncoupling of neuronal and glial metabolism, high extracellular levels of lactate would accumulate, explaining the association between high lactate and poor outcome. Recently, an intravenous exogenous lactate supplementation study in TBI patients revealed evidence for a beneficial effect judged by surrogate endpoints. Here we review the current state of knowledge about glycolysis and lactate in TBI, how it can be measured in patients, and whether it can be modulated to achieve better clinical outcome. PMID:25904838

Carpenter, Keri L. H.; Jalloh, Ibrahim; Hutchinson, Peter J.

2015-01-01

209

The crystal structure of Lactococcus lactis dihydroorotate dehydrogenase A complexed with the enzyme reaction product throws light on its enzymatic function.  

PubMed Central

Dihydroorotate dehydrogenases (DHODs) catalyze the oxidation of (S)-dihydroorotate to orotate, the fourth step and only redox reaction in the de novo biosynthesis of pyrimidine nucleotides. A description is given of the crystal structure of Lactococcus lactis dihydroorotate dehydrogenase A (DHODA) complexed with the product of the enzyme reaction orotate. The structure of the complex to 2.0 A resolution has been compared with the structure of the native enzyme. The active site of DHODA is known to contain a water filled cavity buried beneath a highly conserved and flexible loop. In the complex the orotate displaces the water molecules from the active site and stacks above the DHODA flavin isoalloxazine ring, causing only small movements of the surrounding protein residues. The orotate is completely buried beneath the protein surface, and the orotate binding causes a significant reduction in the mobility of the active site loop. The orotate is bound by four conserved asparagine side chains (Asn 67, Asn 127, Asn 132, and Asn 193), the side chains of Lys 43 and Ser 194, and the main chain NH groups of Met 69, Gly 70, and Leu 71. Of these the Lys 43 side chain makes hydrogen bonds to both the flavin isoalloxazine ring and the carboxylate group of the orotate. Potential interactions with bound dihydroorotate are considered using the orotate complex as a basis for molecular modeling. The role of Cys 130 as the active site base is discussed, and the sequence conservation of the active site residues across the different families of DHODs is reviewed, along with implications for differences in substrate binding and in the catalytic mechanisms between these families. PMID:9655329

Rowland, P.; Björnberg, O.; Nielsen, F. S.; Jensen, K. F.; Larsen, S.

1998-01-01

210

Characterization of electron tunneling and hole hopping reactions between different forms of MauG and methylamine dehydrogenase within a natural protein complex  

PubMed Central

Respiration, photosynthesis and metabolism require the transfer of electrons through and between proteins over relatively long distances. It is critical that this electron transfer (ET) occur with specificity to avoid cellular damage, and at a rate which is sufficient to support the biological activity. A multi-step hole hopping mechanism could, in principle, enhance the efficiency of long range ET through proteins as it does in organic semiconductors. To explore this possibility, two different ET reactions that occur over the same distance within the protein complex of the diheme enzyme MauG and different forms of methylamine dehydrogenase (MADH) were subjected to kinetic and thermodynamic analysis. An ET mechanism of single-step direct electron tunneling from diferrous MauG to the quinone form of MADH is consistent with the data. In contrast, the biosynthetic ET from preMADH, which contains incompletely synthesized tryptophan tryptophylquinone, to the bis-Fe(IV) form of MauG is best described by a two-step hole hopping mechanism. Experimentally-determined values of ET distance matched the distances determined from the crystal structure that would be expected for single-step tunneling and multi-step hopping, respectively. Experimentally-determined relative values of electronic coupling (HAB) for the two reactions correlated well with the relative HAB values predicted from computational analysis of the structure. The rate of the hopping-mediated ET reaction is also ten-fold greater than that of the single-step tunneling reaction despite having a smaller overall driving force for the reaction. These data provide insight into how the intervening protein matrix and redox potentials of the electron donor and acceptor determine whether the ET reaction proceeds via single-step tunneling or multi-step hopping. PMID:22897160

Choi, Moonsung; Shin, Sooim; Davidson, Victor L.

2012-01-01

211

Lactic acid-producing yeast cells having nonfunctional L- or D-lactate:ferricytochrome C oxidoreductase cells  

SciTech Connect

Yeast cells having an exogenous lactate dehydrogenase gene ae modified by reducing L- or D-lactate:ferricytochrome c oxidoreductase activity in the cell. This leads to reduced consumption of lactate by the cell and can increase overall lactate yields in a fermentation process. Cells having the reduced L- or D-lactate:ferricytochrome c oxidoreductase activity can be screened for by resistance to organic acids such as lactic or glycolic acid.

Miller, Matthew (Boston, MA); Suominen, Pirkko (Maple Grove, MN); Aristidou, Aristos (Highland Ranch, CO); Hause, Benjamin Matthew (Currie, MN); Van Hoek, Pim (Camarillo, CA); Dundon, Catherine Asleson (Minneapolis, MN)

2012-03-20

212

The ylo-1 gene encodes an aldehyde dehydrogenase responsible for the last reaction in the Neurospora carotenoid pathway.  

PubMed

The accumulation of the apocarotenoid neurosporaxanthin and its carotene precursors explains the orange pigmentation of the Neurospora surface cultures. Neurosporaxanthin biosynthesis requires the activity of the albino gene products (AL-1, AL-2 and AL-3), which yield the precursor torulene. Recently, we identified the carotenoid oxygenase CAO-2, which cleaves torulene to produce the aldehyde beta-apo-4'-carotenal. This revealed a last missing step in Neurospora carotenogenesis, namely the oxidation of the CAO-2 product to the corresponding acid neurosporaxanthin. The mutant ylo-1, which exhibits a yellow colour, lacks neurosporaxanthin and accumulates several carotenes, but its biochemical basis is unknown. Based on available genetic data, we identified ylo-1 in the Neurospora genome, which encodes an enzyme representing a novel subfamily of aldehyde dehydrogenases, and demonstrated that it is responsible for the yellow phenotype, by sequencing and complementation of mutant alleles. In contrast to the precedent structural genes in the carotenoid pathway, light does not induce the synthesis of ylo-1 mRNA. In vitro incubation of purified YLO-1 protein with beta-apo-4'-carotenal produced neurosporaxanthin through the oxidation of the terminal aldehyde into a carboxyl group. We conclude that YLO-1 completes the set of enzymes needed for the synthesis of this major Neurospora pigment. PMID:18627463

Estrada, Alejandro F; Youssar, Loubna; Scherzinger, Daniel; Al-Babili, Salim; Avalos, Javier

2008-09-01

213

Importance of glutamate 87 and the substrate ?-amine for the reaction catalyzed by d-arginine dehydrogenase.  

PubMed

Pseudomonas aeruginosad-arginine dehydrogenase (PaDADH) catalyzes the oxidation of d-arginine to iminoarginine, which is non-enzymatically hydrolyzed to 2-ketoarginine and ammonia. Here, site-directed mutagenesis and pH effects were used to investigate binding and catalysis of zwitterionic and cationic substrates for the enzyme. An unprotonated group with apparent pKa value ?7.9 is required for binding d-arginine or d-lysine, but not d-methionine or d-leucine. This group is E87, as suggested by its replacement with leucine. An unprotonated group with pKa of 9.5, which persists in the H48F and E87L variants, is required for amine oxidation with all substrates. Since Y53 and Y249 were previously ruled out, the pKa is assigned to the substrate ?-NH3(+) group, which previous QM/MM and Kd pH-profile demonstrated to be protonated for preferred binding to the enzyme. Lack of pH effects on the (D)kred with d-leucine established 9.5 as the intrinsic pKa, and d-leucine as a non-sticky substrate. d-Arginine, d-lysine and d-methionine and their corresponding iminoproducts were significantly stickier than d-leucine, as indicated by apparent pKa values <9.5 in both kcat/Km and kcat. Restricted proton movements in catalysis were established from hollowed kcat pH profiles in wild-type PaDADH with d-lysine and in the H48F and E87L enzymes with d-arginine. PMID:25637657

Ball, Jacob; Bui, Quan V V; Gannavaram, Swathi; Gadda, Giovanni

2015-02-15

214

Modification of metabolic pathways of Saccharomyces cerevisiae by the expression of lactate dehydrogenase and deletion of pyruvate decarboxylase genes for the lactic acid fermentation at low pH value  

Microsoft Academic Search

Extractive lactic acid fermentation has recently been paid a great deal of attention. The problem with such a process is, however, that only undissociated lactate can be extracted. Therefore, lactic acid fermentation at low pH values is desirable. In the present study, we modified the metabolism of yeast (not lactic acid producing bacteria often cultivated at pH of 6–7) by

Eri Adachi; Mikiko Torigoe; Minetaka Sugiyama; Jun-Ichi Nikawa; Kazuyuki Shimizu

1998-01-01

215

Mechanistic and computational studies of the reductive half-reaction of tyrosine to phenylalanine active site variants of D-arginine dehydrogenase.  

PubMed

The flavin-mediated enzymatic oxidation of a CN bond in amino acids can occur through hydride transfer, carbanion, or polar nucleophilic mechanisms. Previous results with D-arginine dehydrogenase from Pseudomonas aeruginosa (PaDADH) using multiple deuterium kinetic isotope effects (KIEs) and computational studies established preferred binding of the substrate protonated on the ?-amino group, with cleavages of the NH and CH bonds occurring in asynchronous fashion, consistent with the three possible mechanisms. The hydroxyl groups of Y53 and Y249 are ?4 Å from the imino and carboxylate groups of the reaction product iminoarginine, suggesting participation in binding and catalysis. In this study, we have investigated the reductive half-reactions of the Y53F and Y249F variants of PaDADH using substrate and solvent deuterium KIEs, solvent viscosity and pH effects, and quantum mechanical/molecular mechanical computational approaches to gain insights into the catalytic roles of the tyrosines and evaluate whether their mutations affect the transition state for substrate oxidation. Both Y53F and Y249F enzymes oxidized D-arginine with steady-state kinetic parameters similar to those of the wild-type enzyme. Rate constants for flavin reduction (k(red)) with D-leucine, a slow substrate amenable to rapid kinetics, were 3-fold smaller than the wild-type value with similar pKa values for an unprotonated group of ?10.0. Similar pKa values were observed for (app)Kd in the variant and wild-type enzymes. However, cleavage of the substrate NH and CH bonds in the enzyme variants occurred in synchronous fashion, as suggested by multiple deuterium KIEs on k(red). These data can be reconciled with a hydride transfer mechanism, but not with carbanion and polar nucleophilic mechanisms. PMID:25243743

Gannavaram, Swathi; Sirin, Sarah; Sherman, Woody; Gadda, Giovanni

2014-10-21

216

Comprehensive review on lactate metabolism in human health.  

PubMed

Metabolic pathways involved in lactate metabolism are important to understand the physiological response to exercise and the pathogenesis of prevalent diseases such as diabetes and cancer. Monocarboxylate transporters are being investigated as potential targets for diagnosis and therapy of these and other disorders. Glucose and alanine produce pyruvate which is reduced to lactate by lactate dehydrogenase in the cytoplasm without oxygen consumption. Lactate removal takes place via its oxidation to pyruvate by lactate dehydrogenase. Pyruvate may be either oxidized to carbon dioxide producing energy or transformed into glucose. Pyruvate oxidation requires oxygen supply and the cooperation of pyruvate dehydrogenase, the tricarboxylic acid cycle, and the mitochondrial respiratory chain. Enzymes of the gluconeogenesis pathway sequentially convert pyruvate into glucose. Congenital or acquired deficiency on gluconeogenesis or pyruvate oxidation, including tissue hypoxia, may induce lactate accumulation. Both obese individuals and patients with diabetes show elevated plasma lactate concentration compared to healthy subjects, but there is no conclusive evidence of hyperlactatemia causing insulin resistance. Available evidence suggests an association between defective mitochondrial oxidative capacity in the pancreatic ?-cells and diminished insulin secretion that may trigger the development of diabetes in patients already affected with insulin resistance. Several mutations in the mitochondrial DNA are associated with diabetes mellitus, although the pathogenesis remains unsettled. Mitochondrial DNA mutations have been detected in a number of human cancers. d-lactate is a lactate enantiomer normally formed during glycolysis. Excess d-lactate is generated in diabetes, particularly during diabetic ketoacidosis. d-lactic acidosis is typically associated with small bowel resection. PMID:24929216

Adeva-Andany, M; López-Ojén, M; Funcasta-Calderón, R; Ameneiros-Rodríguez, E; Donapetry-García, C; Vila-Altesor, M; Rodríguez-Seijas, J

2014-07-01

217

Pyruvate and Lactate Metabolism by Shewanella oneidensis MR-1 under Fermentation, Oxygen Limitation, and Fumarate Respiration Conditions  

SciTech Connect

Shewanella oneidensis MR-1 is a facultative anaerobe growing by coupling organic matter oxidation to reduction of wide range of electron acceptors. Here we quantitatively assessed lactate and pyruvate metabolism of these bacteria under three distinct conditions: electron acceptor limited growth on lactate with O2 and fumarate, and pyruvate fermentation, which does not sustain growth but allows cells to survive for prolonged period. Using physiological and genetic approaches combined with flux balance analysis, we showed that the proportion of ATP produced by substrate-level phosphorylation varied from 33% to 72.5% of all ATP needed for growth depending on the electron acceptor nature and availability. While being indispensible for growth, respiration of fumarate does not contribute much to ATP generation and likely serves to remove formate, a product of pyruvate formate-lyase-catalyzed pyruvate disproportionation. Under both tested respiratory conditions S. oneidensis MR-1 carried out incomplete substrate oxidation, and TCA cycle did not contribute significantly to substrate oxidation. Pyruvate dehydrogenase reaction was not involved in lactate metabolism under O2 limitation, however was important for anaerobic growth probably supplying reducing equivalents for biosynthesis. Unexpectedly, obtained results suggest that pyruvate fermentation by S. oneidensis MR-1 cells represents a combination between substrate-level phosphorylation and a respiratory process, where pyruvate serves as electron donor and electron acceptor. Pyruvate reduction to lactate at the expense of formate oxidation is catalyzed by recently described new type of oxidative NAD(P)H independent D-lactate dehydrogenase (Dld-II). Based on involved enzymes localization we hypothesize that pyruvate reduction coupled to formate oxidation may be accompanied by proton motive force generation.

Pinchuk, Grigoriy E.; Geydebrekht, Oleg V.; Hill, Eric A.; Reed, Jennifer L.; Konopka, Allan; Beliaev, Alex S.; Fredrickson, Jim K.

2011-12-30

218

The aerobic CO dehydrogenase from Oligotropha carboxidovorans.  

PubMed

We review here the recent literature dealing with the molybdenum- and copper-dependent CO dehydrogenase, with particular emphasis on the structure of the enzyme and recent advances in our understanding of the reaction mechanism of the enzyme. PMID:25156151

Hille, Russ; Dingwall, Stephanie; Wilcoxen, Jarett

2015-03-01

219

NEUROENDOCRINOLOGY OF LACTATION  

Microsoft Academic Search

Although other hormones influence the quantity of milk secretion, prolactin and adrenal corticosteroids are essential for initiating and maintaining lactation. During pregnancy high levels of estrogen and progesterone inhibit lactation, and elevation of prolactin and ACTH-adrenal corticosteroid levels result in the initiation of lactation at the end of pregnancy. Suckling maintains lactation through release of prolactin, ACTH-adrenal glucocorticoid, and oxytocin.

Joseph Meites

1974-01-01

220

Localization of multiple human dihydrodiol dehydrogenase (DDH1 and DDH2) and chlordecone reductase (CHDR) genes in chromosome 10 by the polymerase chain reaction and fluorescence in situ hybridization  

SciTech Connect

Multiple human dihydrodiol dehydrogenases and human chlordecone reductase belong to the aldo-keto reductase superfamily. These two enzymes are involved in the metabolism of xenobiotics, such as polycyclic aromatic hydrocarbons and pesticides. Recently we have isolated three closely related genes encoding two dihydrodiol dehydrogenases (DDH1 and DDH2) and the chlordecone reductase (CHDR). Mapping of the location of the genes was performed using the polymerase chain reaction using gene-specific primers to amplify gene sequences in human/hamster hybrid DNA. All three genes were found to be located on chromosome 10. In situ hybridization using a lambda clone as the probe further confirmed regional localization at 10p14-p15. 13 refs., 2 figs.

Khanna, M.; Qin, K.N.; Belkin, S. [Cornell Univ. Medical College, New York, NY (United States)] [and others] [Cornell Univ. Medical College, New York, NY (United States); and others

1995-01-20

221

Immunofluorescent localization of glycogenolytic and glycolytic enzyme proteins and of malate dehydrogenase isozymes in cross-striated skeletal muscle and heart of the rabbit  

Microsoft Academic Search

Specific antisera against glycogen phosphorylase, phosphofructokinase, aldolase, glyceraldehyde-phosphate dehydrogenase, enolase, lactate dehydrogenase, cytosolic and mitochondrial malate dehydrogenase from rabbit muscle were obtained from sheep. The gammaglobulins were used for indirect immunofluorescent localization of the respective enzymes in rabbit skeletal muscle and heart. In stretched skeletal muscle a cross-striation like distribution was observed for all enzymes studied. In the case of

Gottfried Dölken; Elmi Leisner; Dirk Pette

1975-01-01

222

Cis-Chlorobenzene dihydrodiol dehydrogenase (TcbB) from Pseudomonas sp. strain P51, expressed in Escherichia coli DH5{alpha}(pTCB149), catalyzes enantioselective dehydrogenase reactions  

SciTech Connect

cis-Chlorobenzene dihydrodiol dehydrogenase (CDD) from Pseudomonas sp. strain P51, cloned into Escherichia coli DH5{alpha}(pTCB149) was able to oxidize cis-dihydrodihydroxy derivatives (cis-dihydrodiols) of dihydronaphthalene, indene, and four para-substituted toluenes to the corresponding catechols. During the incubation of a nonracemic mixture of cis-1,2-indandiol, only the (+)-cis-(1R,2S) enantiomer was oxidized; the (-)-cis-(S,2R) enantiomer remained unchanged, CDD oxidized both enantiomers of cis-1,2-dihydroxy-1,2,3,4-tetrahydronaphthalene, but oxidation of the (+)-cis-(1S,2R) enantiomer was delayed until the (-)-cis-(1R,2S) enantiomer was completely depleted. When incubated with nonracemic mixtures of para-substituted cis-toluene dihydrodiols, CDD always oxidized the major enantiomer at a higher rate than the minor enantiomer. When incubated with racemic 1-indanol, CDD enantioselectively transformed the (+)-(1S) enatiomer to 1-indanone. This stereoselective transformation shows that CDD also acted as an alcohol dehydrogenase. Additionally, CDD was able to oxidize (+)-cis-(1R,2S)-dihydroxy-1,2-dihydronaphthalene, (+)-cis-monochlorobiphenyl dihydrodiols, and (+)-cis-toluene dihydrodiol to the corresponding catechols.

Raschke, H.; Fleischmann, T.; Meer, J.R. van der; Kohler, H.P.E.

1999-12-01

223

Asparagusate dehydrogenases and lipoyl dehydrogenase from asparagus mitochondria. Physical, chemical, and enzymatic properties.  

PubMed

Asparagusate dehydrogenases I and II and lipoyl dehydrogenase have been obtained in homogeneous state from asparagus mitochondria. They are flavin enzymes with 1 mol of FAD/mol of protein. Asparagusate dehydrogenases I and II and lipoyl dehydrogenase have s20,w of 6.22 S, 6.39 S, and 5.91 S, respectively, and molecular weights of 111,000, 110,000, and 95,000 (sedimentation equilibrium) or 112,000, 112,000, and 92,000 (gel filtration). They are slightly acidic proteins with isoelectric points of 6.75, 5.75, and 6.80. Both asparagusate dehydrogenases catalyzed the reaction Asg(SH)2 + NAD+ equilibrium AsgS2 + NADH + H+ and exhibit lipoyl dehydrogenase and diaphorase activities. Lipoyl dehydrogenase is specific for lipoate and has no asparagusate dehydrogenase activity. NADP cannot replace NAD in any case. Optimum pH for substrate reduction of the three enzymes are near 5.9. Asparagusate dehydrogenases I and II have Km values of 21.5 mM and 20.0 mM for asparagusate and 3.0 mM and 3.3 mM for lipoate, respectively. Lipoyl dehydrogenase activity of asparagusate dehydrogenases is enhanced by NAD and surfactants such as lecithin and Tween 80, but asparagusate dehydrogenase activity is not enhanced. Asparagusate dehydrogenases are strongly inhibited by mercuric ion, p-chloromercuribenzoic acid, and N-ethylmaleimide. Amino acid composition of the three enzymes is presented and discussed. PMID:180003

Yanagawa, H; Egami, F

1976-06-25

224

Multiple Forms of D(--)3-Hydroxybutyrate Dehydrogenase in Rhizobium  

Microsoft Academic Search

SUMMARY 3-Hydroxybutyrate dehydrogenase was studied in 14 Rhizobium strains representing six species. Cell-free extracts of the bacteria were subjected to electrophoresis in starch or polyacrylamide gels and 3-hydroxybutyrate dehydrogenase was located in situ on the gels. Also, the rate of reaction of the enzyme, from different Rhizobium strains, with NAD was compared with its rate of reaction with various analogues

P. F. Fottrell; A. O'Hora

1969-01-01

225

Alcohol and aldehyde dehydrogenase polymorphisms and alcoholism  

Microsoft Academic Search

The alcohol-flush reaction occurs in Asians who inherit the mutantALDH2*2 allele that produces an inactive aldehyde dehydrogenase enzyme. In these individuals, high blood acetaldehyde levels are believed to be the cause of the unpleasant symptoms that follow drinking. We measured the alcohol elimination rates and intensity of flushing in Chinese subjects in whom the alcohol dehydrogenaseADH2 andALDH2 genotypes were determined.

Holly R. Thomasson; David W. Crabb; Howard J. Edenberg; Ting-Kai Li

1993-01-01

226

Stringency of substrate specificity of Escherichia coli malate dehydrogenase.  

SciTech Connect

Malate dehydrogenase and lactate dehydrogenase are members of the structurally and functionally homologous family of 2-ketoacid dehydrogenases. Both enzymes display high specificity for their respective keto substrates, oxaloacetate and pyruvate. Closer analysis of their specificity, however, reveals that the specificity of malate dehydrogenase is much stricter and less malleable than that of lactate dehydrogenase. Site-specific mutagenesis of the two enzymes in an attempt to reverse their specificity has met with contrary results. Conversion of a specific active-site glutamine to arginine in lactate dehydrogenase from Bacillus stearothermophilus generated an enzyme that displayed activity toward oxaloacetate equal to that of the native enzyme toward pyruvate (H. M. Wilks et al. (1988) Science 242, 1541-1544). We have constructed a series of mutants in the mobile, active site loop of the Escherichia coli malate dehydrogenase that incorporate the complementary change, conversion of arginine 81 to glutamine, to evaluate the role of charge distribution and conformational flexibility within this loop in defining the substrate specificity of these enzymes. Mutants incorporating the change R81Q all had reversed specificity, displaying much higher activity toward pyruvate than to the natural substrate, oxaloacetate. In contrast to the mutated lactate dehydrogenase, these reversed-specificity mutants were much less active than the native enzyme. Secondary mutations within the loop of the E. coli enzyme (A80N, A80P, A80P/M85E/D86T) had either no or only moderately beneficial effects on the activity of the mutant enzyme toward pyruvate. The mutation A80P, which can be expected to reduce the overall flexibility of the loop, modestly improved activity toward pyruvate. The possible physiological relevance of the stringent specificity of malate dehydrogenase was investigated. In normal strains of E. coli, fermentative metabolism was not affected by expression of the mutant malate dehydrogenase. However, when expressed in a strain of E. coli unable to ferment glucose, the mutant enzyme restored growth and produced lactic acid as the sole fermentation product.

Boernke, W. E.; Millard, C. S.; Stevens, P. W.; Kakar, S. N.; Stevens, F. J.; Donnelly, M. I.; Nebraska Wesleyan Univ.

1995-09-10

227

Malate dehydrogenase in bovine spermatozoa  

E-print Network

. Freshly ejaculated semen usually appears as a creamy, slightly yellowish or greyish fluid. The volume of the ejaculate, and the concentration I of the spermatozoa in ejaculated semen, vary widely from one species to another. So far as the nutrition... an improved method for demonstra- tion of dehydrogenase after starch gel electrophoresis based on the use of the more sensitive tetrazolium, NBT, I and substitution of phenazine methosulfate (PMS) for exo- genous diaphorase. The reaction sequence...

Lin, Hozong Robert

1973-01-01

228

Involvement of pyruvate dehydrogenase in product formation in pyruvate-limited anaerobic chemostat cultures of Enterococcus faecalis NCTC 775  

Microsoft Academic Search

Enterococcus faecalis NCTC 775 was grown anaerobically in chemostat culture with pyruvate as the energy source. At low culture pH values, high in vivo and in vitro activities were found for both pyruvate dehydrogenase and lactate dehydrogenase. At high culture pH values the carbon flux was shifted towards pyruvate formate lyase. Some mechanisms possibly involved in this metabolic switch are

Jacky L. Snoep; M. Joost Teixeira de Mattos; Pieter W. Postma; Oense M. Neijssel

1990-01-01

229

Structure and Function of Plasmodium falciparum malate dehydrogenase: Role of Critical Amino Acids in C-substrate Binding Procket  

Technology Transfer Automated Retrieval System (TEKTRAN)

Malaria parasite thrives on anaerobic fermentation of glucose for energy. Earlier studies from our lab have demonstrated that a cytosolic malate dehydrogenase (PfMDH) with striking similarity to lactate dehydrogenase (PfLDH) might complement PfLDH function in Plasmodium falciparum. The N-terminal g...

230

Analysis of Quaternary Structure of a [LDH-like] Malate Dehydrogenase of Plasmodium falciparum with Oligomeric Mutants  

Technology Transfer Automated Retrieval System (TEKTRAN)

L-Malate dehydrogenase (PfMDH) from Plasmodium falciparum, the causative agent for the most severe form of malaria, has shown remarkable similarities to L-lactate dehydrogenase (PfLDH). PfMDH is more closely related to [LDH-like] MDHs characterized in archea and other prokaryotes. Initial sequence a...

231

The control of the production of lactate and ethanol by higher plants.  

PubMed

Factors controlling the production of ethanol and lactate have been examined using cell free extracts prepared from pea seeds (Pisum sativum var Alaska) and parsnip roots (Pastinaca sativa). The result suggest that under aerobic conditions pyruvate decarboxylase is inactive. With the onset of anaerobiosis glycolysis leads to an accumulation of lactate with a corresponding fall in pH. The fall in pH activates pyruvate decarboxylase and initiates competition between lactate dehydrogenase and pyruvate decarboxylase for pyruvate. The effect of pyruvate concentration on the partitioning has been analysed in terms of a modified Wegscheider rule and shows that the ratio lactate dehydrogenase activity/pyruvate decarboxylase activity bears an inverse relationship to the pyruvate concentration. The decrease in ratio which occurs when the pyruvate concentration rises is enhanced by the co-operativity which is exhibited by pyruvate decarboxylase. The pH optimum of lactate dehydrogenase is alkaline whilst the pH optimum of pyruvic decarboxylase is acid, thus the two enzymes function as a pH-stat. The possibility of excessive production of lactic acid is further controlled by the response of lactate dehydrogenase to ATP; the enzyme is inhibited by ATP and the inhibition increases as the pH decreases. It is suggested that this mechanism functions to protect the plant from excess production of acid. PMID:24442374

Davies, D D; Grego, S; Kenworthy, P

1974-12-01

232

Modification of aldehyde dehydrogenase with dicyclohexylcarbodiimide: Separation of dehydrogenase from esterase activity  

Microsoft Academic Search

Dehydrogenase activity of the cytoplasmic (E1) isozyme of human liver aldehyde dehydrogenase (EC 1.2.1.3) was almost totally abolished (3% activity remaining) by preincubation with dicyclohexylcarbodiimide (DCC), while esterase activity with p-nitrophenyl acetate as substrate remained intact. The esterase reaction of the modified enzyme exhibited a hysteretic burst prior to achieving steady-state velocity; addition of NAD+ abolished the burst. TheKm for

Darryl P. Abriola; Regina Pietruszko

1992-01-01

233

BACTERIAL EXPRESSION, PURIFICATION, AND CHARACTERIZATION OF ARABIDOPSIS THALIANA PYRUVATE DEHYDROGENASE  

Technology Transfer Automated Retrieval System (TEKTRAN)

The pyruvate dehydrogenase complex (PDC) is a very large multi-component structure that catalyzes decarboxylation of pyruvate, yielding CO2, NADH, and acetyl-CoA as products. The decarboxylation reaction is catalyzed by pyruvate dehydrogenase (E1). The PDC occupies a key position in intermediary met...

234

Cellobiose dehydrogenase, an active agent in cellulose depolymerization  

Microsoft Academic Search

The ability of cellobiose dehydrogenase purified from Phanerochaete chrysosporium to modify a Douglas fir kraft pulp was assessed. Although the addition of cellobiose dehydrogenase alone had little effect, supplemen- tation with cellobiose and iron resulted in a substantial reduction in the degree of polymerization of the pulp cellulose. When the reaction was monitored over time, a progressive depolymerization of the

SHAWN D. MANSFIELD; ED DE JONG; JOHN N. SADDLER

1997-01-01

235

Identification of the Genes That Contribute to Lactate Utilization in Helicobacter pylori  

PubMed Central

Helicobacter pylori are Gram-negative, spiral-shaped microaerophilic bacteria etiologically related to gastric cancer. Lactate utilization has been implicated although no corresponding genes have been identified in the H. pylori genome. Here, we report that gene products of hp0137–0139 (lldEFG), hp0140–0141 (lctP), and hp1222 (dld) contribute to D- and L-lactate utilization in H. pylori. The three-gene unit hp0137–0139 in H. pylori 26695 encodes L-lactate dehydrogenase (LDH) that catalyzes the conversion of lactate to pyruvate in an NAD-dependent manner. Isogenic mutants of these genes were unable to grow on L-lactate-dependent medium. The hp1222 gene product functions as an NAD-independent D-LDH and also contributes to the oxidation of L-lactate; the isogenic mutant of this gene failed to grow on D-lactate-dependent medium. The parallel genes hp0140–0141 encode two nearly identical lactate permeases (LctP) that promote uptake of both D- and L-lactate. Interestingly an alternate route must also exist for lactate transport as the knockout of genes did not completely prevent growth on D- or L-lactate. Gene expression levels of hp0137–0139 and hp1222 were not enhanced by lactate as the carbon source. Expression of hp0140–0141 was slightly suppressed in the presence of L-lactate but not D-lactate. This study identified the genes contributing to the lactate utilization and demonstrated the ability of H. pylori to utilize both D- and L-lactate. PMID:25078575

Iwatani, Shun; Nagashima, Hiroyuki; Reddy, Rita; Shiota, Seiji; Graham, David Y.; Yamaoka, Yoshio

2014-01-01

236

Lactate Dehydrogenase Catalysis: Roles of Keto, Hydrated, and Enol Pyruvate  

ERIC Educational Resources Information Center

Many carbonyl substrates of oxidoreductase enzymes undergo hydration and enolization so that these substrate systems are partitioned between keto, hydrated (gem-diol), and enol forms in aqueous solution. Some oxidoreductase enzymes are subject to inhibition by high concentrations of substrate. For such enzymes, two questions arise pertaining to…

Meany, J. E.

2007-01-01

237

The Partial Purification and Characterization of Lactate Dehydrogenase.  

ERIC Educational Resources Information Center

Offers several advantages over other possibilities as the enzyme of choice for a student's first exposure to a purification scheme. Uses equipment and materials normally found in biochemistry laboratories. Incorporates several important biochemical techniques including spectrophotometry, chromatography, centrifugation, and electrophoresis. (MVL)

Wolf, Edward C.

1988-01-01

238

Physiology of lactation  

Technology Transfer Automated Retrieval System (TEKTRAN)

The breast changes in size, shape, and function during puberty, pregnancy, and lactation. The physiology of lactation is reviewed here. The breast is composed of fat and connective tissue that supports a tubuloalveolar structure. During development, anatomic changes involving new lobule formation an...

239

Water Recycling in Lactation  

Microsoft Academic Search

During lactation, female rodents, dingoes, and kangaroos consume urine and feces excreted by the young. Studies with tritiated water as a tracer for native water showed that roughly one-third of the water secreted as milk was returned to the mother. The results are cogent to studies of water balance of lactation and to current methods used for estimating milk production.

P. Baverstock; B. Green

1975-01-01

240

Direct and Nitroxyl (HNO)-Mediated Reactions of Acyloxy Nitroso Compounds with the Thiol- Containing Proteins Glyceraldehyde 3- Phosphate Dehydrogenase and Alkyl Hydroperoxide Reductase Subunit C  

PubMed Central

Nitroxyl (HNO) reacts with thiols and this reactivity requires the use of donors with 1-nitrosocyclohexyl acetate, pivalate and trifluoroacetate forming a new group. These acyloxy nitroso compounds inhibit glyceraldehyde 3-phosphate dehydrogenase (GAPDH) by forming a reduction reversible active site disulfide and a reduction irreversible sulfinic acid or sulfinamide modification at Cys 244. Addition of these acyloxy nitroso compounds to AhpC C165S yields a sulfinic acid and sulfinamide modification. A potential mechanism for these transformations includes nucleophilic addition of the protein thiol to a nitroso compound to yield an N-hydroxysulfenamide, which reacts with thiol to give disulfide or rearranges to sulfinamides. Known HNO donors produce the un-substituted protein sulfinamide as the major product while the acetate and pivalate give substituted sulfinamides that hydrolyze to sulfinic acids. These results suggest that nitroso compounds form a general class of thiol-modifying compounds allowing their further exploration. PMID:23895568

Mitroka, Susan; Shoman, Mai E.; DuMond, Jenna F.; Bellavia, Landon; Aly, Omar M.; Abdel-Aziz, Mohamed; Kim-Shapiro, Daniel B.; King, S. Bruce

2013-01-01

241

Evidences of Basal Lactate Production in the Main White Adipose Tissue Sites of Rats. Effects of Sex and a Cafeteria Diet  

PubMed Central

Female and male adult Wistar rats were fed standard chow or a simplified cafeteria diet for one month. Then, the rats were killed and the white adipose tissue (WAT) in four sites: perigonadal, retroperitoneal, mesenteric and subcutaneous (inguinal) were sampled and frozen. The complete WAT weight in each site was measured. Gene expression analysis of key lipid and glucose metabolism enzymes were analyzed, as well as tissue and plasma lactate and the activity of lactate dehydrogenase. Lactate gradients between WAT and plasma were estimated. The influence of sex and diet (and indirectly WAT mass) on lactate levels and their relationships with lactate dehydrogenase activity and gene expressions were also measured. A main conclusion is the high production of lactate by WAT, practically irrespective of site, diet or sex. Lactate production is a direct correlate of lactate dehydrogenase activity in the tissue. Furthermore, lactate dehydrogenase activity is again directly correlated with the expression of the genes Ldha and Ldhb for this enzyme. In sum, the ability to produce lactate by WAT is not directly dependent of WAT metabolic state. We postulate that, in WAT, a main function of the lactate dehydrogenase path may be that of converting excess available glucose to 3C fragments, as a way to limit tissue self-utilization as substrate, to help control glycaemia and/or providing short chain substrates for use as energy source elsewhere. More information must be gathered before a conclusive role of WAT in the control of glycaemia, and the full existence of a renewed glucose-lactate-fatty acid cycle is definitely established. PMID:25741703

Arriarán, Sofía; Agnelli, Silvia; Sabater, David; Remesar, Xavier; Fernández-López, José Antonio; Alemany, Marià

2015-01-01

242

Intracellular Shuttle: The Lactate Aerobic Metabolism  

PubMed Central

Lactate is a highly dynamic metabolite that can be used as a fuel by several cells of the human body, particularly during physical exercise. Traditionally, it has been believed that the first step of lactate oxidation occurs in cytosol; however, this idea was recently challenged. A new hypothesis has been presented based on the fact that lactate-to-pyruvate conversion cannot occur in cytosol, because the LDH enzyme characteristics and cytosolic environment do not allow the reaction in this way. Instead, the Intracellular Lactate Shuttle hypothesis states that lactate first enters in mitochondria and only then is metabolized. In several tissues of the human body this idea is well accepted but is quite resistant in skeletal muscle. In this paper, we will present not only the studies which are protagonists in this discussion, but the potential mechanism by which this oxidation occurs and also a link between lactate and mitochondrial proliferation. This new perspective brings some implications and comes to change our understanding of the interaction between the energy systems, because the product of one serves as a substrate for the other. PMID:22593684

Cruz, Rogério Santos de Oliveira; de Aguiar, Rafael Alves; Turnes, Tiago; Penteado Dos Santos, Rafael; Fernandes Mendes de Oliveira, Mariana; Caputo, Fabrizio

2012-01-01

243

Primary Structure and Phylogenetic Relationships of a Malate Dehydrogenase Gene from Giardia lamblia  

Microsoft Academic Search

.   The lactate and malate dehydrogenases comprise a complex protein superfamily with multiple enzyme homologues found in eubacteria,\\u000a archaebacteria, and eukaryotes. In this study we describe the sequence and phylogenetic relationships of a malate dehydrogenase\\u000a (MDH) gene from the amitochondriate diplomonad protist, Giardia lamblia. Parsimony, distance, and maximum-likelihood analyses of the MDH protein family solidly position G. lamblia MDH within

Andrew J. Roger; Hilary G. Morrison; Mitchell L. Sogin

1999-01-01

244

Plant Formate Dehydrogenase  

SciTech Connect

The research in this study identified formate dehydrogenase, an enzyme that plays a metabolic role on the periphery of one-carbon metabolism, has an unusual localization in Arabidopsis thaliana and that the enzyme has an unusual kinetic plasticity. These properties make it possible that this enzyme could be engineered to attempt to engineer plants with an improved photosynthetic efficiency. We have produced transgenic Arabidopsis and tobacco plants with increased expression of the formate dehydrogenase enzyme to initiate further studies.

John Markwell

2005-01-10

245

Inhibition of endogenous lactate turnover with lactate infusion in humans  

SciTech Connect

The extent to which lactate infusion may inhibit endogenous lactate production, though previously considered, has never been critically assessed. To examine this proposition, single injection tracer methodology (U-{sup 14}C Lactate) has been used for the estimation of lactate kinetics in 12 human subjects under basal conditions and with the infusion of sodium lactate. The basal rate of lactate turnover was measured on a day before the study with lactate infusion, and averaged 63.7 + 5.5 mg/kg/h. Six of these individuals received a stable lactate infusion at an approximate rate of 160 mg/kg/h, while the remaining six individuals were infused at the approximate rate of 100 mg/kg/h. It has been found that stable lactate infused at rates approximating 160 mg/kg/h consistently produced a complete inhibition of endogenous lactate production. Infusion of lactate at 100 mg/kg/h caused a lesser and more variable inhibition of endogenous lactate production (12% to 64%). In conclusion, lactate infusion significantly inhibits endogenous lactate production.

Searle, G.L.; Feingold, K.R.; Hsu, F.S.; Clark, O.H.; Gertz, E.W.; Stanley, W.C. (Veterans Administration Medical Center, San Francisco, CA (USA))

1989-11-01

246

Reference values of blood parameters in beef cattle of different ages and stages of lactation.  

PubMed Central

Reference (normal) values for 12 blood serum components were determined for 48 Shorthorn cows (2-10 years old) and their 48 calves, 357 crossbred cows (12-14 years old), 36 feedlot bulls and 36 feedlot steers. In addition, hemoglobin, hematocrit, triiodothyronine, thyroxine and cortisol levels were determined for the crossbred cows, and feedlot bulls and steers. Reference values were tabulated according to sex, age and stage of lactation. Serum concentrations of urea, total protein and bilirubin, and serum activity of aspartate aminotransferase and lactate dehydrogenase increased with age (P less than 0.05), while calcium, phosphorus and alkaline phosphatase decreased with age (P less than 0.05) from birth to the age of ten years. The Shorthorn cows had the highest levels of glucose at parturition (P less than 0.05) with decreasing levels during lactation. Creatinine concentration decreased during lactation and increased during postweaning. Both lactate dehydrogenase and aspartate aminotransferase levels increased (P less than 0.05) during lactation. Urea and uric acid were present at higher concentrations in lactating than nonlactating cows (P less than 0.05). The values reported, based on a wide age range and large number of cattle, could serve as clinical guides and a basis for further research. PMID:3349406

Doornenbal, H; Tong, A K; Murray, N L

1988-01-01

247

The Essential Function of Genes for a Hydratase and an Aldehyde Dehydrogenase for Growth of Pseudomonas sp. Strain Chol1 with the Steroid Compound Cholate Indicates an Aldolytic Reaction Step for Deacetylation of the Side Chain  

PubMed Central

In the bacterial degradation of steroid compounds, the enzymes initiating the breakdown of the steroid rings are well known, while the reactions for degrading steroid side chains attached to C-17 are largely unknown. A recent in vitro analysis with Pseudomonas sp. strain Chol1 has shown that the degradation of the C5 acyl side chain of the C24 steroid compound cholate involves the C22 intermediate 7?,12?-dihydroxy-3-oxopregna-1,4-diene-20S-carbaldehyde (DHOPDCA) with a terminal aldehyde group. In the present study, candidate genes with plausible functions in the formation and degradation of this aldehyde were identified. All deletion mutants were defective in growth with cholate but could transform it into dead-end metabolites. A mutant with a deletion of the shy gene, encoding a putative enoyl coenzyme A (CoA) hydratase, accumulated the C24 steroid (22E)-7?,12?-dihydroxy-3-oxochola-1,4,22-triene-24-oate (DHOCTO). Deletion of the sal gene, formerly annotated as the steroid ketothiolase gene skt, resulted in the accumulation of 7?,12?,22-trihydroxy-3-oxochola-1,4-diene-24-oate (THOCDO). In cell extracts of strain Chol1, THOCDO was converted into DHOPDCA in a coenzyme A- and ATP-dependent reaction. A sad deletion mutant accumulated DHOPDCA, and expression in Escherichia coli revealed that sad encodes an aldehyde dehydrogenase for oxidizing DHOPDCA to the corresponding acid 7?,12?-dihydroxy-3-oxopregna-1,4-diene-20-carboxylate (DHOPDC) with NAD+ as the electron acceptor. These results clearly show that the degradation of the acyl side chain of cholate proceeds via an aldolytic cleavage of an acetyl residue; they exclude a thiolytic cleavage for this reaction step. Based on these results and on sequence alignments with predicted aldolases from other bacteria, we conclude that the enzyme encoded by sal catalyzes this aldolytic cleavage. PMID:23708132

Holert, Johannes; Jagmann, Nina

2013-01-01

248

Asparagusate dehydrogenases and lipoyl dehydrogenase from asparagus mitochondria.  

PubMed

1. Lipoyl dehydrogenase (NADH: lipoamide oxidoreductase, ED 1.6.4.3) and two asparagusate dehydrogenases from asparagus mitochondria were purified by a series of steps, freezing and thawing, sodium dodecylsulfate extraction, and chromatography on Sephadex G-200 and DEAE-cellulose. 2. Lipoyl dehydrogenase was highly specific for alpha-lipoic acid, which could not be replaced at all by asparagusic acid. Each of the asparagusate dehydrogenases was capable of reducing both asparagusic and alpha-lipoic acids by using NADH as hydrogen donor. 3. Reduction of alpha-lipoic cid with NADH by lipoyl dehydrogenase was activated by NAD, but that of asparagusic acid by asparagusate dehydrogenase was inactivated by NAD. 4. Lipoyl dehydrogenase and two asparagusate dehydrogenases differed in electrophoretic mobility on polyacrylamide gels. PMID:1125255

Yanagawa, H; Egami, F

1975-04-19

249

21 CFR 73.165 - Ferrous lactate.  

Code of Federal Regulations, 2011 CFR

...2011-04-01 2011-04-01 false Ferrous lactate. 73.165 Section 73.165 Food...CERTIFICATION Foods § 73.165 Ferrous lactate. (a) Identity. The color additive ferrous lactate is the ferrous lactate defined in §...

2011-04-01

250

21 CFR 73.165 - Ferrous lactate.  

Code of Federal Regulations, 2013 CFR

...2013-04-01 2013-04-01 false Ferrous lactate. 73.165 Section 73.165 Food...CERTIFICATION Foods § 73.165 Ferrous lactate. (a) Identity. The color additive ferrous lactate is the ferrous lactate defined in §...

2013-04-01

251

21 CFR 73.165 - Ferrous lactate.  

Code of Federal Regulations, 2014 CFR

...2014-04-01 2014-04-01 false Ferrous lactate. 73.165 Section 73.165 Food...CERTIFICATION Foods § 73.165 Ferrous lactate. (a) Identity. The color additive ferrous lactate is the ferrous lactate defined in §...

2014-04-01

252

21 CFR 73.165 - Ferrous lactate.  

Code of Federal Regulations, 2012 CFR

...2012-04-01 2012-04-01 false Ferrous lactate. 73.165 Section 73.165 Food...CERTIFICATION Foods § 73.165 Ferrous lactate. (a) Identity. The color additive ferrous lactate is the ferrous lactate defined in §...

2012-04-01

253

21 CFR 73.165 - Ferrous lactate.  

Code of Federal Regulations, 2010 CFR

...2010-04-01 2010-04-01 false Ferrous lactate. 73.165 Section 73.165 Food...CERTIFICATION Foods § 73.165 Ferrous lactate. (a) Identity. The color additive ferrous lactate is the ferrous lactate defined in §...

2010-04-01

254

Interaction of human aldehyde dehydrogenase with aromatic substrates and ligands  

Microsoft Academic Search

The substrate benzaldehyde (but not propionaldehyde) could elute aldehyde dehydrogenase from a p-hydroxyacetophenone-affinity column, and inhibit the esterase activity (Ki=47 ?M), indicating that this simple aromatic aldehyde binds to the free enzyme and possibly in the substrate-binding site. Thus, the kinetic mechanism for aldehyde dehydrogenase might be dependent upon which aldehyde is used in the reaction. Chloramphenicol which also elutes

Abdellah Allali-Hassani; Henry Weiner

2001-01-01

255

Affinity chromatography of nicotinamide–adenine dinucleotide-linked dehydrogenases on immobilized derivatives of the dinucleotide  

PubMed Central

1. Three established methods for immobilization of ligands through primary amino groups promoted little or no attachment of NAD+ through the 6-amino group of the adenine residue. Two of these methods (coupling to CNBr-activated agarose and to carbodi-imide-activated carboxylated agarose derivatives) resulted instead in attachment predominantly through the ribosyl residues. Other immobilized derivatives were prepared by azolinkage of NAD+ (probably through the 8 position of the adenine residue) to a number of different spacer-arm–agarose derivatives. 2. The effectiveness of these derivatives in the affinity chromatography of a variety of NAD-linked dehydrogenases was investigated, applying rigorous criteria to distinguish general or non-specific adsorption effects from truly NAD-specific affinity (bio-affinity). The ribosyl-attached NAD+ derivatives displayed negligible bio-affinity for any of the NAD-linked dehydrogenases tested. The most effective azo-linked derivative displayed strong bio-affinity for glycer-aldehyde 3-phosphate dehydrogenase, weaker bio-affinity for lactate dehydrogenase and none at all for malate dehydrogenase, although these three enzymes have very similar affinities for soluble NAD+. Alcohol dehydrogenase and xanthine dehydrogenase were subject to such strong non-specific interactions with the hydrocarbon spacer-arm assembly that any specific affinity was completely eclipsed. 3. It is concluded that, in practice, the general effectiveness of a general ligand may be considerably distorted and attenuated by the nature of the immobilization linkage. However, this attenuation can result in an increase in specific effectiveness, allowing dehydrogenases to be separated from one another in a manner unlikely to be feasible if the general effectiveness of the ligand remained intact. 4. The bio-affinity of the various derivatives for lactate dehydrogenase is correlated with the known structure of the NAD+-binding site of this enzyme. Problems associated with the use of immobilized derivatives for enzyme binding and mechanistic studies are briefly discussed. PMID:4360246

Barry, Standish; O'Carra, Pádraig

1973-01-01

256

Photo-production of lactate from glyoxylate: how minerals can facilitate energy storage in a prebiotic world  

E-print Network

Photo-production of lactate from glyoxylate: how minerals can facilitate energy storage924179e The reaction of glyoxylate with carbon dioxide to produce lactate is promoted when zinc sulfide (HCOCOO� ) reacts with CO2 to produce the C3 compound lactate (CH3­HCOH­COO� ) in 15% yield through a Zn

257

Mechanism and applications of phosphite dehydrogenase.  

PubMed

Phosphite dehydrogenase catalyzes the NAD+-dependent oxidation of hydrogen phosphonate (common name phosphite) to phosphate in what amounts to a formal phosphoryl transfer reaction from hydride to hydroxide. This review places the enzyme in the context of phosphorus redox metabolism in nature and discusses the results of mechanistic investigations into its reaction mechanism. The potential of the enzyme as a NAD(P)H cofactor regeneration system is discussed as well as efforts to engineer the cofactor specificity of the protein. PMID:15888310

Relyea, Heather A; van der Donk, Wilfred A

2005-06-01

258

Elevated plasma citrulline: look for dihydrolipoamide dehydrogenase deficiency.  

PubMed

The E3 subunit of the pyruvate dehydrogenase complex (dihydrolipoamide dehydrogenase/dihydrolipoyl dehydrogenase/DLD/lipoamide dehydrogenase/LAD), is a mitochondrial matrix enzyme and also a part of the branched-chain ketoacid dehydrogenase and alpha-ketoglutarate dehydrogenase complexes. DLD deficiency (MIM #246900), is relatively frequent in the Ashkenazi Jewish population but occurs in other populations as well. Early diagnosis is important to prevent episodes of metabolic decompensation, liver failure, and encephalopathy. The clinical presentations are varied and may include Reye-like syndrome, hepatic failure, myopathy, and myoglobinuria. Laboratory markers, such as elevated urinary alpha-ketoglutarate, blood pyruvate, lactate, and ammonia, are mostly nonspecific and not always present, making the diagnosis difficult. Since we observed elevated plasma citrulline levels in a number of confirmed cases, we retrospectively examined the value of citrulline as a biochemical marker for DLD deficiency. Data was gathered from the files of 17 pediatric patients with DLD deficiency, confirmed by enzymatic and genetic analysis. The control group included 19 patients in whom urea cycle defects were ruled out but DLD deficiency was suspected. Seven of the DLD-deficient patients presented with elevated plasma citrulline levels (median value 205 ?M, range 59-282 ?M) (normal range 1-45 ?M) while none in the control patient group. In five patients, elevated citrulline was associated with elevated plasma glutamine and metabolic acidosis. Interestingly, elevated plasma citrulline was associated with the common G229C mutation. In conclusion, we suggest that elevated plasma citrulline in the absence of urea cycle defects warrants an investigation for DLD deficiency. PMID:23995961

Haviv, Ruby; Zeharia, Avraham; Belaiche, Corinne; Haimi Cohen, Yishai; Saada, Ann

2014-02-01

259

Uranyl nitrate inhibits lactate gluconeogenesis in isolated human and mouse renal proximal tubules: A {sup 13}C-NMR study  

SciTech Connect

As part of a study on uranium nephrotoxicity, we investigated the effect of uranyl nitrate in isolated human and mouse kidney cortex tubules metabolizing the physiological substrate lactate. In the millimolar range, uranyl nitrate reduced lactate removal and gluconeogenesis and the cellular ATP level in a dose-dependent fashion. After incubation in phosphate-free Krebs-Henseleit medium with 5 mM L-[1-{sup 13}C]-, or L-[2-{sup 13}C]-, or L-[3-{sup 13}C]lactate, substrate utilization and product formation were measured by enzymatic and NMR spectroscopic methods. In the presence of 3 mM uranyl nitrate, glucose production and the intracellular ATP content were significantly reduced in both human and mouse tubules. Combination of enzymatic and NMR measurements with a mathematical model of lactate metabolism revealed an inhibition of fluxes through lactate dehydrogenase and the gluconeogenic enzymes in the presence of 3 mM uranyl nitrate; in human and mouse tubules, fluxes were lowered by 20% and 14% (lactate dehydrogenase), 27% and 32% (pyruvate carboxylase), 35% and 36% (phosphoenolpyruvate carboxykinase), and 39% and 45% (glucose-6-phosphatase), respectively. These results indicate that natural uranium is an inhibitor of renal lactate gluconeogenesis in both humans and mice.

Renault, Sophie; Faiz, Hassan; Gadet, Rudy; Ferrier, Bernard; Martin, Guy; Baverel, Gabriel [Metabolomique et Maladies Metaboliques, Institut National de la Sante et de la recherche Medicale, Unit 820, Faculte de Medecine R.T.H. Laennec, Universite de Lyon, 7-11 rue G. Paradin, 69372 Lyon Cedex 08 (France); Conjard-Duplany, Agnes, E-mail: agnes.duplany@recherche.univ-lyon1.f [Metabolomique et Maladies Metaboliques, Institut National de la Sante et de la recherche Medicale, Unit 820, Faculte de Medecine R.T.H. Laennec, Universite de Lyon, 7-11 rue G. Paradin, 69372 Lyon Cedex 08 (France)

2010-01-01

260

Uranyl nitrate inhibits lactate gluconeogenesis in isolated human and mouse renal proximal tubules: a 13C-NMR study.  

PubMed

As part of a study on uranium nephrotoxicity, we investigated the effect of uranyl nitrate in isolated human and mouse kidney cortex tubules metabolizing the physiological substrate lactate. In the millimolar range, uranyl nitrate reduced lactate removal and gluconeogenesis and the cellular ATP level in a dose-dependent fashion. After incubation in phosphate-free Krebs-Henseleit medium with 5 mM L-[1-13C]-, or L-[2-13C]-, or L-[3-13C]lactate, substrate utilization and product formation were measured by enzymatic and NMR spectroscopic methods. In the presence of 3 mM uranyl nitrate, glucose production and the intracellular ATP content were significantly reduced in both human and mouse tubules. Combination of enzymatic and NMR measurements with a mathematical model of lactate metabolism revealed an inhibition of fluxes through lactate dehydrogenase and the gluconeogenic enzymes in the presence of 3 mM uranyl nitrate; in human and mouse tubules, fluxes were lowered by 20% and 14% (lactate dehydrogenase), 27% and 32% (pyruvate carboxylase), 35% and 36% (phosphoenolpyruvate carboxykinase), and 39% and 45% (glucose-6-phosphatase), respectively. These results indicate that natural uranium is an inhibitor of renal lactate gluconeogenesis in both humans and mice. PMID:19747499

Renault, Sophie; Faiz, Hassan; Gadet, Rudy; Ferrier, Bernard; Martin, Guy; Baverel, Gabriel; Conjard-Duplany, Agnès

2010-01-01

261

Lactating Mother and Psychotropic Drugs  

PubMed Central

Usage of psychotropics during pregnancy and lactation has always been a topic of debate and controversy. The debate stems from the potential adverse effects on the growing fetus or infants due to the transfer of psychotropic drugs through placenta or breast milk of mothers receiving them; and the problem of discontinuing psychotropics in lactating mother considering chances of relapse. However, most of the psychotropics are found to be relatively safe when used cautiously during the lactation phase. This article describes available data on the use of psychotropics in lactating mothers, in particular, in relation to the safety profile of infants. PMID:21327172

Tripathi, B. M.; Majumder, Pradipta

2010-01-01

262

reactions  

Microsoft Academic Search

Gene regulatory, signal transduction and metabolic networks are major areas of interest in the newly emerging field of systems biology. In living cells, stochastic dynamics play an important role; however, the kinetic parameters of biochemical reactions necessary for modelling these processes are often not accessible directly through experiments. The problem of estimating stochastic reaction constants from molecule count data measured,

S. Reinker; R. M. Altman; J. Timmer

263

Characterization of lactate utilization and its implication on the physiology of Haemophilus influenzae  

PubMed Central

Haemophilus influenzae is a Gram-negative bacillus and a frequent commensal of the human nasopharynx. Earlier work demonstrated that in H. influenzae type b, l-lactate metabolism is associated with serum resistance and in vivo survival of the organism. To further gain insight into lactate utilization of the non-typeable (NTHi) isolate 2019 and laboratory prototype strain Rd KW20, deletion mutants of the l-lactate dehydrogenase (lctD) and permease (lctP) were generated and characterized. It is shown, that the apparent KM of l-lactate uptake is 20.1 ?M as determined for strain Rd KW20. Comparison of the COPD isolate NTHi 2019-R with the corresponding lctP knockout strain for survival in human serum revealed no lactate dependent serum resistance. In contrast, we observed a 4-fold attenuation of the mutant strain in a murine model of nasopharyngeal colonization. Characterization of lctP transcriptional control shows that the lactate utilization system in H. influenzae is not an inductor inducible system. Rather negative feedback regulation was observed in the presence of l-lactate and this is dependent on the ArcAB regulatory system. Additionally, for 2019 it was found that lactate may have signaling function leading to increased cell growth in late log phase under conditions where no l-lactate is metabolized. This effect seems to be ArcA independent and was not observed in strain Rd KW20. We conclude that l-lactate is an important carbon-source and may act as host specific signal substrate which fine tunes the globally acting ArcAB regulon and may additionally affect a yet unknown signaling system and thus may contribute to enhanced in vivo survival. PMID:24674911

Lichtenegger, Sabine; Bina, Isabelle; Roier, Sandro; Bauernfeind, Stilla; Keidel, Kristina; Schild, Stefan; Anthony, Mark; Reidl, Joachim

2014-01-01

264

Relationships within the aldehyde dehydrogenase extended family.  

PubMed Central

One hundred-forty-five full-length aldehyde dehydrogenase-related sequences were aligned to determine relationships within the aldehyde dehydrogenase (ALDH) extended family. The alignment reveals only four invariant residues: two glycines, a phenylalanine involved in NAD binding, and a glutamic acid that coordinates the nicotinamide ribose in certain E-NAD binary complex crystal structures, but which may also serve as a general base for the catalytic reaction. The cysteine that provides the catalytic thiol and its closest neighbor in space, an asparagine residue, are conserved in all ALDHs with demonstrated dehydrogenase activity. Sixteen residues are conserved in at least 95% of the sequences; 12 of these cluster into seven sequence motifs conserved in almost all ALDHs. These motifs cluster around the active site of the enzyme. Phylogenetic analysis of these ALDHs indicates at least 13 ALDH families, most of which have previously been identified but not grouped separately by alignment. ALDHs cluster into two main trunks of the phylogenetic tree. The largest, the "Class 3" trunk, contains mostly substrate-specific ALDH families, as well as the class 3 ALDH family itself. The other trunk, the "Class 1/2" trunk, contains mostly variable substrate ALDH families, including the class 1 and 2 ALDH families. Divergence of the substrate-specific ALDHs occurred earlier than the division between ALDHs with broad substrate specificities. A site on the World Wide Web has also been devoted to this alignment project. PMID:10210192

Perozich, J.; Nicholas, H.; Wang, B. C.; Lindahl, R.; Hempel, J.

1999-01-01

265

Genomic reconstruction of Shewanella oneidensis MR-1 metabolism reveals previously uncharacterized machinery for lactate utilization  

SciTech Connect

The ability to utilize lactate as a sole source of carbon and energy is one of the key metabolic signatures of Shewanellae, a diverse group of dissimilatory metal reducing bacteria commonly found in aquatic and sedimentary environments. Nonetheless, homology searches failed to recognize orthologs of previously described bacterial D- or L-lactate oxidizing enzymes (Escherichia coli genes dld and lldD) in any of the 13 analyzed genomes of Shewanella spp. Using comparative genomic techniques, we identified a conserved chromosomal gene cluster in Shewanella oneidensis MR-1 (locus tag: SO1522-SO1518) containing lactate permease and candidate genes for both D- and L-lactate dehydrogenase enzymes. The predicted D-LDH gene (dldD, SO1521) is a distant homolog of FAD-dependent lactate dehydrogenase from yeast, whereas the predicted L-LDH is encoded by three genes with previously unknown functions (lldEGF, SO1520-19-18). Through a combination of genetic and biochemical techniques, we experimentally confirmed the predicted physiological role of these novel genes in S. oneidensis MR-1 and carried out successful functional validation studies in Escherichia coli and Bacillus subtilis. We conclusively showed that dldD and lldEFG encode fully functional D-and L-LDH enzymes, which catalyze the oxidation of the respective lactate stereoisomers to pyruvate. Notably, the S. oneidensis MR-1 LldEFG enzyme is the first described example of a multi-subunit lactate oxidase. Comparative analysis of >400 bacterial species revealed the presence of LldEFG and Dld in a broad range of diverse species accentuating the potential importance of these previously unknown proteins in microbial metabolism.

Pinchuk, Grigoriy E.; Rodionov, Dmitry A.; Yang, Chen; Li, Xiaoqing; Osterman, Andrei L.; Dervyn, Etienne; Geydebrekht, Oleg V.; Reed, Samantha B.; Romine, Margaret F.; Collart, Frank R.; Scott, J.; Fredrickson, Jim K.; Beliaev, Alex S.

2009-02-24

266

Modulation of susceptibility to weight gain and insulin resistance in low birthweight rats by treatment of their mothers with leptin during pregnancy and lactation  

Microsoft Academic Search

OBJECTIVES: To investigate whether administration of leptin to rats during pregnancy and lactation affects placental 11?-hydroxysteroid dehydrogenase (11?-HSD2) activity and the susceptibility of their offspring to weight gain and insulin resistance.DESIGN: Pregnant rats fed on a low-protein diet were administered leptin or saline by subcutaneous minipump from day 14 of gestation and throughout lactation. A further group was fed a

C Stocker; J O'Dowd; N M Morton; E Wargent; M V Sennitt; D Hislop; S Glund; J R Seckl; J R S Arch; M A Cawthorne

2004-01-01

267

Neonatal pyruvate dehydrogenase deficiency with lipoate responsive lactic acidaemia and hyperammonaemia  

Microsoft Academic Search

A 2-day-old girl developed a severe lactic acidosis with a normal lactate\\/pyruvate ratio and hyperammonaemia. Plasma arginine and citrulline levels were below the limit of detection. In muscle total pyruvate dehydrogenase complex (PDHC) and pyruvate decarboxylase (E1) activities were reduced to a fraction of lower control values. The acute neonatal period was bridged with peritoneal dialysis, dichloroacetate therapy, supplements of

D. J. Byrd; H.-P. Krohn; L. Winkler; C. Steinborn; M. Hadam; J. Brodehl; D. H. Hunneman

1989-01-01

268

Synthesis of Triptorelin Lactate Catalyzed by Lipase in Organic Media  

PubMed Central

Triptorelin lactate was successfully synthesized by porcine pancreatic lipase (PPL) in organic solvents. The effects of acyl donor, substrate ratio, organic solvent, temperature, and water activity were investigated. Under the optimum conditions, a yield of 30% for its ester could be achieved in the reaction for about 48 h. PMID:22949842

Zhuang, Hong; Wang, Zhi; Wang, Jiaxin; Zhang, Hong; Xun, Erna; Chen, Ge; Yue, Hong; Tang, Ning; Wang, Lei

2012-01-01

269

Metformin suppresses gluconeogenesis by inhibiting mitochondrial glycerophosphate dehydrogenase.  

PubMed

Metformin is considered to be one of the most effective therapeutics for treating type 2 diabetes because it specifically reduces hepatic gluconeogenesis without increasing insulin secretion, inducing weight gain or posing a risk of hypoglycaemia. For over half a century, this agent has been prescribed to patients with type 2 diabetes worldwide, yet the underlying mechanism by which metformin inhibits hepatic gluconeogenesis remains unknown. Here we show that metformin non-competitively inhibits the redox shuttle enzyme mitochondrial glycerophosphate dehydrogenase, resulting in an altered hepatocellular redox state, reduced conversion of lactate and glycerol to glucose, and decreased hepatic gluconeogenesis. Acute and chronic low-dose metformin treatment effectively reduced endogenous glucose production, while increasing cytosolic redox and decreasing mitochondrial redox states. Antisense oligonucleotide knockdown of hepatic mitochondrial glycerophosphate dehydrogenase in rats resulted in a phenotype akin to chronic metformin treatment, and abrogated metformin-mediated increases in cytosolic redox state, decreases in plasma glucose concentrations, and inhibition of endogenous glucose production. These findings were replicated in whole-body mitochondrial glycerophosphate dehydrogenase knockout mice. These results have significant implications for understanding the mechanism of metformin's blood glucose lowering effects and provide a new therapeutic target for type 2 diabetes. PMID:24847880

Madiraju, Anila K; Erion, Derek M; Rahimi, Yasmeen; Zhang, Xian-Man; Braddock, Demetrios T; Albright, Ronald A; Prigaro, Brett J; Wood, John L; Bhanot, Sanjay; MacDonald, Michael J; Jurczak, Michael J; Camporez, Joao-Paulo; Lee, Hui-Young; Cline, Gary W; Samuel, Varman T; Kibbey, Richard G; Shulman, Gerald I

2014-06-26

270

Lactate and lactate clearance in acute cardiac care patients  

PubMed Central

Hyperlactataemia is commonly used as a diagnostic and prognostic tool in intensive care settings. Recent studies documented that serial lactate measurements over time (or lactate clearance), may be clinically more reliable than lactate absolute value for risk stratification in different pathological conditions. While the negative prognostic role of hyperlactataemia in several critical ill diseases (such as sepsis and trauma) is well established, data in patients with acute cardiac conditions (i.e. acute coronary syndromes) are scarce and controversial. The present paper provides an overview of the current available evidence on the clinical role of lactic acid levels and lactate clearance in acute cardiac settings (acute coronary syndromes, cardiogenic shock, cardiac surgery), focusing on its prognostic role. PMID:24062898

Lazzeri, Chiara; Picariello, Claudio; Dini, Carlotta Sorini; Gensini, Gian Franco; Valente, Serafina

2012-01-01

271

Mechanism of activation of pyruvate dehydrogenase by dichloroacetate and other halogenated carboxylic acids  

PubMed Central

1. Monochloroacetate, dichloroacetate, trichloroacetate, difluoroacetate, 2-chloropropionate, 2,2?-dichloropropionate and 3-chloropropionate were inhibitors of pig heart pyruvate dehydrogenase kinase. Dichloroacetate was also shown to inhibit rat heart pyruvate dehydrogenase kinase. The inhibition was mainly non-competitive with respect to ATP. The concentration required for 50% inhibition was approx. 100?m for the three chloroacetates, difluoroacetate and 2-chloropropionate and 2,2?-dichloropropionate. Dichloroacetamide was not inhibitory. 2. Dichloroacetate had no significant effect on the activity of pyruvate dehydrogenase phosphate phosphatase when this was maximally activated by Ca2+ and Mg2+. 3. Dichloroacetate did not increase the catalytic activity of purified pig heart pyruvate dehydrogenase. 4. Dichloroacetate, difluoroacetate, 2-chloropropionate and 2,2?-dichloropropionate increased the proportion of the active (dephosphorylated) form of pyruvate dehydrogenase in rat heart mitochondria with 2-oxoglutarate and malate as respiratory substrates. Similar effects of dichloroacetate were shown with kidney and fat-cell mitochondria. Glyoxylate, monochloroacetate and dichloroacetamide were inactive. 5. Dichloroacetate increased the proportion of active pyruvate dehydrogenase in the perfused rat heart, isolated rat diaphragm and rat epididymal fat-pads. Difluoroacetate and dichloroacetamide were also active in the perfused heart, but glyoxylate, monochloroacetate and trichloroacetate were inactive. 6. Injection of dichloroacetate into rats starved overnight led within 60 min to activation of pyruvate dehydrogenase in extracts from heart, psoas muscle, adipose tissue, kidney and liver. The blood concentration of lactate fell within 15 min to reach a minimum after 60 min. The blood concentration of glucose fell after 90 min and reached a minimum after 120 min. There was no significant change in plasma glycerol concentration. 7. In epididymal fatpads dichloroacetate inhibited incorporation of 14C from [U-14C]glucose, [U-14C]fructose and from [U-14C]lactate into CO2 and glyceride fatty acid. 8. It is concluded that the inhibition of pyruvate dehydrogenase kinase by dichloroacetate may account for the activation of pyruvate dehydrogenase and pyruvate oxidation which it induces in isolated rat heart and diaphragm muscles, subject to certain assumptions as to the distribution of dichloroacetate across the plasma membrane and the mitochondrial membrane. 9. It is suggested that activation of pyruvate dehydrogenase by dichloroacetate could contribute to its hypoglycaemic effect by interruption of the Cori and alanine cycles. 10. It is suggested that the inhibitory effect of dichloroacetate on fatty acid synthesis in adipose tissue may involve an additional effect or effects of the compound. PMID:4478069

Whitehouse, Sue; Cooper, Ronald H.; Randle, Philip J.

1974-01-01

272

Antibacterial characteristics and activity of water-soluble chitosan derivatives prepared by the Maillard reaction.  

PubMed

The antibacterial activity of water-soluble chitosan derivatives prepared by Maillard reactions against Staphylococcus aureus, Listeria monocytogenes, Bacillus cereus, Escherichia coli, Shigella dysenteriae, and Salmonella typhimurium was examined. Relatively high antibacterial activity against various microorganisms was noted for the chitosan-glucosamine derivative as compared to the acid-soluble chitosan. In addition, it was found that the susceptibility of the test organisms to the water-soluble chitosan derivative was higher in deionized water than in saline solution. Metal ions were also found to reduce the antibacterial activity of the water-soluble chitosan derivative on S. aureus. The marked increase in glucose level, protein content and lactate dehydrogenase (LDH) activity was observed in the cell supernatant of S. aureus exposed to the water-soluble chitosan derivative in deionized water. The results suggest that the water-soluble chitosan produced by Maillard reaction may be a promising commercial substitute for acid-soluble chitosan. PMID:21989311

Chung, Ying-Chien; Yeh, Jan-Ying; Tsai, Cheng-Fang

2011-01-01

273

The metabolism and action of insulin and glucagon in lactating and non-lactating goats  

E-print Network

The metabolism and action of insulin and glucagon in lactating and non-lactating goats J. GRIZARD in the adaptation of metabolism to lactation (Gill and Hart, 1980 ; Burnol et al., 1985) but the mechanisms and their action on blood glucose in lactating and non-lactating goats. Material and methods. Nine Alpine goats 4

Paris-Sud XI, Université de

274

Kinetic mechanism of chicken liver xanthine dehydrogenase.  

PubMed Central

The kinetic behaviour of chicken-liver xanthine dehydrogenase (xanthine/NAD+ oxidoreductase; EC 1.2.1.37) has been studied. Steady-state results, obtained from a wide range of concentrations of substrates and products, were fitted by rational functions of degree 1:1, 1:2, 2:2 and 3:3 with respect to substrates, and 0:1, 1:1, 0:2 and 1:2 with regard to products, using a non-linear regression program which guarantees the fit. The goodness of fit was improved using a computer program that combines model discrimination, parameter refinement and sequential experimental design. The AIC and F tests were also used for model discrimination. For comparative purposes, the xanthine/oxygen oxidoreductase reaction was also studied. From the functions which give the maximum improvement, the complete rate equation was deduced. The significance of the terms was stated by the above methods. It was concluded that xanthine dehydrogenase requires a minimum mechanism of degree 1:1 for xanthine, 2:2 for NAD+, 1:1 for uric acid and 1:2 for NADH in the xanthine/NAD+ oxidoreductase reaction. These are the minimum degrees required but a rate equation of higher degree is not excluded. PMID:3422556

Bruguera, P; Lopez-Cabrera, A; Canela, E I

1988-01-01

275

Molybdenum and tungsten-dependent formate dehydrogenases.  

PubMed

The prokaryotic formate metabolism is considerably diversified. Prokaryotes use formate in the C1 metabolism, but also evolved to exploit the low reduction potential of formate to derive energy, by coupling its oxidation to the reduction of numerous electron acceptors. To fulfil these varied physiological roles, different types of formate dehydrogenase (FDH) enzymes have evolved to catalyse the reversible 2-electron oxidation of formate to carbon dioxide. This review will highlight our present knowledge about the diverse physiological roles of FDH in prokaryotes, their modular structural organisation and active site structures and the mechanistic strategies followed to accomplish the formate oxidation. In addition, the ability of FDH to catalyse the reverse reaction of carbon dioxide reduction, a potentially relevant reaction for carbon dioxide sequestration, will also be addressed. PMID:25476858

Maia, Luisa B; Moura, José J G; Moura, Isabel

2015-03-01

276

l-Lactate Production from Biodiesel-Derived Crude Glycerol by Metabolically Engineered Enterococcus faecalis: Cytotoxic Evaluation of Biodiesel Waste and Development of a Glycerol-Inducible Gene Expression System.  

PubMed

Biodiesel waste is a by-product of the biodiesel production process that contains a large amount of crude glycerol. To reuse the crude glycerol, a novel bioconversion process using Enterococcus faecalis was developed through physiological studies. The E. faecalis strain W11 could use biodiesel waste as a carbon source, although cell growth was significantly inhibited by the oil component in the biodiesel waste, which decreased the cellular NADH/NAD(+) ratio and then induced oxidative stress to cells. When W11 was cultured with glycerol, the maximum culture density (optical density at 600 nm [OD600]) under anaerobic conditions was decreased 8-fold by the oil component compared with that under aerobic conditions. Furthermore, W11 cultured with dihydroxyacetone (DHA) could show slight or no growth in the presence of the oil component with or without oxygen. These results indicated that the DHA kinase reaction in the glycerol metabolic pathway was sensitive to the oil component as an oxidant. The lactate dehydrogenase (Ldh) activity of W11 during anaerobic glycerol metabolism was 4.1-fold lower than that during aerobic glycerol metabolism, which was one of the causes of low l-lactate productivity. The E. faecalis pflB gene disruptant (?pfl mutant) expressing the ldhL1LP gene produced 300 mM l-lactate from glycerol/crude glycerol with a yield of >99% within 48 h and reached a maximum productivity of 18 mM h(-1) (1.6 g liter(-1) h(-1)). Thus, our study demonstrates that metabolically engineered E. faecalis can convert crude glycerol to l-lactate at high conversion efficiency and provides critical information on the recycling process for biodiesel waste. PMID:25576618

Doi, Yuki

2015-03-15

277

Lactate Utilization Is Regulated by the FadR-Type Regulator LldR in Pseudomonas aeruginosa  

PubMed Central

NAD-independent l-lactate dehydrogenase (l-iLDH) and NAD-independent d-lactate dehydrogenase (d-iLDH) activities are induced coordinately by either enantiomer of lactate in Pseudomonas strains. Inspection of the genomic sequences of different Pseudomonas strains revealed that the lldPDE operon comprises 3 genes, lldP (encoding a lactate permease), lldD (encoding an l-iLDH), and lldE (encoding a d-iLDH). Cotranscription of lldP, lldD, and lldE in Pseudomonas aeruginosa strain XMG starts with the base, C, that is located 138 bp upstream of the lldP ATG start codon. The lldPDE operon is located adjacent to lldR (encoding an FadR-type regulator, LldR). The gel mobility shift assays revealed that the purified His-tagged LldR binds to the upstream region of lldP. An XMG mutant strain that constitutively expresses d-iLDH and l-iLDH was found to contain a mutation in lldR that leads to an Ile23-to-serine substitution in the LldR protein. The mutated protein, LldRM, lost its DNA-binding activity. A motif with a hyphenated dyad symmetry (TGGTCTTACCA) was identified as essential for the binding of LldR to the upstream region of lldP by using site-directed mutagenesis. l-Lactate and d-lactate interfered with the DNA-binding activity of LldR. Thus, l-iLDH and d-iLDH were expressed when the operon was induced in the presence of l-lactate or d-lactate. PMID:22408166

Gao, Chao; Hu, Chunhui; Zheng, Zhaojuan; Jiang, Tianyi; Dou, Peipei; Zhang, Wen; Che, Bin; Wang, Yujiao; Lv, Min

2012-01-01

278

Measurement of interstitial lactate during hypoxia-induced dilatation in isolated pressurised porcine coronary arteries  

PubMed Central

Lactate is formed in the coronary arterial wall and in the myocardium as a consequence of ischaemia and infarction. We combined direct measurement of coronary artery diameter and interstitial arterial wall lactate concentration ex vivo in order to ascertain the possible role of lactate in hypoxia-induced vasodilatation. The wall of porcine coronary arteries, precontracted during an intraluminal pressure of 40 mmHg by addition of prostaglandin F2?, was cannulated using a microdialysis catheter, and exposed to hypoxia for 60 min, followed by 45 min of reoxygenation. The exchange fraction of [14C]lactate over the microdialysis membrane increased from 0.38 ± 0.04 to 0.52 ± 0.05 (P < 0.001) during the study period. Coronary artery diameter increased by 15.5 ± 2.0 % (n = 20) during hypoxia (P < 0.001, compared to normoxic controls) and interstitial lactate concentration rose from 1.07 ± 0.21 to 2.50 ± 0.40 mmol l?1 during hypoxia (P < 0.01) and was unchanged in controls. The increase in coronary artery diameter correlated with the increase in interstitial lactate concentration in the period between 30 and 60 min of hypoxia (r = 0.62; P = 0.02). Dichloroacetate (10?5m), an agent that reduces lactate generation by activating pyruvate dehydrogenase, abolished hypoxia-induced lactate production, but caused a further increase in coronary arterial diameter (30.2 ± 4.4 %, n = 9; P < 0.001 vs. hypoxia and no dichloroacetate). Under control conditions, the addition of l-lactate (10?3-10?2m) increased dose-dependently coronary arterial diameter by 22.0 ± 4.2 % (n = 5) and interstitial lactate concentration from 0.52 ± 0.04 to 5.70 ± 0.66 mmol l?1 (P < 0.001). There was a correlation between the increase in coronary artery diameter and interstitial lactate concentration (r = 0.60; P = 0.02). The present observations represent the first direct measurements of metabolites by microdialysis in a blood vessel wall. The lactate concentration may affect, but is not essential for, hypoxia-induced vasodilatation in porcine coronary arteries. PMID:11850519

Frøbert, Ole; Mikkelsen, Erich O; Bagger, Jens P; Gravholt, Claus H

2002-01-01

279

Boosting D-lactate production in engineered cyanobacteria using sterilized anaerobic digestion effluents.  

PubMed

Anaerobic digestion (AD) is an environmentally friendly approach to waste treatment, which can generate N and P-rich effluents that can be used as nutrient sources for microalgal cultivations. Modifications of AD processes to inhibit methanogenesis leads to the accumulation of acetic acid, a carbon source that can promote microalgal biosynthesis. This study tested different AD effluents from municipal wastes on their effect on D-lactate production by an engineered Synechocystis sp. PCC 6803 (carrying a novel lactate dehydrogenase). The results indicate that: (1) AD effluents can be supplemented into the modified BG-11 culture medium (up to 1:4 volume ratio) to reduce N and P cost; (2) acetate-rich AD effluents enhance D-lactate synthesis by ? 40% (1.2g/L of D-lactate in 20 days); and (3) neutral or acidic medium had a deleterious effect on lactate secretion and biomass growth by the engineered strain. This study demonstrates the advantages and guidelines in employing wastewater for photomixotrophic biosynthesis using engineered microalgae. PMID:25084044

Hollinshead, Whitney D; Varman, Arul M; You, Le; Hembree, Zachary; Tang, Yinjie J

2014-10-01

280

Exogenous lactate supply affects lactate kinetics of rainbow trout, not swimming performance.  

PubMed

Intense swimming causes circulatory lactate accumulation in rainbow trout because lactate disposal (Rd) is not stimulated as strongly as lactate appearance (Ra). This mismatch suggests that maximal Rd is limited by tissue capacity to metabolize lactate. This study uses exogenous lactate to investigate what constrains maximal Rd and minimal Ra. Our goals were to determine how exogenous lactate affects: 1) Ra and Rd of lactate under baseline conditions or during graded swimming, and 2) exercise performance (critical swimming speed, Ucrit) and energetics (cost of transport, COT). Results show that exogenous lactate allows swimming trout to boost maximal Rd lactate by 40% and reach impressive rates of 56 ?mol·kg(-1)·min(-1). This shows that the metabolic capacity of tissues for lactate disposal is not responsible for setting the highest Rd normally observed after intense swimming. Baseline endogenous Ra (resting in normoxic water) is not significantly reduced by exogenous lactate supply. Therefore, trout have an obligatory need to produce lactate, either as a fuel for oxidative tissues and/or from organs relying on glycolysis. Exogenous lactate does not affect Ucrit or COT, probably because it acts as a substitute for glucose and lipids rather than extra fuel. We conclude that the observed 40% increase in Rd lactate is made possible by accelerating lactate entry into oxidative tissues via monocarboxylate transporters (MCTs). This observation together with the weak expression of MCTs and the phenomenon of white muscle lactate retention show that lactate metabolism of rainbow trout is significantly constrained by transmembrane transport. PMID:25121611

Omlin, Teye; Langevin, Karolanne; Weber, Jean-Michel

2014-10-15

281

Enzymatic properties of ALDH1L2, a mitochondrial 10-formyltetrahydrofolate dehydrogenase  

Microsoft Academic Search

10-Formyltetrahydrofolate dehydrogenase (FDH, ALDH1L1), an abundant cytosolic enzyme of folate metabolism, shares significant sequence similarity with enzymes of the aldehyde dehydrogenase (ALDH) family. The enzyme converts 10-formyltetrahydrofolate (10-fTHF) to tetrahydrofolate and CO2 in an NADP+-dependent manner. The mechanism of this reaction includes three consecutive steps with the final occurring in an ALDH-homologous domain. We have recently identified a mitochondrial isoform

Kyle C. Strickland; Natalia I. Krupenko; Marianne E. Dubard; Calvin J. Hu; Yaroslav Tsybovsky; Sergey A. Krupenko

2011-01-01

282

Allosteric inhibition of human liver aldehyde dehydrogenase by the isoflavone prunetin  

Microsoft Academic Search

Isoflavonoid derivatives including prunetin (4?,5-dihydroxy-7-methoxyisoflavone) were shown to be potent inhibitors of human aldehyde dehydrogenases (Keung W-M and Vallee BL, Proc Nad Acad Sci USA 90: 1247–1251, 1993). The inhibition reaction was reinvestigated using recombinantly expressed human aldehyde dehydrogenases. The kinetic analyses showed that prunetin inhibits competitively against both NAD and propionaldehyde with the mitochondrial and cytoplasmic enzymes. The Ki

Saifuddin Sheikh; Henry Weiner

1997-01-01

283

Deficiency of dihydrolipoyl dehydrogenase (a component of the pyruvate and alpha-ketoglutarate dehydrogenase complexes): a cause of congenital chronic lactic acidosis in infancy.  

PubMed

A male child died at 7 months of age with progressive neurologic deterioration and persistent metabolic acidosis. Investigations during life showed this child to have elevated blood pyruvate, lactate, and alpha-ketoglutarate as well as elevation of branched chain amino acids and occasional hypoglycemia. Cofactor therapy using either thiamine-HCl (2 g/kg/24 hr) or thiamine tetrahydrofurfuryl disulfide had no measurable effect on the clinical or biochemical status of the patient. Tissue taken postmortem showed normal levels of key gluconeogenic enzymes but a deficiency in the activity of pyruvate dehydrogenase in all tissues tested (liver, brain, kidney, skeletal muscle, and heart). Examination of the individual activities of pyruvate dehydrogenase complex showed pyruvate decarboxylase (E1) to be normal in liver and other tissues. Dihydrolipoyl dehydrogenase (E3), on the other hand, was deficient in all tissues tested. alpha-Ketoglutarate dehydrogenase complex, which depends of E3 for its total activity, was also deficient in all tissues tested. The absence of this enzyme id discussed in relation to the clinical and biochemical status of the patient. PMID:413089

Robinson, B H; Taylor, J; Sherwood, W G

1977-12-01

284

EXPRESSION ARABIDOPSIS THALIANA PYRUVATE DEHYDROGENASE KINASE  

Technology Transfer Automated Retrieval System (TEKTRAN)

Pyruvate dehydrogenase kinase (PDK) is the primary regulator of flux through the mitochondrial pyruvate dehydrogenase complex (PDC). The PDK catalyzes phosphorylation of Ser residues in the alpha subunit of the pyruvate dehydrogenase (E1alpha). Phosphorylation of E1alpha inactivates the PDC. In mamm...

285

High-pressure phase behavior of propyl lactate and butyl lactate in supercritical carbon dioxide  

Microsoft Academic Search

Lactate esters synthesized with lactic acid and ester are used as solvents and reactants in various industries, including agricultural chemistry, pharmaceuticals, electronics, and fine chemicals. Among lactate esters, high purity propyl lactate and butyl lactate are used to produce fine chemicals and in the synthesis of chiral intermediates for use in pesticides and drugs. However, distillation for the removal of

Dong Woo Cho; Jungin Shin; Moon Sam Shin; Won Bae; Hwayong Kim

286

DV Office/Lactation Sub-Committee 11/19/12 Workplace Lactation Policy  

E-print Network

DV Office/Lactation Sub-Committee 11/19/12 Workplace Lactation Policy Effective January 1, 2013 Brookhaven Science Associates, LLC FOR ITS EMPLOYEES AT Brookhaven National Laboratory #12;DV Office/Lactation Sub-Committee 11/19/12 Workplace Lactation Policy Brookhaven National Laboratory (BNL) provides

Ohta, Shigemi

287

Lysine Metabolism by the Mammary Gland of Lactating Goats at Two Stages of Lactation  

E-print Network

Lysine Metabolism by the Mammary Gland of Lactating Goats at Two Stages of Lactation S. J. Mabjeesh lysine and protein metabolism in goats (n = 4) at two stages of lactation (80 ± 17 vs. 233 ± 14 DIM) in response to an i.v. infusion of lysine (Lys) plus methionine (Met). At each stage of lactation [2-15 N

Bequette, Brian J.

288

Lactation performance of mid-Lactation Dairy Cows Fed ruminally Degradable protein at Concentrations Lower  

E-print Network

Lactation performance of mid-Lactation Dairy Cows Fed ruminally Degradable protein, and the apparent efficiency of N utilization by mid-lactation dairy cows. During the covariate period (d 1 to 28), 40 mid-lactation cows (36 Holstein and 4 Jersey Ã? Holstein cross-breds) were fed a common diet

Bequette, Brian J.

289

Is cellobiose dehydrogenase from Phanerochaete chrysosporium a lignin degrading enzyme?  

Microsoft Academic Search

Cellobiose dehydrogenase (CDH) is an extracellular redox enzyme of ping-pong type, i.e. it has separate oxidative and reductive half reactions. Several wood degrading fungi produce CDH, but the biological function of the enzyme is not known with certainty. It can, however, indirectly generate hydroxyl radicals by reducing Fe3+ to Fe2+ and O2 to H2O2. Hydroxyl radicals are then generated by

Gunnar Henriksson; Liming Zhang; Jiebing Li; Pierre Ljungquist; Torbjörn Reitberger; Göran Pettersson; Gunnar Johansson

2000-01-01

290

Lactation/Quiet Rooms Physical Specifications  

E-print Network

Lactation/Quiet Rooms Physical Specifications L/QR in New Construction New construction on the University of Texas at Austin campus will provide for multipurpose private rooms, called Lactation condition. Ideally, Lactation/Quiet rooms will be located around campus so that no user has more than a five

Johnston, Daniel

291

The Occurrence of Glycolate Dehydrogenase and Glycolate Oxidase in Green Plants  

PubMed Central

Homogenates of various lower land plants, aquatic angiosperms, and green algae were assayed for glycolate oxidase, a peroxisomal enzyme present in green leaves of higher plants, and for glycolate dehydrogenase, a functionally analogous enzyme characteristic of certain green algae. Green tissues of all lower land plants examined (including mosses, liverworts, ferns, and fern allies), as well as three freshwater aquatic angiosperms, contained an enzyme resembling glycolate oxidase, in that it oxidized l- but not d-lactate in addition to glycolate, and was insensitive to 2 mm cyanide. Many of the green algae (including Chlorella vulgaris, previously claimed to have glycolate oxidase) contained an enzyme resembling glycolate dehydrogenase, in that it oxidized d- but not l-lactate, and was inhibited by 2 mm cyanide. Other green algae had activity characteristic of glycolate oxidase and, accordingly, showed a substantial glycolate-dependent O2 uptake. It is pointed out that this distribution pattern of glycolate oxidase and glycolate dehydrogenase among the green plants may have phylogenetic significance. Activities of catalase, a marker enzyme for peroxisomes, were also determined and were generally lower in the algae than in the land plants or aquatic angiosperms. Among the algae, however, there were no consistent correlations between levels of catalase and the type of enzyme which oxidized glycolate. PMID:16658555

Frederick, Sue Ellen; Gruber, Peter J.; Tolbert, N. E.

1973-01-01

292

Glucose-6-phosphate dehydrogenase revisited.  

PubMed

Hemolytic diseases associated with drugs have been recognized since antiquity. Many of these anemias have been associated with oxidizing agents and deficiencies in the intraerythrocytic enzyme glucose-6-phosphate dehydrogenase. This paper outlines the discovery, prevalence, and variants of this enzyme. Methods of diagnosis of associated anemias are offered. PMID:6502728

O'Connell, J T; Henderson, A R

1984-11-01

293

CYTOCHEMICAL LOCALIZATION OF TWO GLYCOLYTIC DEHYDROGENASES IN WHITE SKELETAL MUSCLE  

PubMed Central

The cytochemical localization, by conventional methods, of lactate and glyceraldehyde-3-phosphate dehydrogenases is limited, firstly, by the solubility of these enzymes in aqueous media and, secondly, by the dependence of the final electron flow from reduced nicotinamide-adenine dinucleotide (NADH) to the tetrazolium on tissue diaphorase activity: localization is therefore that of the diaphorase, which in rabbit adductor magnus is mitochondrial. NADH has been found to have great affinity to bind in the sarcoplasmic reticulum, and, therefore, if it is generated freely in the incubation media containing 2,2',5,5'-tetra-p-nitrophenyl-3,3'-(3,3'-dimethoxy-4,4'-phenylene)-ditetrazolium chloride (TNBT) and N-methyl phenazonium methyl sulfate (PMS), it can bind there and cause a false staining. Since such a production of NADH can readily occur in the incubation media for glycolytic dehydrogenases due to diffusion of these soluble enzymes from tissue sections, the prevention of enzyme solubilization is extremely important. Fixation in formaldehyde prevented such enzyme diffusion, while at the same time sufficient activity persisted to allow for adequate staining. The incubation media contained PMS, so that the staining system was largely independent of tissue diaphorase activity. Application of these methods to adductor magnus of rabbit revealed by light microscopy, for both enzymes, a fine network which was shown by electron microscopy to represent staining of the sarcoplasmic reticulum. Mitochondria also reacted. These findings add further support for the notion that the sarcoplasmic reticulum is probably involved in glycolytic activity. PMID:4288329

Fahimi, H. Dariush; Karnovsky, Morris J.

1966-01-01

294

Sorbitol dehydrogenase is a zinc enzyme.  

PubMed Central

Evidence is given that tetrameric sorbitol dehydrogenase from sheep liver contains one zinc atom per subunit, most probably located at the active site, and no other specifically bound zinc or iron atom. In alcohol dehydrogenases that are structurally related to sorbitol dehydrogenase, more than one zinc atom per subunit can complicate investigations of zinc atom function. Therefore, sorbitol dehydrogenase will be particularly valuable for defining the precise roles of zinc in alcohol and polyol dehydrogenases, and for establishing correlations of structure and function with other important zinc-containing proteins. PMID:6370679

Jeffery, J; Chesters, J; Mills, C; Sadler, P J; Jörnvall, H

1984-01-01

295

Pyruvate and Lactate Metabolism by Shewanella oneidensis MR-1 under Fermentation, Oxygen Limitation, and Fumarate Respiration Conditions  

SciTech Connect

Shewanella oneidensis MR-1 is a facultative anaerobe that derives energy by coupling organic matter oxidation to the reduction of wide range of electron acceptors. Here, we quantitatively assessed lactate and pyruvate metabolism of MR-1 under three distinct conditions: electron acceptor limited growth on lactate with O2; lactate with fumarate; and pyruvate fermentation. The latter does not support growth but provides energy for cell survival. Using physiological and genetic approaches combined with flux balance analysis, we showed that the proportion of ATP produced by substrate-level phosphorylation varied from 33% to 72.5% of that needed for growth depending on the electron acceptor nature and availability. While being indispensible for growth, respiration of fumarate does not contribute significantly to ATP generation and likely serves to remove formate, a product of pyruvate formate-lyase-catalyzed pyruvate disproportionation. Under both tested respiratory conditions S. oneidensis MR-1 carried out incomplete substrate oxidation, whereby the TCA cycle did not contribute significantly. Pyruvate dehydrogenase was not involved in lactate metabolism under O2 limitation but was required for anaerobic growth likely by supplying reducing equivalents for biosynthesis. The results suggest that pyruvate fermentation by S. oneidensis MR-1 cells represents a combination of substrate-level phosphorylation and respiration, where pyruvate serves as electron donor and electron acceptor. Pyruvate reduction to lactate at the expense of formate oxidation is catalyzed by recently described new type of oxidative NAD(P)H independent D-lactate dehydrogenase (Dld-II). The results further indicate that pyruvate reduction coupled to formate oxidation may be accompanied by proton motive force generation.

Pinchuk, Grigoriy E.; Geydebrekht, Oleg V.; Hill, Eric A.; Reed, Jennifer L.; Konopka, Allan; Beliaev, Alex S.; Fredrickson, Jim K.

2011-12-01

296

Structure of liver alcohol dehydrogenase at 2.9-angstrom resolution.  

PubMed

The conformation of the polypeptide chain in horse liver alcohol dehydrogenase (EC 1.1.1.1), as well as the binding sites for some inhibitor molecules, have been determined from x-ray crystallographic data to a resolution of 2.9 A. Each subunit of the dimeric molecule is organized into two parts unequal in size and separated by a wide and deep active-site cleft. The adenosine moiety of the coenzyme is bound within the smaller region. Interactions between these coenzyme-binding substructures define the subunit contact area of the molecule. The "catalytic" zinc atoms are bound at the bottom of the clefts about 20 A from the surface of the molecule. The coenzyme binding region has a main-chain conformation very similar to a corresponding region in lactate and malate dehydrogenase. It is suggested that this substructure is a general one for binding of nucleotides and, in particular, the coenzyme NAD(+). PMID:4365379

Brändén, C I; Eklund, H; Nordström, B; Boiwe, T; Söderlund, G; Zeppezauer, E; Ohlsson, I; Akeson, A

1973-08-01

297

Polymorphisms of Human Aldehyde Dehydrogenases  

Microsoft Academic Search

Aldehyde dehydrogenases (ALDHs), a superfamily of NAD(P)+-dependent enzymes with similar primary structures, catalyze the oxidation of a wide spectrum of endogenous and exogenous aliphatic and aromatic aldehydes. Thus far, 16 ALDH genes with distinct chromosomal locations have been identified in the human genome. Polymorphism in ALDH2 is associated with altered acetaldehyde metabolism, decreased risk of alcoholism and increased risk of

Vasilis Vasiliou; Aglaia Pappa

2000-01-01

298

Dichloroacetate increases glucose use and decreases lactate in developing rat brain  

SciTech Connect

Dichloroacetate (DCA) activates pyruvate dehydrogenase (PDH) by inhibiting PDH kinase. Neutralized DCA (100 mg/kg) or saline was intravenously administered to 20 to 25-day-old rats (50-75g). Fifteen minutes later a mixture of {sup 6-14}C glucose and {sup 3}H fluorodeoxyglucose (FDG) was administered intravenously and the animals were sacrificed by microwave irradiation (2450 MHz, 8.0 kW, 0.6-0.8 sec) after 2 or 5 min. Brain regional rates of glucose use and metabolite levels were determined. DCA-treated rats had increased rates of glucose use in all regions studied (cortex, thalamus, striatum, and brain stem), with an average increase of 41%. Lactate levels were lower in all regions, by an average of 35%. There were no significant changes in levels of ATP, creatine phosphate, or glycogen in any brain region. Blood levels of lactate did not differ significantly between the DCA- and the saline-treated groups. Blood glucose levels were higher in the DCA group. In rats sacrificed by freeze-blowing, DCA treatment caused lower brain levels of both lactate and pyruvate. These results cannot be explained by any systemic effect of DCA. Rather, it appears that in the immature rat, DCA treatment results in activation of brain PDH, increased metabolism of brain pyruvate and lactate, and a resulting increase in brain glycolytic rate.

Miller, A.L.; Hatch, J.P.; Prihoda, T.J. (Univ. of Texas Health Science Center, San Antonio (USA))

1990-12-01

299

Lactate formation in Caldicellulosiruptor saccharolyticus is regulated by the energy carriers pyrophosphate and ATP.  

PubMed

Caldicellulosiruptor saccharolyticus displays superior H(2) yields on a wide range of carbon sources provided that lactate formation is avoided. Nevertheless, a low lactate flux is initiated as the growth rate declined in the transition to the stationary phase, which coincides with a drastic decrease in the glucose consumption and acetate production fluxes. In addition, the decrease in growth rate was accompanied by a sudden increase and then decrease in NADH levels. The V'(MAX) of the lactate dehydrogenase (LDH) doubled when the cells entered the stationary phase. Kinetic analysis revealed that at the metabolic level LDH activity is regulated through (i) competitive inhibition by pyrophosphate (PPi, k(i)=1.7 mM) and NAD (k(i)=0.43 mM) and (ii) allosteric activation by FBP (300%), ATP (160%) and ADP (140%). From these data a MWC-based model was derived. Simulations with this model could explain the observed lactate shift by displaying how the sensitivity of LDH activity to NADH/NAD ratio varied with different PP(i) concentrations. Moreover, the activation of LDH by ATP indicates that C. saccharolyticus uses LDH as a means to adjusts its flux of ATP and NADH production. To our knowledge, this is the first time PPi is observed as an effector of LDH. PMID:20060925

Willquist, Karin; van Niel, Ed W J

2010-05-01

300

Anilides of (R)-trifluoro-2-hydroxy-2-methylpropionic acid as inhibitors of pyruvate dehydrogenase kinase.  

PubMed

The optimization of a series of anilide derivatives of (R)-3,3, 3-trifluoro-2-hydroxy-2-methylpropionic acid as inhibitors of pyruvate dehydrogenase kinase (PDHK) is described that started from N-phenyl-3,3,3-trifluoro-2-hydroxy-2-methylpropanamide 1 (IC(50) = 35 +/- 1.4 microM). It was found that small electron-withdrawing groups on the ortho position of the anilide, i.e., chloro, acetyl, or bromo, increased potency 20-40-fold. The oral bioavailability of the compounds in this series is optimal (as measured by AUC) when the anilide is substituted at the 4-position with an electron-withdrawing group (i.e., carboxyl, carboxyamide, and sulfoxyamide). N-(2-Chloro-4-isobutylsulfamoylphenyl)-(R)-3,3, 3-trifluoro-2-hydroxy-2-methylpropionamide (10a) inhibits PDHK in the primary enzymatic assay with an IC(50) of 13 +/- 1.5 nM, enhances the oxidation of [(14)C]lactate into (14)CO(2) in human fibroblasts, lowers blood lactate levels significantly 2.5 and 5 h after oral doses as low as 30 micromol/kg, and increases the ex vivo activity of PDH in muscle, kidney, liver, and heart tissues. However, in contrast to sodium dichloroacetate (DCA), these PDHK inhibitors did not lower blood glucose levels. Nevertheless, they are effective at increasing the utilization and disposal of lactate and could be of utility to ameliorate conditions of inappropriate blood lactate elevation. PMID:10841803

Bebernitz, G R; Aicher, T D; Stanton, J L; Gao, J; Shetty, S S; Knorr, D C; Strohschein, R J; Tan, J; Brand, L J; Liu, C; Wang, W H; Vinluan, C C; Kaplan, E L; Dragland, C J; DelGrande, D; Islam, A; Lozito, R J; Liu, X; Maniara, W M; Mann, W R

2000-06-01

301

Tin-containing Silicates: Alkali Salts Improve Methyl Lactate Yield from Sugars.  

PubMed

This study focuses on increasing the selectivity to methyl lactate from sugars using stannosilicates as heterogeneous catalyst. All group?I ions are found to have a promoting effect on the resulting methyl lactate yield. Besides, the alkali ions can be added both during the preparation of the catalyst or directly to the solvent mixture to achieve the highest reported yield of methyl lactate (ca. 75?%) from sucrose at 170?°C in methanol. The beneficial effect of adding alkali to the reaction media applies not only to highly defect-free Sn-Beta prepared through the fluoride route, but also to materials prepared by post-treatment of dealuminated commercial Beta zeolites, as well as ordered mesoporous stannosilicates, in this case Sn-MCM-41 and Sn-SBA-15. These findings open the door to the possibility of using other preparation methods or different Sn-containing silicates with equally high methyl lactate yields as Sn-Beta. PMID:25605624

Tolborg, Søren; Sádaba, Irantzu; Osmundsen, Christian M; Fristrup, Peter; Holm, Martin S; Taarning, Esben

2015-02-01

302

Dehydrogenase activity measurement in yeast fermentation  

Microsoft Academic Search

The dehydrogenase activity was used as a measure of active biomass in preference to other biochemical parameters because of\\u000a the simple, but accurate nature of the dehydrogenase test. After a consierable amount of experimental work on the dehydrogenase\\u000a activity measurement technique and the consideration of utilization of the technique as a measure of the active biomass in\\u000a yeast fermentation systems,

A. E. Ghaly; R. M. Ben-Hassan

1993-01-01

303

Bacterial 2,3-butanediol dehydrogenases  

Microsoft Academic Search

Enterobacter aerogenes, Aeromonas hydrophila, Serratia marcescens and Staphylococcus aureus possessing L(+)-butanediol dehydrogenase produced mainly meso-butanediol and small amounts of optically active butanediol; Acetobacter suboxydans, Bacillus polymyxa and Erwinia carotovora containing D(-)-butanediol dehydrogenase produced more optically active butanediol than meso-butanediol. Resting and growing cells of these organisms oxidized only one enantiomer of racemic butanediol. The D(-)-butanediol dehydrogenase from Bacillus polymyxa was

Hanni Höhn-Bentz; F. Radler

1978-01-01

304

Lactate infusions in patients with bulimia.  

PubMed

We performed lactate infusions in 18 bulimic patients and 11 normal controls. On the basis of blind ratings, bulimic patients appeared to react to the infusion with greater anxiety than controls. The frequency of lactate-induced panic, per se, was lower in bulimic patients than rates reported for panic disorder patients. However, it would be premature to conclude that bulimia is not a heterogeneous syndrome which includes a group of patients who panic with lactate. PMID:3222393

Lindy, D C; Walsh, B T; Gorman, J M; Roose, S P; Gladis, M; Devlin, M J; Glassman, A H

1988-12-01

305

Flavocytochrome b2-Based Enzymatic Method of L-Lactate Assay in Food Products  

PubMed Central

L-lactate, a key metabolite of the anaerobic glycolytic pathway, plays an important role as a biomarker in medicine, in the nutritional sector and food quality control. For these reasons, there is a need for very specific, sensitive, and simple analytical methods for the accurate L-lactate measuring. A new highly selective enzymatic method for L-lactate determination based on the use of flavocytochrome b2 (EC 1.1.2.3; FC b2) isolated from the recombinant strain of the yeast Hansenula polymorpha has been developed. A proposed enzymatic method exploits an enzymatic oxidation of L-lactate to pyruvate coupled with nitrotetrazolium blue (NTZB) reduction to a colored product, formazan. The maximal absorption peak of the colored product is near ? = 525?nm and the linear range is observed in the interval 0.005–0.14?mM of L-lactate. The main advantages of the proposed method when compared to the LDH-based routine approaches are a higher sensitivity (2.0??M of L-lactate), simple procedure of analysis, usage of inexpensive, nontoxic reagents, and small amount of the enzyme. Enzymatic oxidation of L-lactate catalyzed by flavocytochrome b2 and coupled with formazan production from nitrotetrazolium blue was shown to be used for L-lactate assay in food samples. A high correlation between results of the proposed method and reference ones proves the possibility to use flavocytochrome b2-catalysed reaction for enzymatic measurement of L-lactate in biotechnology and food chemistry. PMID:24223505

Smutok, Halyna

2013-01-01

306

The Crystal Structure of a Ternary Complex of Betaine Aldehyde Dehydrogenase from Pseudomonas aeruginosa Provides New Insight Into the Reaction Mechansim and Shows A Novel Binding Mode of the 2'-Phosphate of NADP+ and A Novel Cation Binding Site  

SciTech Connect

In the human pathogen Pseudomonas aeruginosa, the NAD(P)+-dependent betaine aldehyde dehydrogenase (PaBADH) may play the dual role of assimilating carbon and nitrogen from choline or choline precursors-abundant at infection sites-and producing glycine betaine and NADPH, potentially protective against the high-osmolarity and oxidative stresses prevalent in the infected tissues. Disruption of the PaBADH gene negatively affects the growth of bacteria, suggesting that this enzyme could be a target for antibiotic design. PaBADH is one of the few ALDHs that efficiently use NADP+ and one of the even fewer that require K+ ions for stability. Crystals of PaBADH were obtained under aerobic conditions in the presence of 2-mercaptoethanol, glycerol, NADP+ and K+ ions. The three-dimensional structure was determined at 2.1-A resolution. The catalytic cysteine (C286, corresponding to C302 of ALDH2) is oxidized to sulfenic acid or forms a mixed disulfide with 2-mercaptoethanol. The glutamyl residue involved in the deacylation step (E252, corresponding to E268 of ALDH2) is in two conformations, suggesting a proton relay system formed by two well-conserved residues (E464 and K162, corresponding to E476 and K178, respectively, of ALDH2) that connects E252 with the bulk water. In some active sites, a bound glycerol molecule mimics the thiohemiacetal intermediate; its hydroxyl oxygen is hydrogen bonded to the nitrogen of the amide groups of the side chain of the conserved N153 (N169 of ALDH2) and those of the main chain of C286, which form the 'oxyanion hole.' The nicotinamide moiety of the nucleotide is not observed in the crystal, and the adenine moiety binds in the usual way. A salt bridge between E179 (E195 of ALDH2) and R40 (E53 of ALDH2) moves the carboxylate group of the former away from the 2?-phosphate of the NADP+, thus avoiding steric clashes and/or electrostatic repulsion between the two groups. Finally, the crystal shows two K+ binding sites per subunit. One is in an intrasubunit cavity that we found to be present in all known ALDH structures. The othersingle bondnot described before for any ALDH but most likely present in most of themsingle bondis located in between the dimeric unit, helping structure a region involved in coenzyme binding and catalysis. This may explain the effects of K+ ions on the activity and stability of PaBADH.

Gonzalez-Segura, L.; Rudino-Pinera, E; Munoz-Clares, R; Horjales, E

2009-01-01

307

Mechanisms of generation of oxygen radicals and reductive mobilization of ferritin iron by lipoamide dehydrogenase.  

PubMed

The oxidase reaction of lipoamide dehydrogenase with NADH generates superoxide radicals and hydrogen peroxide under aerobic conditions. ESR spin trapping using 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) was applied to characterize the oxygen radical species generated by lipoamide dehydrogenase and the mechanism of their generation. During the oxidase reaction of lipoamide dehydrogenase, DMPO-OOH and DMPO-OH signals were observed. The DMPO-OOH signal disappeared on addition of superoxide dismutase. These results demonstrate that the DMPO-OOH adduct was produced from the superoxide radical generated by lipoamide dehydrogenase. In the presence of dimethyl sulfoxide, a DMPO-CH3 signal appeared at the expense of the DMPO-OH signal, indicating that the DMPO-OH adduct was produced directly from the hydroxyl radical rather than by decomposition of the DMPO-OOH adduct. The DMPO-OH signal decreased on addition of superoxide dismutase, catalase, or diethylenetriaminepentaacetic acid, indicating that the hydroxyl radical was generated via the metal-catalyzed Haber-Weiss reaction from the superoxide radical and hydrogen peroxide. Addition of ferritin to the NADH-lipoamide dehydrogenase system resulted in a decrease of the DMPO-OOH signal, indicating that the superoxide radical interacted with ferritin iron. PMID:1652585

Bando, Y; Aki, K

1991-03-01

308

Amperometric lactate oxidase catheter for real-time lactate monitoring based on thin film technology  

Microsoft Academic Search

An amperometric lactate oxidase catheter has been developed for in vivo application to real-time lactate monitoring. The electrochemical behaviour of the 1 × 3 mm Pt-Ag\\/AgCl thin film electrode is not significantly influenced by lactate oxidase-polyurethane covering. Gamma-irradiation (25 kGy) is suitable for the sterilization procedure. The final lactate catheter is characterized by a linear concentration range between 0·5 and

Sylvio Fischer

1997-01-01

309

Field energetics of free-living, lactating and non-lactating echidnas ( Tachyglossus aculeatus)  

Microsoft Academic Search

We measured daily energy expenditure (DEE) and water turnover rates in lactating and non-lactating short beaked echidnas (Tachyglossus aculeatus) using the doubly labelled water technique during the lactation period in spring. Reproductively inactive echidnas were on average significantly heavier (median: 3354 g; range: 2929–3780 g; N=4) than lactating females (median: 2695 g; range: 2690–2715 g; N=3) during the equivalent time

Jutta Schmid; Niels A. Andersen; John R. Speakman; Stewart C. Nicol

2003-01-01

310

[Pharmaceutical treatment of lactation deficiency lacks evidence.  

PubMed

In this review we have looked at the evidence for the pharmacological treatment of lactation deficiency. Five RCTs (n = 166) of metoclopramide found no effect on lactation and two RCTs (n = 26) of older date and lesser quality found significant effect. One RCT (n = 51) of syntocinon found no effect on lactation and two older RCTs (n = 60) of lesser quality found significant effect. Three RCTs (n = 105) found significant effect of domperidone on lactation. Education on breastfeeding is important to avoid the need for pharmacological treatment. PMID:25350407

Axelsson, Paul Bryde; Bjerrum, Flemming; Løkkegaard, Ellen Christine Leth

2014-02-24

311

Reconstruction of an Acetogenic 2,3-Butanediol Pathway Involving a Novel NADPH-Dependent Primary-Secondary Alcohol Dehydrogenase  

PubMed Central

Acetogenic bacteria use CO and/or CO2 plus H2 as their sole carbon and energy sources. Fermentation processes with these organisms hold promise for producing chemicals and biofuels from abundant waste gas feedstocks while simultaneously reducing industrial greenhouse gas emissions. The acetogen Clostridium autoethanogenum is known to synthesize the pyruvate-derived metabolites lactate and 2,3-butanediol during gas fermentation. Industrially, 2,3-butanediol is valuable for chemical production. Here we identify and characterize the C. autoethanogenum enzymes for lactate and 2,3-butanediol biosynthesis. The putative C. autoethanogenum lactate dehydrogenase was active when expressed in Escherichia coli. The 2,3-butanediol pathway was reconstituted in E. coli by cloning and expressing the candidate genes for acetolactate synthase, acetolactate decarboxylase, and 2,3-butanediol dehydrogenase. Under anaerobic conditions, the resulting E. coli strain produced 1.1 ± 0.2 mM 2R,3R-butanediol (23 ?M h?1 optical density unit?1), which is comparable to the level produced by C. autoethanogenum during growth on CO-containing waste gases. In addition to the 2,3-butanediol dehydrogenase, we identified a strictly NADPH-dependent primary-secondary alcohol dehydrogenase (CaADH) that could reduce acetoin to 2,3-butanediol. Detailed kinetic analysis revealed that CaADH accepts a range of 2-, 3-, and 4-carbon substrates, including the nonphysiological ketones acetone and butanone. The high activity of CaADH toward acetone led us to predict, and confirm experimentally, that C. autoethanogenum can act as a whole-cell biocatalyst for converting exogenous acetone to isopropanol. Together, our results functionally validate the 2,3-butanediol pathway from C. autoethanogenum, identify CaADH as a target for further engineering, and demonstrate the potential of C. autoethanogenum as a platform for sustainable chemical production. PMID:24657865

Köpke, Michael; Gerth, Monica L.; Maddock, Danielle J.; Mueller, Alexander P.; Liew, FungMin

2014-01-01

312

Reconstruction of an acetogenic 2,3-butanediol pathway involving a novel NADPH-dependent primary-secondary alcohol dehydrogenase.  

PubMed

Acetogenic bacteria use CO and/or CO2 plus H2 as their sole carbon and energy sources. Fermentation processes with these organisms hold promise for producing chemicals and biofuels from abundant waste gas feedstocks while simultaneously reducing industrial greenhouse gas emissions. The acetogen Clostridium autoethanogenum is known to synthesize the pyruvate-derived metabolites lactate and 2,3-butanediol during gas fermentation. Industrially, 2,3-butanediol is valuable for chemical production. Here we identify and characterize the C. autoethanogenum enzymes for lactate and 2,3-butanediol biosynthesis. The putative C. autoethanogenum lactate dehydrogenase was active when expressed in Escherichia coli. The 2,3-butanediol pathway was reconstituted in E. coli by cloning and expressing the candidate genes for acetolactate synthase, acetolactate decarboxylase, and 2,3-butanediol dehydrogenase. Under anaerobic conditions, the resulting E. coli strain produced 1.1 ± 0.2 mM 2R,3R-butanediol (23 ?M h(-1) optical density unit(-1)), which is comparable to the level produced by C. autoethanogenum during growth on CO-containing waste gases. In addition to the 2,3-butanediol dehydrogenase, we identified a strictly NADPH-dependent primary-secondary alcohol dehydrogenase (CaADH) that could reduce acetoin to 2,3-butanediol. Detailed kinetic analysis revealed that CaADH accepts a range of 2-, 3-, and 4-carbon substrates, including the nonphysiological ketones acetone and butanone. The high activity of CaADH toward acetone led us to predict, and confirm experimentally, that C. autoethanogenum can act as a whole-cell biocatalyst for converting exogenous acetone to isopropanol. Together, our results functionally validate the 2,3-butanediol pathway from C. autoethanogenum, identify CaADH as a target for further engineering, and demonstrate the potential of C. autoethanogenum as a platform for sustainable chemical production. PMID:24657865

Köpke, Michael; Gerth, Monica L; Maddock, Danielle J; Mueller, Alexander P; Liew, FungMin; Simpson, Séan D; Patrick, Wayne M

2014-06-01

313

GDP-fucose synthetase from Escherichia coli: structure of a unique member of the short-chain dehydrogenase\\/reductase family that catalyzes two distinct reactions at the same active site  

Microsoft Academic Search

Background: In all species examined, GDP-fucose is synthesized from GDP-mannose in a three-step reaction catalyzed by two enzymes, GDP-mannose 4,6 dehydratase and a dual function 3,5-epimerase-4-reductase named GDP-fucose synthetase. In this latter aspect fucose biosynthesis differs from that of other deoxy and dideoxy sugars, in which the epimerase and reductase activities are present as separate enzymes. Defects in GDP-fucose biosynthesis

William S Somers; Mark L Stahl; Francis X Sullivan

1998-01-01

314

Improved Production of 2,3-Butanediol in Bacillus amyloliquefaciens by Over-Expression of Glyceraldehyde-3-Phosphate Dehydrogenase and 2,3-butanediol Dehydrogenase  

PubMed Central

Background Previously, a safe strain, Bacillus amyloliquefaciens B10-127 was identified as an excellent candidate for industrial-scale microbial fermentation of 2,3-butanediol (2,3-BD). However, B. amyloliquefaciens fermentation yields large quantities of acetoin, lactate and succinate as by-products, and the 2,3-BD yield remains prohibitively low for commercial production. Methodology/Principal Findings In the 2,3-butanediol metabolic pathway, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) catalyzes the conversion of 3-phosphate glyceraldehyde to 1,3-bisphosphoglycerate, with concomitant reduction of NAD+ to NADH. In the same pathway, 2,3-BD dehydrogenase (BDH) catalyzes the conversion of acetoin to 2,3-BD with concomitant oxidation of NADH to NAD+. In this study, to improve 2,3-BD production, we first over-produced NAD+-dependent GAPDH and NADH-dependent BDH in B. amyloliquefaciens. Excess GAPDH reduced the fermentation time, increased the 2,3-BD yield by 12.7%, and decreased the acetoin titer by 44.3%. However, the process also enhanced lactate and succinate production. Excess BDH increased the 2,3-BD yield by 16.6% while decreasing acetoin, lactate and succinate production, but prolonged the fermentation time. When BDH and GAPDH were co-overproduced in B. amyloliquefaciens, the fermentation time was reduced. Furthermore, in the NADH-dependent pathways, the molar yield of 2,3-BD was increased by 22.7%, while those of acetoin, lactate and succinate were reduced by 80.8%, 33.3% and 39.5%, relative to the parent strain. In fed-batch fermentations, the 2,3-BD concentration was maximized at 132.9 g/l after 45 h, with a productivity of 2.95 g/l·h. Conclusions/Significance Co-overexpression of bdh and gapA genes proved an effective method for enhancing 2,3-BD production and inhibiting the accumulation of unwanted by-products (acetoin, lactate and succinate). To our knowledge, we have attained the highest 2,3-BD fermentation yield thus far reported for safe microorganisms. PMID:24098433

Yang, Taowei; Rao, Zhiming; Zhang, Xian; Xu, Meijuan; Xu, Zhenghong; Yang, Shang-Tian

2013-01-01

315

Cellobiose dehydrogenase in cellulose degradation  

SciTech Connect

Cellobiose dehydrogenase is produced by a variety of fungi. Although it was already discovered during the 70`s, it`s role in cellulose and lignin degradation is yet ambiguous. The enzyme contains both heme and FAD as prosthetic groups, and seems to have a domain specifically designed to bind the enzyme to cellulose. It`s affinity to amorphous cellulose is higher than to crystalline cellulose. We will report on the binding behavior of the enzyme, its usefulness in elucidation of cellulose structures and also, possibilities for applications such as its use in measuring individual and synergistic mechanisms for cellulose degradation by endo- and exo-glucanases.

Eriksson, L. [Univ. of Georgia, Athens, GA (United States); Igarashi, Kiyohiko; Samejima, Masahiro [Univ. of Tokyo (Japan)

1996-10-01

316

Reducing lactate secretion by ldhA Deletion in L-glutamate- producing strain Corynebacterium glutamicum GDK-9  

PubMed Central

L-lactate is one of main byproducts excreted in to the fermentation medium. To improve L-glutamate production and reduce L-lactate accumulation, L-lactate dehydrogenase-encoding gene ldhA was knocked out from L-glutamate producing strain Corynebacterium glutamicum GDK-9, designated GDK-9?ldhA. GDK-9?ldhA produced approximately 10.1% more L-glutamate than the GDK-9, and yielded lower levels of such by-products as ?-ketoglutarate, L-lactate and L-alanine. Since dissolved oxygen (DO) is one of main factors affecting L-lactate formation during L-glutamate fermentation, we investigated the effect of ldhA deletion from GDK-9 under different DO conditions. Under both oxygen-deficient and high oxygen conditions, L-glutamate production by GDK-9?ldhA was not higher than that of the GDK-9. However, under micro-aerobic conditions, GDK-9?ldhA exhibited 11.61% higher L-glutamate and 58.50% lower L-alanine production than GDK-9. Taken together, it is demonstrated that deletion of ldhA can enhance L-glutamate production and lower the unwanted by-products concentration, especially under micro-aerobic conditions. PMID:25763057

Zhang, Dalong; Guan, Dan; Liang, Jingbo; Guo, Chunqian; Xie, Xixian; Zhang, Chenglin; Xu, Qingyang; Chen, Ning

2014-01-01

317

Structural Basis for "Flip-Flop" Action of Human Pyruvate Dehydrogenase  

NASA Technical Reports Server (NTRS)

The derivative of vitamin B1, thiamin pyrophosphate is a cofactor of pyruvate dehydrogenase, a component enzyme of the mitochondrial pyruvate dehydrogenase multienzyme complex that plays a major role in directing energy metabolism in the cell. This cofactor is used to cleave the C(sup alpha)-C(=O) bond of pyruvate followed by reductive acetyl transfer to lipoyl-dihydrolipoamide acetyltransferase. In alpha(sub 2)beta(sub 2)-tetrameric human pyruvate dehydrogenase, there are two cofactor binding sites, each of them being a center of independently conducted, although highly coordinated enzymatic reactions. The dynamic nonequivalence of two, otherwise chemically equivalent, catalytic sites can now be understood based on the recently determined crystal structure of the holo-form of human pyruvate dehydrogenase at 1.95A resolution. The structure of pyruvate dehydrogenase was determined using a combination of MAD phasing and molecular replacement followed by rounds of torsion-angles molecular-dynamics simulated-annealing refinement. The final pyruvate dehydrogenase structure included coordinates for all protein amino acids two cofactor molecules, two magnesium and two potassium ions, and 742 water molecules. The structure was refined to R = 0.202 and R(sub free) = 0.244. Our structural analysis of the enzyme folding and domain assembly identified a simple mechanism of this protein motion required for the conduct of catalytic action.

Ciszak, Ewa; Korotchkina, Lioubov; Dominiak, Paulina; Sidhu, Sukhdeep; Patel, Mulchand

2003-01-01

318

21 CFR 522.1698 - Pentazocine lactate injection.  

Code of Federal Regulations, 2012 CFR

... 2012-04-01 false Pentazocine lactate injection. 522.1698 Section 522...ANIMAL DRUGS § 522.1698 Pentazocine lactate injection. (a) Specifications...sterile aqueous solution contains pentazocine lactate equivalent to 30 milligrams of...

2012-04-01

319

21 CFR 522.1698 - Pentazocine lactate injection.  

Code of Federal Regulations, 2011 CFR

... 2011-04-01 false Pentazocine lactate injection. 522.1698 Section 522...ANIMAL DRUGS § 522.1698 Pentazocine lactate injection. (a) Specifications...sterile aqueous solution contains pentazocine lactate equivalent to 30 milligrams of...

2011-04-01

320

Empirical and Theoretical Constraints on the Evolution of Lactation  

E-print Network

Empirical and Theoretical Constraints on the Evolution of Lactation V. HAYSSEN Department, length of gestation, basal metabolism, and neona- tal development. The primary influence on lactation length is female mass, but phylogenetic constraints are important. Thus, lactation can be characterized

Hayssen, Virginia

321

21 CFR 522.1698 - Pentazocine lactate injection.  

Code of Federal Regulations, 2013 CFR

... 2013-04-01 false Pentazocine lactate injection. 522.1698 Section 522...ANIMAL DRUGS § 522.1698 Pentazocine lactate injection. (a) Specifications...sterile aqueous solution contains pentazocine lactate equivalent to 30 milligrams of...

2013-04-01

322

21 CFR 522.1698 - Pentazocine lactate injection.  

Code of Federal Regulations, 2010 CFR

... 2010-04-01 false Pentazocine lactate injection. 522.1698 Section 522...ANIMAL DRUGS § 522.1698 Pentazocine lactate injection. (a) Specifications...sterile aqueous solution contains pentazocine lactate equivalent to 30 milligrams of...

2010-04-01

323

Recovery of renal lactate dehydrogenase (LDH) isoenzyme pattern after obstruction relief in experimental hydronephrosis.  

PubMed

The release of ureteral occlusion leads to a progressive recovery in LDH isoenzyme pattern with gradual increase of anodic fractions and decrease of middle and cathodic ones. Our findings demonstrate that the recovery is accomplished on the 10-14th day, in agreement with morphological and metabolic observation. PMID:467572

Emanuelli, G; Anfossi, G; Camussi, G; Calcamuggi, G; Cestonaro, G; Gatti, G

1979-06-15

324

Lactate dehydrogenase isoenzyme patterns upon chronic exposure to cigarette smoke: Protective effect of bacoside A  

Microsoft Academic Search

Despite a strong association between cigarette smoking and alarming increase in mortality rate from smoking-related diseases, around 35–40% of the world's population continues to smoke and many more are being exposed to environmental tobacco smoke. Since the role of free radicals and oxidative damage in the pathogenesis of smoking-related diseases has been suggested, bacoside A, a potent antioxidant was tested

Kothandapani Anbarasi; Kuruvimalai Ekambaram Sabitha; Chennam Srinivasulu Shyamala Devi

2005-01-01

325

Affinity Chromatography of Lactate Dehydrogenase: An Experiment for the Undergraduate Biochemistry Laboratory.  

ERIC Educational Resources Information Center

Discusses a laboratory technique of enzyme purification by affinity chromatography as part of an undergraduate biochemical methodology course. Provides preparation details of the rat muscle homogenate and reagents. Proposes column requirements and assaying information. (MVL)

Anderson, Alexander J.

1988-01-01

326

Effect of a Marathon Run on Serum Lipoproteins, Creatine Kinase, and Lactate Dehydrogenase in Recreational Runners  

ERIC Educational Resources Information Center

The objective of this study was to determine the effect of a marathon run on serum lipid and lipoprotein concentrations and serum muscle enzyme activities and follow their recovery after the run. These blood concentrations were measured before, immediately after, and serially after a marathon run in 15 male recreational runners. The triglyceride…

Kobayashi, Yoshio; Takeuchi, Toshiko; Hosoi, Teruo; Yoshizaki, Hidekiyo; Loeppky, Jack A.

2005-01-01

327

Maintenance of Quaternary Structure in the Frozen State Stabilizes Lactate Dehydrogenase during Freeze–Drying  

Microsoft Academic Search

Sugars inhibit protein unfolding during the drying step of lyophilization by replacing hydrogen bonds to the protein lost upon removal of water. In many cases, polymers fail to inhibit dehydration-induced damage to proteins because steric hindrance prevents effective hydrogen bonding of the polymer to the protein's surface. However, in certain cases, polymers have been shown to stabilize multimeric enzymes during

Thomas J. Anchordoquy; Ken-Ichi Izutsu; Theodore W. Randolph; John F. Carpenter

2001-01-01

328

Activities of malate dehydrogenase, 3-hydroxyacyl-CoA dehydrogenase and fructose-1,6-diphosphatase with regard to metabolic subpopulations of fast- and slow-twitch fibres in rabbit muscles  

Microsoft Academic Search

Activities of malate dehydrogenase (MDH), 3-hydroxyacyl-CoA dehydrogenase (HAD) and fructose-1,6-diphosphatase (FDPase) were determined in single fibres dissected from freeze-dried rabbit psoas and soleus muscles. Slow-twitch fibres as determined by qualitative ATPase reaction represent a rather uniform population with regard to HAD and MDH activities. In these fibres the two enzymes are in constant proportions. FDPase is found at extremely low

Cornelia Spamer; D. Pette

1979-01-01

329

THE HISTOCHEMICAL DEMONSTRATION OF GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE ACTIVITY  

PubMed Central

A histochemical method for demonstration of glyceraldehyde-3-phosphate dehydrogenation by tissues is described. The method utilizes Nitro BT as an indicator, glyceraldehyde-3-phosphate obtained from hydrolysis of commercially obtainable glyceraldehyde-3-phosphate diethylacetal (monobarium salt) as substrate, and (ethylenediamine)tetraacetic acid acid disodium as an activating agent in a medium buffered to pH 7.2 by 0.2 M sodium phosphate. The heat lability, substrate and coenzyme specificity, and sulfhydryl and phosphate dependence of the tissue component catalyzing this reaction indicate that glyceraldehyde-3-phosphate dehydrogenase activity is being demonstrated. The disparity between the known pH optimum of this enzyme and that determined histochemically, and the anomalous histochemical localization to mitochondria of this enzyme which has been found in the soluble fraction by differential centrifugation, are thought to result from the diaphorase dependence of the tetrazolium methods and to emphasize the need for caution in the interpretation of histochemically determined intracellular localization of dehydrogenating enzymes. The evidence gathered by previous workers concerning the feasibility of demonstrating specific dehydrogenases with Nitro BT, and the correspondence of the distribution of glyceraldehyde-3-phosphate dehydrogenase determined histochemically with available quantitative data, suggest that at the cellular level the histochemical results accurately reflect the distribution of this enzyme. PMID:13714413

Himmelhoch, S. Ralph; Karnovsky, Morris J.

1961-01-01

330

Hybridizability of gamma-irradiated lactic dehydrogenase  

Microsoft Academic Search

The hybridizabilities of the gamma-irradiated chicken heart and pig muscle lactic dehydrogenases were estimated by hybridizing the irradiated enzymes with the unirradiated pig heart lactic dehydrogenase. The disc gel electrophoretic patterns of the inter- and intraspecific hybrids showed that the LDH activity of the pig heart isozyme band increased as a function of dose. This observation was analyzed upon the

1976-01-01

331

Calcium deficiency, pregnancy, and lactation in rats  

Microsoft Academic Search

Summary The calcium homeostatic mechanism was challenged in adult female rats by feeding them a calcium-deficient diet containing oxalate, and by subjecting them to pregnancy and lactation. The regimen caused a substantial weight loss, especially in those animals which reared their young well. Severe hypocalcaemia was observed in the lactating rats. Serum-P was slightly elevated. The content of hydroxyproline in

P. Rasmussen

1977-01-01

332

Net Energy Value of Feeds for Lactation  

Microsoft Academic Search

Results of 543 energy balance trials with lactating cows were summarized to partition the energy required by lac- tating cows into maintenance and pro- duction components and to determine the irtfluence of energy source on the efficiency with which dietary energy is used for milk production. The-total energy requirement for lactating cows was expressed in net energy for milk production

P. W. Moe; W. P. Flatt; H. F. TYRRELL

1972-01-01

333

21 CFR 184.1768 - Sodium lactate.  

Code of Federal Regulations, 2011 CFR

...1768 Sodium lactate. (a) Sodium lactate (C3 H5 O3 Na , CAS Reg. No. 72-17-3) is the sodium salt of lactic acid. It is prepared...neutralization of lactic acid with sodium hydroxide. (b) The ingredient...

2011-04-01

334

21 CFR 184.1768 - Sodium lactate.  

Code of Federal Regulations, 2012 CFR

...1768 Sodium lactate. (a) Sodium lactate (C3 H5 O3 Na , CAS Reg. No. 72-17-3) is the sodium salt of lactic acid. It is prepared...neutralization of lactic acid with sodium hydroxide. (b) The ingredient...

2012-04-01

335

21 CFR 184.1768 - Sodium lactate.  

Code of Federal Regulations, 2014 CFR

...1768 Sodium lactate. (a) Sodium lactate (C3 H5 O3 Na , CAS Reg. No. 72-17-3) is the sodium salt of lactic acid. It is prepared...neutralization of lactic acid with sodium hydroxide. (b) The ingredient...

2014-04-01

336

21 CFR 184.1768 - Sodium lactate.  

Code of Federal Regulations, 2013 CFR

...1768 Sodium lactate. (a) Sodium lactate (C3 H5 O3 Na , CAS Reg. No. 72-17-3) is the sodium salt of lactic acid. It is prepared...neutralization of lactic acid with sodium hydroxide. (b) The ingredient...

2013-04-01

337

Is brain lactate increased in Huntington's disease?  

Microsoft Academic Search

Impaired brain energy metabolism with increased regional brain lactate may play a role in the pathogenesis of Huntington’s disease (HD). Magnetic resonance spectroscopy (MRS) has provided conflicting evidence, however, regarding metabolic changes. Our objective was to evaluate the potential contribution of CSF lactate to the changes observed with MRS in HD. We performed single voxel MRS at 3 T in 23

W. R. Wayne Martin; Marguerite Wieler; Christopher C. Hanstock

2007-01-01

338

Identification and expression of a cDNA from Daucus carota encoding a bifunctional aspartokinase-homoserine dehydrogenase  

Microsoft Academic Search

Aspartokinase (EC 2.7.2.4) and homoserine dehydrogenase (EC 1.1.1.3) catalyze steps in the pathway for the synthesis of lysine, threonine, and methionine from aspartate. Homoserine dehydrogenase was purified from carrot (Daucus carota L.) cell cultures and portions of it were subjected to amino acid sequencing. Oligonucleotides deduced from the amino acid sequences were used as primers in a polymerase chain reaction

Jane M. Weisemann; Benjamin F. Matthews

1993-01-01

339

A bioluminescence assay for aldehyde dehydrogenase activity.  

PubMed

The aldehyde dehydrogenase (ALDH) family of enzymes is critical for cell survival and adaptation to cellular and environmental stress. These enzymes are of interest as therapeutic targets and as biomarkers of stem cells. This article describes a novel, homogeneous bioluminescence assay to study the activity of the ALDH enzymes. The assay is based on a proluciferin-aldehyde substrate that is recognized and utilized by multiple ALDH enzyme isoforms to generate luciferin. A detection reagent is added to inactivate ALDH and generate light from the luciferin product. The luminescent signal is dependent on the ALDH enzyme concentration and the incubation time in the ALDH reaction; moreover, the luminescent signal generated with the detection reagent is stable for greater than 2 h. This assay provides many advantages over standard NADH fluorescence assays. It is more sensitive and the signal stability provided allows convenient assay setup in batch mode-based high-throughput screens. The assay also shows an accurate pharmacological response for a common ALDH inhibitor and is robust, with a large assay window (S/B=64) and Z'=0.75. PMID:23219557

Duellman, Sarah J; Valley, Michael P; Kotraiah, Vinayaka; Vidugiriene, Jolanta; Zhou, Wenhui; Bernad, Laurent; Osterman, Jean; Kimball, Joshua J; Meisenheimer, Poncho; Cali, James J

2013-03-15

340

Energy substrate metabolism in pyruvate dehydrogenase complex deficiency.  

PubMed

Pyruvate dehydrogenase (PDH) deficiency is an inherited disorder of carbohydrate metabolism, resulting in lactic acidosis and neurological dysfunction. In order to provide energy for the brain, a ketogenic diet has been tried. Both the disorder and the ketogenic therapy may influence energy production. The aim of the study was to assess hepatic glucose production, lipolysis and resting energy expenditure (REE) in an infant, given a ketogenic diet due to neonatal onset of the disease. Lipolysis and glucose production were determined for two consecutive time periods by constant-rate infusions of [1,1,2,3,3-²H?]-glycerol and [6,6-²H²]-glucose. The boy had been fasting for 2.5 h at the start of the sampling periods. REE was estimated by indirect calorimetry. Rates of glucose production and lipolysis were increased compared with those of term neonates. REE corresponded to 60% of normal values. Respiratory quotient (RQ) was increased, indicating a predominance of glucose oxidation. Blood lactate was within the normal range. Several mechanisms may underlie the increased rates of glucose production and lipolysis. A ketogenic diet will result in a low insulin secretion and reduced peripheral and hepatic insulin sensitivity, leading to increased production of glucose and decreased peripheral glucose uptake. Surprisingly, RQ was high, indicating active glucose oxidation, which may reflect a residual enzyme activity, sufficient during rest. Considering this, a strict ketogenic diet might not be the optimal choice for patients with PDH deficiency. We propose an individualised diet for this group of patients aiming at the highest glucose intake that each patient will tolerate without elevated lactate levels. PMID:24914713

Stenlid, Maria Halldin; Ahlsson, Fredrik; Forslund, Anders; von Döbeln, Ulrika; Gustafsson, Jan

2014-11-01

341

Orbital Contributions to CO Oxidation in Mo-Cu Carbon Monoxide Dehydrogenase  

PubMed Central

A molecular orbital analysis provides new insight into the mechanism of Mo/Cu carbon monoxide dehydrogenase, and reveals electronic structure contributions to reactivity that are remarkably similar to the structurally related molybdenum hydroxylases. A calculated reaction barrier of ~12 kcal/mol is in excellent agreement with experiment. PMID:24322538

Stein, Benjamin W.; Kirk, Martin L.

2014-01-01

342

Environmental pollutants and hydroxysteroid dehydrogenases.  

PubMed

Hydroxysteroid dehydrogenases (HSD) are a group of steroidogenic enzymes that are involved in the steroid biosynthesis and metabolism. Four classes of HSDs, namely, 3?-, 11?-, 17?-, and 20?-HSDs, are discussed. 3?-HSDs catalyze the conversion of pregnenolone, 17?-hydroxypregnenolone, and dehydroepiandrosterone to progesterone, 17?-hydroxyprogesterone, and androstenedione, respectively. 11?-HSDs catalyze the interconversion between active cortisol and inactive cortisone. 17?-HSDs catalyze the interconversion between 17?-hydroxyl steroids and 17-ketoandrogens and estrogens. 20?-HSDs catalyze the conversion of progesterone into 20?-hydroxyprogesterone. Many environmental pollutants directly inhibit one or more enzymes of these HSDs, thus interfering with endogenous active steroid hormone levels. These chemicals include industrial materials (perfluoroalkyl compounds, phthalates, bisphenol A, and benzophenone), pesticides/biocides (methoxychlor, organotins, 1,2-dibromo-3-chloropropane, and prochloraz), and plant constituents (genistein, gossypol, and licorice). This chapter reviews these inhibitors targeting on HSDs. PMID:24388197

Ye, Leping; Guo, Jingjing; Ge, Ren-Shan

2014-01-01

343

In vitro effects of bicarbonate and bicarbonate-lactate buffered peritoneal dialysis solutions on mesothelial and neutrophil function.  

PubMed

The inclusion of bicarbonate in the formulation of peritoneal dialysis solutions may avoid the in vitro impairment of certain cell functions seen with acidic lactate-based fluids. The supranormal physiological levels of HCO3- and PCO2 inherent in such formulations may, however, not be biocompatible. This study compared the in vitro biocompatibility of a pH 5.2 lactate-based formulation with formulations containing either 40 mM lactate at pH 7.4, 38 mM HCO3- at pH 6.8 (PCO2 at approximately 240 mm Hg) or 7.4 (PCO2 at approximately 60 mm Hg), and 25 mM HCO3- plus 15 mM lactate at pH 6.8 (PCO2 at approximately 160 mm Hg) or 7.4 (PCO2 at approximately 40 mm Hg). Significant release of lactate dehydrogenase or decreases in ATP content by human peritoneal mesothelial cells (HPMC) and human peripheral polymorphonuclear leukocytes (PMN) after a 30-min exposure to each test solution was only seen with the pH 5.2 lactate-based fluid. The ATP content of HPMC exposed to this fluid returned to control levels after 30 min of recovery in M199 control medium but showed a trend toward decreasing ATP content at 240 min. Similarly, interleukin (IL)-1 beta-induced IL-6 synthesis by HPMC was also only significantly reduced by the pH 5.2 lactate solution. PMN chemiluminescence was unaffected by 30-min exposure to all test solutions except for the pH 5.2 lactate formulation. Staphylococcus epidermidis phagocytosis was reduced to between 46 to 57% of control with all test solutions except the pH 5.2 lactate solution, which further suppressed the chemiluminescence response to 17% of control. These data suggest that short exposure to supranormal physiological levels of HCO3- and PCO2 does not impair HPMC or PMN viability and function. Furthermore, neutral pH lactate-containing solutions show equivalent biocompatibility to bicarbonate-based ones. PMID:8785390

Topley, N; Kaur, D; Petersen, M M; Jörres, A; Williams, J D; Faict, D; Holmes, C J

1996-02-01

344

Etiology and therapeutic approach to elevated lactate  

PubMed Central

Lactate levels are commonly evaluated in acutely ill patients. Although most commonly used in the context of evaluating shock, lactate can be elevated for many reasons. While tissue hypoperfusion is probably the most common cause of elevation, many other etiologies or contributing factors exist. Clinicians need to be aware of the many potential causes of lactate elevation as the clinical and prognostic importance of an elevated lactate varies widely by disease state. Moreover, specific therapy may need to be tailored to the underlying cause of elevation. The current review is based on a comprehensive PubMed search and contains an overview of the pathophysiology of lactate elevation followed by an in-depth look at the varied etiologies, including medication-related causes. The strengths and weaknesses of lactate as a diagnostic/prognostic tool and its potential use as a clinical endpoint of resuscitation will be discussed. The review ends with some general recommendations on management of patients with elevated lactate. PMID:24079682

Andersen, Lars W.; Mackenhauer, Julie; Roberts, Jonathan C.; Berg, Katherine M.; Cocchi, Michael N.; Donnino, Michael W.

2014-01-01

345

Pyruvate dehydrogenase complex activity controls metabolic and malignant phenotype in cancer cells.  

PubMed

High lactate generation and low glucose oxidation, despite normal oxygen conditions, are commonly seen in cancer cells and tumors. Historically known as the Warburg effect, this altered metabolic phenotype has long been correlated with malignant progression and poor clinical outcome. However, the mechanistic relationship between altered glucose metabolism and malignancy remains poorly understood. Here we show that inhibition of pyruvate dehydrogenase complex (PDC) activity contributes to the Warburg metabolic and malignant phenotype in human head and neck squamous cell carcinoma. PDC inhibition occurs via enhanced expression of pyruvate dehydrogenase kinase-1 (PDK-1), which results in inhibitory phosphorylation of the pyruvate dehydrogenase alpha (PDHalpha) subunit. We also demonstrate that PDC inhibition in cancer cells is associated with normoxic stabilization of the malignancy-promoting transcription factor hypoxia-inducible factor-1alpha (HIF-1alpha) by glycolytic metabolites. Knockdown of PDK-1 via short hairpin RNA lowers PDHalpha phosphorylation, restores PDC activity, reverts the Warburg metabolic phenotype, decreases normoxic HIF-1alpha expression, lowers hypoxic cell survival, decreases invasiveness, and inhibits tumor growth. PDK-1 is an HIF-1-regulated gene, and these data suggest that the buildup of glycolytic metabolites, resulting from high PDK-1 expression, may in turn promote HIF-1 activation, thus sustaining a feed-forward loop for malignant progression. In addition to providing anabolic support for cancer cells, altered fuel metabolism thus supports a malignant phenotype. Correction of metabolic abnormalities offers unique opportunities for cancer treatment and may potentially synergize with other cancer therapies. PMID:18541534

McFate, Thomas; Mohyeldin, Ahmed; Lu, Huasheng; Thakar, Jay; Henriques, Jeremy; Halim, Nader D; Wu, Hong; Schell, Michael J; Tsang, Tsz Mon; Teahan, Orla; Zhou, Shaoyu; Califano, Joseph A; Jeoung, Nam Ho; Harris, Robert A; Verma, Ajay

2008-08-15

346

The TyrA family of aromatic-pathway dehydrogenases in phylogenetic context  

Microsoft Academic Search

BACKGROUND: The TyrA protein family includes members that catalyze two dehydrogenase reactions in distinct pathways leading to L-tyrosine and a third reaction that is not part of tyrosine biosynthesis. Family members share a catalytic core region of about 30 kDa, where inhibitors operate competitively by acting as substrate mimics. This protein family typifies many that are challenging for bioinformatic analysis

Jian Song; Carol A Bonner; Murray Wolinsky; Roy A Jensen

2005-01-01

347

Lactate biosensors: current status and outlook.  

PubMed

Many research efforts over the last few decades have been devoted to sensing lactate as an important analytical target in clinical care, sport medicine, and food processing. Therefore, research in designing lactate sensors is no longer in its infancy and now is more directed toward viable sensors for direct applications. In this review, we provide an overview of the most immediate and relevant developments toward this end, and we discuss and assess common transduction approaches. Further, we critically describe the pros and cons of current commercial lactate sensors and envision how future sensing design may benefit from emerging new technologies. PMID:24037614

Rassaei, Liza; Olthuis, Wouter; Tsujimura, Seiya; Sudhölter, Ernst J R; van den Berg, Albert

2014-01-01

348

Cotton fabric-based electrochemical device for lactate measurement in saliva.  

PubMed

Lactate measurement is vital in clinical diagnostics especially among trauma and sepsis patients. In recent years, it has been shown that saliva samples are an excellent applicable alternative for non-invasive measurement of lactate. In this study, we describe a method for the determination of lactate concentration in saliva samples by using a simple and low-cost cotton fabric-based electrochemical device (FED). The device was fabricated using template method for patterning the electrodes and wax-patterning technique for creating the sample placement/reaction zone. Lactate oxidase (LOx) enzyme was immobilised at the reaction zone using a simple entrapment method. The LOx enzymatic reaction product, hydrogen peroxide (H2O2) was measured using chronoamperometric measurements at the optimal detection potential (-0.2 V vs. Ag/AgCl), in which the device exhibited a linear working range between 0.1 to 5 mM, sensitivity (slope) of 0.3169 ?A mM(-1) and detection limit of 0.3 mM. The low detection limit and wide linear range were suitable to measure salivary lactate (SL) concentration, thus saliva samples obtained under fasting conditions and after meals were evaluated using the FED. The measured SL varied among subjects and increased after meals randomly. The proposed device provides a suitable analytical alternative for rapid and non-invasive determination of lactate in saliva samples. The device can also be adapted to a variety of other assays that requires simplicity, low-cost, portability and flexibility. PMID:24776756

Malon, Radha S P; Chua, K Y; Wicaksono, Dedy H B; Córcoles, Emma P

2014-06-21

349

Metformin suppresses gluconeogenesis by inhibiting mitochondrial glycerophosphate dehydrogenase  

PubMed Central

Metformin is considered to be one of the most effective therapeutics for the treatment of type 2 diabetes (T2D) since it specifically reduces hepatic gluconeogenesis without increasing insulin secretion, inducing weight gain, or posing a risk of hypoglycemia1,2. For over half a century, this agent has been prescribed to T2D patients worldwide, yet the underlying mechanism by which metformin inhibits hepatic gluconeogenesis remains unknown. Here we show that metformin non-competitively inhibits the redox shuttle enzyme mitochondrial glycerophosphate dehydrogenase (mGPD), resulting in an altered hepatocellular redox state, reduced conversion of lactate and glycerol to glucose, and decreased hepatic gluconeogenesis. Acute and chronic low-dose metformin treatment effectively reduced endogenous glucose production (EGP), while increasing cytosolic redox and decreasing mitochondrial redox states. Antisense oligonucleotide (ASO) knockdown of hepatic mGPD in rats resulted in a phenotype akin to chronic metformin treatment, and abrogated metformin-mediated increases in cytosolic redox state, decrease in plasma glucose concentrations and inhibition of EGP. These findings were replicated in whole-body mGPD knockout mice. These results have significant implications for understanding the mechanism of metformin’s blood glucose lowering effects and provide a novel therapeutic target for T2D. PMID:24847880

Madiraju, Anila K.; Erion, Derek M.; Rahimi, Yasmeen; Zhang, Xian-Man; Braddock, Demetrios; Albright, Ronald A.; Prigaro, Brett J.; Wood, John L.; Bhanot, Sanjay; MacDonald, Michael J.; Jurczak, Michael; Camporez, Joao-Paulo; Lee, Hui-Young; Cline, Gary W.; Samuel, Varman T.; Kibbey, Richard G.; Shulman, Gerald I.

2014-01-01

350

Not only students can express alcohol dehydrogenase: goldfish can too!  

PubMed

This article describes a novel approach to study the metabolic regulation of the respiratory system in vertebrates that suits physiology lessons for undergraduate students. It consists of an experimental demonstration of the goldfish's (Carassius auratus) adaptations to anoxia. The goldfish is one of the few vertebrates showing strong enzymatic plasticity for the expression of alcohol dehydrogenase (ADH), which allows it to survive long periods of severe anoxia. Therefore, we propose two simple laboratory exercises in which students are first asked to characterize the distribution of ADH isozymes in the goldfish by performing cellulose acetate electrophoresis. The second part of this laboratory lesson is the determination of liver glycogen. To further student comprehension, an interspecies comparative component is integrated, in which the same subjects are studied in an anoxia-sensitive species, the brook charr (Salvelinus fontinalis). ADH in goldfish is restricted to skeletal muscles, where it catalyzes alcoholic fermentation, permitting ethanol excretion through the gills and therefore preventing lactate acidosis caused by sustained glycolysis during anoxia. Electrophoresis also reveals the occurrence of a liver isozyme in the brook charr, which ADH catalyzes in the opposite pathway, allowing the usual ethanol degradation. As for the liver glycogen assay, it shows largely superior content in the goldfish liver compared with the brook charr, providing goldfish with a sustained energy supply during anoxia. The results of this laboratory exercise clearly demonstrate several physiological strategies developed by goldfish to cope with such a crucial environmental challenge as oxygen depletion. PMID:21098391

Chamberland, Valérie; Rioux, Pierre

2010-12-01

351

Not only students can express alcohol dehydrogenase: goldfish can too!  

NSDL National Science Digital Library

This article describes a novel approach to study the metabolic regulation of the respiratory system in vertebrates that suits physiology lessons for undergraduate students. It consists of an experimental demonstration of the goldfish's (Carassius auratus) adaptations to anoxia. The goldfish is one of the few vertebrates showing strong enzymatic plasticity for the expression of alcohol dehydrogenase (ADH), which allows it to survive long periods of severe anoxia. Therefore, we propose two simple laboratory exercises in which students are first asked to characterize the distribution of ADH isozymes in the goldfish by performing cellulose acetate electrophoresis. The second part of this laboratory lesson is the determination of liver glycogen. To further student comprehension, an interspecies comparative component is integrated, in which the same subjects are studied in an anoxia-sensitive species, the brook charr (Salvelinus fontinalis). ADH in goldfish is restricted to skeletal muscles, where it catalyzes alcoholic fermentation, permitting ethanol excretion through the gills and therefore preventing lactate acidosis caused by sustained glycolysis during anoxia. Electrophoresis also reveals the occurrence of a liver isozyme in the brook charr, which ADH catalyzes in the opposite pathway, allowing the usual ethanol degradation. As for the liver glycogen assay, it shows largely superior content in the goldfish liver compared with the brook charr, providing goldfish with a sustained energy supply during anoxia. The results of this laboratory exercise clearly demonstrate several physiological strategies developed by goldfish to cope with such a crucial environmental challenge as oxygen depletion.

Dr. Pierre Rioux (Universite du Quebec a Rimouski Dept de Biologie Chimie et des Sciences de la Sante)

2010-10-01

352

Original article Insulin and/or dexamethasone regulation of lactate  

E-print Network

Original article Insulin and/or dexamethasone regulation of lactate production and its relationship the hormonal regulation of lactate production and the lactate production/glucose utilization ratio in sheepM) were measured in perirenal AT of adult non- lactating non-pregnant cows (n = 5) or ewes (n = 5) given

Paris-Sud XI, Université de

353

Thiol-catalyzed formation of lactate and glycerate from glyceraldehyde. [significance in molecular evolution  

NASA Technical Reports Server (NTRS)

The rate of lactate formation from glyceraldehyde, catalyzed by N-acetyl-cysteine at ambient temperature in aqueous sodium phosphate (pH 7.0), is more rapid at higher sodium phosphate concentrations and remains essentially the same in the presence and absence of oxygen. The dramatic increase in the rate of glycerate formation that is brought about by this thiol, N-acetylcysteine, is accompanied by commensurate decreases in the rates of glycolate and formate production. It is suggested that the thiol-dependent formation of lactate and glycerate occurs by way of their respective thioesters. Attention is given to the significance of these reactions in the context of molecular evolution.

Weber, A. L.

1983-01-01

354

Alteration in substrate specificity of horse liver alcohol dehydrogenase by an acyclic nicotinamide analog of NAD(+).  

PubMed

A new, acyclic NAD-analog, acycloNAD(+) has been synthesized where the nicotinamide ribosyl moiety has been replaced by the nicotinamide (2-hydroxyethoxy)methyl moiety. The chemical properties of this analog are comparable to those of ?-NAD(+) with a redox potential of -324mV and a 341nm ?max for the reduced form. Both yeast alcohol dehydrogenase (YADH) and horse liver alcohol dehydrogenase (HLADH) catalyze the reduction of acycloNAD(+) by primary alcohols. With HLADH 1-butanol has the highest Vmax at 49% that of ?-NAD(+). The primary deuterium kinetic isotope effect is greater than 3 indicating a significant contribution to the rate limiting step from cleavage of the carbon-hydrogen bond. The stereochemistry of the hydride transfer in the oxidation of stereospecifically deuterium labeled n-butanol is identical to that for the reaction with ?-NAD(+). In contrast to the activity toward primary alcohols there is no detectable reduction of acycloNAD(+) by secondary alcohols with HLADH although these alcohols serve as competitive inhibitors. The net effect is that acycloNAD(+) has converted horse liver ADH from a broad spectrum alcohol dehydrogenase, capable of utilizing either primary or secondary alcohols, into an exclusively primary alcohol dehydrogenase. This is the first example of an NAD analog that alters the substrate specificity of a dehydrogenase and, like site-directed mutagenesis of proteins, establishes that modifications of the coenzyme distance from the active site can be used to alter enzyme function and substrate specificity. These and other results, including the activity with ?-NADH, clearly demonstrate the promiscuity of the binding interactions between dehydrogenases and the riboside phosphate of the nicotinamide moiety, thus greatly expanding the possibilities for the design of analogs and inhibitors of specific dehydrogenases. PMID:25280628

Malver, Olaf; Sebastian, Mina J; Oppenheimer, Norman J

2014-11-01

355

Isocitrate Dehydrogenase Parameters of Enzyme Activity  

NSDL National Science Digital Library

One of four biology laboratories where students research the properties of a model enzyme, isocitrate dehydrogenase, by using scientifitic method, molecular biology enzyme assay techniques and data analysis using a computer graphing program.

John H. Williamson (Davidson College; )

1999-01-01

356

Lactate shuttling and lactate use as fuel after traumatic brain injury: metabolic considerations.  

PubMed

Lactate is proposed to be generated by astrocytes during glutamatergic neurotransmission and shuttled to neurons as 'preferred' oxidative fuel. However, a large body of evidence demonstrates that metabolic changes during activation of living brain disprove essential components of the astrocyte-neuron lactate shuttle model. For example, some glutamate is oxidized to generate ATP after its uptake into astrocytes and neuronal glucose phosphorylation rises during activation and provides pyruvate for oxidation. Extension of the notion that lactate is a preferential fuel into the traumatic brain injury (TBI) field has important clinical implications, and the concept must, therefore, be carefully evaluated before implementation into patient care. Microdialysis studies in TBI patients demonstrate that lactate and pyruvate levels and lactate/pyruvate ratios, along with other data, have important diagnostic value to distinguish between ischemia and mitochondrial dysfunction. Results show that lactate release from human brain to blood predominates over its uptake after TBI, and strong evidence for lactate metabolism is lacking; mitochondrial dysfunction may inhibit lactate oxidation. Claims that exogenous lactate infusion is energetically beneficial for TBI patients are not based on metabolic assays and data are incorrectly interpreted. PMID:25204393

Dienel, Gerald A

2014-11-01

357

Inducing Lactation: Breastfeeding for Adoptive Moms  

MedlinePLUS

... for Adoptive Moms Family Life Listen Inducing Lactation: Breastfeeding for Adoptive Moms Article Body A growing number ... a breastfeeding relationship while further stimulating milk production. Nursing Supplement While there is no way to predict ...

358

Regulation of bone mineral loss during lactation  

NASA Technical Reports Server (NTRS)

The effects of varyng dietary calcium and phosphorous levels, vitamin D deficiency, oophorectomy, adrenalectomy, and simultaneous pregnancy on bone mineral loss during lactation in rats are studied. The experimental procedures and evaluations are described. The femur ash weight of lactating and nonlactating rats are calculated. The data reveals that a decrease in dietary calcium of 0.02 percent results in an increased loss of bone mineral, an increase in calcium to 1.4 percent does not lessen bone mineral loss, and bone mineral loss in vitamin D deficient rats is independent of calcium levels. It is observed that changes in dietary phosphorous level, oophorectomy, adrenalectomy, and simultaneous pragnancy do not reduce bone mineral loss during lactation. The analysis of various hormones to determine the mechanism that triggers bone mineral loss during lactation is presented.

Brommage, R.; Deluca, H. F.

1985-01-01

359

Glutamic Dehydrogenase of Mung Bean Mitochondria  

Microsoft Academic Search

GLUTAMIC dehydrogenase has been demonstrated in several higher plants1 and is associated with mitochondria of pea2 and oat3. Mitochondria isolated from mung bean (Phaseolus aureus) seedlings4 were found capable of oxidizing glutamic acid (90 &µl. oxygen\\/hr.\\/mgm. (nitrogen). The assay of glutamic dehydrogenase in intact mitochondria is limited by a permeability barrier to pyridine nucleotides. Reduced diphosphopyridine nucleotide was oxidized by

D. H. Bone

1959-01-01

360

IMP Dehydrogenase: Structural Schizophrenia and an Unusual Base  

SciTech Connect

Textbooks describe enzymes as relatively rigid templates for the transition state of a chemical reaction, and indeed an enzyme such as chymotrypsin, which catalyzes a relatively simple hydrolysis reaction, is reasonably well described by this model. Inosine monophosphate dehydrogenase (IMPDH) undergoes a remarkable array of conformational transitions in the course of a complicated catalytic cycle, offering a dramatic counterexample to this view. IMPDH displays several other unusual mechanistic features, including an Arg residue that may act as a general base catalyst and a dynamic monovalent cation site. Further, IMPDH appears to be involved in 'moon-lighting' functions that may require additional conformational states. How the balance between conformational states is maintained and how the various conformational states interconvert is only beginning to be understood.

Hedstrom,L.; Gan, L.

2006-01-01

361

Phosphorylation site on yeast pyruvate dehydrogenase complex  

SciTech Connect

The pyruvate dehydrogenase complex was purified to homogeneity from baker's yeast (Saccharomyces cerevisiae). Yeast cells were disrupted in a Manton-Gaulin laboratory homogenizer. The pyruvate dehydrogenase complex was purified by fractionation with polyethylene glycol, isoelectric precipitation, ultracentrifugation and chromatography on hydroxylapatite. Final purification of the yeast pyruvate dehydrogenase complex was achieved by cation-exchange high pressure liquid chromatography (HPLC). No endogenous pyruvate dehydrogenase kinase activity was detected during the purification. However, the yeast pyruvate dehydrogenase complex was phosphorylated and inactivated with purified pyruvate dehydrogenase kinase from bovine kidney. Tryptic digestion of the /sup 32/P-labeled complex yielded a single phosphopeptide which was purified to homogeniety. The tryptic digest was subjected to chromatography on a C-18 reverse phase HPLC column with a linear gradient of acetonitrile. Radioactive fractions were pooled, concentrated, and subjected to anion-exchange HPLC. The column was developed with a linear gradient of ammonium acetate. Final purification of the phosphopeptide was achieved by chromatography on a C-18 reverse phase HPLC column developed with a linear gradient of acetonitrile. The amino acid sequence of the homogeneous peptide was determined by manual modified Edman degradation.

Uhlinger, D.J.

1986-01-01

362

Structural and Kinetic Basis for Substrate Selectivity in Populus tremuloides Sinapyl Alcohol Dehydrogenase  

PubMed Central

We describe the three-dimensional structure of sinapyl alcohol dehydrogenase (SAD) from Populus tremuloides (aspen), a member of the NADP(H)-dependent dehydrogenase family that catalyzes the last reductive step in the formation of monolignols. The active site topology revealed by the crystal structure substantiates kinetic results indicating that SAD maintains highest specificity for the substrate sinapaldehyde. We also report substantial substrate inhibition kinetics for the SAD-catalyzed reduction of hydroxycinnamaldehydes. Although SAD and classical cinnamyl alcohol dehydrogenases (CADs) catalyze the same reaction and share some sequence identity, the active site topology of SAD is strikingly different from that predicted for classical CADs. Kinetic analyses of wild-type SAD and several active site mutants demonstrate the complexity of defining determinants of substrate specificity in these enzymes. These results, along with a phylogenetic analysis, support the inclusion of SAD in a plant alcohol dehydrogenase subfamily that includes cinnamaldehyde and benzaldehyde dehydrogenases. We used the SAD three-dimensional structure to model several of these SAD-like enzymes, and although their active site topologies largely mirror that of SAD, we describe a correlation between substrate specificity and amino acid substitution patterns in their active sites. The SAD structure thus provides a framework for understanding substrate specificity in this family of enzymes and for engineering new enzyme specificities. PMID:15829607

Bomati, Erin K.; Noel, Joseph P.

2005-01-01

363

Kinetics of lactate transport into rat liver in vivo  

SciTech Connect

Lactate clearance by liver plays an important role in lactate homeostasis and in the development of lactic acidosis. The role of lactate delivery to liver as a limiting factor in hepatic uptake of lactate is unclear. Lactate delivery of mechanisms could be important if rates of lactate transport approximate rates of lactate metabolism by liver. The rates of lactate transport into liver have been determined in vitro with isolated liver cells and the results have been conflicting. Therefore, the present studies measure the rate of transport of (14C)-L-lactate, and its poorly metabolizeable stereoisomer, (14C)-D-lactate, into rat liver in vivo using a portal vein injection technique. The transport of (3H)-water and of (14C)-sucrose, an extracellular reference compound, were also studied. Portal blood flow was determined from the kinetics of (3H)-water efflux in liver and was 1.93 +/- 0.22 mL/min/g. The volumes of distribution of (14C)-L-lactate, and (14C)-sucrose were 1.31 +/- 0.22, 0.71 +/- 0.07, and 0.22 +/- 0.07 mL/g, respectively. The extraction of unidirectional influx of (14C)-L-lactate and (14C)-D-lactate by rat liver was 93% +/- 10% and 91% +/- 9%, respectively. The rate of lactate transport into rat liver in vivo, 1.8 mumols.min-1.g-1, is approximately twofold greater than the rate of lactate metabolism by rat liver reported in the literature. Therefore, lactate uptake by liver may not be limited by transport under normal conditions. However, conditions such as decreased portal blood flow, which slow lactate delivery to liver by 50% or more, could cause lactate uptake by liver to be limited by transport of circulating lactate.

Lupo, M.A.; Cefalu, W.T.; Pardridge, W.M.

1990-04-01

364

PRODUCTIVE LIFE INCLUDING ALL LACTATIONS AND LONGER LACTATIONS WITH DIMINISHING CREDITS  

Technology Transfer Automated Retrieval System (TEKTRAN)

Alternative measures of productive life (PL) were compared and life expectancy factors were updated to replace estimates from 1993. Alternatives were proposed with extra credits for lactations longer than 10 mo and beyond 84 mo of age, and for each calf produced so that an extremely long lactation w...

365

Coniferyl aldehyde dimers in dehydrogenative polymerization: model of abnormal lignin formation in cinnamyl alcohol dehydrogenase-deficient plants  

Microsoft Academic Search

The enzymatically dehydrogenative polymerization of coniferyl aldehyde and coniferyl alcohol was studied to understand lignins\\u000a in cinnamyl alcohol dehydrogenase (CAD)-downregulated plants. The sample dimers were prepared by polymerization under three\\u000a reaction systems (coniferyl alcohol, coniferyl aldehyde, and their combination) with horseradish peroxidase\\/H2O2 under the conditions of limited reaction time. In addition, the residual amount of substrate in each reaction was

Takashi Ito; Reiko Hayase; Shingo Kawai; Hideo Ohashi; Takayoshi Higuchi

2002-01-01

366

Effect of major hepatectomy on glucose and lactate metabolism.  

PubMed Central

BACKGROUND: The liver plays an important role in glucose and lactate metabolism. Major hepatectomy may therefore be suspected to cause alterations of glucose and lactate homeostasis. METHODS: Thirteen subjects were studied: six patients after major hepatectomy and seven healthy subjects who had fasted overnight. Glucose turnover was measured with 6,6(2)H glucose. Lactate metabolism was assessed using two complementary approaches: 13C-glucose synthesis and 13CO2 production from an exogenous 13C-labeled lactate load infused over 15 minutes were measured, then the plasma lactate concentrations observed over 185 minutes after lactate load were fitted using a biexponential model to calculate lactate clearance, endogenous production, and half-lives. RESULTS: Three to five liver segments were excised. Compared to healthy controls, the following results were observed in the patients: 1) normal endogenous glucose production; 2) unchanged 13C-lactate oxidation and transformation into glucose; 3) similar basal plasma lactate concentration, lactate clearance, and lactate endogenous production; 4) decreased plasma lactate half-life 1 and increased half-life 2. CONCLUSIONS: Glucose and lactate metabolism are well maintained in patients after major hepatectomy, demonstrating a large liver functional reserve. Reduction in the size of normal liver parenchyma does not lead to hyperlactatemia. The use of a pharmacokinetic model, however, allows the detection of subtle alterations of lactate metabolism. PMID:10203083

Chioléro, R; Tappy, L; Gillet, M; Revelly, J P; Roth, H; Cayeux, C; Schneiter, P; Leverve, X

1999-01-01

367

Flexible graphene bio-nanosensor for lactate.  

PubMed

The development of a flexible nanosensor for detecting lactate could expand opportunities for using graphene, both in fundamental studies for a variety of device platforms and in practical applications. Graphene is a delicate single-layer, two-dimensional network of carbon atoms with ultrasensitive sensing capabilities. Lactic acid is important for clinical analysis, sports medicine, and the food industry. Recently, wearable and flexible bioelectronics on plastics have attracted great interest for healthcare, sports and defense applications due to their advantages of being light-weight, bendable, or stretchable. Here, we demonstrate for the first time the development of a flexible graphene-based bio-nanosensor to detect lactate. Our results show that flexible lactate biosensors can be fabricated on a variety of plastic substrates. The sensor can detect lactate sensitively from 0.08 ?M to 20 ?M with a fast steady-state measuring time of 2s. The sensor can also detect lactate under different mechanical bending conditions, the sensor response decreased as the bending angle and number of bending repetitions increased. We anticipate that these results could open exciting opportunities for fundamental studies of flexible graphene bioelectronics by using other bioreceptors, as well as a variety of wearable, implantable, real-time, or on-site applications in fields ranging from clinical analysis to defense. PMID:22954527

Labroo, Pratima; Cui, Yue

2013-03-15

368

High-performance liquid chromatography of proteins on deformed nonporous agarose beads. Affinity chromatography of dehydrogenases based on cibacron blue-derivatized agarose.  

PubMed

Nonporous agarose beads, prepared by shrinkage and cross-linking in organic solvents, were derivatized with Cibacron Blue F3G-A. A compressed bed of these beads was used for purification of dehydrogenases (glucose-6-phosphate dehydrogenase, lactate dehydrogenase and alcohol dehydrogenase). The chromatographic conditions for the purification of glucose-6-phosphate dehydrogenase were optimized by varying the pH of the buffer; the concentrations of eluting agents, i.e. NADP (specific elution) and sodium chloride (nonspecific elution); flow rate; residence time of the protein on the column bed; and protein load. Specific elution with NADP (2 mM in 0.025 M Tris-HCl, pH 8.0) gave the highest recovery (140%) and highest purification factor (200-fold) of the enzyme. The ability of the compressed bed of nonporous agarose beads to tolerate high flow rates was essential, since the recovery of the enzyme activity increased with an increase in flow rate. PMID:2235911

Li, J P; Eriksson, K O; Hjertén, S

1990-01-01

369

Conversion of oral glucose to lactate in dogs. Primary site and relative contribution to blood lactate  

SciTech Connect

The authors evaluated the relative contribution of oral glucose to arterial lactate and the relative role of the splanchnic bed in converting glucose to lactate in dogs. After an oral glucose load (1.2 g/kg) spiked with (U-14C)glucose (16.9 muCi/kg; protocol 1, n = 7), arterial blood lactate increased from 0.43 {plus minus} 0.03 mM at basal to a peak of 1.04 {plus minus} 0.07 mM at 45 min and then slowly decreased to 0.47 {plus minus} 0.07 mM at 240 min. Arterial blood {sup 14}Clactate peaked at 60 min and then decreased to {approximately} 35% of the peak at 4 h. When arterial blood lactate peaked at 45 min, the proportion of arterial lactate that was derived from oral glucose was 34 {plus minus} 3%. The integrated area under the curve of lactate derived from exogenous glucose was 40 {plus minus} 2% of that of total lactate. The splanchnic bed released lactate and {sup 14}Clactate during the initial 2 h after oral {sup 14}Cglucose. Thus, the splanchnic bed apparently contributed to the conversion of exogenous glucose to lactate. In the matched experiments (protocol 2, n = 5), dogs were given the same amount of oral glucose but no {sup 14}Cglucose, and (U-14C)lactate was infused into the right atrium to match the splanchnic {sup 14}Clactate release from the first experiment. Despite a well-matched splanchnic {sup 14}Clactate contribution, arterial concentrations of {sup 14}Clactate were markedly lower in protocol 2 compared with protocol 1. The integrated area under the {sup 14}Clactate profile in protocol 2 was only 11 {plus minus} 1% of that in protocol 1. These results indicate that the splanchnic bed is responsible for only 11% of arterial blood lactate that was derived from oral glucose. They concluded that (1) after oral glucose loading, a major portion of circulating lactate has its origin not in exogenous glucose but in endogenous sources, and (2) the splanchnic bed is not the major site of oral glucose conversion to lactate after glucose ingestion.

Youn, J.H.; Bergman, R.N. (Department of Physiology and Biophysics, University of Southern California Medical School, Los Angeles (USA))

1991-06-01

370

Inhibition of Horse Liver Alcohol Dehydrogenase by Methyltin Compounds  

PubMed Central

The study of inorganic tin (SnCl2, SnCl4) and methyltin compounds (MeSnCl3, Me2SnCI2, Me3SnCl) effects on the enzymatic activity of alcohol dehydrogenase (ADH) in the reaction of ethanol oxidation has been carried out. The experimental results of the study show that inorganic tin and methyltin substances induce slight inhibition of the catalytic activity of horse liver alcohol dehydrogenase (HLADH), unable to be improved during pre-incubation with the enzyme. The conditions for carrying out the kinetic investigation of the mentioned phenomenon were optimized and as it turned out the mechanism of methyltin trichloride action, as the most effective methyltin inhibitor, is more complex than the proposed interaction of the metal atom with SH-groups of the enzyme protein. It was demonstrated that the tin compounds act in the same manner as methylmercury compounds and might serve as oxidative agents towards the co-enzyme NADH. Kinetic data on MeSnCl3 were calculated. Data acquired on NAD-dependent ADH from horse liver and those regarding NAD-dependent LDH from sturgeon liver were compared. PMID:18365099

Bychkov, Pavel V.; Shekhovtsova, Tatyana N.; Milaeva, Elena R.

2005-01-01

371

Alcohol and aldehyde dehydrogenase polymorphisms in Chinese and Indian populations.  

PubMed

The association between two functional polymorphisms in alcohol dehydrogenase (ADH2/ADH1B) and aldehyde dehydrogenase (ALDH2) genes and alcohol dependence was examined in 182 Chinese and Indian patients undergoing treatment for alcohol dependence and 184 screened control subjects from Singapore. All subjects were screened by the Alcohol Use Disorders Identification Test (AUDIT). Patients were also administered the Severity of Alcohol Dependence Questionnaire (SADQ). Polymorphisms were genotyped by allele-specific polymerase chain reaction and selected genotypes confirmed by DNA sequencing or restriction fragment length polymorphism. Our results showed that frequencies of ADH1B*2 and ALDH2*2 were higher in controls compared to alcohol-dependent subjects for both Chinese and Indians. Frequencies of these two alleles were also higher in the 104 Chinese controls compared to the 80 Indian controls. None of the eight Chinese who were homozygous for both protective alleles was alcohol dependent. The higher frequencies of the protective alleles could explain the lower rate of alcohol dependence in Chinese. PMID:20025435

Tan, Ene-Choo; Lim, Leslie; Leong, Jern-Yi; Lim, Jing-Yan; Lee, Arthur; Yang, Jun; Tan, Chay-Hoon; Winslow, Munidasa

2010-01-01

372

The lactose synthetase particles of lactating bovine mammary gland. Preparation of particles with intact lactose synthetase.  

PubMed

1. The particulate form of lactating bovine mammary lactose synthetase activity is shown to be more highly organized than previously reported. 2. A novel method of shattering frozen mammary tissue with effective cell disruption is described. 3. The apparent subcellular distribution of lactose synthetase was shown to reflect the method of homogenization. 4. After mild homogenization particles associated with a high content of intact lactose synthetase activity sedimented in the lysosome size range between 5x10(4) and 3x10(5)g-min. 5. Lactose synthetase was dissociated and solubilized by VirTis homogenization and ultrasonic treatment. The activities and behaviour of UDP-galactose hydrolase, succinate dehydrogenase, beta-glucuronidase and phosphodiesterase I were compared. 6. Inhibition of UDP-galactose hydrolase by UTP and alpha-lactalbumin was observed. PMID:4300506

Coffey, R G; Reithel, F J

1968-09-01

373

NAD + -dependent Formate Dehydrogenase from Plants  

PubMed Central

NAD+-dependent formate dehydrogenase (FDH, EC 1.2.1.2) widely occurs in nature. FDH consists of two identical subunits and contains neither prosthetic groups nor metal ions. This type of FDH was found in different microorganisms (including pathogenic ones), such as bacteria, yeasts, fungi, and plants. As opposed to microbiological FDHs functioning in cytoplasm, plant FDHs localize in mitochondria. Formate dehydrogenase activity was first discovered as early as in 1921 in plant; however, until the past decade FDHs from plants had been considerably less studied than the enzymes from microorganisms. This review summarizes the recent results on studying the physiological role, properties, structure, and protein engineering of plant formate dehydrogenases. PMID:22649703

Alekseeva, A.A.; Savin, S.S.; Tishkov, V.I.

2011-01-01

374

Comparison of selected lactate threshold parameters with maximal lactate steady state in cycling.  

PubMed

The aim of the present investigation was to compare power at "onset of blood lactate accumulation" (OBLA), "individual anaerobic threshold" (IAT) and "+1.5?mmol???l(-1) lactate model" with power in maximal lactate steady state (MLSS) in cycling. However, there is a lack of studies concerning the absolute individual differences between different lactate parameters and MLSS.A total of 57 male participants performed several 30-min constant-load tests to determine MLSS by measuring blood lactate concentration (BLC). Depending on BLC, power was increased or decreased by 10?W in the following 30-min test. For detecting power at different threshold parameters, an incremental test was performed that began at 40?W and increased by 40?W every 4?min.Highly significant correlations were found between OBLA and MLSS: r=0.89 (mean difference -7.4?W); IAT and MLSS: r=0.83 (mean difference 12.4W), "+1.5?mmol???l(-1) lactate model" and MLSS: r=0.88 (mean difference -37.4W). On average, the parameters of OBLA and IAT approximate MLSS with no significant differences. The "+1.5?mmol???l(-1) lactate model" underestimates MLSS significantly.Based on Bland-and-Altman, the comparison of power of all threshold parameters with power in MLSS shows great individual differences despite the high regression coefficients and low mean differences between these methods. PMID:24227122

Hauser, T; Adam, J; Schulz, H

2014-06-01

375

Neural pathways underlying lactate-induced panic.  

PubMed

Panic disorder is a severe anxiety disorder characterized by susceptibility to induction of panic attacks by subthreshold interoceptive stimuli such as 0.5 M sodium lactate infusions. Although studied for four decades, the mechanism of lactate sensitivity in panic disorder has not been understood. The dorsomedial hypothalamus/perifornical region (DMH/PeF) coordinates rapid mobilization of behavioral, autonomic, respiratory and endocrine responses to stress, and rats with disrupted GABA inhibition in the DMH/PeF exhibit panic-like responses to lactate, similar to panic disorder patients. Utilizing a variety of anatomical and pharmacological methods, we provide evidence that lactate, via osmosensitive periventricular pathways, activates neurons in the compromised DMH/PeF, which relays this signal to forebrain limbic structures such as the bed nucleus of the stria terminalis to mediate anxiety responses, and specific brainstem sympathetic and parasympathetic pathways to mediate the respiratory and cardiovascular components of the panic-like response. Acutely restoring local GABAergic tone in the DMH/PeF blocked lactate-induced panic-like responses. Autonomic panic-like responses appear to be a result of DMH/PeF-mediated mobilization of sympathetic responses (verified with atenolol) and resetting of the parasympathetically mediated baroreflex. Based on our findings, DMH/PeF efferent targets such as the C1 adrenergic neurons, paraventricular hypothalamus, and the central amygdala are implicated in sympathetic mobilization; the nucleus of the solitary tract is implicated in baroreflex resetting; and the parabrachial nucleus is implicated in respiratory responses. These results elucidate neural circuits underlying lactate-induced panic-like responses and the involvement of both sympathetic and parasympathetic systems. PMID:18059441

Johnson, Philip L; Truitt, William A; Fitz, Stephanie D; Lowry, Christopher A; Shekhar, Anantha

2008-08-01

376

21 CFR 862.1670 - Sorbitol dehydrogenase test system.  

Code of Federal Regulations, 2010 CFR

...FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL...dehydrogenase test system is a device intended to measure the activity of the enzyme sorbitol dehydrogenase in serum. Measurements...

2010-04-01

377

Breast diseases during pregnancy and lactation  

PubMed Central

Breast is a typical female sexual physiologic organ that is influenced by steroid hormone from menarche until menopause. Therefore various diseases can be developed by continuous action of estrogen and progesterone. Breast diseases are mainly categorized as benign and malignant. It is very important to distinguish the malignancy from breast diseases. However, it is very difficult to diagnose malignancy in pregnant and lactating women even though the same breast diseases took place. Therefore, we will review breast diseases such as breast carcinoma during pregnancy and lactation. PMID:24327995

Yu, Ji Hoon; Kim, Min Jeong; Cho, Hyonil; Liu, Hyun Ju; Han, Sei-Jun

2013-01-01

378

D-Lactate altered mitochondrial energy production in rat brain and heart but not liver  

PubMed Central

Background Substantially elevated blood D-lactate (DLA) concentrations are associated with neurocardiac toxicity in humans and animals. The neurological symptoms are similar to inherited or acquired abnormalities of pyruvate metabolism. We hypothesized that DLA interferes with mitochondrial utilization of L-lactate and pyruvate in brain and heart. Methods Respiration rates in rat brain, heart and liver mitochondria were measured using DLA, LLA and pyruvate independently and in combination. Results In brain mitochondria, state 3 respiration was 53% and 75% lower with DLA as substrate when compared with LLA and pyruvate, respectively (p < 0.05). Similarly in heart mitochondria, state 3 respiration was 39% and 86% lower with DLA as substrate when compared with LLA or pyruvate, respectively (p < 0.05). However, state 3 respiration rates were similar between DLA, LLA and pyruvate in liver mitochondria. Combined incubation of DLA with LLA or pyruvate markedly impaired state 3 respiration rates in brain and heart mitochondria (p < 0.05) but not in liver mitochondria. DLA dehydrogenase activities were 61% and 51% lower in brain and heart mitochondria compared to liver, respectively, whereas LLA dehydrogenase activities were similar across all three tissues. An LDH inhibitor blocked state 3 respiration with LLA as substrate in all three tissues. A monocarboxylate transporter inhibitor blocked respiration with all three substrates. Conclusions DLA was a poor respiratory substrate in brain and heart mitochondria and inhibited LLA and pyruvate usage in these tissues. Further studies are warranted to evaluate whether these findings support, in part, the possible neurological and cardiac toxicity caused by high DLA levels. PMID:22296683

2012-01-01

379

Alpha-Glycerophosphate dehydrogenase (insoluble) and lactic dehydrogenase activities in the skeletal muscles of two insects.  

PubMed

The flight muscle preparations of the dragonfly Pantala flavescens and the aquatic beetle Cybister confusus showed extremely low levels of lactic dehydrogenase activity and high levels of alpha-glycerophosphate dehydrogenase (insoluble) activity. The activities of these two enzymes in the leg muscle of the beetle were approximately the same (1:1), but lactic dehydrogenase activity was several times higher than that in the flight muscles of both Insects. These results have been interpreted as indicating the high energy-yielding demands of the flight muscles during continuous sustained activity, while the leg muscles of the beetle which are involved in swimming activity derive their energy predominantly through anaerobic glycolysis. PMID:54055

Kallapur, V L; George, C J

1975-05-01

380

Drosophila melanogaster alcohol dehydrogenase: mechanism of aldehyde oxidation and dismutation.  

PubMed Central

Drosophila alcohol dehydrogenase (Adh) catalyses the oxidation of both alcohols and aldehydes. In the latter case, the oxidation is followed by a reduction of the aldehyde, i.e. a dismutation reaction. At high pH, dismutation is accompanied by a small release of NADH, which is not observed at neutral pH. Previously it has been emphasized that kinetic coefficients obtained by measuring the increase in A340, i.e. the release of NADH at high pH is not a direct measure of the aldehyde oxidation reaction and these values cannot be compared with those for alcohol dehydrogenation. In this article we demonstrate that this is not entirely true, and that the coefficients phiB and phiAB, where B is the aldehyde and A is NAD+, are the same for a dismutation reaction and a simple aldehyde dehydrogenase reaction. Thus the substrate specificity of the aldehyde oxidation reaction can be determined by simply measuring the NADH release. The coefficients for oxidation and dehydrogenation reactions (phi0d and phiAd respectively) are complex and involve the constants for the dismutation reaction. However, dead-end inhibitors can be used to determine the quantitative contribution of the kinetic constants for the aldehyde oxidation and reduction pathways to the phi0d and phiAd coefficients. The combination of dead-end and product inhibitors can be used to determine the reaction mechanism for the aldehyde oxidation pathway. Previously, we showed that with Drosophila Adh, the interconversion between alcohols and aldehydes followed a strictly compulsory ordered pathway, although aldehydes and ketones formed binary complexes with the enzyme. This raised the question regarding the reaction mechanism for the oxidation of aldehydes, i.e. whether a random ordered pathway was followed. In the present work, the mechanism for the oxidation of different aldehydes and the accompanying dismutation reaction with the slow alleloenzyme (AdhS) from Drosophila melanogaster has been studied. To obtain reliable results for the liberation of NADH during the initial-rate phase, the reaction was measured with a sensitive recording filter fluorimeter, and the complexes formed with the different dead-end and product inhibitors have been interpreted on the basis of a full dismutation reaction. The results are only consistent with a compulsory ordered reaction mechanism, with the formation of a dead-end binary enzyme-aldehyde complex. Under initial-velocity conditions, the rate of acetate release was calculated to be larger than 2.5 s-1, which is more than ten times that of NADH. The substrate specificity constant (kcat/Km or 1/phiB) with respect to the oxidation of substrates was propan-2-ol>ethanol>acetaldehyde>trimethylacetaldehyde. PMID:9445383

Winberg, J O; McKinley-McKee, J S

1998-01-01

381

Application of isotopes to the study of lactate metabolism  

SciTech Connect

The use of /sup 14/C as tracer to measure lactate turnover and oxidation and its role in gluconeogenesis are discussed. Lactate is formed as well as utilized in many cells, and most of it in the body is present within cells so that interpretation of /sup 14/C data from labelled lactate is more complex and more difficult than that of compounds present largely extracellularly, such as glucose. Apparent uptake of (/sup 14/C)lactate may occur in the absence of net lactate utilization, and /sup 14/CO/sub 2/ production does not provide a measure of true lactate oxidation. In vivo sites of tracer administration and sampling of blood are of critical significance for evaluation of lactate turnover, lactate space, its incorporation into glucose, and oxidation.

Katz, J.

1986-06-01

382

Enzymatic reaction paths as determined by transition path sampling  

NASA Astrophysics Data System (ADS)

Enzymes are biological catalysts capable of enhancing the rates of chemical reactions by many orders of magnitude as compared to solution chemistry. Since the catalytic power of enzymes routinely exceeds that of the best artificial catalysts available, there is much interest in understanding the complete nature of chemical barrier crossing in enzymatic reactions. Two specific questions pertaining to the source of enzymatic rate enhancements are investigated in this work. The first is the issue of how fast protein motions of an enzyme contribute to chemical barrier crossing. Our group has previously identified sub-picosecond protein motions, termed promoting vibrations (PVs), that dynamically modulate chemical transformation in several enzymes. In the case of human heart lactate dehydrogenase (hhLDH), prior studies have shown that a specific axis of residues undergoes a compressional fluctuation towards the active site, decreasing a hydride and a proton donor--acceptor distance on a sub-picosecond timescale to promote particle transfer. To more thoroughly understand the contribution of this dynamic motion to the enzymatic reaction coordinate of hhLDH, we conducted transition path sampling (TPS) using four versions of the enzymatic system: a wild type enzyme with natural isotopic abundance; a heavy enzyme where all the carbons, nitrogens, and non-exchangeable hydrogens were replaced with heavy isotopes; and two versions of the enzyme with mutations in the axis of PV residues. We generated four separate ensembles of reaction paths and analyzed each in terms of the reaction mechanism, time of barrier crossing, dynamics of the PV, and residues involved in the enzymatic reaction coordinate. We found that heavy isotopic substitution of hhLDH altered the sub-picosecond dynamics of the PV, changed the favored reaction mechanism, dramatically increased the time of barrier crossing, but did not have an effect on the specific residues involved in the PV. In the mutant systems, we observed changes in the reaction mechanism and altered contributions of the mutated residues to the enzymatic reaction coordinate, but we did not detect a substantial change in the time of barrier crossing. These results confirm the importance of maintaining the dynamics and structural scaffolding of the hhLDH PV in order to facilitate facile barrier passage. We also utilized TPS to investigate the possible role of fast protein dynamics in the enzymatic reaction coordinate of human dihydrofolate reductase (hsDHFR). We found that sub-picosecond dynamics of hsDHFR do contribute to the reaction coordinate, whereas this is not the case in the E. coli version of the enzyme. This result indicates a shift in the DHFR family to a more dynamic version of catalysis. The second inquiry we addressed in this thesis regarding enzymatic barrier passage concerns the variability of paths through reactive phase space for a given enzymatic reaction. We further investigated the hhLDH-catalyzed reaction using a high-perturbation TPS algorithm. Though we saw that alternate reaction paths were possible, the dominant reaction path we observed corresponded to that previously elucidated in prior hhLDH TPS studies. Since the additional reaction paths we observed were likely high-energy, these results indicate that only the dominant reaction path contributes significantly to the overall reaction rate. In conclusion, we show that the enzymes hhLDH and hsDHFR exhibit paths through reactive phase space where fast protein motions are involved in the enzymatic reaction coordinate and exhibit a non-negligible contribution to chemical barrier crossing.

Masterson, Jean Emily

383

Calculations of hydrogen tunnelling and enzyme catalysis: a comparison of liver alcohol dehydrogenase, methylamine dehydrogenase and soybean lipoxygenase  

NASA Astrophysics Data System (ADS)

Although the potential energy barrier for hydrogen transfer is similar for the enzymes liver alcohol dehydrogenase, methylamine dehydrogenase and soybean lipoxygenase, the degree of tunnelling is predicted to differ greatly, and is reflected by their primary kinetic isotope effects.

Tresadern, Gary; McNamara, Jonathan P.; Mohr, Matthias; Wang, Hong; Burton, Neil A.; Hillier, Ian H.

2002-06-01

384

Mechanistic enzymology of CO dehydrogenase from Clostridium thermoaceticum  

SciTech Connect

The final steps in acetyl-CoA biosynthesis by anaerobic bacteria are performed by carbon monoxide dehydrogenase (CODH), a nickel/iron-sulfur protein. An important achievement was to establish conditions under which acetyl-CoA synthesis by purified enzymes equals the in vivo rate of acetate synthesis. Under these optimized conditions we established that the rate limiting step in the synthesis of acetyl-CoA from methyl-H[sub 4]folate, CO and CoA is likely to be the methylation of CODH by the methylated corrinoid/iron-sulfur protein. We then focused on stopped flow studies of this rate limiting transmethylation reaction and established its mechanism. We have studied the carbonylation of CODH by infrared and resonance Raman spectroscopy and determined that the [Ni-Fe[sup 3-4]S[sub 4

Ragsdale, S.W.

1992-01-01

385

Fabricating polystyrene fiber-dehydrogenase assemble as a functional biocatalyst.  

PubMed

Immobilization of the enzymes on nano-structured materials is a promising approach to enhance enzyme stabilization, activation and reusability. This study aimed to develop polystyrene fiber-enzyme assembles to catalyze model formaldehyde to methanol dehydrogenation reaction, which is an essential step for bioconversion of CO2 to a renewable bioenergy. We fabricated and modified electrospun polystyrene fibers, which showed high capability to immobilize dehydrogenase for the fiber-enzyme assembles. Results from evaluation of biochemical activities of the fiber-enzyme assemble showed that nitriation with the nitric/sulfuric acid ratio (v/v, 10:1) and silanization treatment delivered desirable enzyme activity and long-term storage stability, showing great promising toward future large-scale applications. PMID:25435501

An, Hongjie; Jin, Bo; Dai, Sheng

2015-01-01

386

Structurally Integrated Photoluminescence-Based Lactate Sensor Using Organic Light Emitting Devices (OLEDs) as the Light Source  

SciTech Connect

Multianalyte bio(chemical) sensors are extensively researched for monitoring analytes in complex systems, such as blood serum. As a step towards developing such multianalyte sensors, we studied a novel, structurally integrated, organic light emitting device (OLED)-based sensing platform for detection of lactate. Lactate biosensors have attracted numerous research efforts, due to their wide applications in clinical diagnosis, athletic training and food industry. The OLED-based sensor is based on monitoring the oxidation reaction of lactate, which is catalyzed by the lactate oxidase (LOX) enzyme. The sensing component is based on an oxygen-sensitive dye, Platinum octaethyl porphyrin (PtOEP), whose photoluminescence (PL) lifetime {tau} decreases as the oxygen level increases. The PtOEP dye was embedded in a thin film polystyrene (PS) matrix; the LOX was dissolved in solution or immobilized in a sol-gel matrix. {tau} was measured as a function of the lactate concentration; as the lactate concentration increases, {tau} increases due to increased oxygen consumption. The sensors performance is discussed in terms of the detection sensitivity, dynamic range, and response time. A response time of {approx}32 sec was achieved when the LOX was dissolved in solution and kept in a closed cell. Steps towards development of a multianalyte sensor array using an array of individually addressable OLED pixels were also presented.

Chengliang Qian

2006-08-09

387

Identification of a Cytosolically Directed NADH Dehydrogenase in Mitochondria of Saccharomyces cerevisiae  

Microsoft Academic Search

The reoxidation of NADH generated in reactions within the mitochondrial matrix of Saccharomyces cerevisiae is catalyzed by an NADH dehydrogenase designated Ndi1p (C. A. M. Marres, S. de Vries, and L. A. Grivell, Eur. J. Biochem. 195:857-862, 1991). Gene disruption analysis was used to examine possible metabolic functions of two proteins encoded by open reading frames having significant primary sequence

W. CURTIS SMALL; LEE MCALISTER-HENN

1998-01-01

388

Differential regulation of the oxidative 11?-hydroxysteroid dehydrogenase activity in testis and liver  

Microsoft Academic Search

11?-hydroxysteroid dehydrogenase (11?-HSD) Type I enzyme is found in testis and liver. In Leydig cell cultures, 11?-HSD activity is reported to be primarily oxidative while another report concluded that is primarily reductive. Hepatic 11?-HSD preferentially catalyzes reduction and the reaction direction is unaffected by the external factors. Recent analysis of testicular 11?-HSD revealed two kinetically distinct components. In the present

Khatiza H. H. Nwe; Abdul Hamid; Paden B. Morat; B. A. K. Khalid

2000-01-01

389

The quinohaemoprotein alcohol dehydrogenase from Gluconacetobacter xylinus : molecular and catalytic properties  

Microsoft Academic Search

Gluconacetobacter xylinus possesses a constitutive membrane-bound oxidase system for the use of ethanol. Its alcohol dehydrogenase complex (ADH) was\\u000a purified to homogeneity and characterized. It is a 119-kDa heterodimer (68 and 41 kDa subunits). The peroxidase reaction confirmed\\u000a the presence of haem C in both subunits. Four cytochromes c per enzyme were determined by pyridine hemochrome spectroscopy. Redox titrations of the

J. L. Chávez-Pacheco; M. Contreras-Zentella; J. Membrillo-Hernández; R. Arreguín-Espinoza; G. Mendoza-Hernández; S. Gómez-Manzo; J. E. Escamilla

2010-01-01

390

Proximal tubular lactate transport in rat kidney: A micropuncture study  

Microsoft Academic Search

Proximal tubular lactate transport in rat kidney: A micropuncture study. Micropuncture studies of renal lactate behavior in the rat showed free filtration across the glomerular membrane. Under free-flow conditions, 95% of the filtered load was reabsorbed by the proximal tubule, thus generating a transtubular concentration gradient. In the absence of volume changes, an intratubular steady-state concentration of lactate was established

Bernhard Höhmann; Peter P Frohnert; Rolf Kinne; Karl Baumann; F Papavassiliou; M Wagner

1974-01-01

391

LACTATION PERSISTENCY: INSIGHTS FROM MAMMARY CELL PROLIFERATION STUDIES  

Technology Transfer Automated Retrieval System (TEKTRAN)

A persistent lactation is dependent upon maintaining number and activity of milk secreting cells with advancing lactation. When dairy cows are milked twice daily, the increase in milk yield from parturition to peak lactation is due to increased secretory activity per cell, rather than to accretion ...

392

Myocardial metabolism during hypoxia: Maintained lactate oxidation during increased glycolysis  

SciTech Connect

In the intact animal, myocardial lactate utilization and oxidation during hypoxia are not well understood. Nine dogs were chronically instrumented with flow probes on the left anterior descending coronary artery and with a coronary sinus sampling catheter. ({sup 14}C)lactate and ({sup 13}C)glucose tracers, or ({sup 13}C)lactate and ({sup 14}C)glucose were administered to quantitate lactate and glucose oxidation, lactate conversion to glucose, and simultaneous lactate extraction and release. The animals were anesthetized and exposed to 90 minutes of severe hypoxia (PO2 = 25 +/- 4 torr). Hypoxia resulted in significant increases in heart rate, cardiac output and myocardial blood flow, but no significant change in myocardial oxygen consumption. The arterial/coronary sinus differences for glucose and lactate did not change from normoxia to hypoxia; however, the rate of glucose uptake increased significantly due to the increase in myocardial blood flow. Tracer-measured lactate extraction did not decrease with hypoxia, despite a 250% increase in lactate release. During hypoxia, 90% +/- 4% of the extracted {sup 14}C-lactate was accounted for by the appearance of {sup 14}CO{sub 2} in the coronary sinus, compared with 88% +/- 4% during normoxia. Thus, in addition to the expected increase in glucose uptake and lactate production, we observed an increase in lactate oxidation during hypoxia.

Mazer, C.D.; Stanley, W.C.; Hickey, R.F.; Neese, R.A.; Cason, B.A.; Demas, K.A.; Wisneski, J.A.; Gertz, E.W. (Univ. of California, San Francisco (USA))

1990-09-01

393

EFFECTS OF LACTATE, PYRUVATE, BUTYRATE AND AMMONIA ON GLUCONEOGENESIS  

E-print Network

EFFECTS OF LACTATE, PYRUVATE, BUTYRATE AND AMMONIA ON GLUCONEOGENESIS FROM PROPIONATE BY ISOLATED cedex, France Résumé EFFET DU LACTATE, DU PYRUVATE, DU BUTYRATE ET DES IONS AMMONIUM SUR LA à partir du propionate, du lactate, du pyruvate utilisés à différentes concentrations, seuls ou en

Paris-Sud XI, Université de

394

Shaking up glycolysis: Sustained, high lactate flux during aerobic rattling  

E-print Network

Shaking up glycolysis: Sustained, high lactate flux during aerobic rattling William F. Kemper in vertebrate muscle. An alternative hypothesis is that the lactate generated during contraction reflects by comparing intracellular glycolysis during anoxia to lactate efflux from muscle during sustained, aerobic

Lindstedt, Stan

395

Original article Influence of feed restriction in primiparous lactating  

E-print Network

Original article Influence of feed restriction in primiparous lactating sows on body condition 2.3 mm) during lactation than H sows. On day W-1, L sows had higher mean concentrations of NEFA (P sows (P lactating primiparous sows alter secretion of metabolic hormones

Paris-Sud XI, Université de

396

Original article Effects of simultaneous gestation and lactation  

E-print Network

Original article Effects of simultaneous gestation and lactation in rabbit does on muscular the influence of concurrent gestation and lactation in rabbit does on the post-natal growth and muscular simultaneously pregnant and lactating does (PL group) or from only pregnant does (P group). There were

Boyer, Edmond

397

Milking lactating mares using oxytocin : milk volume and composition  

E-print Network

Milking lactating mares using oxytocin : milk volume and composition M. DOREAU, Sylviane BOULOT W. MARTIN-ROSSET H. DUBROEUCQ Station des Productions bovines et chevalines, Laboratoire de la Lactation, (*) Unité Elevage et Alimentation du Cheval, l. N. R. A., Theix, 63122 Ceyrat France. Summary. Ten lactating

Boyer, Edmond

398

Technical Note Temporal Dynamics of Lactate Concentration in the  

E-print Network

Technical Note Temporal Dynamics of Lactate Concentration in the Human Brain During Acute the feasibility of measuring the temporal dynamics of cerebral lactate concentration and examine these dynamics with 10-minute normoxic recovery was used, throughout which lactate-edited MRS was per- formed

Mitsis, Georgios

399

Microbial production of lactate-containing polyesters  

PubMed Central

Due to our increasing concerns on environmental problems and limited fossil resources, biobased production of chemicals and materials through biorefinery has been attracting much attention. Optimization of the metabolic performance of microorganisms, the key biocatalysts for the efficient production of the desired target bioproducts, has been achieved by metabolic engineering. Metabolic engineering allowed more efficient production of polyhydroxyalkanoates, a family of microbial polyesters. More recently, non-natural polyesters containing lactate as a monomer have also been produced by one-step fermentation of engineered bacteria. Systems metabolic engineering integrating traditional metabolic engineering with systems biology, synthetic biology, protein/enzyme engineering through directed evolution and structural design, and evolutionary engineering, enabled microorganisms to efficiently produce natural and non-natural products. Here, we review the strategies for the metabolic engineering of microorganisms for the in vivo biosynthesis of lactate-containing polyesters and for the optimization of whole cell metabolism to efficiently produce lactate-containing polyesters. Also, major problems to be solved to further enhance the production of lactate-containing polyesters are discussed. PMID:23718266

Yang, Jung Eun; Choi, So Young; Shin, Jae Ho; Park, Si Jae; Lee, Sang Yup

2013-01-01

400

The origin and evolution of lactation  

PubMed Central

The presence of mammary glands is the defining morphological feature of mammals. The recent assembly of the bovine genome and a report in Genome Biology that links the milk and lactation data of bovine and other mammalian genomes will help biologists investigate this economically and medically important feature. PMID:19439024

Capuco, Anthony V; Akers, R Michael

2009-01-01

401

Biomonitoring Polybrominated Diphenyl Ethers in Lactating Women  

Technology Transfer Automated Retrieval System (TEKTRAN)

Breast milk is a valuable biological specimen for biomonitoring lipid-soluble polybrominated diphenyl ethers (PBDEs). The goal of this project was to determine the levels of PBDEs in breast milk of lactating women from the Seacoast region of New Hampshire and to examine potential relationships betw...

402

Studying Reliability Using Identical Handheld Lactate Analyzers  

ERIC Educational Resources Information Center

Accusport analyzers were used to generate lactate performance curves in an investigative laboratory activity emphasizing the importance of reliable instrumentation. Both the calibration and testing phases of the exercise provided students with a hands-on opportunity to use laboratory-grade instrumentation while allowing for meaningful connections…

Stewart, Mark T.; Stavrianeas, Stasinos

2008-01-01

403

ENERGY REQUIREMENTS DURING PREGNANCY AND LACTATION  

Technology Transfer Automated Retrieval System (TEKTRAN)

Objective: To estimate the energy requirements of pregnant and lactating women consistent with optimal pregnancy outcome and adequate milk production. Design: Total energy cost of pregnancy was estimated using the factorial approach from pregnancy-induced increments in basal metabolic rate measured...

404

Alternaria alternata mannitol dehydrogenase, a fungal allergen  

Microsoft Academic Search

RationaleMannitol dehydrogenase (MtDH) has previously been shown to be the major allergen of Cladosporium herbarum. Since cross-reactivity has been demonstrated for several fungal allergens we have cloned the homolog protein of Alternaria alternata and tested it in respect to its IgE-reactivity.

P. B. Schneider; U. Denk; C. Ebner; M. Breitenbach; B. Simon-Nobbe

2004-01-01

405

Serum Dihydrolipoamide Dehydrogenase Is a Labile Enzyme  

PubMed Central

Dihydrolipoamide dehydrogenase (DLDH) is a multifunctional oxidoreductase and is well known as an essential component of four mammalian mitochondrial multienzyme complexes: pyruvate dehydrogenase, ?-ketoglutarate dehydrogenase, branched chain ?-keto acid dehydrogenase, and the glycine cleavage system. However, existence of extracellular DLDH in mammals, if any, has not been clearly defined. The present article reports identification and biochemical characterization of serum DLDH. Proteomic analysis of rat serum using blue native polyacrylamide gel electrophoresis (BN-PAGE) and mass spectrometry peptide sequencing led to generation of 6 tryptic peptides in one band that matched to mitochondrial DLDH, indicating the existence of DLDH in rat serum. Measurement of enzymatic activity also indicated the existence of DLDH in human and mouse serum. Further biochemical analysis of rat serum DLDH revealed that this enzyme lacked diaphorase activity and could not be detected on Western blots probed with antibodies that recognized mitochondrial DLDH. Moreover, both ammonium sulfate fractioning and gel filtration of serum samples rendered a great loss in DLDH activity, indicating that the enzyme activity of this serum protein, unlike that of mitochondrial DLDH, is very labile. When DTT was supplemented in the buffer used for gel filtration, DLDH activity was found to be largely preserved; indicating that serum DLDH is susceptible to air-implicated inactivation. Results of the present study indicate that serum DLDH differs from mitochondrial DLDH in that it is a very labile enzyme. PMID:23646291

Yan, Liang-Jun; Thangthaeng, Nopporn; Sumien, Nathalie; Forster, Michael J.

2013-01-01

406

Short-chain acyl-coenzyme A dehydrogenase deficiency. Clinical and biochemical studies in two patients.  

PubMed Central

We describe two patients with short-chain acyl-coenzyme A (CoA) dehydrogenase (SCADH) deficiency. Neonate I excreted large amounts of ethylmalonate and methylsuccinate; ethylmalonate excretion increased after a medium-chain triglyceride load. Neonate II died postnatally and excreted ethylmalonate, butyrate, 3-hydroxybutyrate, adipate, and lactate. Both neonates' fibroblasts catabolized [1-14C]butyrate poorly (29-64% of control). Neonate I had moderately decreased [1-14C]octanoate catabolism (43-60% of control), while neonate II oxidized this substrate normally; both catabolized radiolabeled palmitate, succinate, and/or leucine normally. Cell sonicates from neonates I and II dehydrogenated [2,3-3H]butyryl-CoA poorly (41 and 53% of control) and [2,3-3H]octanoyl-CoA more effectively (59 and 95% of control). Mitochondrial acyl-CoA dehydrogenase (ADH) activities with butyryl- and octanoyl-CoAs were 37 and 56% of control in neonate I, and 47 and 81% of control in neonate II, respectively. Monospecific medium-chain ADH (MCADH) antisera inhibited MCADH activity towards both butyryl- and octanoyl-CoAs, revealing SCADH activities to be 1 and 11% of control for neonates I and II, respectively. Fibroblast SCADH and MCADH activities were normal in an adult female with muscular SCADH deficiency. PMID:3571488

Amendt, B A; Greene, C; Sweetman, L; Cloherty, J; Shih, V; Moon, A; Teel, L; Rhead, W J

1987-01-01

407

Pyruvate dehydrogenase kinase expression and metabolic changes following dichloroacetate exposure in anoxic human colorectal cancer cells.  

PubMed

Dichloroacetate (DCA) is a small molecule that inhibits pyruvate dehydrogenase kinase (PDK) to constrain the aerobic glycolytic pathway observed in many cancer cells and effectively kill them with limited cytotoxicity on normal cells. We previously showed that DCA induced a cytoprotective effect in different human colorectal cancer (CRC) cell lines under anoxic conditions. In this study, we investigated the molecular and metabolic changes that may be providing this cytoprotection. The expression profiles of PDK isoforms in SW480 and LS174T cells along with subsequent changes in pyruvate dehydrogenase (PDH) phosphorylation were assessed following DCA exposure. Changes in mitochondrial activity and subsequent glucose consumption and lactate production were then examined. We show evidence of differential regulation in PDH phosphorylation between different human CRC cells leading to differences in mitochondrial activity following DCA exposure. However, these effects did not lead to significant changes in cellular metabolism nor growth. In conclusion, DCA may only be beneficial in treating a subset of tumor types based on their molecular profiles of different PDK isoforms. PMID:25536473

Ho, Nelson; Coomber, Brenda L

2015-02-01

408

Succinate dehydrogenase subunit B mutations modify human neuroblastoma cell metabolism and proliferation.  

PubMed

Paragangliomas (PGLs) are rare neuroendocrine tumours. About 30-40 % of these tumours are mutated in one of the different susceptibility genes, including those encoding the different subunits of the succinate dehydrogenase, a complex involved both in the tricarboxylic acid cycle and in the oxygen transport chain. The aim of this work was to investigate whether SDHB mutations may account for alterations in cell metabolism and functions. Since human PGL cell lines are not available, we used the neuroblastoma cell line (SK-N-AS) stably transfected with the wild-type human SDHB or different SDHB-mutated constructs carrying some significant mutations found in our patients affected by PGLs. Similarly to succinate dehydrogenase (SDH)-mutated tumour cells, mutated SK-N-AS clones showed reduced SDH enzyme activity. All clones showed normal citrate synthase activity, reduced oxygen consumption and reduced carbonic anhydride production, thus demonstrating a decreased in mitochondrial metabolism. In two of the three mutated SK-N-AS, we also found an increase in HIF1? expression. Surprisingly and unexpectedly, in all the SDHB-mutated clones, we found a significant decrease in glucose uptake and in lactate culture medium concentration, suggesting also a decrease of cytosolic metabolism. Finally, we found that these energetic changes were associated to an increase in cell proliferation and migration. Overall, these data demonstrate that although SDHB mutations significantly downregulate both mitochondrial and cytoplasmic cellular metabolism, these mutations are associated to an upregulation of some cellular functions, such as growth rate and invasiveness. PMID:24595825

Rapizzi, Elena; Ercolino, Tonino; Fucci, Rossella; Zampetti, Benedetta; Felici, Roberta; Guasti, Daniele; Morandi, Andrea; Giannoni, Elisa; Giaché, Valentino; Bani, Daniele; Chiarugi, Alberto; Mannelli, Massimo

2014-06-01

409

Development of an NADPH-dependent homophenylalanine dehydrogenase by protein engineering.  

PubMed

l-Homophenylalanine is a nonproteinogenic amino acid and can be used as a versatile pharmaceutical intermediate. Production of l-homophenylalanine involves amination of the keto acid precursor 2-oxo-4-phenylbutyric acid (2-OPBA), which can be accomplished by bioenzymatic processes. Current biocatalysts for this reaction include transaminases and NADH-dependent phenylalanine dehydrogenases, which are not optimal for metabolic engineering of whole-cell biocatalysis. Here, we report the development of an NADPH-dependent homophenylalanine dehydrogenase by engineering the NADPH-dependent glutamate dehydrogenase (GDH) from Escherichia coli, which provides a new tool for in vitro catalysis and in vivo metabolic engineering. We took a stepwise substrate walking strategy: the first round directed evolution switched GDH's substrate specificity from its natural substrate 2-ketoglutarate to the intermediate target phenylpyruvate, which has similar structure as 2-OPBA; and the second round further improved the enzyme's catalytic efficiency toward the final target 2-OPBA. Compared to wild type GDH, the catalytic efficiency (kcat/Km) of the final mutant was ?100 fold higher for 2-OPBA and ?3000 fold lower for the original substrate 2-ketoglutarate. When overexpressed in E. coli, the engineered GDH aminated 2-OPBA to l-homophenylalanine more effectively than the transaminases and NADH-dependent phenylalanine dehydrogenase, possibly because it utilizes the strong anabolic driving force NADPH under aerobic condition. PMID:24053171

Li, Han; Liao, James C

2014-01-17

410

Hormonal and behavioral responses to stress in lactating and non-lactating female common marmosets (Callithrix jacchus)  

E-print Network

Hormonal and behavioral responses to stress in lactating and non-lactating female common marmosets hormone Cortisol Lactation Maternal care Parental care Stress In several mammalian species, hypothalamic preventing stress-induced disruptions of maternal care. Experimental elevations of HPA axis hormones have

Saltzman, Wendy

411

Exposure to Mother's Pregnancy and Lactation in Infancy is Associated with Sexual Attraction to Pregnancy and Lactation  

E-print Network

Exposure to Mother's Pregnancy and Lactation in Infancy is Associated with Sexual Attraction to Pregnancy and Lactation in Adulthoodjsm_2065 140..147 Magnus Enquist, PhD,* Hanna Aronsson, B.Sc.,* Stefano from individuals with a sexual preference for pregnant and/or lactating women, under the hypothesis

412

Microphotometric measurement of initial maximum reaction rates in quantitative enzyme histochemistry in situ  

Microsoft Academic Search

Summary  Final reaction product formation was recorded microphotometrically for succinate dehydrogenase in cross-sectioned muscle fibres at initial rate conditions and during prolonged incubations. Incubations with gel films and aqueous reaction medium both showed a decline of reaction rates. Maximum reaction rates could only be determined at initial rate conditions during the first minute of the incubation. Reaction rates recorded in different

Dirk Pette; Universitfit Konstanz

1981-01-01

413

Identification of clinical, epidemiological and laboratory risk factors for leprosy reactions during and after multidrug therapy  

PubMed Central

This cross-sectional retrospective study evaluated 440 leprosy patients; 57% (251/440) had leprosy reactions during and/or after multidrug therapy, 80.5% (202/251) of whom presented with multibacillary leprosy. At diagnosis, positive bacterial index (BI) [odds ratio (OR) = 6.39; 95% confidence interval (CI): 4.1-10.1)] or polymerase chain reaction (PCR) (OR = 9.15; 95% CI: 5.4-15.5) in skin smears, anti-phenolic glycolipid-1 (anti-PGL-1) ELISA (OR = 4.77; 95% CI: 2.9-7.9), leucocytosis (OR = 9.97; 95% CI: 3.9-25.7), thrombocytopenia (OR = 5.72; 95% CI: 2.3-14.0) and elevated lactate dehydrogenase (OR = 2.38; 95% CI: 1.4-4.0) were potential markers for the development of reactions during treatment. After treatment, positive BI (OR = 8.47; 95% CI: 4.7-15.3) and PCR (OR = 6.46; 95% CI: 3.4-12.3) in skin smears, anti-PGL-1 ELISA (OR = 2.25; 95% CI: 1.3-3.9), anaemia (OR = 2.36; 95% CI: 1.2-4.5), leucocytosis (OR = 4.14; 95% CI: 1.5-11.6) and thrombocytopenia (OR = 3.70; 95% CI: 1.3-2.2) were risk factors for the occurrence of reactions during the study period. The identification of groups with an increased risk for developing reactions will allow for the timely development of a treatment plan to prevent nerve damage and, therefore, the appearance of the disabling sequelae associated with the stigma of leprosy. PMID:24271045

Antunes, Douglas Eulálio; Araujo, Sergio; Ferreira, Gabriela Porto; da Cunha, Ana Carolina Sousa Rodrigues; da Costa, Adeilson Vieira; Gonçalves, Maria Aparecida; Goulart, Isabela Maria Bernardes

2013-01-01

414

Effect of lactation number, year, and season of initiation of lactation on milk yield of cows hormonally induced into lactation and treated with recombinant bovine somatotropin.  

PubMed

Records representing data from 1,500 barren Holstein cows over an 8-yr period from a large commercial dairy farm in northern Mexico were analyzed to determine the effects of lactation number and season and year of initiation of lactation on milk production of cows induced hormonally into lactation and treated with recombinant bovine somatotropin (rbST) throughout lactation. Peak and 305-d milk yields were also assessed as predictors of total milk yield in cows induced into lactation. A significant quadratic relationship was found between 305-d milk yield and number of lactation [7,607±145 and 9,548±181 kg for first- and ?6-lactation cows, respectively; mean ± standard error of the mean (SEM)] with the highest production occurring in the fifth lactation. Total milk yields of cows with ?2 lactations were approximately 4,500 kg less than milk yields of adult cows (the overall average ± standard milk yield was 13,544±5,491 kg per lactation and the average lactation length was 454±154 d). Moreover, 305-d milk production was depressed in cows induced into lactation in spring (8,804±153 kg; mean ± SEM) and summer (8,724±163 kg) than in fall (9,079±151 kg) and winter (9,085±143 kg). Partial regression coefficients for 305-d milk yield and peak milk yield indicated an increment of 157 kg of milk per lactation per 1-kg increase in peak milk yield (r(2)=0.69). Neither peak milk yield (r(2)=0.18) nor 305-d milk yield (r(2)=0.29) was accurate for predicting total milk yield per lactation. Year, parity, and season effects had significant influence on milk yield of cows induced into lactation and treated with rbST throughout lactation, and peak milk yield can assist in the prediction of 305-d milk yield but not total milk yield. This study also showed that hormonal induction of lactation in barren high-yielding cows is a reliable, practical, and affordable technique in countries where rbST treatment and prolonged steroid administration of dairy cows are legally permitted. PMID:21854924

Mellado, M; Antonio-Chirino, E; Meza-Herrera, C; Veliz, F G; Arevalo, J R; Mellado, J; de Santiago, A

2011-09-01

415

Long-chain acyl-CoA dehydrogenase is a key enzyme in the mitochondrial ?-oxidation of unsaturated fatty acids  

Microsoft Academic Search

The first reaction of mitochondrial ?-oxidation, which is catalyzed by acyl-CoA dehydrogenases, was studied with unsaturated fatty acids that have a double bond either at the 4,5 or 5,6 position. The CoA thioesters of docosahexaenoic acid, arachidonic acid, 4,7,10-cis-hexadecatrienoic acid, 5-cis-tetradecenoic acid, and 4-cis-decenoic acid were effectively dehydrogenated by both rat and human long-chain acyl-CoA dehydrogenases (LCAD), whereas they were

Weiping Le; Azfar S. Abbas; Howard Sprecher; Jerry Vockley; Horst Schulz

2000-01-01

416

Channel-mediated lactate release by k+-stimulated astrocytes.  

PubMed

Excitatory synaptic transmission is accompanied by a local surge in interstitial lactate that occurs despite adequate oxygen availability, a puzzling phenomenon termed aerobic glycolysis. In addition to its role as an energy substrate, recent studies have shown that lactate modulates neuronal excitability acting through various targets, including NMDA receptors and G-protein-coupled receptors specific for lactate, but little is known about the cellular and molecular mechanisms responsible for the increase in interstitial lactate. Using a panel of genetically encoded fluorescence nanosensors for energy metabolites, we show here that mouse astrocytes in culture, in cortical slices, and in vivo maintain a steady-state reservoir of lactate. The reservoir was released to the extracellular space immediately after exposure of astrocytes to a physiological rise in extracellular K(+) or cell depolarization. Cell-attached patch-clamp analysis of cultured astrocytes revealed a 37 pS lactate-permeable ion channel activated by cell depolarization. The channel was modulated by lactate itself, resulting in a positive feedback loop for lactate release. A rapid fall in intracellular lactate levels was also observed in cortical astrocytes of anesthetized mice in response to local field stimulation. The existence of an astrocytic lactate reservoir and its quick mobilization via an ion channel in response to a neuronal cue provides fresh support to lactate roles in neuronal fueling and in gliotransmission. PMID:25762664

Sotelo-Hitschfeld, Tamara; Niemeyer, María I; Mächler, Philipp; Ruminot, Iván; Lerchundi, Rodrigo; Wyss, Matthias T; Stobart, Jillian; Fernández-Moncada, Ignacio; Valdebenito, Rocío; Garrido-Gerter, Pamela; Contreras-Baeza, Yasna; Schneider, Bernard L; Aebischer, Patrick; Lengacher, Sylvain; San Martín, Alejandro; Le Douce, Juliette; Bonvento, Gilles; Magistretti, Pierre J; Sepúlveda, Francisco V; Weber, Bruno; Barros, L Felipe

2015-03-11

417

Inhibition kinetics of catabolic dehydrogenases by elevated moieties of ATP and ADP--implication for a new regulation mechanism in Lactococcus lactis.  

PubMed

ATP and ADP inhibit, in varying degrees, several dehydrogenases of the central carbon metabolism of Lactococcus lactis ATCC 19435 in vitro, i.e. glyceraldehyde-3-phosphate dehydrogenase (GAPDH), lactate dehydrogenase (LDH) and alcohol dehydrogenase (ADH). Here we demonstrate mixed inhibition for GAPDH and competitive inhibition for LDH and ADH by adenine nucleotides in single inhibition studies. The nonlinear negative co-operativity was best modelled with Hill-type kinetics, showing greater flexibility than the usual parabolic inhibition equation. Because these natural inhibitors are present simultaneously in the cytoplasm, multiple inhibition kinetics was determined for each dehydrogenase. For ADH and LDH, the inhibitor combinations ATP plus NAD and ADP plus NAD are indifferent to each other. Model discrimination suggested that the weak allosteric inhibition of GAPDH had no relevance when multiple inhibitors are present. Interestingly, with ADH and GAPDH the combination of ATP and ADP exhibits lower dissociation constants than with either inhibitor alone. Moreover, the concerted inhibition of ADH and GAPDH, but not of LDH, shows synergy between the two nucleotides. Similar kinetics, but without synergies, were found for horse liver and yeast ADHs, indicating that dehydrogenases can be modulated by these nucleotides in a nonlinear manner in many organisms. The action of an elevated pool of ATP and ADP may effectively inactivate lactococcal ADH, but not GAPDH and LDH, providing leverage for the observed metabolic shift to homolactic acid formation in lactococcal resting cells on maltose. Therefore, we interpret these results as a regulation mechanism contributing to readjusting the flux of ATP production in L. lactis. PMID:20193044

Cao, Rong; Zeidan, Ahmad A; Rådström, Peter; van Niel, Ed W J

2010-04-01

418

Relation between Neonatal Icter and Gilbert Syndrome in Gloucose-6-Phosphate Dehydrogenase Deficient Subjects  

PubMed Central

Background and Aim: The pathogenesis of neonatal hyperbilirubinemia hasn’t been completely defined in Gloucose-6-Phosphate Dehydrogenase (G6PD) deficient newborns. The aim of this study was to detect the relationship between Gilbert’s syndrome and hyperbilirubinemia in Gloucose-6-Phosphate Dehydrogenase (G6PD) deficient neonates. Materials and Methods: This case-control study was conducted in Amirkola pediatrics teaching hospital, Babol, Iran. A total number of one hundred four infants were included in the study (51 infants with neonatal jaundice and Gloucose-6-Phosphate Dehydrogenase (G6PD) deficiency admitted to phototherapy or transfusion were selected as the case group and 53 infants with Gloucose-6-Phosphate Dehydrogenase (G6PD) deficiency admitted for other reasons than jaundice were selected as the control group). Exclusion criteria were ABO or Rh incompatibility or other reasons that made Coombs test positive, sepsis, hepatosplenomegaly, metabolic diseases, medical treatment and phototherapy. The promoter and coding regions of Uridine diphosphate Glucuronosyl Transferase 1A1 (UGT1A1) of genomic DNA were amplified by polymerase chain reaction (PCR) isolated from leukocytes. We used chi-square test and t-test to compare cases and controls. Results: Distribution of Gilbert genome was not significantly different between the two groups; among cases, 33.3% were homozygote, 35.3% heterozygote, and 31.4% normal. Among controls, 22.6% were homozygote, 34% heterozygote, and 43.4% normal (p-value=xxx). Hyperbilirubinemia family history didn’t differ significantly between these two groups. Conclusions: We showed that in Gloucose-6-Phosphate Dehydrogenase (G6PD) deficient neonates, there was no significant association between Gilbert’s syndrome (promoter polymorphism) and hyperbilirubinemia. PMID:24783083

Zahedpasha, Yadollah; Ahmadpour, Mousa; Niaki, Haleh Akhavan; Alaee, Ehsan

2014-01-01

419

Studying reliability using identical handheld lactate analyzers  

NSDL National Science Digital Library

Accusport analyzers were used to generate lactate performance curves in an investigative laboratory activity emphasizing the importance of reliable instrumentation. Both the calibration and testing phases of the exercise provided students with a hands-on opportunity to use laboratory-grade instrumentation while allowing for meaningful connections to be made between data collection and analysis. Pairs of student teams tested individual aerobically trained participants exercising to voluntary exhaustion on a cycle ergometer. The analysis of four volunteers' postexercise blood samples revealed lactate data that, although highly correlated, showed small but statistically significant differences between devices. This laboratory activity provides a useful platform for introducing students to the reliability of instrumentation, in particular noting its relevance to designs employing repeated measures.

Mark T Stewart (Willamette University Psychology)

2008-03-24

420

Calcium deficiency, pregnancy, and lactation in rats  

Microsoft Academic Search

Summary Adult female rats were subjected to a calcium-depriving regimen (calcium-deficient diet containing oxalate+pregnancy+lactation) to obtain maximum bone mineral mobilization in as short a time as possible. The femur and tibia were investigated by histological, microradiographic and fluorescent microscopic methods. The regimen caused osteoporosis, which varied in severity with the degree of calcium deprivation. Most of the bone tissue removed

P. Rasmussen

1977-01-01

421

Extended lactation in dairy cows: effects of milking frequency, calving season and nutrition on lactation persistency and milk quality.  

PubMed

Twelve spring-calving and twelve winter-calving cows were managed for extended lactation cycles of 18-months duration, with the former group then completing a second extended lactation. Half of the cows were fed according to standard management practice for the herd; the other half received supplementary concentrate from week 9 of lactation onwards. Commencing at the same time, half of the udder of each cow was subjected to increased milking frequency (thrice daily rather than twice daily). Lactation persistency (and hence total milk yield) was significantly increased by frequent milking. Winter calving cows and supplemented cows also exhibited better persistency, but this was only evident up until the point of re-breeding, at around lactation week 33. Milk composition was measured in the spring-calving cows in both their first and second extended lactations. Composition altered during the course of the lactation, protein and fat percentages increasing and lactose percentage decreasing, irrespective of treatment. The quality of the milk for processing into cheese, fermented products, heat-treated products and cream liqueurs was assessed by calculation of casein number (casein protein as a proportion of total protein). Processing quality declined across the course of lactation in those groups that showed poor persistency but not in those that maintained a persistent lactation. Milk hygienic quality (somatic cell counts) showed parallel changes. Body condition score increased during the course of lactation but was not affected by supplementation; none of the cows became excessively fat. All cows remained healthy throughout the extended lactations and the majority (33/36) re-bred successfully. By demonstrating that lactation persistency is plastic and can be improved by simple management interventions, the results lend support to the economic arguments in favour of extended lactation cycles. The likely welfare benefits of extended lactation are also discussed. PMID:18226299

Sorensen, Annette; Muir, D Donald; Knight, Christopher H

2008-02-01

422

Increased Brain Lactate Concentrations Without Increased Lactate Oxidation During Hypoglycemia in Type 1 Diabetic Individuals  

PubMed Central

Previous studies have reported that brain metabolism of acetate is increased more than twofold during hypoglycemia in type 1 diabetic (T1D) subjects with hypoglycemia unawareness. These data support the hypothesis that upregulation of blood-brain barrier monocarboxylic acid (MCA) transport may contribute to the maintenance of brain energetics during hypoglycemia in subjects with hypoglycemia unawareness. Plasma lactate concentrations are ?10-fold higher than acetate concentrations, making lactate the most likely alternative MCA as brain fuel. We therefore examined transport of [3-13C]lactate across the blood-brain barrier and its metabolism in the brains of T1D patients and nondiabetic control subjects during a hypoglycemic clamp using 13C magnetic resonance spectroscopy. Brain lactate concentrations were more than fivefold higher (P < 0.05) during hypoglycemia in the T1D subjects compared with the control subjects. Surprisingly, we observed no increase in the oxidation of blood-borne lactate in the T1D subjects, as reflected by similar 13C fractional enrichments in brain glutamate and glutamine. Taken together, these data suggest that in addition to increased MCA transport at the blood-brain barrier, there may be additional metabolic adaptations that contribute to hypoglycemia unawareness in patients with T1D. PMID:23715622

De Feyter, Henk M.; Mason, Graeme F.; Shulman, Gerald I.; Rothman, Douglas L.; Petersen, Kitt Falk

2013-01-01

423

Sorbit-Dehydrogenase in menschlicher Epidermis  

Microsoft Academic Search

The presence of the enzyme Sorbit-Dehydrogenase (SDH) was proved in the homogen of humanate epidermis. The concentration of SDH in the epidermis of the corpse was roughly with 287 m.U. in the vehicle twice as much higher as in the amputated skin with 105 m.U. It appears that the general disease has no influence on the activity of the enzyme

U. Ganzer; G. Weber

1967-01-01

424

Active site of human liver aldehyde dehydrogenase  

Microsoft Academic Search

Bromoacetophenone (2-bromo-1-phenylethanone) functions as an affinity reagent for human aldehyde dehydrogenase (EC 1.2.1.3) and has been found specifically to label a unique tryptic peptide in the enzyme. Amino-terminal sequence analysis of the labeled peptide after purification by two different procedures revealed the following sequence: Val-Thr-Leu-Glu-Leu-Gly-Gly-Lys. Radioactivity was found to be associated with the glutamate residue, which was identified as Glu-268

Darryl P. Abriola; Robert Fields; Stanley Stein; Alexander D. MacKerell; Regina Pietruszko

1987-01-01

425

Lactic Dehydrogenase Activity in Bovine Semen1  

Microsoft Academic Search

SUMMARY Lactic dehydrogenase (LD) activity has been determined in 43 samples of bovine semen from 23 bulls of the Aberdeen-Angus, Holstein-Friesian, Guernsey, Jersey, and Milking Shorthorn breeds. The mean activity value expressed as Berger-Broida units\\/ milliliter of bovine semen was 2,026 ± 151 units. Correlation coefficient between LD units\\/milliliter of semen and 30-60 day nonreturn conception data was --.414 (P

O. T. Stallcup; J. S. Hayden

1960-01-01

426

Cloning, sequence analysis, and expression in Escherichia coli of gene encoding N-Benzyl-3-pyrrolidinol dehydrogenase from Geotrichum capitatum.  

PubMed

The gene encoding N-benzyl-3-pyrrolidinol dehydrogenase (DDBJ/EMBL/GenBank accession no. AB294179), a useful b