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1

Optimum Reaction Conditions for Human Lactate Dehydrogenase lsoenzymes as They Affect Total Lactate Dehydrogenase Activity  

Microsoft Academic Search

Optimum reaction conditions at 30#{176} ± 0.5#{176} for two continuous spectrophotometric assayprocedures,lactate to pyruvate (L- P) and pyruvate to lactate (P- L), were determined with respect to pH at 30#{176} (pH30) substrate concentration, and coenzyme concentration for the human LDH isoenxymes. For the P - L procedure, broad pH30optima were within the range of 7.20-7.40 for all the LDH isoenzymes.The

Royal J. Gay; Robert B. McComb; George N. Bowers

2

Effect of osmolytes on protein dynamics in the lactate dehydrogenase-catalyzed reaction.  

PubMed

Laser-induced temperature jump relaxation spectroscopy was used to probe the effect of osmolytes on the microscopic rate constants of the lactate dehydrogenase-catalyzed reaction. NADH fluorescence and absorption relaxation kinetics were measured for the lactate dehydrogenase (LDH) reaction system in the presence of varying amounts of trimethylamine N-oxide (TMAO), a protein-stabilizing osmolyte, or urea, a protein-destabilizing osmolyte. Trimethylamine N-oxide (TMAO) at a concentration of 1 M strongly increases the rate of hydride transfer, nearly nullifies its activation energy, and also slightly increases the enthalpy of hydride transfer. In 1 M urea, the hydride transfer enthalpy is almost nullified, but the activation energy of the step is not affected significantly. TMAO increases the preference of the closed conformation of the active site loop in the LDH·NAD(+)·lactate complex; urea decreases it. The loop opening rate in the LDH·NADH·pyruvate complex changes its temperature dependence to inverse Arrhenius with TMAO. In this complex, urea accelerates the loop motion, without changing the loop opening enthalpy. A strong, non-Arrhenius decrease in the pyruvate binding rate in the presence of TMAO offers a decrease in the fraction of the open loop, pyruvate binding competent form at higher temperatures. The pyruvate off rate is not affected by urea but decreases with TMAO. Thus, the osmolytes strongly affect the rates and thermodynamics of specific events along the LDH-catalyzed reaction: binding of substrates, loop closure, and the chemical event. Qualitatively, these results can be understood as an osmolyte-induced change in the energy landscape of the protein complexes, shifting the conformational nature of functional substates within the protein ensemble. PMID:21306147

Zhadin, Nickolay; Callender, Robert

2011-02-09

3

Use of pyruvate oxidase to overcome pyruvate inhibition during the lactate to pyruvate reaction for assaying lactate dehydrogenase in serum.  

PubMed

Automated assays of lactate dehydrogenase (LD) in serum are based on measuring the rate of NADH produced in a reverse LD reaction using lactate and NAD. The observed nonlinearity of LD reaction used in earlier assays performed in phosphate buffers has generally been attributed to the formation of a ternary complex of NAD, pyruvate, and phosphate. this is not satisfactory to explain the course of assay reaction carried out in organic buffers. Investigation of the possible causes of nonlinearity during the course of the reverse LD reaction during LD assays performed in Tris or other organic buffers indicated that inhibition of LD activity by pyruvate may be chiefly responsible for the observed effects, especially in serum exhibiting abnormally high LD enzyme activity. Most of the LD activity in serum was inhibited by 5 mMoles/L pyruvate. By contrast, the LD isoenzyme activities were inhibited partially at 0.5 mMole/L pyruvate, LD1 being the most and LD4 the least susceptible. In assays of serum samples with abnormally high LD and PYR concentration using LD reagent containing Tris buffer, pH 9.3, the inclusion of a bacterial pyruvate oxidase (PO) enabled the removal of pyruvate accumulating in situ, making it possible to assay LD activity in the absence of inhibitory concentration of pyruvate. The inclusion of 10 U/L of PO in our routine LD reagent was sufficient to overcome pyruvate inhibition, thus permitting the assay of serum exhibiting high LD activity, hence the extension of the upper limits of linearity of LD assay without compromising assay performance. PMID:7562238

Murthy, V V

1995-01-01

4

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

5

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

2005-08-08

6

[Malate dehydrogenase and lactate dehydrogenase in trematodes and turbellarians].  

PubMed

Studies have been made on the activity and properties of malate and lactate dehydrogenases from the cattle rumen trematodes Eurytrema pancreaticum, Calicophoron ijimai and the turbellarian Phagocata sibirica which has a common free-living ancestor with the trematodes. All the species studied have a highly active malate dehydrogenase, its activity in the reaction of reducing oxaloacetate being 6-14 times higher than in the reaction of malate oxidation. The affinity of malate dehydrogenase to oxaloacetate was found to be higher than that to malate. The activity of lactate dehydrogenase (reducing the pyruvate) was lower than the activity of malate dehydrogenase, the difference being 50 times for C. ijimai, 4 times for E. pancreaticum and 10 times for P. sibirica. PMID:3962529

Vykhrestiuk, N P; Burenina, E A; Iarygina, G V

7

The lactate dehydrogenase catalyzed pyruvate adduct reaction: simultaneous general acid-base catalysis involving an enzyme and an external catalyst.  

PubMed

The pH dependence of the reaction catalyzed by lactate dehydrogenase, where pyruvate adds covalently to NAD to yield a NAD-Pyr adduct, together with published data on the pH dependence of parameters in the normal redox reaction suggests similar binding modes for enolpyruvate and lactate in their complexes with E X NAD (where E is one-fourth of the tetramer), for ketopyruvate in its complexes with the protonated species, E X H X NAD and E X H X NADH, and for the NAD--Pyr adduct and NADH plus pyruvate in their complexes with E X H. These similarities, together with previous data, suggest a reaction scheme for the formation of the enzyme-adduct complex that includes the relevant proton-transfer steps. Seven different amine chloride buffers were used in a study of the reverse adduct reaction, i.e., the decomposition of E X H X NAD--Pyr. These act with varying efficiencies as external general acid catalysts; the enzyme apparently acts as a (internal) general base. The involvement of the amine chloride buffers as external general catalysts is supported by the concentration dependence of the buffer effect, by a Brönsted plot, and by solvent deuterium isotope effects. The involvement of the enzyme as an internal general catalyst is inferred from the pH dependence of the reaction and the identities of the nearby groups in the E X H X NAD--Pyr complex (from crystallographic studies). The dependence of the adduct reaction on chloride concentration indicates the presence of dead-end inhibitor complexes of E X H X Cl and E X H X NAD X Cl. Chloride also accelerates the decomposition of the adduct in the complex E X H X NAD--Pyr by binding to this complex. PMID:6477888

Burgner, J W; Ray, W J

1984-07-31

8

Drosophila lactate dehydrogenase: Developmental aspects  

Microsoft Academic Search

Partially purified lactate dehydrogenase (LDH) from third-instar larvae displays two bands (one major and one minor) on polyacrylamide\\u000a gels. Analogous preparations from pupae and adults exhibit three LDH-staining bands (one major and two minor) in a similar\\u000a pattern. The migration of the major band is similar for larvae, pupae, and adults, while the two minor LDH bands of pupae\\u000a and

S. N. Alahiotis; A. Onoufriou; M. Fotaki; M. Pelecanos

1983-01-01

9

21 CFR 862.1445 - Lactate dehydrogenase isoenzymes test system.  

Code of Federal Regulations, 2010 CFR

... 2009-04-01 false Lactate dehydrogenase isoenzymes test system...Systems § 862.1445 Lactate dehydrogenase isoenzymes test system. (a) Identification. A lactate dehydrogenase isoenzymes test system...

2009-04-01

10

21 CFR 862.1445 - Lactate dehydrogenase isoenzymes test system.  

Code of Federal Regulations, 2010 CFR

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

2010-04-01

11

21 CFR 862.1445 - Lactate dehydrogenase isoenzymes test system.  

Code of Federal Regulations, 2013 CFR

...lactate dehydrogenase isoenzymes test system is a device intended to measure the activity of lactate dehydrogenase isoenzymes (a group of enzymes with similar biological activity) in serum. Measurements of lactate dehydrogenase isoenzymes are...

2013-04-01

12

Structural Adaptations of Lactate Dehydrogenase Isozymes  

Microsoft Academic Search

The three-dimensional structures of dogfish M4 (muscle) and pig H4 (heart) lactate dehydrogenase (L-lactate:NAD+ oxidoreductase, EC 1.1.1.27) have been determined and correlated with the amino acid sequences of the dogfish M4, pig M4, pig H4, chicken M4, and chicken H4 lactate dehydrogenase isozymes. These results have been related to the known differences of physicochemical properties between the M and H

William Eventoff; Michael G. Rossmann; Susan S. Taylor; Hans-Joachim Torff; Helmut Meyer; Walter Keil; Hans-Hermann Kiltz

1977-01-01

13

Quantitative Electrophoretic Determination of Lactate Dehydrogenase Isoenzymes.  

National Technical Information Service (NTIS)

Lactate dehydrogenase isoenzymes of human serum and tissue extracts were separated by agar gel electrophoresis on a microscope slide. The isoenzyme activity was identified by a procedure utilizing p-iodonitrotetrazolium and quantitated by densitometry. Fi...

N. M. Papadopoulos J. A. Kintzios

1966-01-01

14

Convergent Evolution of Trichomonas vaginalis Lactate Dehydrogenase from Malate Dehydrogenase  

Microsoft Academic Search

Lactate dehydrogenase (LDH) is present in the amitochondriate parasitic protist Trichomonas vaginalis and some but not all other trichomonad species. The derived amino acid sequence of T. vaginalis LDH (TvLDH) was found to be more closely related to the cytosolic malate dehydrogenase (MDH) of the same species than to any other LDH. A key difference between the two T. vaginalis

Gang Wu; Andras Foser; Benno Ter Kuile; Andrej Sali; Miklos Muller

1999-01-01

15

Role of Mitochondrial Lactate Dehydrogenase and Lactate Oxidation in the Intracellular Lactate Shuttle  

Microsoft Academic Search

To evaluate the potential role of mitochondrial lactate dehydrogenase (LDH) in tissue lactate clearance and oxidation in vivo, isolated rat liver, cardiac, and skeletal muscle mitochondria were incubated with lactate, pyruvate, glutamate, and succinate. As well, alpha -cyano-4-hydroxycinamate (CINN), a known monocarboxylate transport inhibitor, and oxamate, a known LDH inhibitor were used. Mitochondria readily oxidized pyruvate and lactate, with similar

George A. Brooks; Herve Dubouchaud; Marcia Brown; James P. Sicurello; C. Eric Butz

1999-01-01

16

Structure-Function Relationships in Lactate Dehydrogenase  

Microsoft Academic Search

The binding of coenzyme and substrate are considered in relation to the known primary and tertiary structure of lactate dehydrogenase (EC 1.1.1.27). The adenine binds in a hydrophobic crevice, and the two coenzyme phosphates are oriented by interactions with the protein. The positively charged guanidinium group of arginine 101 then folds over the negatively charged phosphates, collapsing the loop region

Margaret J. Adams; Manfred Buehner; K. Chandrasekhar; Geoffrey C. Ford; Marvin L. Hackert; Anders Liljas; Michael G. Rossmann; Ira E. Smiley; William S. Allison; Johannes Everse; Nathan O. Kaplan; Susan S. Taylor

1973-01-01

17

Amperometric immunosensor for lactate dehydrogenase LD-1.  

PubMed

An amperometric immunosensor for the detection of the lactate dehydrogenase isoenzyme LD-1 has been developed. Polyclonal antibodies for LD-1 have been covalently immobilised onto a preactivated Immunodyne ABC membrane, reacted with standard LD-1 solutions and placed onto a platinum working electrode polarised at +600 mV vs. Ag/AgCl. Lactate dehydrogenase activity has been measured by detection of the oxidation of NADH at the electrode surface. A calibration curve for LDH in the 0.005-0.12 U range has been obtained with a repeatability of 3% and a reproducibility of 10%. The probe exhibited good selectivity (response of LD-2 was 18% of LD-1) which was further improved using thermal treatment of the membranes. Measurement of LD-1 content in human control serum with the developed procedure gave a LD-1 concentration in the reported assigned range. PMID:9597734

Kelly, S; Compagnone, D; Guilbault, G

1998-02-01

18

Insights into substrate binding by D -2-ketoacid dehydrogenases from the structure of Lactobacillus pentosus D -lactate dehydrogenase  

Microsoft Academic Search

Background:D-Lactate dehydrogenases (D-LDHs) and L-lactate dehydrogenases (L-LDHs) catalyze a reaction differing only in the chirality of the product. Both enzymes utilize the same kind of amino acid side chains in substrate binding and catalysis. Models based on D-LDH-related enzymes propose that these side chains assume identical roles in both enzymes with their active sites related by a simple geometrical relationship

Vincent S Stoll; Matthew S Kimber; Emil F Pai

1996-01-01

19

Substrate and cofactor specificity and selective inhibition of lactate dehydrogenase from the malarial parasite P. falciparum  

Microsoft Academic Search

Lactate dehydrogenase from the malarial parasite Plasmodium falciparum has many amino acid residues that are unique compared to any other known lactate dehydrogenase. This includes residues that define the substrate and cofactor binding sites. Nevertheless, parasite lactate dehydrogenase exhibits high specificity for pyruvic acid, even more restricted than the specificity of human lactate dehydrogenases M4 and H4. Parasite lactate dehydrogenase

Manuel S Gomez; Robert C Piper; Lucy A Hunsaker; Robert E Royer; Lorraine M Deck; Michael T Makler; David L Vander Jagt

1997-01-01

20

21 CFR 862.1440 - Lactate dehydrogenase test system.  

Code of Federal Regulations, 2013 CFR

...dehydrogenase in serum. Lactate dehydrogenase measurements are used in the diagnosis and treatment of liver diseases such as acute viral hepatitis, cirrhosis, and metastatic carcinoma of the liver, cardiac diseases such as myocardial infarction, and tumors of the...

2013-04-01

21

Catecholamine regulation of lactate dehydrogenase in rat brain cell culture  

SciTech Connect

The mechanism of catecholamine induction of the soluble cytoplasmic enzyme lactate dehydrogenase (EC 1.1.1.27) was studied in the rat glial tumor cell line, C6. Lactate dehydrogenase was partially purified from extracts of (/sup 3/H)leucine-labeled cells by affinity gel chromatography and quantitatively immunoprecipitated with anti-lactate dehydrogenase-5 IgG and with antilactate dehydrogenase-1 IgG. The immunoprecipitates were dissociated and electrophoresed on sodium dodecyl sulfate polyacrylamide gels. Using this methodology, the increased enzyme activity of lactate dehydrogenase in norepinephrine-treated C6 cells was observed to be concomitant with the increased synthesis of enzyme molecules. Despite the continued presence of norepinephrine, the specific increase in the rate of synthesis of lactate dehydrogenase was transient. It was first detected at 4 h, was maximum at 9 h, and returned to basal levels by 24 h. The half-life of lactate dehydrogenase enzyme activity was 36 h during the induction and 40 h during deinduction. The half-life for decay of /sup 3/H-labeled lactate dehydrogenase was 41 h. These observations suggest that the increase in lactate dehydrogenase activity in norepinephrine-treated cells does not involve any change in the rate of degradation. Norepinephrine increased the specific rate of synthesis of both lactate dehydrogenase-5 (a tetramer of four M subunits) and lactate dehydrogenase-1 (a tetramer of four H subunits), although to different extents. Since these subunits are coded for by two separate genes on separate chromosomes, it suggests that the regulatory mechanism involves at least two separate sites of action.

Kumar, S.; McGinnis, J.F.; de Vellis, J.

1980-03-25

22

Lactate Dehydrogenase in Testis: Dissociation and Recombination of Subunits  

Microsoft Academic Search

Electrophoretic resolution of lactate dehydrogenase in mature testes from a variety of animals revealed one or more unusual isozymes in addition to the usual five forms. Dissociation of the enzyme and recombination of the polypeptide subunits led to the formation of new isozymes and to a redistribution of activity among those normally present, indicating that lactate dehydrogenase synthesis in postpubertal

William H. Zinkham; Antonio Blanco; Luba Kupchyk

1963-01-01

23

Lactate Dehydrogenase Isozymes: Further Kinetic Studies at High Enzyme Concentration  

Microsoft Academic Search

By competition with lactate dehydrogenase (LDH) for nicotinamide adenine dinucleotide (NAD), commonly occurring intracellular proteins, such as glyceraldehyde-3-phosphate dehydrogenase, malate dehydrogenase, and albumin, can protect LDH-1 and LDH-5 from inhibition and ternary complex formation with NAD and pyruvate. The existence of intracellular proteins that compete with LDH for NAD renders unphysiological a model for estimating the extent of intracellular LDH

Thomas Wuntch; Raymond F. Chen; Elliot S. Vesell

1970-01-01

24

NAD(H) recycling activity of an engineered bifunctional enzyme galactose dehydrogenase\\/lactate dehydrogenase  

Microsoft Academic Search

A chimeric bifunctional enzyme composing of galactose dehydrogenase (galDH; from Pseudomonas fluorescens) and lactate dehydrogenase (LDH; from Bacillus stearothermophilus) was successfully constructed. The chimeric galDH\\/LDH possessed dual characteristics of both galactose dehydrogenase and lactate dehydrogenase activities while exhibiting hexameric rearrangement with a molecular weight of approximately 400 kDa. In vitro observations showed that the chimeric enzyme was able to recycle

Virapong Prachayasittikul; Sarah Ljung; Chartchalerm Isarankura-Na-Ayudhya; Leif Bülow

2006-01-01

25

Plasmodium falciparum and Plasmodium vivax: Lactate-Dehydrogenase Activity and Its Application for in Vitro Drug Susceptibility Assay  

Microsoft Academic Search

Lactate dehydrogenase, the terminal enzyme of anerobic Embden-Meyerhoff glycolysis, plays an important role in the carbohydrate metabolism of human malaria parasites. Based on the ability of malarial lactate dehydrogenase to use 3-acetylpyridine NAD as a coenzyme in a reaction leading to the formation of pyruvate from L-lactate, the enzymatic activity of fresh clinical isolates of Plasmodium falciparum and Plasmodium vivax

L. K. Basco; F. Marquet; M. M. Makler; J. Lebras

1995-01-01

26

Determination of lactate dehydrogenase in human erythrocytes by capillary electrophoresis with electrochemical detection  

Microsoft Academic Search

Capillary electrophoresis (CE) was employed to analyze lactate dehydrogenase (LDH) in human erythrocytes using an amperometric detector with a carbon fiber micro-disk bundle electrode. LDH activity was measured by determining the amount of NADH generated by LDH through a enzyme-catalyzed reaction between NAD+ and lithium lactate. The factors influencing the enzyme-catalyzed reaction, separation and detection were examined and optimized. The

Wei Wang; Xuemei Sun; Wenrui Jin

2003-01-01

27

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

NASA Astrophysics Data System (ADS)

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 enzyme "substrate" interactions: (i) which form of the substrate system serves as the preferential substrate and (ii) which form acts to inhibit the enzyme? Thus the relative concentrations of the forms of these substrate systems (keto, hydrated, enol) may provide a form of metabolic control. In this light, the present article considers the reduction of pyruvate by lactate dehydrogenase in the presence of NADH. This reaction is inhibited by relatively high concentrations of pyruvate and the physiological significance of this inhibition has been a subject of controversy for many years. Summarized in this article are data from the literature pertaining to the interactions of keto, hydrated, and enol pyruvate with lactate dehydrogenase. Biochemistry instructors and their students are invited to review such pertinent articles so that they also may evaluate the possibility that the "substrate" inhibition of the isoenzymes in the heart muscle may be, under certain conditions, relevant as a form of metabolic control.

Meany, J. E.

2007-09-01

28

In vivo alterations in lactate dehydrogenase (LDH) and LDH isoenzymes patterns by acute carbofuran intoxication  

Microsoft Academic Search

Intoxication with an acute dose of carbofuran (1.5 mg\\/kg, sc) in male Sprague-Dawley rats evoked severe toxic manifestations characteristic of hypercholinergic preponderance with profound muscle fasciculations and convulsions during 30–60 min, lasting for about 2 h. Lactate dehydrogenase, a “biomarker” cytoplasmic enzyme catalyzing the reversible reaction of lactate-pyruvate, was represented by five electrophoretically distinct isoenzymes in the serum and tissues.

Ramesh C. Gupta; John T. Goad; Wade L. Kadel

1991-01-01

29

Genetic Variation and Relative Catalytic Efficiencies: Lactate Dehydrogenase B Allozymes of Fundulus Heteroclitus  

Microsoft Academic Search

In order to evaluate whether functional differences exist between allelic variants of a B type lactate dehydrogenase (LDH; L-lactate:NAD+ oxidoreductase, EC 1.1.1.27) in the teleost fish Fundulus heteroclitus (Linnaeus), the kinetic properties of pyruvate reduction were examined. While the pH dependence and the temperature dependence for maximal catalysis were indistinguishable among the allozymes, reaction velocities at low pyruvate concentrations were

Dennis A. Powers

1979-01-01

30

Molecular cloning and characterization of a novel lactate dehydrogenase gene from Clonorchis sinensis  

Microsoft Academic Search

From a Clonorchis sinensis adult worm cDNA library, we isolated a cDNA clone encoding a novel lactate dehydrogenase (LDH) gene which encoded a putative protein with a predicted molecular weight of 35.6 kDa. The optimum pH and temperature for the enzyme were 7.5 and 50°C in the pyruvate reduction while 11 and 80°C in the lactate oxidation reaction, respectively. CsLDH showed

Guang Yang; Chunxia Jing; Peixian Zhu; Xuchu Hu; Jin Xu; Zhongdao Wu; Xinbing Yu

2006-01-01

31

Kinetic Parameters and Lactate Dehydrogenase Isozyme Activities Support Possible Lactate Utilization by Neurons  

Microsoft Academic Search

Lactate is potentially a major energy source in brain, particularly following hypoxia\\/ischemia; however, the regulation of\\u000a brain lactate metabolism is not well understood. Lactate dehydrogenase (LDH) isozymes in cytosol from primary cultures of\\u000a neurons and astrocytes, and freshly isolated synaptic terminals (synaptosomes) from adult rat brain were separated by electrophoresis,\\u000a visualized with an activity-based stain, and quantified. The activity and

Janet O’Brien; Koffi M. Kla; Irene B. Hopkins; Elise A. Malecki; Mary C. McKenna

2007-01-01

32

Genetic Analysis with Man-Mouse Somatic Cell Hybrids: Linkage between Human Lactate Dehydrogenase B and Peptidase B Genes  

Microsoft Academic Search

Evidence of an active linkage between the human genes that control lactate dehydrogenase B and peptidase B is presented. It is also concluded that there is no link between the genes for lactate dehydrogenase A and lactate dehydrogenase B.

A. Silvana Santachiara; M. Nabholz; V. Miggiano; A. J. Darlington; W. Bodmer

1970-01-01

33

In vitro effect of some anthelmintics on lactate dehydrogenase activity of Cotylophoron cotylophorum (Digenea: Paramphistomidae)  

Microsoft Academic Search

Effects of praziquantel (PZQ), levamisole (LEV), mebendazole (MBZ), fenbendazole (FBZ) and albendazole (ABZ) on the lactate dehydrogenase (LDH) activity of Cotylophoron cotylophorum were studied in vitro. Maximum levels of inhibition of LDH catalysing both oxidation and reduction reactions were observed in PZQ- and LEV-treated worms. Similarly, benzimidazoles — MBZ, FBZ and ABZ — have also significantly inhibited the activity of

L Veerakumari; N Munuswamy

2000-01-01

34

Conformational preferences of the substrates of lactate dehydrogenase  

Microsoft Academic Search

The substrates of lactate dehydrogenase (pyruvate and lactate) as well as an inhibitor (oxamate) are characterized using ab initio methods employing the basis sets 6-31+G(d) and 6-31+G(d,p) at both RHF and MP2 levels. The first aim of these studies is to explore the conformational preferences of these molecules and to compare the minimum energy conformation with that found in crystallographic

R. K. Schmidt; J. E. Gready

2000-01-01

35

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

36

The diverse Michaelis constants and maximum velocities of lactate dehydrogenase in situ in various types of cell  

Microsoft Academic Search

Summary  The kinetics of lactate dehydrogenase in mouse cardiac muscle fibres, skeletal muscle fibres, gastric parietal cells, parotid gland ductal and acinar cells, oocytes and mouse and human hepatocytes were studied as a function of substrate concentration in sections of unfixed mouse and human tissues incubated at 37°C on lactate agarose gel films. The absorbances of the final reaction products deposited

Yoshiko Nakae; Peter J. Stoward

1994-01-01

37

Interaction Between Lactate Dehydrogenase and Tween 80 in Aqueous Solution  

Microsoft Academic Search

Purpose. The weak aqueous interaction between the protein lactate dehydrogenase (LDH) and the nonionic surfactant Tween 80 has been investigated, because weak protein-amphiphile interactions are of significant importance in pharmaceutical formulations, but are experimentally hard to determine. The system LDH\\/sodium dodecyl sulphate (SDS) was used as reference because SDS, by its strong protein binding, denatures LDH completely.

Anna Hillgren; Hans Evertsson; Maggie Aldén

2002-01-01

38

Oxygen Induced Inhibition of Mouse Brain Lactate Dehydrogenase.  

National Technical Information Service (NTIS)

The lactate dehydrogenase (LDH) activity of mouse brain homogenates was examined after exposure to hyperbaric oxygen (5763.8 mm Hg PO2) and compared to room air controls (158.8 mm Hg PO2). The effect of reduced glutathione on LDH activity after hyperbaric...

D. A. Baeyens

1975-01-01

39

New insights into salivary lactate dehydrogenase of human subjects  

Microsoft Academic Search

Lactate dehydrogenase (LDH) is a ubiquitous enzyme that plays a significant role in the clinical diagnosis of pathologic processes. The purpose of the current study was to examine LDH activity and isoenzyme profile of whole saliva and to compare it with the LDH activity of salivary glands and plasma before and after exposure to cigarette smoke (CS). The range of

Rafael M. Nagler; Sophie Lischinsky; Eric Diamond; Ifat Klein; Abraham Z. Reznick

2001-01-01

40

Selective Distribution of Lactate Dehydrogenase Isoenzymes in Neurons and Astrocytes of Human Brain  

Microsoft Academic Search

In vertebrates, the interconversion of lactate and pyruvate is catalyzed by the enzyme lactate dehydrogenase. Two distinct subunits combine to form the five tetrameric isoenzymes of lactate dehydrogenase. The LDH-5 subunit (muscle type) has higher maximal velocity (Vmax) and is present in glycolytic tissues, favoring the formation of lactate from pyruvate. The LDH-1 subunit (heart type) is inhibited by pyruvate

Philippe G. Bittar; Yves Charnay; Luc Pellerin; Constantin Bouras; Pierre J. Magistretti

1996-01-01

41

D(-)Lactic Acid Formation and D(-)Lactate Dehydrogenase in Octopus Spermatozoa  

Microsoft Academic Search

Spermatozoa of Octopus dofleini martini produce D(-)lactic acid in the course of their metabolism, and they also possess a highly active D(-)-lactate dehydrogenase. This distinguishes the octopus spermatozoa from mammalian spermatozoa which form L(+)lactic acid and have a L(+)-lactate dehydrogenase. The D(-)lactate dehydrogenase has been extracted from the octopus testis. It was shown to be a D(-)lactate:NAD oxidoreductase with electrophoretic

A. W. Martin; R. Jones; T. Mann

1976-01-01

42

Evolution of D-lactate dehydrogenase activity from glycerol dehydrogenase and its utility for D-lactate production from lignocellulose.  

PubMed

Lactic acid, an attractive, renewable chemical for production of biobased plastics (polylactic acid, PLA), is currently commercially produced from food-based sources of sugar. Pure optical isomers of lactate needed for PLA are typically produced by microbial fermentation of sugars at temperatures below 40?°C. Bacillus coagulans produces L(+)-lactate as a primary fermentation product and grows optimally at 50?°C and pH 5, conditions that are optimal for activity of commercial fungal cellulases. This strain was engineered to produce D(-)-lactate by deleting the native ldh (L-lactate dehydrogenase) and alsS (acetolactate synthase) genes to impede anaerobic growth, followed by growth-based selection to isolate suppressor mutants that restored growth. One of these, strain QZ19, produced about 90 g L(-1) of optically pure D(-)-lactic acid from glucose in < 48 h. The new source of D-lactate dehydrogenase (D-LDH) activity was identified as a mutated form of glycerol dehydrogenase (GlyDH; D121N and F245S) that was produced at high levels as a result of a third mutation (insertion sequence). Although the native GlyDH had no detectable activity with pyruvate, the mutated GlyDH had a D-LDH specific activity of 0.8 ?moles min(-1) (mg protein)(-1). By using QZ19 for simultaneous saccharification and fermentation of cellulose to D-lactate (50?°C and pH 5.0), the cellulase usage could be reduced to 1/3 that required for equivalent fermentations by mesophilic lactic acid bacteria. Together, the native B. coagulans and the QZ19 derivative can be used to produce either L(+) or D(-) optical isomers of lactic acid (respectively) at high titers and yields from nonfood carbohydrates. PMID:22065761

Wang, Qingzhao; Ingram, Lonnie O; Shanmugam, K T

2011-11-07

43

Phylogenetic Analysis of Vertebrate Lactate Dehydrogenase (LDH) Multigene Families  

Microsoft Academic Search

.   In this paper we analyzed 49 lactate dehydrogenase (LDH) sequences, mostly from vertebrates. The amino acid sequence differences\\u000a were found to be larger for a human–killifish pair than a human–lamprey pair. This indicates that some protein sequence convergence\\u000a may occur and reduce the sequence differences in distantly related species. We also examined transitions and transversions\\u000a separately for several species

Yi-Ju Li; Stephen C.-M. Tsoi

2002-01-01

44

Ligand Binding and Protein Dynamics in Lactate Dehydrogenase  

Microsoft Academic Search

Recent experimental studies suggest that lactate dehydrogenase (LDH) binds its substrate via the formation of a LDH\\/NADH·substrate encounter complex through a select-fit mechanism, whereby only a minority population of LDH\\/NADH is binding-competent. In this study, we perform molecular dynamics calculations to explore the variations in structure accessible to the binary complex with a focus on identifying structures that seem likely

J. R. Exequiel T. Pineda; Robert Callender; Steven D. Schwartz

2007-01-01

45

Immunochemical study of immunoglobulins bound to lactate dehydrogenase  

Microsoft Academic Search

Immunochemical analysis of ? type Bence Jones protein (BJP; designated as Suzuki-BJP) and IgG-? type M-protein (designated as Miki-IgG), ? type BJP (designated as Miki-BJP) which showed non-specific binding with lactate dehydrogenase (LD, EC 1.1.1.27) was carried out in two cases. When the purified LD mixed with NADH was eluted through the CNBr-Sepharose 4B coupled to Suzuki-BJP or Miki-IgG, the

Kiyotaka Fujita

1997-01-01

46

The Murine Antibody Response to Lactate Dehydrogenase-elevating Virus  

Microsoft Academic Search

SUMMARY Mice infected with lactate dehydrogenase-elevating virus (LDV) were found to produce high titres of IgG anti-LDV antibodies that remained elevated for more than l year. This response, which was T-dependent, showed a striking preponderance of IgG2a with, from one strain to another, variable proportions of IgG2b and IgG3 but always very little IgG1. The binding of these antibodies to

J. P. Coutelier; E. van Roost; P. Lambotte; J. van Snick

1986-01-01

47

Lactate dehydrogenase from adult Setaria digitata (Nematoda: Onchocercidae).  

PubMed

Lactate dehydrogenase of Setaria digitata exhibited an optimum pH of 7.0-8.0 and showed resistance to high temperature. The inhibition/activation of various anions differed in both the forward and backward directions. Filarin (a drug used in Siddha medicine) and diethylcarbamazine (DEC) inhibited pyruvate reduction rather than lactate oxidation. High pyruvate reduction:lactate oxidation at Vmax and Vmax/Km favoured pyruvate reduction in vivo. The enzyme exists as isozymes (four in the female and three in the male) and their separation depended on the percentage of gel and on pH. The mobility of the 700 X g supernatant fraction in the gel was less than that of the 10,000 X g supernatant. PMID:2781717

Banu, M J; Nellaiappan, K; Dhandayuthapani, S

1989-08-01

48

The Envelope Proteins of Lactate Dehydrogenase-Elevating Virus and Their Membrane Topography  

Microsoft Academic Search

We have studied the membrane topography and N-glycosylation of the envelope proteins of lactate dehydrogenase-elevating virus (LDV, strain P). Transcripts of open reading frames (ORFs) 2, 5, and 6 were in vitro translated in the absence and presence of microsomal membranes, and the products analyzed for molecular weight, sensitivity to endoglycosidase F\\/N-glycosidase F and proteinases, and reaction with anti-LDV antibodies.

Kay S. Faaberg; Peter G. W. Plagemann

1995-01-01

49

Use of PurifiedLyophilized HumanLactateDehydrogenase Isoenzymesin a Studyof the Measurement of LactateDehydrogenase Activity  

Microsoft Academic Search

LD-1 but poorfor LD-2 and LD-3. We suggestthat prepara- tions of human LD-1 be further investigatedas a possible referencematerial. AddItIonalKeyphrases:enzyme activity potential reference material We recently described the purification, with good yield and specific activity, of lactate dehydrogenase (iAacta- te:NAD oxidoreductase, EC 1.1.1.27; LD) isoenzymes 1, 2, and 3 from human erythrocytes (1). We suggested that these LD isoenzymes might

Deborah A. Smith; Godfrey C. Moses; A. Ralph Henderson

50

Immunological study of lactate dehydrogenase from Streptococcus mutans and evidence of common antigenic domains with lactate dehydrogenases from lactic bacteria.  

PubMed Central

Rabbit polyclonal antibodies directed against purified Streptococcus mutans L-(+)-lactate dehydrogenase reacted with the purified enzyme, giving a marked deviation of its kinetic parameters. The enzyme affinity for pyruvate or NADH decreased in the presence of antibody, the affinity for fructose 1,6-diphosphate (FDP) appeared to be slightly affected, and the cooperativity of the ligand binding was lowered. A partial protective effect was observed when the enzyme was preincubated with FDP prior to the antibody adjunction. An enzyme-linked immunosorbent assay allowed detection of a 30% decrease in enzyme-antibody fixation when FDP was added. The protective effect observed with FDP could be correlated with a conformational change induced by the activator. A decrease of antibody binding in the presence of FDP was also obtained with S. sanguis, Actinomyces viscosus, and Lactobacillus casei lactate dehydrogenases, which reflects a similar mechanism of activation among lactic bacteria. NADH did not offer any protection against antibody inhibition or fixation, and the coenzyme affinity decrease could be attributed to an indirect mechanism. On the contrary, pyruvate and the immunoglobulins apparently could compete for specific binding sites. A decrease of antibody binding was also obtained with three heterologous lactic bacterial lactate dehydrogenases, indicating a conservation of antigenic determinants implicated in the substrate binding. Images

Sommer, P; Klein, J P; Ogier, J A; Frank, R M

1986-01-01

51

Self-assembled monolayers with biospecific affinity for lactate dehydrogenase for the electroenzymatic oxidation of lactate  

Microsoft Academic Search

Surface modified gold electrodes with high biospecific affinity for NAD(H)-dependent lactate dehydrogenase have been prepared by covalent attachment of several traizine dyes to stepwise functionalized mixed alkanethiol self-assembled monolayers. The biospecific affinity of such ligand-anchored monolayers to bind submonolayer amounts of enzyme was demonstrated from the course of the protein adsorption events monitored by surface plasmon resonance. Electroenzymatic activity measurements

Daniela D. Schlereth; Rob P. H. Kooyman

1997-01-01

52

Lactate Dehydrogenase C and Energy Metabolism in Mouse Sperm1  

PubMed Central

We demonstrated previously that disruption of the germ cell-specific lactate dehydrogenase C gene (Ldhc) led to male infertility due to defects in sperm function, including a rapid decline in sperm ATP levels, a decrease in progressive motility, and a failure to develop hyperactivated motility. We hypothesized that lack of LDHC disrupts glycolysis by feedback inhibition, either by causing a defect in renewal of the NAD+ cofactor essential for activity of glyceraldehyde 3-phosphate dehydrogenase, sperm (GAPDHS), or an accumulation of pyruvate. To test these hypotheses, nuclear magnetic resonance analysis was used to follow the utilization of labeled substrates in real time. We found that in sperm lacking LDHC, glucose consumption was disrupted, but the NAD:NADH ratio and pyruvate levels were unchanged, and pyruvate was rapidly metabolized to lactate. Moreover, the metabolic disorder induced by treatment with the lactate dehydrogenase (LDH) inhibitor sodium oxamate was different from that caused by lack of LDHC. This supported our earlier conclusion that LDHA, an LDH isozyme present in the principal piece of the flagellum, is responsible for the residual LDH activity in sperm lacking LDHC, but suggested that LDHC has an additional role in the maintenance of energy metabolism in sperm. By coimmunoprecipitation coupled with mass spectrometry, we identified 27 proteins associated with LDHC. A majority of these proteins are implicated in ATP synthesis, utilization, transport, and/or sequestration. This led us to hypothesize that in addition to its role in glycolysis, LDHC is part of a complex involved in ATP homeostasis that is disrupted in sperm lacking LDHC.

Odet, Fanny; Gabel, Scott A.; Williams, Jason; London, Robert E.; Goldberg, Erwin; Eddy, Edward M.

2011-01-01

53

Isotope effects on binding of NAD+ to lactate dehydrogenase  

SciTech Connect

The isotope effect on binding (4-/sup 2/H)NAD+ and (4-/sup 3/H)NAD+ to lactate dehydrogenase has been shown to be 1.10 +/- 0.03 by whole molecule isotope ratio mass spectrometry and 1.085 +/- 0.01 by /sup 3/H//sup 14/C scintillation counting. These values demonstrate that specific interactions of the nicotinamide ring with the enzyme make the C-H bond at C-4 less stiff in the binary complex.

LaReau, R.D.; Wan, W.; Anderson, V.E.

1989-04-18

54

High brain lactate is a hallmark of aging and caused by a shift in the lactate dehydrogenase A/B ratio  

PubMed Central

At present, there are few means to track symptomatic stages of CNS aging. Thus, although metabolic changes are implicated in mtDNA mutation-driven aging, the manifestations remain unclear. Here, we used normally aging and prematurely aging mtDNA mutator mice to establish a molecular link between mitochondrial dysfunction and abnormal metabolism in the aging process. Using proton magnetic resonance spectroscopy and HPLC, we found that brain lactate levels were increased twofold in both normally and prematurely aging mice during aging. To correlate the striking increase in lactate with tissue pathology, we investigated the respiratory chain enzymes and detected mitochondrial failure in key brain areas from both normally and prematurely aging mice. We used in situ hybridization to show that increased brain lactate levels were caused by a shift in transcriptional activities of the lactate dehydrogenases to promote pyruvate to lactate conversion. Separation of the five tetrameric lactate dehydrogenase (LDH) isoenzymes revealed an increase of those dominated by the Ldh-A product and a decrease of those rich in the Ldh-B product, which, in turn, increases pyruvate to lactate conversion. Spectrophotometric assays measuring LDH activity from the pyruvate and lactate sides of the reaction showed a higher pyruvate ? lactate activity in the brain. We argue for the use of lactate proton magnetic resonance spectroscopy as a noninvasive strategy for monitoring this hallmark of the aging process. The mtDNA mutator mouse allows us to conclude that the increased LDH-A/LDH-B ratio causes high brain lactate levels, which, in turn, are predictive of aging phenotypes.

Ross, Jaime M.; Oberg, Johanna; Brene, Stefan; Coppotelli, Giuseppe; Terzioglu, Mugen; Pernold, Karin; Goiny, Michel; Sitnikov, Rouslan; Kehr, Jan; Trifunovic, Aleksandra; Larsson, Nils-Goran; Hoffer, Barry J.; Olson, Lars

2010-01-01

55

Amino Acid Sequence Homology among the 2-hydroxy Acid Dehydrogenases: Mitochondrial and Cytoplasmic Malate Dehydrogenases Form a Homologous System with Lactate Dehydrogenase  

Microsoft Academic Search

The amino acid sequence of porcine heart mitochondrial malate dehydrogenase (mMDH; L-malate:NAD+ oxidoreductase, EC 1.1.1.37) has been compared with the sequences of six different lactate dehydrogenases (LDH; L-lactate:NAD+ oxidoreductase, EC 1.1.1.27) and with the ``x-ray'' sequence of cytoplasmic malate dehydrogenase (sMDH). The main points are that (i) all thre enzymes are homologous; (ii) invariant residues in the catalytic center of

Jens J. Birktoft; Ross T. Fernley; Ralph A. Bradshaw; Leonard J. Banaszak

1982-01-01

56

D(-)Lactic Acid and D(-)Lactate Dehydrogenase in Octopus Spermatozoa  

Microsoft Academic Search

The spermatozoa of Octopus dofleini martini produce anaerobically D(-)-lactic acid and possess a very active D(-)-lactate dehydrogenase. In this respect, while resembling certain microorganisms, they differ strikingly from mammalian spermatozoa which produce L(+)-lactic acid and contain L(+)-lactate dehydrogenase.

Thaddeus Mann; Arthur W. Martin; John B. Thiersch; Cecilia Lutwak-Mann; David E. Brooks; Robert Jones

1974-01-01

57

Method of Identifying an Individual Homozygous or Heterozygous for Lactate Dehydrogenase-A Deficiency.  

National Technical Information Service (NTIS)

The invention relates to a method of identifying an individual homozygous or heterozygous for lactate dehydrogenase-A deficiency comprising amplifying a DNA segment of exon 6 of the lactate dehydrogenase-A gene of the individual and analyzing products of ...

S. S. Li M. Maekawa T. Kanno K. Sudo

1991-01-01

58

L-Lactate dehydrogenase in the longitudinal muscle of the sea cucumber Sclerodactyla briareus (Echinodermata: Holothuroidea)  

Microsoft Academic Search

The infaunal holothurian Sclerodactyla briareus (Thyone briareus) is able to tolerate exposure to hypoxic conditions for over 2 days. Since the in vitro anaerobic degradation of glucose-U-C14 by longitudinal muscle preparations leads to an accumulation of labeled lactic acid, it is apparent that lactate dehydrogenase plays a key role during anoxia. Disc electrophoresis resolved one major band of lactate dehydrogenase

W. R. Ellington

1976-01-01

59

Characterization of potato (Solanum tuberosum) lactate dehydrogenase isoenzymes by affinity chromatography and hybridization  

PubMed Central

The five isoenzymes of potato (Solanum tuberasum) lactate dehydrogenase have been resolved by affinity chromatography. Mixtures of isoenzymes LDH-1 and LDH-5 dissociate and reassociate during freezing and thawing to produce five isoenzymes. These results indicate that potato lactate dehydrogenase isoenzymes are primary isoenzymes of the vertebrate type, which are composed of two subunit types.

Jervis, L.

1981-01-01

60

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

61

Isolation and Expression of Lactate Dehydrogenase Genes from Rhizopus oryzae  

PubMed Central

Rhizopus oryzae is used for industrial production of lactic acid, yet little is known about the genetics of this fungus. In this study I cloned two genes, ldhA and ldhB, which code for NAD+-dependent l-lactate dehydrogenases (LDH) (EC 1.1.1.27), from a lactic acid-producing strain of R. oryzae. These genes are similar to each other and exhibit more than 90% nucleotide sequence identity and they contain no introns. This is the first description of ldh genes in a fungus, and sequence comparisons revealed that these genes are distinct from previously isolated prokaryotic and eukaryotic ldh genes. Protein sequencing of the LDH isolated from R. oryzae during lactic acid production confirmed that ldhA codes for a 36-kDa protein that converts pyruvate to lactate. Production of LdhA was greatest when glucose was the carbon source, followed by xylose and trehalose; all of these sugars could be fermented to lactic acid. Transcripts from ldhB were not detected when R. oryzae was grown on any of these sugars but were present when R. oryzae was grown on glycerol, ethanol, and lactate. I hypothesize that ldhB encodes a second NAD+-dependent LDH that is capable of converting l-lactate to pyruvate and is produced by cultures grown on these nonfermentable substrates. Both ldhA and ldhB restored fermentative growth to Escherichia coli (ldhA pfl) mutants so that they grew anaerobically and produced lactic acid.

Skory, Christopher D.

2000-01-01

62

Site-directed mutagenesis reveals role of mobile arginine residue in lactate dehydrogenase catalysis  

Microsoft Academic Search

The binding of substrates to lactate dehydrogenases induces a marked rearrangement of the protein structure in which a `loop' of polypeptide (residues 98-110) closes over the active site of the enzyme1,2. In this rearrangement, arginine 109 (a basic residue conserved in all known lactate dehydrogenase sequences and in the homologous malate dehydrogenases3) moves 0.8 nm from a position in the

Anthony R. Clarke; Dale B. Wigley; William N. Chia; David Barstow; Tony Atkinson; J. John Holbrook

1986-01-01

63

Studies on the active center of D- and L-lactate dehydrogenases using oxamate-diaminohexyl-Sepharose affinity chromatography.  

PubMed Central

Vertebrate and invertebrate L-lactate dehydrogenases (L-lactate:NAD+ oxidoreductase, EC 1.1.1.27) are effectively bound to oxamate-diaminohexyl-Sepharose, whereas several D-lactate dehydrogenases (D-lactate:NAD+ oxidoreductase, EC 1.1.1.28) do not bind to the same Sepharose. One explanation for our findings is that the enzymes' substrate is oriented in a reversed manner in the active center of the D- and L-lactate dehydrogenases.

Tuengler, P; Stein, T N; Long, G L

1980-01-01

64

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 190g/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

2013-08-27

65

Lactate dehydrogenase and glutamate dehydrogenase activities in the circumventricular organs of rat brain following neonatal monosodium glutamate  

Microsoft Academic Search

Glutamate (glu) an excitatory neurotransmitter amino acid, is present in high concentrations in the mammalian central nervous system and is the most abundant amino acid in our daily diet. In the present study the activities of lactate dehydrogenase (LDH) and glutamate dehydrogenase (GDH) were evaluated in the circumventricular organs (CVO) of the brain in 25-day-old rats following MSG administration at

M. Bawari; G. N. Babu; M. M. Ali; U. K. Misra; S. V. Chandra

1993-01-01

66

Quinone-dependent D-lactate dehydrogenase Dld (Cg1027) is essential for growth of Corynebacterium glutamicum on D-lactate  

Microsoft Academic Search

BACKGROUND: Corynebacterium glutamicum is able to grow with lactate as sole or combined carbon and energy source. Quinone-dependent L-lactate dehydrogenase LldD is known to be essential for utilization of L-lactate by C. glutamicum. D-lactate also serves as sole carbon source for C. glutamicum ATCC 13032. RESULTS: Here, the gene cg1027 was shown to encode the quinone-dependent D-lactate dehydrogenase (Dld) by

Osamu Kato; Jung-Won Youn; K Corinna Stansen; Daisuke Matsui; Tadao Oikawa; Volker F Wendisch

2010-01-01

67

D- and L-lactate dehydrogenases during invertebrate evolution  

PubMed Central

Background The L-lactate and D-lactate dehydrogenases, which are involved in the reduction of pyruvate to L(-)-lactate and D(+)-lactate, belong to evolutionarily unrelated enzyme families. The genes encoding L-LDH have been used as a model for gene duplication due to the multiple paralogs found in eubacteria, archaebacteria, and eukaryotes. Phylogenetic studies have suggested that several gene duplication events led to the main isozymes of this gene family in chordates, but little is known about the evolution of L-Ldh in invertebrates. While most invertebrates preferentially oxidize L-lactic acid, several species of mollusks, a few arthropods and polychaetes were found to have exclusively D-LDH enzymatic activity. Therefore, it has been suggested that L-LDH and D-LDH are mutually exclusive. However, recent characterization of putative mammalian D-LDH with significant similarity to yeast proteins showing D-LDH activity suggests that at least mammals have the two naturally occurring forms of LDH specific to L- and D-lactate. This study describes the phylogenetic relationships of invertebrate L-LDH and D-LDH with special emphasis on crustaceans, and discusses gene duplication events during the evolution of L-Ldh. Results Our phylogenetic analyses of L-LDH in vertebrates are consistent with the general view that the main isozymes (LDH-A, LDH-B and LDH-C) evolved through a series of gene duplications after the vertebrates diverged from tunicates. We report several gene duplication events in the crustacean, Daphnia pulex, and the leech, Helobdella robusta. Several amino acid sequences with strong similarity to putative mammalian D-LDH and to yeast DLD1 with D-LDH activity were found in both vertebrates and invertebrates. Conclusion The presence of both L-Ldh and D-Ldh genes in several chordates and invertebrates suggests that the two enzymatic forms are not necessarily mutually exclusive. Although, the evolution of L-Ldh has been punctuated by multiple events of gene duplication in both vertebrates and invertebrates, a shared evolutionary history of this gene in the two groups is apparent. Moreover, the high degree of sequence similarity among D-LDH amino acid sequences suggests that they share a common evolutionary history.

2008-01-01

68

Fabrication of lactate biosensor based on lactate dehydrogenase immobilized on cerium oxide nanoparticles.  

PubMed

An electrochemical biosensor was developed to determine lactate that plays an important role in clinical diagnosis, fermentation and food quality analysis. Abnormal concentration of lactate has been related to diseases such as hypoxia, acute heart disorders, lactic acidosis, muscle fatigue and meningitis. Also, lactate concentration in blood helps to evaluate the athletic performance in sports. The main aim of the work is to fabricate NADH/LDH/Nano-CeO2/GCE bio-electrode for sensing lactate in human blood samples. Toward this, CeO2 nanoparticles were synthesized by a hydroxide mediated approach using cerium nitrate hexahydrate (Ce(NO3)3?6H2O) and NaOH as precursors. X-ray diffraction (XRD) and Field Emission Scanning Electron Microscopy (FE-SEM) studies were carried out to determine the structural and morphological characteristics of CeO2 nanoparticles. XRD pattern indicated the formation of highly crystalline CeO2 nanoparticles with face centered cubic structure. The FE-SEM studies revealed the formation of nanospherical particles of size 29.73±2.59nm. The working electrode was fabricated by immobilizing nicotinamide adenine dinucleotide (NADH) and lactate dehydrogenase (LDH) on GCE surface with CeO2 nanoparticles as an interface. Electrochemical studies were carried out through cyclic voltammetry using a three electrode system with NADH/LDH/NanoCeO2/GCE as a working electrode, Ag/AgCl saturated with 0.1M KCl as a reference electrode and Pt wire as a counter electrode. From the amperometric study, the linearity was found to be in the range of 0.2-2mM with the response time of less than 4s. PMID:24034216

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

2013-08-22

69

NAD-Independent L-Lactate Dehydrogenase Is Required for L-Lactate Utilization in Pseudomonas stutzeri SDM  

PubMed Central

Background Various Pseudomonas strains can use l-lactate as their sole carbon source for growth. However, the l-lactate-utilizing enzymes in Pseudomonas have never been identified and further studied. Methodology/Principal Findings An NAD-independent l-lactate dehydrogenase (l-iLDH) was purified from the membrane fraction of Pseudomonas stutzeri SDM. The enzyme catalyzes the oxidation of l-lactate to pyruvate by using FMN as cofactor. After cloning its encoding gene (lldD), l-iLDH was successfully expressed, purified from a recombinant Escherichia coli strain, and characterized. An lldD mutant of P. stutzeri SDM was constructed by gene knockout technology. This mutant was unable to grow on l-lactate, but retained the ability to grow on pyruvate. Conclusions/Significance It is proposed that l-iLDH plays an indispensable function in Pseudomonas l-lactate utilization by catalyzing the conversion of l-lactate into pyruvate.

Dou, Peipei; Ma, Cuiqing; Li, Lixiang; Kong, Jian; Xu, Ping

2012-01-01

70

Isolation and Partial Characterization of Lactate Dehydrogenase Isozymes from Normal and Variant Rainbow Trout Livers.  

National Technical Information Service (NTIS)

The homotetrameric lactate dehydrogenase (LDH) isozymes, B2 prime and B2 double prime, from normal and variant rainbow trout livers have been purified to homogeneity by affinity chromatography using a Sepharose-linked oxamate ligand. The two isozymes have...

Y. H. J. Kao

1977-01-01

71

The Genetics of Lactate Dehydrogenase Polymorphism in Rainbow Trout, 'Salmo gairdneri'.  

National Technical Information Service (NTIS)

Two electrophoretic forms (phenotypes) of lactate dehydrogenase (LDH) were discovered in blood serum of a group of Beity strain domestic rainbow trout. Experimental crosses were made to determine the genetics of phenotypes. Results of crosses showed that ...

G. A. Stillings

1974-01-01

72

Purification and biochemical studies of lactate dehydrogenase-X from mouse  

Microsoft Academic Search

Lactate dehydrogenase-X from testes of several rodent species was purified to homogeneity by an 8-(6-aminohexyl)-amino-AMP-Sepharose affinity column. In the case of mouse, the testicle extracts was first heated to 60° for fifteen minutes before the passage through the affinity column. A biospecific elution with reduced NAD+-pyruvate adduct resulted in a homogeneous preparation of lactate dehydrogenase-X. A similar procedure was also

Chi-Yu Lee; Bruna Pegoraro; Joyce L. Topping; James H. Yuan

1977-01-01

73

The Approach to the Michaelis Complex in Lactate Dehydrogenase: The Substrate Binding Pathway  

Microsoft Academic Search

We examine here the dynamics of forming the Michaelis complex of the enzyme lactate dehydrogenase by characterizing the binding kinetics and thermodynamics of oxamate (a substrate mimic) to the binary lactate dehydrogenase\\/NADH complex over multiple timescales, from nanoseconds to tens of milliseconds. To access such a wide time range, we employ standard stopped-flow kinetic approaches (slower than 1ms) and laser-induced

Sebastian McClendon; Nick Zhadin; Robert Callender

2005-01-01

74

Usefulness of lactate dehydrogenase and its isoenzymes as indicators of lung damage or inflammation  

Microsoft Academic Search

Usefulness of lactate dehydrogenase and its isoenzymes as indicators of lung damage or inflammation. M. Drent, N.A.M. Cobben, R.F. Henderson, E.F.M. Wouters, M. van Dieijen- Visser. ?ERS Journals Ltd 1996. ABSTRACT: This review describes the usefulness of monitoring the activity level of lactate dehydrogenase (LDH) and its isoenzyme pattern as indicators of patho- logical conditions in the lungs, such as

M. Drent; N. A. M. Cobben; R. F. Henderson; E. F. M. Wouters; M. van Dieijen-Visser

1996-01-01

75

Pressure-Adaptive Differences in Lactate Dehydrogenases of Congeneric Fishes Living at Different Depths  

Microsoft Academic Search

The muscle-type (M4) lactate dehydrogenases of Sebastolobus altivelis, a deep-water scorpaenid, and S. alascanus, a shallower species, are electrophoretically indistinguishable, yet differ in pressure sensitivities. The lactate dehydrogenase of S. altivelis exhibits lower pressure sensitivities of substrate and coenzyme binding and catalytic rate. Such apparently pressure-adaptive kinetic properties may be important for establishing species depth zonation patterns in the ocean.

Joseph Siebenaller; George N. Somero

1978-01-01

76

The rapid purification of lactate dehydrogenase from alcaligenes eutrophus in a two-step procedure  

Microsoft Academic Search

A NAD-linked L(+)-lactate dehydrogenase (EC 1.1.1.27) was isolated from Alcaligenes eutrophus N9A. During purification advantage was taken of the high affinity of MatrexTM Gel Green A for this enzyme in crude extracts. One ml of this medium adsorbed 2660 U lactate dehydrogenase if 129 ml crude extract containing 2,480 mg protein were applied onto the column, as determined by frontal

Alexander Steinbfichel; Hans G. Schlegel

1983-01-01

77

Plasmodium falciparum: Enhanced soluble expression, purification and biochemical characterization of lactate dehydrogenase  

Microsoft Academic Search

Plasmodium lactate dehydrogenase (pLDH), owing to unique structural and kinetic properties, is a well known target for antimalarial compounds. To explore a new approach for high level soluble expression of Plasmodium falciparum lactate dehydrogenase (PfLDH) in E. coli, PfLDH encoding sequence was cloned into pQE-30 Xa vector. When transformed E. coli SG13009 cells were induced at 37°C with 0.5mM isopropyl

Ritu Berwal; Natarajan Gopalan; Kshitij Chandel; G. B. K. S. Prasad; Shri Prakash

2008-01-01

78

Affinity isolation of a cold-adapted enzyme: lactate dehydrogenase from Bacillus psychrosaccharolyticus  

Microsoft Academic Search

A simple, economical and rapid affinity chromatography procedure with dyes as the ligand has been described for the one-step purification of a cold-adapted lactate dehydrogenase. Non-specific elution of Procion blue H-ERD-modified Sepharose yielded homogeneous preparations of lactate dehydrogenase both in column based procedures and in batch wise operations. Low operational temperatures resulted in the enhanced binding of the enzyme to

R. Nandakumar; B. Mattiasson

1998-01-01

79

Are cell redox or lactate dehydrogenase kinetics responsible for the absence of gluconeogenesis from lactate in sea raven, hepatocytes?  

Microsoft Academic Search

Previous studies have reported very low rates of gluconeogenesis from lactate in sea raven (Hemitripterus americanus) hepatocytes compared to other teleosts studied. This study examines whether hepatic cell redox or lactate dehydrogenase\\u000a (LDH) characteristics may explain this observation. Sea raven hepatic optimal LDH activities (pyruvate reductase direction)\\u000a were more than 40 times less compared with rainbow trout liver values (40

Glen D. Foster; J. Zhang; T. W. Moon

1994-01-01

80

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

PubMed

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, potentially leading to lower cell growth and lower therapeutic protein production during manufacturing. Lactate dehydrogenase (LDH) is an enzyme that catalyzes the conversion of the substrate, pyruvate, into lactate and many factors including pyruvate concentration modulate LDH activity. Alternately, pyruvate can be converted to acetyl-CoA by pyruvate dehydrogenases (PDHs), to be metabolized in the TCA cycle. PDH activity is inhibited when phosphorylated by pyruvate dehydrogenase kinases (PDHKs). In this study, we knocked down the gene expression of lactate dehydrogenase A (LDHa) and PDHKs to investigate the effect on lactate metabolism and protein production. We found that LDHa and PDHKs can be successfully downregulated simultaneously using a single targeting vector carrying small inhibitory RNAs (siRNA) for LDHa and PDHKs. Moreover, our fed-batch shake flask evaluation data using siRNA-mediated LDHa/PDHKs knockdown clones showed that downregulating LDHa and PDHKs in CHO cells expressing a therapeutic monoclonal antibody reduced lactate production, increased specific productivity and volumetric antibody production by approximately 90%, 75% and 68%, respectively, without appreciable impact on cell growth. Similar trends of lower lactate level and higher antibody productivity on average in siRNA clones were also observed from evaluations performed in bioreactors. PMID:21392546

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

2011-03-30

81

Ligand binding and protein dynamics in lactate dehydrogenase.  

PubMed

Recent experimental studies suggest that lactate dehydrogenase (LDH) binds its substrate via the formation of a LDH/NADH.substrate encounter complex through a select-fit mechanism, whereby only a minority population of LDH/NADH is binding-competent. In this study, we perform molecular dynamics calculations to explore the variations in structure accessible to the binary complex with a focus on identifying structures that seem likely to be binding-competent and which are in accord with the known experimental characterization of forming binding-competent species. We find that LDH/NADH samples quite a range of protein conformations within our 2.148 ns calculations, some of which yield quite facile access of solvent to the active site. The results suggest that the mobile loop of LDH is perhaps just partially open in these conformations and that multiple open conformations, yielding multiple binding pathways, are likely. These open conformations do not require large-scale unfolding/melting of the binary complex. Rather, open versus closed conformations are due to subtle protein and water rearrangements. Nevertheless, the large heat capacity change observed between binding-competent and binding-incompetent can be explained by changes in solvation and an internal rearrangement of hydrogen bonds. We speculate that such a strategy for binding may be necessary to get a ligand efficiently to a binding pocket that is located fairly deep within the protein's interior. PMID:17483170

Pineda, J R Exequiel T; Callender, Robert; Schwartz, Steven D

2007-05-04

82

Cloning and polymorphisms of yak lactate dehydrogenase B gene.  

PubMed

The main objective of this work was to study the unique polymorphisms of the lactate dehydrogenase-1 (LDH1) gene in yak (Bos grunniens). Native polyacrylamide gel electrophoresis revealed three phenotypes of LDH1 (a tetramer of H subunit) in yak heart and longissimus muscle extracts. The corresponding gene, ldhb, encoding H subunits of three LDH1 phenotypes was obtained by RT-PCR. A total of six nucleotide differences were detected in yak ldhb compared with that of cattle, of which five mutations cause amino acid substitutions. Sequence analysis shows that the G896A and C689A, mutations of ldhb gene, result in alterations of differently charged amino acids, and create the three phenotypes (F, M, and S) of yak LDH1. Molecular modeling of the H subunit of LDH indicates that the substituted amino acids are not located within NAD+ or substrate binding sites. PCR-RFLP examination of G896A mutation demonstrated that most LDH1-F samples are actually heterozygote at this site. These results help to elucidate the molecular basis and genetic characteristic of the three unique LDH1 phenotypes in yak. PMID:23739677

Wang, Guosheng; Zhao, Xingbo; Zhong, Juming; Cao, Meng; He, Qinghua; Liu, Zhengxin; Lin, Yaqiu; Xu, Yaou; Zheng, Yucai

2013-06-05

83

Regulation of murine lactate dehydrogenase C (Ldhc) gene expression.  

PubMed

Expression of Ldhc begins with the onset of meiosis in male germ cells and continues throughout spermatogenesis. Transcriptional regulatory mechanisms, especially in primary spermatocytes, are poorly described because of the lack of a reliable cell culture system. We constructed mouse transgenics and transfected germ cells in situ to study expression of the testis-specific isozyme of lactate dehydrogenase (LDH). From previous work, we determined that a 100-bp Ldhc core promoter contained potential cis regulatory elements, including a palindrome (-21 to +10), GC box (-70 to -65), and cAMP-responsive element (CRE) sites (-53 to -49, -39 to -35). We provide here the demonstration of a functional role for these sequences by expression of mutated transgenes in vivo. Our results reveal for the first time that mutation of the GC box does not abolish promoter activity, which remains testis-specific. Mutation of GC box or CRE sites resulted in a 73% and 74% reduction in promoter activity, respectively, in a transient transfection of germ cells in vivo by electroporation; the combination of GC box and CRE site mutations eliminates promoter activity. Therefore, we conclude that simultaneous occupancy of the GC box and CRE sites in the core promoter is necessary for full expression of Ldhc in the testis. PMID:18057313

Tang, HuangHui; Kung, Aisha; Goldberg, Erwin

2007-12-05

84

Multiple molecular forms of lactate dehydrogenase and glucose 6-phosphate dehydrogenase in normal and abnormal human breast tissues.  

PubMed

Normal and abnormal breast samples of women were analyzed for multiple molecular forms of lactate dehydrogenase and glucose 6-phosphate dehydrogenase, using acrylamide disc gel electrophoresis and specific enzyme staining techniques for separation and quantitation. Infiltrating ductal carcinomas demonstrated a significant increase (three to six-fold) in the proportion of LDH-5 compared to samples of normal breast, fibrocystic disease and fibroadenoma, indicative of a shift toward the muscle-type lactate dehydrogenase in neoplasia. For glucose 6-phosphate dehydrogenase, carcinomas were found to contain increased proportions of the fastest migrating species, G6PD-I. Total enzyme activity/mg DNA was elevated in meoplastic tissues. Little or no alteration in isoenzyme profiles could be related to menopausal status of the patient. PMID:177177

Hilf, R; Rector, W D; Orlando, R A

1976-04-01

85

Purification and Kinetic Characterisation of Lactate Dehydrogenase from Dioscorea cayenensis Tuber  

Microsoft Academic Search

Lactate dehydrogenase from yellow yam tuber (Dioscorea cayenensis Lam.) was isolated and purified using various chromatographic methods and electrophoresis. Only one form of the enzyme obtained, which obeyed Michaelis-Menten kinetics, was activated by Mg2+ and Ca2+ and inhibited by nucleotides and PEP. AMP, which activated the enzyme in the direction of pyruvate reduction, inhibited it in the direction of lactate

U. Oluoha

2001-01-01

86

Malate\\/lactate dehydrogenase in mollicutes: evidence for a multienzyme protein  

Microsoft Academic Search

The malate (MDH) and lactate (LDH) dehydrogenases belong to the homologous class of 2-ketoacid dehydrogenases. The specificity for their respective substrates depends on residues differing at two or three regions within each molecule. Theoretical peptide-mass fingerprinting and PROSITE analysis of nine MDH and six LDH molecules were used to describe conserved sites related to function. A unique LDH is described

Stuart J Cordwell; David J Basseal; J. Dennis Pollack; Ian Humphery-Smith

1997-01-01

87

Genetic variation and relative catalytic efficiencies: lactate dehydrogenase B allozymes of Fundulus heteroclitus.  

PubMed Central

In order to evaluate whether functional differences exist between allelic variants of a B type lactate dehydrogenase (LDH; L-lactate:NAD+ oxidoreductase, EC 1.1.1.27) in the teleost fish Fundulus heteroclitus (Linnaeus), the kinetic properties of pyruvate reduction were examined. While the pH dependence and the temperature dependence for maximal catalysis were indistinguishable among the allozymes, reaction velocities at low pyruvate concentrations were significantly different. At pH values below 8.00, the LDH-BbBb allozyme showed a greater reaction rate at lower temperatures (e.g., 10 degrees C) than LDH-BaBa. The phenomenon was reversed at higher temperatures (e.g., greater than 25 degrees C) for pH values between 6.50 and 7.00. The rates for the heterozygous phenotype, LDH-BaBb, were not the arithmetic average of the two homotetrameric allozymes. When reaction rates were compared at constant relative alkalinity, that is, a constant [OH-]/[H+] ratio, the findings were similar. The differences in the temperature dependence and the pH dependence for pyruvate reduction found between the LDH-B allozymes may reflect a selective adaptation and help explain the geographical variation in the Ldh-B gene frequencies of F. heteroclitus.

Place, A R; Powers, D A

1979-01-01

88

Overexpression of monocarboxylate transporter and lactate dehydrogenase alters insulin secretory responses to pyruvate and lactate in ? cells  

PubMed Central

Previous investigations revealed low activities of lactate dehydrogenase (LDH) and plasma membrane monocarboxylate transporters (MCT) in the pancreatic ? cell. In this study the significance of these characteristics was explored by overexpressing type A LDH (LDH-A) and/or type 1 MCT (MCT-1) in the clonal INS-1 ? cells and isolated rat islets. Inducible overexpression of LDH-A resulted in an 87-fold increase in LDH activity in INS-1 cells. Adenovirus-mediated overexpression of MCT-1 increased lactate transport activity 3.7-fold in INS-1 cells. Although overexpression of LDH-A, and/or MCT-1 did not affect glucose-stimulated insulin secretion, LDH-A overexpression resulted in stimulation of insulin secretion even at a low lactate concentration with a concomitant increase in its oxidation in INS-1 cells regardless of MCT-1 co-overexpression. Adenovirus-mediated overexpression of MCT-1 caused an increase in pyruvate oxidation and conferred pyruvate-stimulated insulin release to isolated rat islets. Although lactate did not stimulate insulin secretion from control or MCT-1–overexpressing islets, co-overexpression of LDH-A and MCT-1 evoked lactate-stimulated insulin secretion with a concomitant increase in lactate oxidation in rat islets. These results suggest that low expression of MCT and LDH is requisite to the specificity of glucose in insulin secretion, protecting the organism from undesired hypoglycemic actions of pyruvate and lactate during exercise and other catabolic states. J. Clin. Invest. 104:1621–1629 (1999).

Ishihara, Hisamitsu; Wang, Haiyan; Drewes, Lester R.; Wollheim, Claes B.

1999-01-01

89

Some Lactobacillus L-Lactate Dehydrogenases Exhibit Comparable Catalytic Activities for Pyruvate and Oxaloacetate  

Microsoft Academic Search

L-Lactate dehydrogenase (L-LDH; EC 1.1.1.27) and L- malate dehydrogenase (L-MDH; EC 1.1.1.37) are similar with respect to both protein structure and catalytic machinery, and they catalyze oxidation-reduction of common 2-ketoacids and L-2-hydroxyacids with NAD as the coenzyme (1, 19, 26). Nev- ertheless, unless artificially modified, most L-LDHs and L- MDHs strictly discriminate their own substrates, pyruvate (L-lactate) and oxaloacetate (L-malate),

KAZUHITO ARAI; TAKEO KAMATA; HIROYUKI UCHIKOBA; SHINYA FUSHINOBU; HIROSHI MATSUZAWA; HAYAO TAGUCHI

2001-01-01

90

Molecular Evolution Within the L-Malate and L-Lactate Dehydrogenase SuperFamily  

Microsoft Academic Search

.   The NAD(P)-dependent malate (L-MalDH) and NAD-dependent lactate (L-LDH) form a large super-family that has been characterized\\u000a in organisms belonging to the three domains of life. In the first part of this study, the group of [LDH-like] L-MalDH, which\\u000a are malate dehydrogenases resembling lactate dehydrogenase, were analyzed and clearly defined with respect to the other enzymes.\\u000a In the second part,

Dominique Madern

2002-01-01

91

Domain Closure, Substrate Specificity and Catalysis of d-Lactate Dehydrogenase from Lactobacillus bulgaricus  

Microsoft Academic Search

NAD-dependent Lactobacillus bulgaricusd-Lactate dehydrogenase (d-LDHb) catalyses the reversible conversion of pyruvate into d-lactate. Crystals of d-LDHb complexed with NADH were grown and X-ray data collected to 2.2Å. The structure of d-LDHb was solved by molecular replacement using the dimeric Lactobacillus helveticusd-LDH as a model and was refined to an R-factor of 20.7%. The two subunits of the enzyme display strong

Adelia Razeto; Sunil Kochhar; Herbert Hottinger; Miroslava Dauter; Keith S Wilson; Victor S Lamzin

2002-01-01

92

Identification and characterization of a hypoxically induced maize lactate dehydrogenase gene  

Microsoft Academic Search

In cereal root tissue, hypoxia induces the enzyme lactate dehydrogenase (LDH); (S)-lactate:NADH oxidoreductase, EC 1.1.1.27). In barley, both biochemical and genetic data indicate that five isozymes are induced under hypoxia. These isozymes are tetramers and arise from the random association of the products of two Ldh genes. The induction of LDH activity in root tissue has been shown to be

Allen G. Good; David H. Paetkau

1992-01-01

93

Toxoplasma gondii expresses two distinct lactate dehydrogenase homologous genes during its life cycle in intermediate hosts  

Microsoft Academic Search

Two Toxoplasma gondii genes were characterized that are differentially expressed during the parasite's life cycle. The genes named LDH1 and LDH2, respectively, encode polypeptides similar to the enzyme lactate dehydrogenase (LDH; l-lactate:NAD+ oxidoreductase, EC 1.1.1.27) from a variety of organisms. They show 64.0% nucleotide identity in the coding region and both have an intron at the same relative position. The

Shumin Yang; Stephen F Parmley

1997-01-01

94

Hypoxically Inducible Barley Lactate Dehydrogenase: cDNA Cloning and Molecular Analysis  

Microsoft Academic Search

In the roots of barley and other cereals, hypoxia induces a set of five isozymes of L-lactate dehydrogenase [LDH; (S)-lactate:NADH oxidoreductase, EC 1.1.1.27]. Biochemical and genetic data indicate that the five LDH isozymes are tetramers that arise from random association of the products of two Ldh loci. To investigate this system, cDNA clones of LDH were isolated from a lambdagt11

David Hondred; Andrew D. Hanson

1990-01-01

95

The Archaeoglobus fulgidus D-Lactate Dehydrogenase Is a Zn21 Flavoprotein  

Microsoft Academic Search

Archaeoglobus fulgidus, a hyperthermophilic, archaeal sulfate reducer, is one of the few organisms that can utilize D-lactate as a sole source for both carbon and electrons. The A. fulgidus open reading frame, AF0394, which is predicted to encode a D-(2)-lactate dehydrogenase (Dld), was cloned, and its product was expressed in Escherichia coli as a fusion with the maltose binding protein

DAVID W. REED; PATRICIA L. HARTZELL

1999-01-01

96

Differentiation-dependent Increases in Lactate Dehydrogenase Activity and Isoenzyme Expression in Rabbit Corneal Epithelial Cells  

Microsoft Academic Search

Lactate dehydrogenase (LDH) and glucose-6-phosphate dehydrogenase (G-6-PDH) activities were studied during corneal epithelial growth and differentiation in cell culture. LDH and G-6-PDH activities increased up to 60 and 150-fold, respectively, when corneal epithelial cells constituted a differentiated four to five layered epithelium; these increases showed a similar time-course to the expression of K3 keratin. Immunostaining experiments showed that in growing

Miriam Hernández-Quintero; Refugio Garc??a-Villegas; Federico Castro-Muñozledo

2002-01-01

97

Large scale production of d-lactate dehydrogenase for the stereospecific reduction of pyruvate and phenylpyruvate  

Microsoft Academic Search

To initiate studies of the stereospecific reduction of pyruvate and phenylpyruvate to the corresponding d-2-hydroxyacids a limited screening was carried out for microorganisms possessing a high NADH-dependet d-lactate dehydrogenase activity. Lactobacillus confusus was found to produce the desired dehydrogenase, which showed also relatively high activity towards phenylpyruvate, so this strain was selected for large scale production of the enzyme.

Werner Hummel; Horst Schütte; Maria-Regina Kula

1983-01-01

98

Purification, properties and DNA sequence of the D-lactate dehydrogenase from Leuconostoc mesenteroides subsp. cremoris  

Microsoft Academic Search

The complete sequence of the D-lactate dehydrogenase (D-ldh) gene from Leuconostoc mesenteroides subsp. cremoris, cloned in Escherichia coli, were determined. The deduced amino acid sequence showed homologies with all members of the D-specific-2-hydroxyacid dehydrogenase family. Furthermore, the essential residues detected so far as being involved in catalysis were also conserved. Purification of the enzyme revealed physico-chemical properties corresponding to those

V Dartois; V Phalip; P Schmitt; C Diviès

1995-01-01

99

Immobilization of lactate dehydrogenase on electrochemically prepared polypyrrole–polyvinylsulphonate composite films for application to lactate biosensors  

Microsoft Academic Search

The immobilization of lactate dehydrogenase (LDH) on electrochemically polymerized polypyrrole–polyvinylsulphonate (PPY–PVS) films has been accomplished via cross-linking technique using glutaraldehyde. The characterization of the LDH-immobilized PPY–PVS films has been carried out using FTIR and cyclic voltammetry. These PPY–PVS–LDH electrodes are shown to have a detection limit of 1×10?4 M, a response time of about 40 s, and a shelf-life of

Asha Chaubey; Manju Gerard; Rahul Singhal; V. S Singh; B. D Malhotra

2001-01-01

100

Kinetic properties of two transaminases and lactate dehydrogenase of Biomphalaria alexandrina snails, intermediate hosts of Schistosoma mansoni.  

PubMed

Aspartate (AST) and alanine (ALT) aminotransferase together with lactate dehydrogenase (LD) from the tissue homogenate of the Biomphalaria alexandrina snails, were partially characterized by measuring the Michaelis constant (km) and the maximum velocity (Vmax). The isoenzymatic pattern of lactate dehydrogenase was investigated through polyacrylamide gel electrophoresis. PMID:2279260

Nabih, I; el Dardiri, Z; el Ansary, A; Ahmed, S A

1990-01-01

101

Functional Replacement of the Escherichia colid-(-)-Lactate Dehydrogenase Gene (ldhA) with the l-(+)-Lactate Dehydrogenase Gene (ldhL) from Pediococcus acidilactici†  

PubMed Central

The microbial production of l-(+)-lactic acid is rapidly expanding to allow increased production of polylactic acid (PLA), a renewable, biodegradable plastic. The physical properties of PLA can be tailored for specific applications by controlling the ratio of l-(+) and d-(?) isomers. For most uses of PLA, the l-(+) isomer is more abundant. As an approach to reduce costs associated with biocatalysis (complex nutrients, antibiotics, aeration, product purification, and waste disposal), a recombinant derivative of Escherichia coli W3110 was developed that contains five chromosomal deletions (focA-pflB frdBC adhE ackA ldhA). This strain was constructed from a d-(?)-lactic acid-producing strain, SZ63 (focA-pflB frdBC adhE ackA), by replacing part of the chromosomal ldhA coding region with Pediococcus acidilactici ldhL encoding an l-lactate dehydrogenase. Although the initial strain (SZ79) grew and fermented poorly, a mutant (SZ85) was readily isolated by selecting for improved growth. SZ85 exhibited a 30-fold increase in l-lactate dehydrogenase activity in comparison to SZ79, functionally replacing the native d-lactate dehydrogenase activity. Sequencing revealed mutations in the upstream, coding, and terminator regions of ldhL in SZ85, which are presumed to be responsible for increased l-lactate dehydrogenase activity. SZ85 produced l-lactic acid in M9 mineral salts medium containing glucose or xylose with a yield of 93 to 95%, a purity of 98% (based on total fermentation products), and an optical purity greater than 99%. Unlike other recombinant biocatalysts for l-lactic acid, SZ85 remained prototrophic and is devoid of plasmids and antibiotic resistance genes.

Zhou, Shengde; Shanmugam, K. T.; Ingram, L. O.

2003-01-01

102

[Lactate dehydrogenase isoenzymes in the eye, cardiac and skeletal muscles of several decapods].  

PubMed

Properties of lactate dehydrogenase (LDH) in the eye, heart and muscles of Hemigrapsus sanguineus, Paralithodes camtschatica, Erimacrus isenbeckii, Pandalus latirastrus, Pagurus brachiomastus have been studied with acrylamide gel electrophoresis and kinetics analysis. LDH in all the tissues of all the representatives studied was found to be specific for L-pyruvate and lactate; it migrated in electrophoresis as a single band revealing low mobility towards anode. The isoenzyme from P. camtschatica and P. latirastrus differed from the isoenzymes of other animals studied by higher mobility towards anode that reflected higher negative value of its total charge. The LDH isoenzymes in all the animals studied resembled the A4 (LDH5) of the vertebrates being unstable to the denaturing action of high temperature and being unaffected by high concentrations of pyruvate up to 1.0.10-3M. On the other hand, in conrast to the A4 of mammals, the LDH in question displayed enhancement of the reaction rate and decrease of the Km values upon increase in the NAD+ and NAD.H concentrations both in the presence of high or low lactate and pyruvate concentrations. The isoenzymes displayed catalytic activity also in the presence of NADP, the Km values for pyruvate in the presence of equimolar (2.25 mM) concentrations of NAD.H or NADP.H were practically identical and were found to be within the limits of 14-26.10-5 M. Molecular weight of the LDH studied assessed by the gel filtration method was found to be 130-140,000. It is suggested that the LDH isoenzyme from the representatives of the decapod crayfish studied is homologous in its certain properties to the homotetrameric A4 form of the vertebrates. PMID:1020551

Cherny?, V G; Chizhevich, E P; Shukoliukov, S A

103

INFLUENCE OF STEROID IMPLANTATION AND SUPPLEMENTATION ON PERFORMANCE AND LACTATE DEHYDROGENASE ACTIVITY IN STEERS GRAZING BERMUDAGRASS  

Technology Transfer Automated Retrieval System (TEKTRAN)

Forty-five steers (BW = 246 ± 5.4 kg) were randomly allocated to one of three paddocks of bermudagrass [Cynodon dactylon (L.) Pers] to determine the effects of timing of steroid implantation and supplementation on average daily gain and lactate dehydrogenase (LDH) activity. Steers received either n...

104

Ontogeny of lactate dehydrogenase isozymes in chicken-quail hybrid embryos  

Microsoft Academic Search

The ontogeny of lactate dehydrogenase (LDH) isozymes was examined in avian hybrids and compared with the isozyme patterns of the parental species. Hybrids were obtained by crossing female Japanese quail (Coturnix coturnix japonica) with male domestic chickens (Gallus gallus domesticus). By use of starch gel electrophoresis and an enzyme-specific stain, traces of embryonic paternally derived LDH were detected in unincubated

Peter G. Meyerhof; Leslie E. Haley

1975-01-01

105

Semiempirical calculations of the oxygen equilibrium isotope effect on binding of oxamate to lactate dehydrogenase  

Microsoft Academic Search

Semiempirical methods have been used in an attempt to predict theoretically the experimentally observed value of 0.9840 for the oxygen isotope effect on binding of oxamate to lactate dehydrogenase. The overall strategy involved vibrational analysis of oxamate in two different environments; that of the active site residues and in aqueous solution. The comparison of calculated values with the experimentally determined

Ewa Gawlita; Vernon E. Anderson; Piotr Paneth

1994-01-01

106

Docking Studies on the Binding of Quinoline Derivatives and Hematin to Plasmodium Falciparum Lactate Dehydrogenase  

Microsoft Academic Search

The literature has reported that ferriprotoporphyrin IX (hematin) intoxicates the malarial parasite through competition with NADH for the active site of the enzyme lactate dehydrogenase (LDH). In order to avoid this, the parasite polymerizes hematin to hemozoin. The quinoline derivatives are believed to form complexes with dimeric hematin, avoiding the formation of hemozoin and still inhibiting LDH. In order to

Wilian A. Cortopassi; Aline A. Oliveira; Ana P. Guimarães; Magdalena N. Rennó; Antoniana U. Krettli; Tanos C. C. França

2011-01-01

107

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

2012-10-12

108

On the Pathway of Forming Enzymatically Productive Ligand-Protein Complexes in Lactate Dehydrogenase  

Microsoft Academic Search

We have carried out a series of studies on the binding of a substrate mimic to the enzyme lactate dehydrogenase (LDH) using advanced kinetic approaches, which begin to provide a molecular picture of the dynamics of ligand binding for this protein. Binding proceeds via a binding-competent subpopulation of the nonligated form of the protein (the LDH\\/NADH binary complex) to form

Hua Deng; Scott Brewer; Dung M. Vu; Keith Clinch; Robert Callender; R. Brian Dyer

2008-01-01

109

The Mechanism of Recruitment of the Lactate Dehydrogenase-B\\/?-crystallin Gene by the Duck Lens  

Microsoft Academic Search

In duck, the housekeeping enzyme lactate dehydrogenase B (LDH-B) and the lens structural protein ?-crystallin are encoded by the same single copy gene. Transcription of the gene is initiated from two closely spaced start sites, at ?28 and +1. The usage of the downstream site is greatly enhanced in lens. Deletion mapping of the promoter shows that the region ?70\\/+18

Gerd A. Brunekreef; Harry J. Kraft; John G. G. Schoenmakers; Nicolette H. Lubsen

1996-01-01

110

Using Molecular Beacons To Probe Molecular Interactions between Lactate Dehydrogenase and Single-Stranded DNA  

Microsoft Academic Search

The interactions between two key macromolecular spe- cies, nucleic acids and proteins, control many important biological processes. There have been limited effective methodologies to study these interactions in real time. In this work, we have applied a newly developed molecular beacon (MB) DNA probe for the analysis of an enzyme, lactate dehydrogenase (LDH), and for the investigation of its properties

Xiaohong Fang; Jianwei Jeff Li; Weihong Tan

2000-01-01

111

Increasing the Heme-Dependent Respiratory Efficiency of Lactococcus lactis by Inhibition of Lactate Dehydrogenase  

PubMed Central

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.

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

2013-01-01

112

Influence of pH on the allosteric properties of lactate dehydrogenase activity of Phycomyces blakesleeanus.  

PubMed Central

1. Lactate dehydrogenase from mycelium of Phycomyces blakesleeanus showed positive homotropic interactions with NADH at all pH values studied (pH 5.0-7.7). The calculated values for the first and last intrinsic association constants remained unaltered with pH, in contrast with the Hill coefficient value, which varied significantly, reaching its maximum values at pH 6.0 and 7.7. This suggests the hypothesis that pH regulates these homotropic effects by changes in the value of the intermediate intrinsic association constants. 2. From pH 7.2 to 7.7 lactate dehydrogenase exhibited, likewise, positive homotropic interactions with pyruvate. There were practically no changes in the first and last intrinsic association constants and in Hill coefficient values with pH. At pH values below 7.2 (pH 5.0-6.8) the enzyme showed high substrate inhibition, which was highly dependent on pH, NADH concentration and temperature. By way of substrate inhibition pH regulates, primarily, lactate dehydrogenase activity towards pyruvate, since the homotropic effects appear not to be dependent on pH. 3. Fructose 1,6-bisphosphate is a true allosteric effector of lactate dehydrogenase of Phycomyces blakesleeanus. it decreases positive co-operativity with NADH, and on the other hand pyruvate co-operativity turns into mixed co-operativity. In addition, the effector decreases the inhibitory effect caused by pyruvate.

De Arriaga, D; Soler, J; Cadenas, E

1982-01-01

113

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

114

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...

115

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 ...

116

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...

117

Haptoglobin and lactate dehydrogenase measurements in milk for the identification of subclinically diseased udder quarters  

Microsoft Academic Search

Diagnosis of subclinical mastitis is of increasing importance and appropriate detection methods are needed. Both haptoglobin (Hp), an acute phase protein in cattle, as well as lactate dehydrogenase (LDH), an ubiquitous enzyme, can be successfully used to detect clinical mastitis. The present paper describes quantifica - tion of Hp and LDH in milk samples from healthy and subclinically diseased udder

S. Hiss; U. Mueller; A. Neu-Zahren; H. Sauerwein

118

Hypoxia and cobalt stimulate lactate dehydrogenase (LDH) activity in vascular smooth muscle cells  

Microsoft Academic Search

O2 plays a dominant role in the metabolism and viability of cells; changes in O2 supply lead to many physiological responses in the cell. Recent reports have shown that hypoxia induces the transcription of a number of genes, among them those for the glycolytic enzymes. We have investigated signalling events that may lead to enhanced activity of lactate dehydrogenase (LDH)

Hugo H. Marti; Hans H. Jung; Josef Pfeilschifter; Christian Bauer

1994-01-01

119

Design and Synthesis of New Enzymes Based on the Lactate Dehydrogenase Framework  

Microsoft Academic Search

Analysis of the mechanism and structure of lactate dehydrogenases is summarized in a map of the catalytic pathway. Chemical probes, single tryptophan residues inserted at specific sitcs and a crystal structure reveal slow movements of the protein framework that discriminate between closely related small substrates. Only small and correctly charged substrates allow the protein to engulf the substrate in an

C. R. Dunn; H. M. Wilks; D. J. Halsall; T. Atkinson; A. R. Clarke; H. Muirhead; J. J. Holbrook

1991-01-01

120

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

121

Histamine and lactate dehydrogenase (LDH) release in ischemic myocardium of the guinea-pig  

Microsoft Academic Search

Histamine has been proved to be released during myocardial infarction and ischemic arrhythmias in dogs. The aim of the present experiments was to evaluate if ischemia and reperfusion modify histamine and lactate dehydrogenase (LDH) release in isolated guinea-pig heart. The results obtained show a steady increase of LDH release both in the ischemic and reperfusion phases. The release of histamine

E. Masini; E. Giannella; S. Bianchi; P. F. Mannaioni

1987-01-01

122

A Rapid Electrophoretic Method for the Determination of the Isoenzymes of Serum Lactate Dehydrogenase  

Microsoft Academic Search

A convenient electrophoretic procedure for the separation and quantitation of lactate dehydrogenase (LDH) isoenzymes is described. The system uses polyacetate Sep- raphore Ill strips.* The areas of activity are shown by incubation with an LDH sub- strate combined with tetra-nitro-blue-tetrazolium. The reduction of the latter to the purple formazan is quantitatively related to the enzyme activity. Quantitative determination of the

Albert W. Opher; Charles S. Collier; Joseph M. Miller

123

Lactate dehydrogenase a expression is necessary to sustain rapid angiogenesis of pulmonary microvascular endothelium.  

PubMed

Angiogenesis is a fundamental property of endothelium, yet not all endothelial cells display equivalent angiogenic responses; pulmonary microvascular endothelial cells undergo rapid angiogenesis when compared to endothelial cells isolated from conduit vessels. At present it is not clear how pulmonary microvascular endothelial cells fulfill the bioenergetic demands that are necessary to sustain such rapid blood vessel formation. We have previously established that pulmonary microvascular endothelial cells utilize aerobic glycolysis to generate ATP during growth, a process that requires the expression of lactate dehydrogenase A to convert pyruvate to lactate. Here, we test the hypothesis that lactate dehydrogenase A is required for pulmonary microvascular endothelial cells to sustain rapid angiogenesis. To test this hypothesis, Tet-On and Tet-Off conditional expression systems were developed in pulmonary microvascular endothelial cells, where doxycycline is utilized to induce lactate dehydrogenase A shRNA expression. Expression of LDH-A shRNA induced a time-dependent decrease in LDH-A protein, which corresponded with a decrease in glucose consumption from the media, lactate production and cell growth; re-expression of LDH-A rescued each of these parameters. LDH-A silencing greatly reduced network formation on Matrigel in vitro, and decreased blood vessel formation in Matrigel in vivo. These findings demonstrate that LDH-A is critically important for sustaining the rapid angiogenesis of pulmonary microvascular endothelial cells. PMID:24086675

Parra-Bonilla, Glenda; Alvarez, Diego F; Alexeyev, Mikhail; Vasauskas, Audrey; Stevens, Troy

2013-09-26

124

Lactate Dehydrogenase A Expression Is Necessary to Sustain Rapid Angiogenesis of Pulmonary Microvascular Endothelium  

PubMed Central

Angiogenesis is a fundamental property of endothelium, yet not all endothelial cells display equivalent angiogenic responses; pulmonary microvascular endothelial cells undergo rapid angiogenesis when compared to endothelial cells isolated from conduit vessels. At present it is not clear how pulmonary microvascular endothelial cells fulfill the bioenergetic demands that are necessary to sustain such rapid blood vessel formation. We have previously established that pulmonary microvascular endothelial cells utilize aerobic glycolysis to generate ATP during growth, a process that requires the expression of lactate dehydrogenase A to convert pyruvate to lactate. Here, we test the hypothesis that lactate dehydrogenase A is required for pulmonary microvascular endothelial cells to sustain rapid angiogenesis. To test this hypothesis, Tet-On and Tet-Off conditional expression systems were developed in pulmonary microvascular endothelial cells, where doxycycline is utilized to induce lactate dehydrogenase A shRNA expression. Expression of LDH-A shRNA induced a time-dependent decrease in LDH-A protein, which corresponded with a decrease in glucose consumption from the media, lactate production and cell growth; re-expression of LDH-A rescued each of these parameters. LDH-A silencing greatly reduced network formation on Matrigel in vitro, and decreased blood vessel formation in Matrigel in vivo. These findings demonstrate that LDH-A is critically important for sustaining the rapid angiogenesis of pulmonary microvascular endothelial cells.

Parra-Bonilla, Glenda; Alvarez, Diego F.; Alexeyev, Mikhail; Vasauskas, Audrey; Stevens, Troy

2013-01-01

125

The reversibility of cytosolic dehydrogenase reactions in hepatocytes from starved and fed rats. Effect of fructose.  

PubMed Central

The metabolism of [2-3H]lactate was studied in isolated hepatocytes from fed and starved rats metabolizing ethanol and lactate in the absence and presence of fructose. The yields of 3H in ethanol, water, glucose and glycerol were determined. The rate of ethanol oxidation (3 mumol/min per g wet wt.) was the same for fed and starved rats with and without fructose. From the detritiation of labelled lactate and the labelling pattern of ethanol and glucose, we calculated the rate of reoxidation of NADH catalysed by lactate dehydrogenase, alcohol dehydrogenase and triosephosphate dehydrogenase. The calculated flux of reducing equivalents from NADH to pyruvate was of the same order of magnitude as previously found with [3H]ethanol or [3H]xylitol as the labelled substrate [Vind & Grunnet (1982) Biochim. Biophys. Acta 720, 295-302]. The results suggest that the cytoplasm can be regarded as a single compartment with respect to NAD(H). The rate of reduction of acetaldehyde and pyruvate was correlated with the concentration of these metabolites and NADH, and was highest in fed rats and during fructose metabolism. The rate of reoxidation of NADH catalysed by lactate dehydrogenase was only a few per cent of the maximal activity of the enzymes, but the rate of reoxidation of NADH catalysed by alcohol dehydrogenase was equal to or higher than the maximal activity as measured in vitro, suggesting that the dissociation of enzyme-bound NAD+ as well as NADH may be rate-limiting steps in the alcohol dehydrogenase reaction.

Vind, C; Grunnet, N

1984-01-01

126

The reversibility of cytosolic dehydrogenase reactions in hepatocytes from starved and fed rats. Effect of fructose.  

PubMed

The metabolism of [2-3H]lactate was studied in isolated hepatocytes from fed and starved rats metabolizing ethanol and lactate in the absence and presence of fructose. The yields of 3H in ethanol, water, glucose and glycerol were determined. The rate of ethanol oxidation (3 mumol/min per g wet wt.) was the same for fed and starved rats with and without fructose. From the detritiation of labelled lactate and the labelling pattern of ethanol and glucose, we calculated the rate of reoxidation of NADH catalysed by lactate dehydrogenase, alcohol dehydrogenase and triosephosphate dehydrogenase. The calculated flux of reducing equivalents from NADH to pyruvate was of the same order of magnitude as previously found with [3H]ethanol or [3H]xylitol as the labelled substrate [Vind & Grunnet (1982) Biochim. Biophys. Acta 720, 295-302]. The results suggest that the cytoplasm can be regarded as a single compartment with respect to NAD(H). The rate of reduction of acetaldehyde and pyruvate was correlated with the concentration of these metabolites and NADH, and was highest in fed rats and during fructose metabolism. The rate of reoxidation of NADH catalysed by lactate dehydrogenase was only a few per cent of the maximal activity of the enzymes, but the rate of reoxidation of NADH catalysed by alcohol dehydrogenase was equal to or higher than the maximal activity as measured in vitro, suggesting that the dissociation of enzyme-bound NAD+ as well as NADH may be rate-limiting steps in the alcohol dehydrogenase reaction. PMID:6477525

Vind, C; Grunnet, N

1984-09-01

127

A radiotracer probe to study metal interaction with human lactate dehydrogenase isoenzymes  

SciTech Connect

The electrophilic Ag+ ion was found to destroy completely the enzymatic activity of lactate dehydrogenase isoenzyme LDH-1 while other transition metal ions reduced its activity in varying degrees. A radiotracer probe involving 110mAg-labeled silver ion was used to understand the mechanism of denaturation of LDH and also to determine the number of active sites, if any, for substrate binding with the enzyme. Purified LDH-l was reacted with 110mAg-labeled silver ion and the mixture was passed through the sephadex G-75-120 gel to separate the 110mAg-LDH complex that might be formed during the reaction. The resulting elution curve revealed that a stable complex was formed. From the total radioactivity of 110mAg bound LDH, the specific activity of labeled Ag+ and the amount of LDH used the ratio of the number of moles of Ag+ reacted with 1 mol of LDH was computed. This was found to be approximately 4.0, indicating that there are four binding sites in LDD, probably one on each subunit. Kinetic studies of LDH catalysis of L-P reaction in the presence and absence of Ag+ ion suggest that silver ion is involved in competitive inhibition and that the interaction conforms to the lock-and-key model. The inhibition of catalysis by other metals is presumably of a noncompetitive type.

Menon, M.P.; Wright, C.E. (Savannah State College, Savannah, GA (USA))

1989-12-01

128

Activities of enzymes in the malate–aspartate shuttle and the isoenzyme pattern of lactate dehydrogenase in plasma and peripheral leukocytes of lactating Holstein cows and riding horses  

Microsoft Academic Search

The activities of the enzymes involved in the malate–aspartate shuttle and lactate dehydrogenase (LDH) and the pattern of the isoenzymes of LDH were determined in plasma and peripheral leukocytes of lactating Holstein cows and thoroughbred riding horses as representative herbivorous animals. In the horse plasma, LDH activities were significantly lower and AST activities were significantly higher than those in the

T. Arai; A. Inoue; Y. Uematsu; T. Sako; N. Kimura

2003-01-01

129

Hypoxically inducible barley lactate dehydrogenase: cDNA cloning and molecular analysis  

SciTech Connect

In the roots of barley and other cereals, hypoxia induces a set of five isozymes of L-lactate dehydrogenase (LDH; (S)-lactate:NADH oxidoreductase, EC 1.1.1.27). Biochemical and genetic data indicate that the five LDH isozymes are tetramers that arise from random association of the products of two Ldh loci. To investigate this system, cDNA clones of LDH were isolated from a {lambda}gt11 cDNA library derived from hypoxically treated barley roots. The library was screened with antiserum raised against barley LDH purified {approx}3,000-fold by an improved three-step procedure. Immunopositive clones were rescreened with a cDNA probe synthesized by the polymerase chain reaction using primers modeled from the amino acid sequences of two tryptic LDH peptides. Two types of LDH clones were found. Nucleotide sequence analysis of one representative insert of each type (respectively, 1,305 and 1,166 base pairs) revealed open reading framed encoding 10 peptide fragments of LDH. The 1,305-base-pair insert included the entire coding region of a 356-residue LDH monomer. The nucleotide sequences of the two LDH cDNAs were 92% identical in the coding region, but highly divergent in the 3{prime} noncoding region, and thus probably correspond to the two postulated Ldh loci. The deduced amino acid sequences of the two barley LDHs were 96% identical to each other and very similar to those from vertebrate and bacterial LDHs. RNA blot hybridization showed a single mRNA band of 1.5 kilobases whose level rose about 8-fold in roots during hypoxic induction, as did the level of translatable LDH message.

Hondred, D. (Michigan State Univ., East Lansing (USA)); Hanson, A.D. (Michigan State Univ., East Lansing (USA) Univ. de Montreal, Quebec (Canada))

1990-09-01

130

Characterization of lactate dehydrogenase enzyme in seminal plasma of Japanese quail (Coturnix coturnix japonica).  

PubMed

Lactate dehydrogenase enzyme present in quail seminal plasma has been characterized. Polyacrylamide gel electrophoresis and subsequently with LDH specific staining of seminal plasma revealed a single isozyme in quail semen. Studies on substrate inhibition, pH for optimum activity and inhibitor (urea) indicated the isozyme present in the quail semen has catalytic properties like LDH-1 viz. H-type. Furthermore, unlike other mammalian species, electrophoretic and kinetic investigations did not support the existence of semen specific LDH-X isozyme in quail semen. The effect of exogenous lactate and pyruvate on sperm metabolic activity was also studied. The addition of 1 mM lactate or pyruvate to quail semen increased sperm metabolic activity. Our results suggested that both pyruvate and lactate could be used by quail spermatozoa to maintain their basic functions. Since the H-type isozyme is important for conversion of lactate to pyruvate under anaerobic conditions it was postulated that exogenous lactate being converted into pyruvate via LDH present in semen may be used by sperm mitochondria to generate ATP. During conversion of lactate to pyruvate NADH is being generated that may be useful for maintaining sperm mitochondrial membrane potential. PMID:21074838

Singh, R P; Sastry, K V H; Pandey, N K; Shit, N; Agrawal, R; Singh, K B; Mohan, Jag; Saxena, V K; Moudgal, R P

2010-11-12

131

Galloflavin (CAS 568-80-9): a novel inhibitor of lactate dehydrogenase.  

PubMed

One of the most prominent alterations in cancer cells is their strict dependence on the glycolytic pathway for ATP generation. This observation led to the evaluation of glycolysis inhibitors as potential anticancer agents. The inhibition of lactate dehydrogenase (LDH) is a promising way to inhibit tumor cell glucose metabolism without affecting the energetic balance of normal tissues. However, the success of this approach depends chiefly on the availability of inhibitors that display good selectivity. We identified a compound (galloflavin, CAS 568-80-9) which, in contrast to other inhibitors of human LDH, hinders both the A and B isoforms of the enzyme. To determine the mechanism of action, we collected LDH-A and -B inhibition data in competition reactions with pyruvate or NADH and evaluated the results using software for enzyme kinetics analysis. We found that galloflavin inhibits both human LDH isoforms by preferentially binding the free enzyme, without competing with the substrate or cofactor. The calculated Ki values for pyruvate were 5.46??M (LDH-A) and 15.06??M (LDH-B). In cultured tumor cells, galloflavin blocked aerobic glycolysis at micromolar concentrations, did not interfere with cell respiration, and induced cell death by triggering apoptosis. To our knowledge, the inhibition of LDH is, to date, the only biochemical effect described for galloflavin. Because galloflavin is not commercially available, we also describe herein a procedure for its synthesis and report its first full chemical characterization. PMID:22052811

Manerba, Marcella; Vettraino, Marina; Fiume, Luigi; Di Stefano, Giuseppina; Sartini, Andrea; Giacomini, Elisa; Buonfiglio, Rosa; Roberti, Marinella; Recanatini, Maurizio

2011-11-04

132

Regulation of Cyclic GMP, Cyclic amp and Lactate Dehydrogenase by Putative Neutrotransmitters in the C6 Rat Glioma Cell Line.  

National Technical Information Service (NTIS)

In C6 cells norepinephrine and dopamine caused transient increases in cyclic GMP and cyclic AMP, as well as an induction of lactate dehydrogenase. All of these responses were blocked by 1-propranolol, suggesting mediation by a beta -receptor. Phentolamine...

J. E. Bottenstein J. de Vellis

1977-01-01

133

Creatine Kinase and Lactate Dehydrogenase in the Muscles Encountered during Median Sternotomy and the Myocardium of the Cardiac Chambers.  

National Technical Information Service (NTIS)

The validity of using kinase MB and lactate dehydrogenase, serum isoenzymes to confirm the diagnosis of perioperative m ycocardial infarction in patients who have had cardiac operations has been questioned, since both have been detected in sketetal muscle...

G. M. Graeber

1985-01-01

134

Metabolic Engineering of Bacillus subtilis for Ethanol Production: Lactate Dehydrogenase Plays a Key Role in Fermentative Metabolism  

Microsoft Academic Search

Received 19 March 2007\\/Accepted 1 June 2007 Wild-type Bacillus subtilis ferments 20 g\\/liter glucose in 48 h, producing lactate and butanediol, but not ethanol or acetate. To construct an ethanologenic B. subtilis strain, homologous recombination was used to disrupt the native lactate dehydrogenase (LDH) gene (ldh) by chromosomal insertion of the Zymomonas mobilis pyruvate decarboxylase gene (pdc) and alcohol dehydrogenase

Susana Romero; Enrique Merino; F. Bolivar; Guillermo Gosset; Alfredo Martinez

2007-01-01

135

Alkaline-induced unfolding and salt-induced folding of pig heart lactate dehydrogenase under high pH conditions  

Microsoft Academic Search

The alkaline-induced unfolding and the salt-induced folding of pig heart lactate dehydrogenase under high pH conditions have been followed by fluorescence emission spectra and circular dichroism spectra. The results for alkaline-induced denaturation of lactate dehydrogenase show that at low ionic strength, increasing the pH value increased the extent of unfolding of the enzyme to the maximum ultimate unfolded conformation at

Ji-Hong Bai; Hao-Jing Wang; Hai-Meng Zhou

1998-01-01

136

Effect of the Buffer Concentration on the Separation of Lactate Dehydrogenase Isoenzymes from Different Sources by Electrophoresis  

Microsoft Academic Search

The separation of lactate dehydrogenase isoenzymes by zone electrophoresis using cellulose acetate strips as support was dependent on the concentration of the buffer used (5 mM and 50 mM, pH 7.4) and on the source of the material (chicken liver or guinea-pig liver).In three different 5 mM buffer systems, pH 7.4 (phosphate, veronal and Tris-HC1) the four lactate dehydrogenase isoenzymes

S. Imperial; J. Li Gelpí; M. Busquets; A. Mazo; A. Cortés

1991-01-01

137

Ethanol production by anaerobic thermophilic bacteria: regulation of lactate dehydrogenase activity in Clostridium thermohydrosulfuricum  

Microsoft Academic Search

The enzyme lactate dehydrogenase (LDH) in Clostridium thermohydrosulfuricum is controlled by the type and the concentration of the substrate. In batch fermentations an increase of the initial concentration of glucose leads to an increase in the activity of LDH. This increase in activity is related to the accumulation of fructose 1,6-diphosphate (F 1,6-DP), an intermediate of the Embden-Meyerhof-Parnas (EMP) pathway,

Pierre Germain; Fatiou Toukourou; Luiz Donaduzzi

1986-01-01

138

Cryoprotection mechanisms of polyethylene glycols on lactate dehydrogenase during freeze-thawing  

Microsoft Academic Search

The purpose of this study was to explore the cryoprotection mechanisms of high molecular weight polyethylene glycols (PEGs)\\u000a (eg, PEG 4000 and PEG 8000) on lactate dehydrogenase (LDH). Ultraviolet activity assays, circular dichroism (CD) spectroscopy,\\u000a gel filtration, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE),14C-PEG 4000 labeling and binding, and cryostage microscopic study were conducted. Different molecular weights and concentrations\\u000a of

Yanli Mi; George Wood; Laura Thoma

2004-01-01

139

Lactate dehydrogenase levels during MACOP-B chemotherapy for non-Hodgkin’s lymphoma  

Microsoft Academic Search

Lactate dehydrogenase (LD) levels rose consistently during MACOP-B chemotherapy for intermediate and high-grade non-Hodgkin’s\\u000a lymphoma (NHL). Levels peaked at week nine and fell to normal within six weeks of completion of therapy. Isoenzyme patterns,\\u000a studied prospectively in seven patients, showed a parallel rise in LD1 and LD2 suggesting a source other than tumour tissue\\u000a for the rise in total LD.

B. McAdam; T. Smith; W. C. Love; M. Murphy; P. A. Daly

1993-01-01

140

Determination of lactate dehydrogenase isoenzymes in single rat glioma cells by capillary electrophoresis with electrochemical detection  

Microsoft Academic Search

A method for determination of lactate dehydrogenase (LDH) isoenzymes in single rat glioma cells (C6) was developed. In this method, a whole cell was electrokinetically injected into the front end of the separation capillary. After that, the cell was lysed by ultrasonication and the isoenzymes in the cell were pre-separated at 20kV for 5min and then incubated for 2min with

Wei Wang; Shuaibing Han; Wenrui Jin

2006-01-01

141

Antiplasmodial studies of Eurycoma longifolia Jack using the lactate dehydrogenase assay of Plasmodium falciparum  

Microsoft Academic Search

The roots of Eurycoma longifolia Jack have been used as traditional medicine to treat malaria. A systematic bioactivity-guided fractionation of this plant was conducted involving the determination of the effect of its various extracts and their chemical constituents on the lactate dehydrogenase activity of in vitro chloroquine-resistant Gombak A isolate and chloroquine-sensitive D10 strain of Plasmodium falciparum parasites. Their antiplasmodial

Kit-Lam Chan; Chee-Yan Choo; Noor Rain Abdullah; Zakiah Ismail

2004-01-01

142

Null alleles at two lactate dehydrogenase loci in rainbow trout are associated with decreased developmental stability  

Microsoft Academic Search

Previous studies with rainbow trout (Oncorhynchus mykiss) have shown that allozymic heterozygotes have increased developmental stability, as measured by reduced fluctuating bilateral\\u000a asymmetry. In this paper, we examine the phenotypic effects of null alleles at two lactate dehydrogenase (LDH) loci. If the\\u000a association between allozymic heterozygosity and developmental stability is due largely to linked chromosomal segments, then\\u000a we would expect

Robb F. Leary; Fred W. Allendorf; Kathy L. Knudsen

1993-01-01

143

Evolution of l-lactate dehydrogenase gene expression in non-avian reptiles  

Microsoft Academic Search

Isozyme characters, exclusive of allozymes (alleles), are potentially useful in phylogenetic investigations. Such data have been derived from the multilocus l-lactate dehydrogenase (LDH) enzyme system. Among non-avian reptiles, the classes of potential characters include (1) and number of loci encoding LDH, (2) heterotramer patterns, (3) tissue-specific patterns of gene expression, (4) tissue-specific posttranslational modification and (5) reversed relative mobility of

Robert W. Murphy

1999-01-01

144

Identification of yak lactate dehydrogenase B gene variants by gene cloning  

Microsoft Academic Search

Native polyacrylamide gel electrophoresis showed that two types of lactate dehydrogenase (LDH) existed in yaks. Based on the\\u000a electrophoresis characteristics of LDH isoenzymes, yak LDH variants were speculated to be the gene mutation on H subunit encoded\\u000a by B gene. According to the mobility in electrophoresis, the fast-band LDH type was named LDH-Hf and the slow-band LDH type\\u000a LDH-Hs. In

YuCai Zheng; XingBo Zhao; Jing Zhou; Ying Piao; SuYu Jin; QingHua He; Jian Hong; Ning Li; ChangXin Wu

2008-01-01

145

A coupled enzyme nylon tube reactor pyruvate kinase-lactate dehydrogenase system  

Microsoft Academic Search

A coupled enzyme nylon tube reactor has been made by simultaneously immobilizing rabbit muscle pyruvate kinase (EC 2.7.1.40)\\u000a and lactate dehydrogenase (EC 1.1.1.27) onto the inside surface of a nylon tube initially derivatized with poly (scl)-lysine\\u000a to serve as a spacer molecule. The enzymes were covalently linked to the amino groups of the poly lysine spacer molecule by\\u000a cross-linking with

P. V. Sundaram

1978-01-01

146

Plasmodium falciparum: Stage specific effects of a selective inhibitor of lactate dehydrogenase  

Microsoft Academic Search

Plasmodium falciparum lactate dehydrogenase (PfLDH) is essential for ATP generation. Based on structural differences within the active site between P. falciparum and human LDH, we have identified a series of heterocyclic azole-based inhibitors that selectively bind within the PfLDH but not the human LDH (hLDH) active site and showed anti-malarial activity in vitro and in vivo. Here we expand on

Livia Vivas; Anna Easton; Howard Kendrick; Angus Cameron; Jose-Luis Lavandera; David Barros; Federico Gomez de las Heras; R. Leo Brady; Simon L. Croft

2005-01-01

147

Mapping the binding site for gossypol-like inhibitors of Plasmodium falciparum lactate dehydrogenase  

Microsoft Academic Search

Gossypol is a di-sesquiterpene natural-product in the form of a functionalised binaphthyl and is isolated from cotton plants. The compound has long been known to exhibit anti-malarial and other biological activities. Previous studies have indicated that compounds of this type target Plasmodium falciparum lactate dehydrogenase (pfLDH), an essential enzyme for energy generation within the parasite. In this study, we report

Rebecca Conners; Felix Schambach; Jon Read; Angus Cameron; Richard B. Sessions; Livia Vivas; Anna Easton; Simon L. Croft; R. Leo Brady

2005-01-01

148

Molecular Properties of Pyruvate Bound to Lactate Dehydrogenase: A Raman Spectroscopic Study  

Microsoft Academic Search

Lactate dehydrogenase (LDH; EC 1.1.1.27) catalyzes the addition of pyruvate to the four position of the nicotinamide ring of bound NAD+ this NAD-pyruvate adduct is bound tightly to the enzyme. We have used the adduct as a model for pyruvate in a competent ternary complex by comparing the Raman spectrum of the bound adduct with that for unliganded pyruvate. To

Hua Deng; Jie Zheng; John Burgner; Robert Callender

1989-01-01

149

Kinetic Characterization of the Inhibition of Purified Cynomolgus Monkey Lactate Dehydrogenase Isozymes by Gossypol  

Microsoft Academic Search

This report describes the results of the first step in a sequence of experiments designed to test the hypothesis that the sperm-specific isozyme of lactate dehydrogenase (LDH-C4), is a site of action of the potential male contra- ceptive agent gossypol. Cynomolgus monkey LDH-A4, LDH-B4 and LDH-C4 were purified and kinetically char- acterized. LDH-A4 and LDH-B4 exhibited \\

DONALD T. STEPHENS; KEVIN J. WHALEY; NANETTE M. KLIMKOW; PAULINE GOH; DALE D. HOSKINS

150

Preparation of gold surfaces with biospecific affinity for NAD(H)-dependent lactate dehydrogenase  

Microsoft Academic Search

Gold surfaces with biospecific affinity for NAD(H)-dependent lactate dehydrogenase have been prepared by covalent attachment of several triazine dyes to chemisorbed functionalized alkanethiol self-assembled monolayers. The dyes which work as coenzyme analoga build a site-specific affinity complex with the enzyme by binding through its NAD+-binding pocket. This immobilization method implies a biospecific recognition of the enzyme which is expected to

Daniela D. Schlereth

1997-01-01

151

Regulation of the ldhA gene, encoding the fermentative lactate dehydrogenase of Escherichia coli  

Microsoft Academic Search

The fermentative lactate dehydrogenase (LDH) of Escherichia coli is induced by low pH under anaerobic conditions. Both translational and transcriptional gene fusions to ldhA, which encodes the fermentative LDH, have now been made. Both types of ldhA-lacZ fusion were induced by low pH, but only in the absence of air. However, the translational fusions were consistently expressed at a five-

Gene Ruijun Jiang; Sonia Nikolova; David P. Clark

2001-01-01

152

Antimalarial Activity of Potential Inhibitors of Plasmodium falciparum Lactate Dehydrogenase Enzyme Selected by Docking Studies  

Microsoft Academic Search

The Plasmodium falciparum lactate dehydrogenase enzyme (PfLDH) has been considered as a potential molecular target for antimalarials due to this parasite's dependence on glycolysis for energy production. Because the LDH enzymes found in P. vivax, P. malariae and P. ovale (pLDH) all exhibit ?90% identity to PfLDH, it would be desirable to have new anti-pLDH drugs, particularly ones that are

Julia Penna-Coutinho; Wilian Augusto Cortopassi; Aline Alves Oliveira; Tanos Celmar Costa França; Antoniana Ursine Krettli

2011-01-01

153

Enthalpy-entropy compensation of oxamate binding by homologous lactate dehydrogenases  

Microsoft Academic Search

STUDIES1,2 of the amino acid sequence and crystal structure of M4 lactate dehydrogenase (EC 1.1.1.27) indicate that all binding interactions are of the weak, non-covalent type (proceeding with only minor free energy changes). Because critical regions of the substrate and coenzyme-binding site seem highly conserved3, and because non-covalent interactions are highly, if differentially, sensitive to the physical environment4, we wondered

P. W. Hochachka; C. Norberg; J. Baldwin; J. H. A. Fields

1976-01-01

154

A Kinetic Analysis of the Endogenous Lactate Dehydrogenase Activity of Duck Lens ?-Crystallin in Reverse Micelles  

Microsoft Academic Search

?-Crystallin is a structural protein in duck lenses with endogenous lactate dehydrogenase (LDH) activity. When entrapped in an aerosol-OT (AOT)\\/isooctane\\/H2O reverse micellar system, ?-crystallin preserves this endogenous enzymatic activity. The catalytic constant (kcat) of ?-crystallin exhibited multiple peaks at varying degrees of system hydration ([H2O]\\/[AOT]), thereby suggesting that ?-crystallin exists as various oligomers in reverse micelles and that each oligomer

Hwei-Jen Lee; Gu-Gang Chang

1998-01-01

155

The Effects of Lanthanoid on the Structure–Function of Lactate Dehydrogenase from Mice Heart  

Microsoft Academic Search

The activity of lactate dehydrogenase (LDH, EC1.1.1.27) is often changed upon inflammatory responses in animals. Lanthanoid\\u000a (Ln) was shown to provoke various inflammatory responses both in rats and mice; however, the molecular mechanism by which\\u000a Ln3+ exert its toxicity has not been completely understood, especially that we know little about the mechanism of the interaction\\u000a between Ln with 4f electron

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

2009-01-01

156

Cloning and nucleotide sequence of the Lactobacillus casei lactate dehydrogenase gene.  

PubMed Central

An allosteric L-(+)-lactate dehydrogenase gene of Lactobacillus casei ATCC 393 was cloned in Escherichia coli, and the nucleotide sequence of the gene was determined. The gene was composed of an open reading frame of 981 bp, starting with a GTG codon and ending with a TAA codon. The sequences for the promoter and ribosome binding site were identified, and a sequence for a structure resembling a rho-independent transcription terminator was also found. Images

Kim, S F; Baek, S J; Pack, M Y

1991-01-01

157

Biomimetic dye affinity chromatography for the purification of bovine heart lactate dehydrogenase  

Microsoft Academic Search

Three biomimetic dye ligands bearing as a triazine-linked terminal moiety a carboxylated structure, which mimics substrates and inhibitors of l-lactate dehydrogenase (LDH), were immobilized on cross-linked agarose Ultrogel A6R. These biomimetic dyes are purpose-designed analogues of commercial monochlorotriazine Cibacron Blue 3GA (CB3GA) and parent dichlorotriazine Vilmafix Blue A–R (VBAR). The corresponding biomimetic adsorbents, along with non-biomimetic adsorbents bearing CB3GA and

N. E. Labrou; Y. D. Clonis

1995-01-01

158

Variation of transaminases and lactate dehydrogenase in irradiated Biomphalaria alexandrina snails.  

PubMed

Effect of ultraviolet and gamma radiations on the activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactate dehydrogenase (LD) in Biomphalaria alexandrina snails, the specific intermediate host of schistosomiasis, was investigated. Changes in the electrophoretic pattern of LD in the species under study were also taken as a measured parameter and the effect of gamma-irradiation on the glutathione content in the haemolymph of the snails have been included. PMID:1934013

Nabih, I; el-Ansary, A

1991-01-01

159

Inhibition of lactate dehydrogenase isoenzyme associated with anaerobic respiration in schistosomiasis intermediate host snails.  

PubMed

Lactate dehydrogenase isoenzyme (LD5) which is associated with anaerobic respiration was inhibited to a certain degree in Biomphalaria alexandrina snails, the intermediate host for Schistosoma mansoni. Urea and thiourea were used as inhibitors. The effect of LD5 inhibition on the mortality rate of infected Biomphalaria alexandrina snails and on the susceptibility of the snails to the trematode infection was also studied. PMID:1905584

Nabih, I; el Dardiri, Z; el-Ansary, A

1991-01-01

160

Partial Characterization,Properties,and Clinical Significanceof a Lactate Dehydrogenase-ImmunoglobulinAKComplexin Serum  

Microsoft Academic Search

The existence of a lactate dehydrogenase-immunoglobulin A,, complex was demonstratedas a marked increase of lactatedehydrogenase activityat the lactatedehydrogenase- 3 band in the agar-agarose gel-electrophoresispattern of serafromsevenpatientsfrom an unselectedgroupof 21 800 patients.The complexwas isolated,in an almostpureform, by gel filtration and affinitychromatography, from the serum of a patientwith circulatinghepatitisB surfaceantigen.The complexhada relativemolecularmass(Mr) of approximately 445 000 as determinedby gel filtration. Electrophoresisin sodium

R. N. M. Weljers; J. Mulder

161

Structural basis for discriminatory recognition of Plasmodium lactate dehydrogenase by a DNA aptamer.  

PubMed

DNA aptamers have significant potential as diagnostic and therapeutic agents, but the paucity of DNA aptamer-target structures limits understanding of their molecular binding mechanisms. Here, we report a distorted hairpin structure of a DNA aptamer in complex with an important diagnostic target for malaria: Plasmodium falciparum lactate dehydrogenase (PfLDH). Aptamers selected from a DNA library were highly specific and discriminatory for Plasmodium as opposed to human lactate dehydrogenase because of a counterselection strategy used during selection. Isothermal titration calorimetry revealed aptamer binding to PfLDH with a dissociation constant of 42 nM and 2:1 protein:aptamer molar stoichiometry. Dissociation constants derived from electrophoretic mobility shift assays and surface plasmon resonance experiments were consistent. The aptamer:protein complex crystal structure was solved at 2.1-Å resolution, revealing two aptamers bind per PfLDH tetramer. The aptamers showed a unique distorted hairpin structure in complex with PfLDH, displaying a Watson-Crick base-paired stem together with two distinct loops each with one base flipped out by specific interactions with PfLDH. Aptamer binding specificity is dictated by extensive interactions of one of the aptamer loops with a PfLDH loop that is absent in human lactate dehydrogenase. We conjugated the aptamer to gold nanoparticles and demonstrated specificity of colorimetric detection of PfLDH over human lactate dehydrogenase. This unique distorted hairpin aptamer complex provides a perspective on aptamer-mediated molecular recognition and may guide rational design of better aptamers for malaria diagnostics. PMID:24043813

Cheung, Yee-Wai; Kwok, Jane; Law, Alan W L; Watt, Rory M; Kotaka, Masayo; Tanner, Julian A

2013-09-16

162

Lactic acid production by Saccharomyces cerevisiae expressing a Rhizopus oryzae lactate dehydrogenase gene  

Microsoft Academic Search

.   This work demonstrates the first example of a fungal lactate dehydrogenase (LDH) expressed in yeast. A L(+)-LDH gene, ldhA, from the filamentous fungus Rhizopus oryzae was modified to be expressed under control of the Saccharomyces cerevisiae adh1 promoter and terminator and then placed in a 2µ-containing yeast-replicating plasmid. The resulting construct, pLdhA68X,\\u000a was transformed and tested by fermentation analyses

Christopher D. Skory

2003-01-01

163

Rapid evaluation of nickel binding properties of His-tagged lactate dehydrogenases using surface plasmon resonance  

Microsoft Academic Search

The use of surface plasmon resonance (SPR), for the comparison of metal binding properties of polyhistidine tags, was evaluated. Six different tags containing various number of histidines, either none (tags n and t), three (tags H3A3 and HA2HA2H) or six (tags H6 and His6), were genetically fused to the N-terminal of lactate dehydrogenase (LDH). The binding ability of these constructs

Florent Bernaudat; Leif Bülow

2005-01-01

164

Differential expression of two tomato lactate dehydrogenase genes in response to oxygen deficit  

Microsoft Academic Search

Two different cDNAs encoding lactate dehydrogenase (LDH) were isolated from a library of hypoxically treated tomato roots and sequenced. The use of gene-specific probes on northern blots showed that Ldh2 mRNA was predominant in well-oxygenated roots and levels remained stable upon oxygen deficit; in contrast, Ldh1 mRNA accumulated to high levels within 2 h of hypoxia or anoxia. Immunoblot analyses

Véronique Germain; Philippe Raymond; Bérénice Ricard

1997-01-01

165

Immobilization of glucose oxidase and lactate dehydrogenase onto magnetic nanoparticles for bioprocess monitoring system  

Microsoft Academic Search

Glucose oxidase (GOD) and lactate dehydrogenase (LDH) were immobilized onto magnetic nanoparticles, viz. Fe3O4, via carbodiimide and glutaraldehyde. The immobilization efficiency was largely dependent upon the immobilization time and\\u000a concentration of glutaraldehyde. The magnetic nanoparticles had a mean diameter of 9.3 nm and were superparamagnetic. The\\u000a immobilization of GOD and LDH on the nanoparticles slightly decreased their saturation magnetization. However,

Ok-Jae Sohn; Chun-Kwang Kim; Jong Il Rhee

2008-01-01

166

Inactivation and conformational changes of lactate dehydrogenase from porcine heart in sodium dodecyl sulfate solutions  

Microsoft Academic Search

The inactivation and conformational changes of porcine heart lactate dehydrogenase (LDH) have been studied in sodium dodecyl sulfate (SDS) solutions. Increasing SDS concentration led to a quick and concentration-dependent inhibition of the enzyme, with complete inactivation within 5 min in the presence of 1.0 mM SDS. Meanwhile, fluorescence emission and circular dichroism spectra were used to follow the conformational changes

Yan-bin Zheng; Fan-guo Meng; Bao-yu Chen; Xi-cheng Wang

2002-01-01

167

Identification of 2-amino-5-aryl-pyrazines as inhibitors of human lactate dehydrogenase.  

PubMed

A 2-amino-5-aryl-pyrazine was identified as an inhibitor of human lactate dehydrogenase A (LDHA) via a biochemical screening campaign. Biochemical and biophysical experiments demonstrated that the compound specifically interacted with human LDHA. Structural variation of the screening hit resulted in improvements in LDHA biochemical inhibition and pharmacokinetic properties. A crystal structure of an improved compound bound to human LDHA was also obtained and it explained many of the observed structure-activity relationships. PMID:24012183

Fauber, Benjamin P; Dragovich, Peter S; Chen, Jinhua; Corson, Laura B; Ding, Charles Z; Eigenbrot, Charles; Giannetti, Anthony M; Hunsaker, Thomas; Labadie, Sharada; Liu, Yichin; Liu, Yingchun; Malek, Shiva; Peterson, David; Pitts, Keith; Sideris, Steve; Ultsch, Mark; VanderPorten, Erica; Wang, Jing; Wei, BinQing; Yen, Ivana; Yue, Qin

2013-08-22

168

Molecular cloning, characterization, and immunolocalization of two lactate dehydrogenase homologous genes from Taenia solium  

Microsoft Academic Search

Two novel genes encoding lactate dehydrogenase A (LDHA) and B (LDHB) homologues, respectively, were identified from the cDNA\\u000a libraries of adult Taenia solium (T. solium). The two deduced amino acid sequences both show more than 50% identity to the homologues for Danio rerio, Xenopus laevis, Schistosoma japonicum, Sus scrofa, Homo sapiens, et al. The identity of the amino acid sequence

Wuying Du; Fengyu Hu; Yabo Yang; Dong Hu; Xuchu Hu; Xinbing Yu; Jin Xu; Jialin Dai; Xinjiang Liao; Jiang Huang

169

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

Microsoft Academic Search

Background  Evidence for historical, demographic and selective factors affecting enzyme evolution can be obtained by examining nucleotide\\u000a 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

Teresa J Crease; Robin Floyd; Melania E Cristescu; David Innes

2011-01-01

170

Mode of neutralization of lactate dehydrogenase-elevating virus by polyclonal and monoclonal antibodies  

Microsoft Academic Search

Summary Neutralization of the infectivity of [3H]uridine-labeled lactate dehydrogenase-elevating virus (LDV) by polyclonal mouse or rabbit antibodies to the envelope glycoprotein of LDV, VP-3, or by neutralizing monoclonal antibodies (mAb) that recognize a different epitope on VP-3 than the polyclonal antibodies correlated with an increase in the sedimentation rate of LDV from 230 S to ?270 S. Incubation of LDV

P. G. W. Plagemann; J. T. Harty; C. Even

1992-01-01

171

Fluorimetric quantification of intracellular lactate dehydrogenase during fermentation using flow injection analysis  

Microsoft Academic Search

A flow injection system for the on-line detection of the intracellular enzyme lactate dehydrogenase (LDH) during fermentation has been developed. The system is comprised of an on-line cell disintegration part, an immobilised dye based expanded bed column for the affinity capture of LDH and a fluorimetric detection unit. The system with a linearity of 0.1–5.4 U LDH ml-1 was applied

M. P. Nandakumar; R. Nandakumar; B. Mattiasson

2000-01-01

172

Liquid—liquid extraction of lactate dehydrogenase from muscle using polymer-bound triazine dyes  

Microsoft Academic Search

An extract from porcine muscle containing soluble enzymes has been partitioned between the two liquid phases of an aqueous,\\u000a biphasic system consisting of dextran, polyethylene glycol, and water. The influence of polymer-bound triazine dyes (Cibacron\\u000a blue F3G-A and Procion yellow HE-3G) on the partition of lactate dehydrogenase and total protein was studied. The effects\\u000a of pH and concentrations of polymers

G. Johansson; M. Joelsson

1986-01-01

173

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

Microsoft Academic Search

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

E. D. Wieben

1981-01-01

174

Evaluation of genetically attached histidine affinity tails for purification of lactate dehydrogenase from transgenic tobacco  

Microsoft Academic Search

Lactate dehydrogenase (Bacillus stearothermophilus) has been expressed in transgenic tobacco. To facilitate purification polyhistidine tails were fused to the 5?-end of the gene. Two different tails, His6 and His-X3-His-X3-His, were compared regarding their effect on LDH gene expression and metal ion specificity. His6 exhibited strong binding to all of the tested transition metals (Zn2+, Co2+, Ni2+ and Cu2+) while the

Malin Mejàre; Gösta Lilius; Leif Bülow

1998-01-01

175

Purification and partial characterization of a d (?)-lactate dehydrogenase from Desulfovibrio desulfuricans (ATCC 7757)  

Microsoft Academic Search

Summary  A membrane-boundd(–)-lactate dehydrogenase (LDH), an important enzyme in carbon and energy metabolism in sulfate-reducing bacteria of the genusDesulfovibrio, was solubilized from the membrane fraction ofDesulfovibrio desulfuricans (ATCC 7757). The enzyme was purified 84-fold to a final specific activity of 525 nmol DCPIP-reduced\\/min\\/mg protein by ammonium sulfate precipitation, chloroform extraction, gel filtration with Sephadex G-150, and hydrophobic column chromatography withN-octylamine Sepharose

M. H. Czechowski; H. W. Rossmoore

1990-01-01

176

The reaction of trimethylamine dehydrogenase with trimethylamine.  

PubMed

The reductive half-reaction of trimethylamine dehydrogenase with its physiological substrate trimethylamine has been examined by stopped-flow spectroscopy over the pH range 6.0-11.0, with attention focusing on the fastest of the three kinetic phases of the reaction, the flavin reduction/substrate oxidation process. As in previous work with the slow substrate diethylmethylamine, the reaction is found to consist of three well resolved kinetic phases. The observed rate constant for the fast phase exhibits hyperbolic dependence on the substrate concentration with an extrapolated limiting rate constant (klim) greater than 1000 s-1 at pH above 8.5, 10 degrees C. The kinetic parameter klim/Kd for the fast phase exhibits a bell-shaped pH dependence, with two pKa values of 9.3 +/- 0.1 and 10. 0 +/- 0.1 attributed to a basic residue in the enzyme active site and the ionization of the free substrate, respectively. The sigmoidal pH profile for klim gives a single pKa value of 7.1 +/- 0. 2. The observed rate constants for both the intermediate and slow phases are found to decrease as the substrate concentration is increased. The steady-state kinetic behavior of trimethylamine dehydrogenase with trimethylamine has also been examined, and is found to be adequately described without invoking a second, inhibitory substrate-binding site. The present results demonstrate that: (a) substrate must be protonated in order to bind to the enzyme; (b) an ionization group on the enzyme is involved in substrate binding; (c) an active site general base is involved, but not strictly required, in the oxidation of substrate; (d) the fast phase of the reaction with native enzyme is considerably faster than observed with enzyme isolated from Methylophilus methylotrophus that has been grown up on dimethylamine; and (e) a discrete inhibitory substrate-binding site is not required to account for excess substrate inhibition, the kinetic behavior of trimethylamine dehydrogenase can be readily explained in the context of the known properties of the enzyme. PMID:10224069

Jang, M H; Basran, J; Scrutton, N S; Hille, R

1999-05-01

177

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.

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

1995-01-01

178

Physical and functional association of lactate dehydrogenase (LDH) with skeletal muscle mitochondria.  

PubMed

The intracellular lactate shuttle hypothesis posits that lactate generated in the cytosol is oxidized by mitochondrial lactate dehydrogenase (LDH) of the same cell. To examine whether skeletal muscle mitochondria oxidize lactate, mitochondrial respiratory oxygen flux (JO2) was measured during the sequential addition of various substrates and cofactors onto permeabilized rat gastrocnemius muscle fibers, as well as isolated mitochondrial subpopulations. Addition of lactate did not alter JO2. However, subsequent addition of NAD(+) significantly increased JO2, and was abolished by the inhibitor of mitochondrial pyruvate transport, ?-cyano-4-hydroxycinnamate. In experiments with isolated subsarcolemmal and intermyofibrillar mitochondrial subpopulations, only subsarcolemmal exhibited NAD(+)-dependent lactate oxidation. To further investigate the details of the physical association of LDH with mitochondria in muscle, immunofluorescence/confocal microscopy and immunoblotting approaches were used. LDH clearly colocalized with mitochondria in intact, as well as permeabilized fibers. LDH is likely localized inside the outer mitochondrial membrane, but not in the mitochondrial matrix. Collectively, these results suggest that extra-matrix LDH is strategically positioned within skeletal muscle fibers to functionally interact with mitochondria. PMID:23873936

Elustondo, Pia A; White, Adrienne E; Hughes, Meghan E; Brebner, Karen; Pavlov, Evgeny; Kane, Daniel A

2013-07-20

179

Lysine-5 acetylation negatively regulates lactate dehydrogenase A and is decreased in pancreatic cancer.  

PubMed

Tumor cells commonly have increased glucose uptake and lactate accumulation. Lactate is produced from pyruvate by lactate dehydrogenase A (LDH-A), which is frequently overexpressed in tumor cells and is important for cell growth. Elevated transcription by c-Myc or HIF1? may contribute to increased LDH-A in some cancer types. Here, we show that LDH-A is acetylated at lysine 5 (K5) and that this acetylation inhibits LDH-A activity. Furthermore, the K5-acetylated LDH-A is recognized by the HSC70 chaperone and delivered to lysosomes for degradation. Replacement of endogenous LDH-A with an acetylation mimetic mutant decreases cell proliferation and migration. Importantly, K5 acetylation of LDH-A is reduced in human pancreatic cancers. Our study reveals a mechanism of LDH-A upregulation in pancreatic cancers. PMID:23523103

Zhao, Di; Zou, Shao-Wu; Liu, Ying; Zhou, Xin; Mo, Yan; Wang, Ping; Xu, Yan-Hui; Dong, Bo; Xiong, Yue; Lei, Qun-Ying; Guan, Kun-Liang

2013-03-21

180

Large-Scale Production, Biochemical and Structural Characterization of Lactate Dehydrogenase for the Discovery of Novel Small Molecule Inhibitors  

Microsoft Academic Search

Lactate Dehydrogenase (LDH) catalyzes the interconversion of pyruvate and lactate in the final step of anaerobic glycolysis. Many invasive tumors depend on an increased rate of glycolysis in order to maximize their growth potential under hypoxic environments, a phenomenon known as the \\

Erol Jahja

2011-01-01

181

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

Microsoft Academic Search

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

Ming Zhou; Yuhua Zhao; Yan Ding; Hao Liu; Zixing Liu; Oystein Fodstad; Adam I Riker; Sushama Kamarajugadda; Jianrong Lu; Laurie B Owen; Susan P Ledoux; Ming Tan

2010-01-01

182

In-situ monitoring of the interaction of lactate dehydrogenase with NAD on a gold electrode by FT-SERS  

Microsoft Academic Search

The FT-SERS method was used to observe directly and monitor the combination of dehydrogenases with NAD pre-adsorbed onto the surface of a roughened gold electrode. At negative electrode potentials, the SERS spectra of NAD exhibit significant changes upon the addition of lactate dehydrogenase. The enzyme active site may approach the electrode closely enough to produce several strong bands attributable to

Yi-Jin Xiao; Xiao-Xia Gao; John Markwell

1999-01-01

183

Lactate dehydrogenase ontogeny, paternal gene activation, and tetramer assembly in embryos of brook trout, lake trout, and their hybrids  

Microsoft Academic Search

Measurement of lactate dehydrogenase in reciprocal hybrids of trout during development revealed that a maternal effect was involved in the regulation of enzyme levels until resorption of the yolk sac was completed. Malate dehydrogenase specific activities were the same in these embryos and larvae. The more negatively charged B subunits of LDH predominated during early stages of embryogenesis in lake

Erwin Goldberg; J. P. Cuerrier; J. C. Ward

1969-01-01

184

On the pathway of forming enzymatically productive ligand-protein complexes in lactate dehydrogenase.  

PubMed

We have carried out a series of studies on the binding of a substrate mimic to the enzyme lactate dehydrogenase (LDH) using advanced kinetic approaches, which begin to provide a molecular picture of the dynamics of ligand binding for this protein. Binding proceeds via a binding-competent subpopulation of the nonligated form of the protein (the LDH/NADH binary complex) to form a protein-ligand encounter complex. The work here describes the collapse of the encounter complex to form the catalytically competent Michaelis complex. Isotope-edited static Fourier transform infrared studies on the bound oxamate protein complex reveal two kinds of oxamate environments: 1), a major populated structure wherein all significant hydrogen-bonding patterns are formed at the active site between protein and bound ligand necessary for the catalytically productive Michaelis complex and 2), a minor structure in a configuration of the active site that is unfavorable to carry out catalyzed chemistry. This latter structure likely simulates a dead-end complex in the reaction mixture. Temperature jump isotope-edited transient infrared studies on the binding of oxamate with LDH/NADH suggest that the evolution of the encounter complex between LDH/NADH and oxamate collapses via a branched reaction pathway to form the major and minor bound species. The production of the catalytically competent protein-substrate complex has strong similarities to kinetic pathways found in two-state protein folding processes. Once the encounter complex is formed between LDH/NADH and substrate, the ternary protein-ligand complex appears to "fold" to form a compact productive complex in an all or nothing like fashion with all the important molecular interactions coming together at the same time. PMID:18390601

Deng, Hua; Brewer, Scott; Vu, Dung M; Clinch, Keith; Callender, Robert; Dyer, R Brian

2008-04-04

185

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

186

About the pKa of the active-site histidine in flavocytochrome b2 (yeast L-lactate dehydrogenase).  

PubMed Central

Flavocytochrome b2 or L-lactate dehydrogenase from yeasts catalyzes the oxidation of L-lactate at the expense of monoelectronic acceptors such as cytochrome c, its physiological partner. When incubated in the presence of both L-lactate and a keto acid, the enzyme catalyzes a transhydrogenation reaction wherein only the flavin is involved. During this reaction, the substrate alpha-hydrogen is transferred not only to the solvent but also in part to the keto acid, which acts as reverse substrate. Thus, when bound to the reduced enzyme, this hydrogen is sticky. In the context of a carbanion mechanism, it resides on Nepsilon of His373, the active site base. We have shown before that a correlation between the amount of intermolecular hydrogen transfer from [2-3H] lactate and the keto acid reverse substrate concentration enables the determination of the first-order rate constant, kHe, for exchange of the substrate-derived protein-bound hydrogen with bulk solvent (Urban P, Lederer F, 1985, J Biol Chem 260:11115-11122). In this work, we show that the exchange with the solvent appears to be independent of the phosphate buffer concentration in the range from 40 to 500 mM. It is thus probable that exchange occurs directly with water molecules. The second-order rate constant for exchange is then 0.16 (+/-0.03) M(-1) s(-1). Using the Eigen equation, this figure yields a pKa of 9.1+/-0.1 for His373 in the reduced enzyme, compared to a probable value of 6.0 or less in the oxidized enzyme (Suzuki H, Ogura YC, 1970, J Biochem 67:291-295). The mechanistic significance of these results is discussed.

Rao, K. S.; Lederer, F.

1998-01-01

187

Functional characterization of an alternative [lactate dehydrogenase-like] malate dehydrogenase in Plasmodium falciparum  

Microsoft Academic Search

The catalysis of malate dehydrogenase (MDH) in Plasmodium falciparum (pfMDH) which involves NAD\\/NADH coupling is crucial for the parasite’s pathogenicity. Primers were designed based on the P. falciparum genome resource, and these facilitated the cloning of a gene coding for pfMDH from a local clinical isolate. The DNA sequence of the cloned gene revealed an open-reading frame that encodes a

M. Chan; T. S. Sim

2004-01-01

188

Deletion of lactate dehydrogenase in Enterobacter aerogenes to enhance 2,3-butanediol production.  

PubMed

2,3-Butanediol is an important bio-based chemical product, because it can be converted into several C4 industrial chemicals. In this study, a lactate dehydrogenase-deleted mutant was constructed to improve 2,3-butanediol productivity in Enterobacter aerogenes. To delete the gene encoding lactate dehydrogenase, ? Red recombination method was successfully adapted for E. aerogenes. The resulting strain produced a very small amount of lactate and 16.7% more 2,3-butanediol than that of the wild-type strain in batch fermentation. The mutant and its parental strain were then cultured with six different carbon sources, and the mutant showed higher carbon source consumption and microbial growth rates in all media. The 2,3-butanediol titer reached 69.5 g/l in 54 h during fed-batch fermentation with the mutant,which was 27.4% higher than that with the parental strain.With further optimization of the medium and aeration conditions,118.05 g/l 2,3-butanediol was produced in 54 h during fed-batch fermentation with the mutant. This is by far the highest titer of 2,3-butanediol with E. aerogenes achieved by metabolic pathway engineering. PMID:22297429

Jung, Moo-Young; Ng, Chiam Yu; Song, Hyohak; Lee, Jinwon; Oh, Min-Kyu

2012-07-01

189

The reaction of trimethylamine dehydrogenase with diethylmethylamine.  

PubMed

The reductive half-reaction of trimethylamine dehydrogenase has been studied using the substrate diethylmethylamine over the pH range 6-10. It is found that the reaction occurs with three distinct and, under most conditions, fully resolved kinetic phases. The hyperbolic substrate concentration dependence of the observed rate constant for the fast kinetic phase is consistent with a two-step scheme in which free enzyme and substrate are in rapid equilibrium with an enzyme-substrate Michaelis complex, which then reacts to produce reduced flavin. The pH dependence of the limiting rate constant for the fast phase of the reaction (klim) exhibits a pKa value of 7.9, whereas klim/Kd exhibits a pKa value of 8.7. The rate constant for the intermediate kinetic phase, which reflects intramolecular electron transfer, is controlled by an ionizable group exhibiting a pKa value of 7.3. The equilibrium distribution of reducing equivalents between the flavin and iron-sulfur center of the enzyme at the end of the intermediate phase is controlled by a different ionizable group exhibiting a pKa value of approximately 6. An overall reductive half-reaction kinetic mechanism is proposed involving formation and decay of a covalent substrate-flavin intermediate, with intrinsically rapid intramolecular electron transfer limited by the rate of decay of the covalent adduct. Under conditions of excess substrate, product release is followed by binding of a second substrate molecule, which results in full development of the spectral properties diagnostic of the spin-interacting state. PMID:7983019

Rohlfs, R J; Hille, R

1994-12-01

190

Changes in antibody specificities and cytokine release after infection with lactate dehydrogenase-elevating virus.  

PubMed

Lactate dehydrogenase-elevating virus (LDV) is an apparently innocuous and persistent virus that can modify mouse immune reactions. We have shown that LDV-infected mice immunized with human growth hormone (hGH) showed a deep modification of the specificity of the anti-hGH antibodies (Ab) in CBA/Ht mice but not BALB/c animals. The aim of this work was to extend the previous observations to another mouse strain, C57BL/6, as well as to an antigen unrelated to hGH, ovalbumin (OVA), and to explore at the same time the production of various cytokines at serum and cellular levels. The amount of Ab directed to hGH or OVA native antigenic determinants versus the concentration of Ab to cryptic epitopes was evaluated by ELISA competition experiments. Results indicated that LDV infection affected Ab specificity solely in CBA/Ht mice. In CBA/Ht the virus infection was associated with a reduction of the Ab titers to hGH native epitopes and with a decrease of IL-13 and IL-17 serum levels, but Ab to native OVA epitopes were increased with a simultaneous increase of IL-17. Accordingly, only lymph node cells from infected CBA/Ht mice immunized with OVA were found to produce INF-?, IL-13 and IL-17. Thus, a correlation of cytokine production with a change in Ab specificity after a viral infection was found, although this phenomenon was restricted to a given antigen and to the genetic background of immunized animals. These observations suggest that an apparent harmless virus can affect some immunological mechanisms, which could lead, for example, to inflammatory or autoimmune disorders. PMID:23391715

Aparicio, José L; Saxena, Anubha; Coutelier, Jean-Paul; Van Snick, Jacques; Retegui, Lilia A

2013-02-05

191

Quinone-dependent D-lactate dehydrogenase Dld (Cg1027) is essential for growth of Corynebacterium glutamicum on D-lactate  

PubMed Central

Background Corynebacterium glutamicum is able to grow with lactate as sole or combined carbon and energy source. Quinone-dependent L-lactate dehydrogenase LldD is known to be essential for utilization of L-lactate by C. glutamicum. D-lactate also serves as sole carbon source for C. glutamicum ATCC 13032. Results Here, the gene cg1027 was shown to encode the quinone-dependent D-lactate dehydrogenase (Dld) by enzymatic analysis of the protein purified from recombinant E. coli. The absorption spectrum of purified Dld indicated the presence of FAD as bound cofactor. Inactivation of dld resulted in the loss of the ability to grow with D-lactate, which could be restored by plasmid-borne expression of dld. Heterologous expression of dld from C. glutamicum ATCC 13032 in C. efficiens enabled this species to grow with D-lactate as sole carbon source. Homologs of dld of C. glutamicum ATCC 13032 are not encoded in the sequenced genomes of other corynebacteria and mycobacteria. However, the dld locus of C. glutamicum ATCC 13032 shares 2367 bp of 2372 bp identical nucleotides with the dld locus of Propionibacterium freudenreichii subsp. shermanii, a bacterium used in Swiss-type cheese making. Both loci are flanked by insertion sequences of the same family suggesting a possible event of horizontal gene transfer. Conclusions Cg1067 encodes quinone-dependent D-lactate dehydrogenase Dld of Corynebacterium glutamicum. Dld is essential for growth with D-lactate as sole carbon source. The genomic region of dld likely has been acquired by horizontal gene transfer.

2010-01-01

192

Simultaneous purification of L-malate dehydrogenase and L-lactate dehydrogenase from bovine heart by biomimetic-dye affinity chromatography  

Microsoft Academic Search

Two commercially important enzymes, L-lactate dehydrogenase (LDH) and L-malate dehydrogenase (MDH) were purified simultaneously from bovine heart, on an agarose affinity adsorbent. This adsorbent bears a dye-ligand composed of an anthraquinone chlorotriazine chromophore linked to a biomimetic terminal 4-aminophenyloxanylic acid moiety. The purification protocol exploited the biomimetic affinity adsorbent, in combination with a cross-linked agarose DEAE anion-exchanger. The procedure comprised

N. E. Labrou; Y. D. Clonis

1997-01-01

193

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.

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

2010-01-01

194

Isolation and characterization of two cDNA clones of anaerobically induced lactate dehydrogenase from barley roots  

SciTech Connect

In barley roots during hypoxia, five lactate dehydrogenase (LDH) isozymes accumulate with a concomitant increase in enzyme activity ({approximately}20-fold). These isozymes are thought to be tetramers resulting from the random association of the products of two Ldh loci. To investigate this system, cDNA clones of LDH have been isolated from a {lambda}gt11 library using antiserum raised against barley LDH purified {approximately}3,000-fold and using nucleic acid probes synthesized by the polymerase chain reaction. Two cDNA clones were obtained (1,305 and 1,166 bp). The deduced amino acid sequences of the two barley LDHs are 96% identical to each other, and 50% and 40% identical to vertebrate and bacterial LDHs, respectively. Northern blots identified a single mRNA band ({approximately}1.5 kb) whose level rose 8-fold during hypoxia.

Hondred, D.; Hanson, A.D. (Michigan State Univ., East Lansing (USA))

1990-05-01

195

Cloning of a lactate dehydrogenase gene from Clostridium acetobutylicum B643 and expression in Escherichia coli  

SciTech Connect

A lactate dehydrogenase (LDH) gene of Clostridium acetobutylicum B643 was cloned on two recombinant plasmids. pPC37 and pPC58, that were selected by complementation of Escherichia coli PRC436 (acd), a fermentation-defective mutant that does not grown anaerobically on glucose. E. coli PRC436(pPC37) and PRC436(pPC58) grew anaerobically and fermented glucose to mostly lactate. When pPC37 and pPC58 were transformed into E. coli FMJ39 (ldh pfl), an LDH-deficient strain, the resulting strains grew anaerobically on glucose and produced lactate. Crude extracts of E. coli FMJ39(pPC37) and FMJ39(pPC58) contained high LDH activity only when assayed for pyruvate reduction to lactate, and the LDH activity was activated 15- to 30-fold by the addition of fructose 1,6-diphosphate (FDP). E. coli FMJ39 had no detectable LDH activity, and E. coli LDH from a wild-type strain was not activated by FDP. Maxicell analysis showed that both plasmids pPC37 and pPC58 expressed a protein with an apparent M{sub r} of 38,000 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Restriction endonuclease mapping of pPC37 and pPC58 and DNA hybridization studies indicated that a 2.1-kb region of these two clones of C. acetobutylicum DNA encodes the FDP-activated LDH.

Contag, P.R.; Williams, M.G.; Rogers, P. (Univ. of Minnesota, Minneapolis (USA))

1990-12-01

196

Conformational stability of leukocyte lactate dehydrogenase in healthy men of different age.  

PubMed

Comparative analysis of cytosolic fraction distribution and conformational stability in lactate dehydrogenase (LDH) isozymes in healthy men of different age revealed significant differences indicating age-related characteristics of the enzyme. Activity of cytosolic fraction of LDH-1 and conformational stability of LDH-2 and LDH-3 in 40-year-old men were higher than in 20-year-olds. These data suggest age-related determination of conformational stability of individual LDH isozymes and functional heterogeneity of the enzyme within each isoform. PMID:23330087

Fedorenko, O Yu; Bokhan, N A; Ivanova, S A

2012-11-01

197

Photoaffinity labelling of lactate dehydrogenase from pig heart with a bifunctional NAD +-analogue  

Microsoft Academic Search

P1-N6-(4-azidophenylethyl)adenosine-P2-4-(3-azidopyridinio)butyl diphosphate was synthesized with an [8-14C]adenine label. This bifunctional photoaffinity labelling reagent inactivates lactate dehydrogenase from pig heart upon irradiation with light of wavelength 300–380 nm. Stoichiometry of binding and enzymatic parameters suggest that the analogue is bound to the coenzyme binding site and that adjacent residues are modified. Four radioactive peptides were isolated by reverse-phase HPLC after tryptic

Susanne Becker; Tomas Bergman; Lars Hjelmqvist; Reinhard Jeck; Hans Jörnvall; Hanno Leibrock; Christoph Woenckhaus

1996-01-01

198

Molecular, genetic and biochemical characterization of lactate dehydrogenase-A enzyme activity mutations in Mus musculus  

Microsoft Academic Search

Four independent heterozygous lactate dehydrogenase (LDH) mutations with approximately 60% of wild-type enzyme activity in\\u000a whole blood have been recovered. The mutant line Ldh1\\u000a a2Neu proved to be homozygous lethal, whereas for the three lines Ldh1\\u000a a7Neu, Ldh1\\u000a a11Neu, and Ldh1\\u000a a12Neu homozygous mutants with about 20% residual activity occurred in the progeny of heterozygous inter se matings. However, the

Walter Pretsch; Bimal Chatterjee; Jack Favor; Siegbert Merkle; Rodica Sandulache

1998-01-01

199

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

200

Antibody-Independent Thrombocytopenia in Lactate Dehydrogenase-Elevating Virus-Infected Mice  

PubMed Central

Previously we demonstrated that antibody-mediated thrombocytopenia is strongly enhanced by lactate dehydrogenase-elevating virus (LDV) infection. Here we report that mice infected with LDV develop a moderate thrombocytopenia, even in the absence of immunoglobulins or Fc receptors. A similar decrease of platelet counts was observed after mouse hepatitis virus infection. LDV-induced type I interferon-independent thrombocytopenia was partly suppressed by treatment with clodronate-containing liposomes. Therefore, we conclude that the thrombocytopenia results from increased phagocytosis of nonopsonized platelets by macrophages.

Su, Dan; Musaji, Andrei; Legrain, Sarah; Detalle, Laurent; van der Kaa, Jos; Verbeek, J. Sjef; Ryffel, Bernhard

2012-01-01

201

Antibody-independent thrombocytopenia in lactate dehydrogenase-elevating virus-infected mice.  

PubMed

Previously we demonstrated that antibody-mediated thrombocytopenia is strongly enhanced by lactate dehydrogenase-elevating virus (LDV) infection. Here we report that mice infected with LDV develop a moderate thrombocytopenia, even in the absence of immunoglobulins or Fc receptors. A similar decrease of platelet counts was observed after mouse hepatitis virus infection. LDV-induced type I interferon-independent thrombocytopenia was partly suppressed by treatment with clodronate-containing liposomes. Therefore, we conclude that the thrombocytopenia results from increased phagocytosis of nonopsonized platelets by macrophages. PMID:22933286

Su, Dan; Musaji, Andrei; Legrain, Sarah; Detalle, Laurent; van der Kaa, Jos; Verbeek, J Sjef; Ryffel, Bernhard; Coutelier, Jean-Paul

2012-08-29

202

Characterization of D-lactate dehydrogenase from Pediococcus acidilactici that converts phenylpyruvic acid into phenyllactic acid.  

PubMed

The gene coding for D-lactate dehydrogenase (D-LDH) from Pediococcus acidilactici DSM 20284 was cloned and expressed in E. coli. The recombinant enzyme was purified by nickel-affinity chromatography. It converted phenylpyruvic acid (PPA) to 3-phenyllactic acid maximally at 30°C and pH 5.5 with a specific activity of 140 and 422 U/mg for PPA and pyruvate, respectively. The K(m), turnover number (k(cat)), and catalytic efficiency (k(cat)/K(m)) for PPA were 2.9 mM, 305 s(-1), and 105 mM(-1) s(-1), respectively. PMID:22261863

Mu, Wanmeng; Yu, Shuhuai; Jiang, Bo; Li, Xingfeng

2012-01-20

203

The influence of oxygen on radiation-induced structural and functional changes in glyceraldehyde-3-phosphate dehydrogenase and lactate dehydrogenase  

NASA Astrophysics Data System (ADS)

Proteins are major targets for oxidative damage due to their abundance in cells and high reactivity with free radicals. In the present study we examined the influence of oxygen on radiation-induced inactivation and structural changes of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and lactate dehydrogenase (LDH). We chose these two enzymes because they occur at high concentrations and participate in the most important processes in organisms; furthermore, they show considerable similarity in their structure.Protein solutions were irradiated with X-rays in doses ranging from 0.1 to 0.7 kGy, in air and N2O. The much higher radiation inactivation of GAPDH as compared to LDH is correlated with substantially greater structural changes in this protein, mainly involving the loss of free thiol groups (-SH). Of lesser importance in the differentiation of the radiosensitivity of the studied enzymes are tryptophan residues. Molecular oxygen, present during irradiation, increased to a significantly greater extent the inactivation and structural changes of GAPDH than that of LDH. The results suggest that the greater effect of oxygen on GAPDH is due to the higher efficiency of the superoxide radical, the higher amount of hydroperoxides generated, and the higher degree of unfolding of this protein.

Rodacka, Aleksandra; Serafin, Eligiusz; Bubinski, Michal; Krokosz, Anita; Puchala, Mieczyslaw

2012-07-01

204

Regulation of lactate dehydrogenase (LDH) and alcohol dehydrogenase (ADH) synthesis in liver nuclei, following their transfer into oocytes.  

PubMed

The regulatory effect of oocyte cytoplasm on the synthetic activity of transferred somatic cell nuclei was studied using an interspecific hybrid combination of Ambystoma texanum and Ambystoma mexicanum (axolotl). The enzymes lactate dehydrogenase (LDH) and alcohol dehydrogenase (ADH) were used as markers of gene activity. In both species of salamanders, LDH is synthesized in the liver and oocytes, while ADH is tissue-specific being synthesized in the liver but not oocytes. Both LDH and ADH show species-specific patterns on starch gels which permit detection of enzymes synthesized by texanum liver nuclei following their transfer into axolotl oocytes. Analysis of recipient oocytes after 1-3 weeks in culture reveals the presence of newly synthesized texanum LDH but not ADH. These results indicate that the transferred texanum liver cell nuclei continue to synthesize a product (LDH) found in both liver cells and oocytes, but fail to synthesize the liver-specific product (ADH) which is normally absent in oocytes. Thus, in the case of ADH and LDH the oocyte cytoplasm appears to be able to regulate the synthetic activity of the transferred somatic cell nuclei so as to conform to the oocytes' normal synthetic output. PMID:12194433

Etkin, L D

1976-09-01

205

The distribution of lactate dehydrogenase (Lactate tetrazolium reductase) in the hippocamal region of the rat. A reinvestigation with the polyvinyl alcohol method  

Microsoft Academic Search

The histochemical distribution of lactate dehydrogenase (LDH) in the hippocampal region of the rat was studied using a viscous incubation medium with polyvinyl alcohol (PVA) to reduce diffusion from fresh frozen sections. A relatively high concentration of Nitro blue tetrazolium was used, and the sections were incubated with phenazine methosulfate (PMS) to render the LDH demonstration independent of endogenous diaphorase

Svein Ivar Mellgren

1971-01-01

206

Mitochondrial Lactate Dehydrogenase Is Involved in Oxidative-Energy Metabolism in Human Astrocytoma Cells (CCF-STTG1)  

PubMed Central

Lactate has long been regarded as an end product of anaerobic energy production and its fate in cerebral metabolism has not been precisely delineated. In this report, we demonstrate, for the first time, the ability of a human astrocytic cell line (CCF-STTG1) to consume lactate and to generate ATP via oxidative phosphorylation. 13C-NMR and HPLC analyses aided in the identification of tricarboxylic acid (TCA) cyle metabolites and ATP in the astrocytic mitochondria incubated with lactate. Oxamate, an inhibitor of lactate dehydrogenase (LDH), abolished mitochondrial lactate consumption. Electrophoretic and fluorescence microscopic analyses helped localize LDH in the mitochondria. Taken together, this study implicates lactate as an important contributor to ATP metabolism in the brain, a finding that may significantly change our notion of how this important organ manipulates its energy budget.

Lemire, Joseph; Mailloux, Ryan J.; Appanna, Vasu D.

2008-01-01

207

Catabolism of circulating enzymes: plasma clearance, endocytosis, and breakdown of lactate dehydrogenase-1 in rabbits  

SciTech Connect

Lactate dehydrogenase-1, intravenously injected into rabbits, was cleared with first-order kinetics (half-life 27 min), until at least 80% of the injected activity had disappeared from plasma. Radioactivity from injected SVI-labeled enzyme disappeared at this same rate. Trichloroacetic-acid-soluble breakdown products started to appear in the circulation shortly after injection of the labeled enzyme. Body scans of the rabbits for 80 min after injection of T I-labeled enzyme revealed rapid accumulation of label in the liver, peaking 10-20 min after injection. Subsequently, activity in the liver declined and radioactivity (probably labeled breakdown products of low molecular mass) steadily accumulated in the bladder. Tissue fractionation of liver, 19 min after injection of labeled enzyme, indicated that the radioactivity was present both in endosomes and in lysosomes, suggesting uptake by endocytosis, followed by breakdown in the lysosomes. Measurements of radioactivity in liver and plasma suggest that the liver is responsible for the breakdown of at least 75% of the injected enzyme. Radioautography of tissue sections of liver and spleen showed accumulated radioactivity in sinusoidal liver cells and red pulpa, respectively. These results are very similar to those for lactate dehydrogenase-5, creatine kinase MM, and several other enzymes that we have previously studied in rats.

Smit, M.J.; Beekhuis, H.; Duursma, A.M.; Bouma, J.M.; Gruber, M.

1988-12-01

208

Catabolism of circulating enzymes: plasma clearance, endocytosis, and breakdown of lactate dehydrogenase-1 in rabbits.  

PubMed

Lactate dehydrogenase-1 (EC 1.1.1.27), intravenously injected into rabbits, was cleared with first-order kinetics (half-life 27 min), until at least 80% of the injected activity had disappeared from plasma. Radioactivity from injected 125I-labeled enzyme disappeared at this same rate. Trichloroacetic-acid-soluble breakdown products started to appear in the circulation shortly after injection of the labeled enzyme. Body scans of the rabbits for 80 min after injection of 131I-labeled enzyme revealed rapid accumulation of label in the liver, peaking 10-20 min after injection. Subsequently, activity in the liver declined and radioactivity (probably labeled breakdown products of low molecular mass) steadily accumulated in the bladder. Tissue fractionation of liver, 19 min after injection of labeled enzyme, indicated that the radioactivity was present both in endosomes and in lysosomes, suggesting uptake by endocytosis, followed by breakdown in the lysosomes. Measurements of radioactivity in liver and plasma suggest that the liver is responsible for the breakdown of at least 75% of the injected enzyme. Radioautography of tissue sections of liver and spleen showed accumulated radioactivity in sinusoidal liver cells and red pulpa, respectively. These results are very similar to those for lactate dehydrogenase-5, creatine kinase MM, and several other enzymes that we have previously studied in rats. PMID:3197286

Smit, M J; Beekhuis, H; Duursma, A M; Bouma, J M; Gruber, M

1988-12-01

209

The topological structure and function of Echinococcus granulosus lactate dehydrogenase, a tegumental transmembrane protein.  

PubMed

Lactate dehydrogenase (LDH), a terminal glycolytic enzyme, is generally considered as a cytosolic protein. We cloned lactate dehydrogenase from Echinococcus granulosus (EgLDH) and predicted it may be a membrane protein with two transmembrane regions through bioinformatics analysis. Intact worm immunofluorescence with antibodies prepared against linear B cell epitopes predicted in the region inside or outside of the membrane demonstrated that EgLDH spans the tegumental membrane twice, with the N terminal and C terminal all outside, just consistent with the putative topological structure. Then, the enzymatic characteristics and kinetic parameters of recombinant EgLDH were surveyed and the results suggested that EgLDH is responsible for catalyzing the reduction of pyruvic acid into lactic acid under physiological conditions. The enzymatic activity of the recombinant protein was inhibited by antibodies directed against the intact protein or against epitopes that contain key residues in the catalytic center or substrate binding sites. EgLDH is a potential target for drugs and vaccines against E. granulosus. PMID:22542488

Gan, Wenjia; Zhang, Zhaoping; Lv, Gang; Xu, Hongxu; Zeng, Suxiang; Li, Yuzhe; Wu, Weiping; Hu, Xuchu

2012-04-20

210

The expression of lactate dehydrogenase is important for the cell cycle of Toxoplasma gondii.  

PubMed

In Toxoplasma gondii, lactate dehydrogenase is encoded by two independent and developmentally regulated genes LDH1 and LDH2. These genes and their products have been implicated in the control of a metabolic flux during parasite differentiation. To investigate the significance of LDH1 and LDH2 in this process, we generated stable transgenic parasite lines in which the expression of these two expressed isoforms of lactate dehydrogenase was knocked down in a stage-specific manner. These LDH knockdown parasites exhibited variable growth rates in either the tachyzoite or the bradyzoite stage, as compared with the parental parasites. Their differentiation processes were impaired when the parasites were grown under in vitro conditions. In vivo studies in a murine model system revealed that tachyzoites of these parasite lines were unable to form significant numbers of tissue cysts and to establish a chronic infection. Most importantly, all mice that were initially infected with tachyzoites of either of the four LDH knockdown lines survived a subsequent challenge with tachyzoites of the parental parasites (10(4)), a dose that usually causes 100% mortality, suggesting that live vaccination of mice with the LDH knockdown tachyzoites can confer protection against T. gondii. Thus, we conclude that LDH expression is essential for parasite differentiation. The knockdown of LDH1 and LDH2 expression gave rise to virulence-attenuated parasites that were unable to exhibit a significant brain cyst burden in a murine model of chronic infection. PMID:15459194

Al-Anouti, Fatme; Tomavo, Stanislas; Parmley, Stephen; Ananvoranich, Sirinart

2004-09-30

211

Attempting to remove the substrate inhibition of L -lactate dehydrogenase from Bacillus stearothermophilus by site-directed mutagenesis  

Microsoft Academic Search

L-lactate dehydrogenase (LDH) catalyzes the interconversion of an oxoacid (pyruvate) and hydroxy-acid (lactate) using the NADH\\/NAD+ pair as a redox cofactor. The enzyme has a commercial significance, as it can be used to produce chiral building blocks for\\u000a the synthesis of key pharmaceuticals and agrochemicals. However, the substrate inhibition which is due to an abortive NAD+-pyruvate complex reducing the steady

Bari? Binay; Nevin Gül Karagüler

2007-01-01

212

Cloning and sequence analysis of a partial L-lactate dehydrogenase gene of Bacillus coagulans Tl4v  

Microsoft Academic Search

Bacillus coagulans TL-4 strain is a thermophilic bacterium for high L-lactate production. We reported here that its 779-bp partial NAD+-dependent L-lactate dehydrogenase (LDH) gene sequence (GenBank accession no. GU354320) encoding a deduced 259 amino acid peptide was cloned by using one pair of designed degenerate primers. The amino acid sequence alignment and the built phylogenetic tree revealed that this partial

Binzhe Wang; Qiang Gao; Bing Liu; Kunyu Jiang; Zhe Su

2010-01-01

213

Diagnosing melanoma patients entering American Joint Committee on Cancer stage IV, C-reactive protein in serum is superior to lactate dehydrogenase  

Microsoft Academic Search

Lactate dehydrogenase (LDH) in serum has recently been introduced into the American Joint Committee on Cancer (AJCC) staging system for cutaneous melanoma because of its prognostic value. We hypothesised LDH to be of value in discriminating melanoma patients entering AJCC stage IV from patients staying in AJCC stages I, II or III. Lactate dehydrogenase was compared to the acute phase

M Deichmann; B Kahle; K Moser; J Wacker; K Wüst

2004-01-01

214

Evidence for the nonhydrophobic interaction of aromatic, nitrogen-containing compounds with the coenzyme binding site of a NAD-linked D-lactate dehydrogenase  

Microsoft Academic Search

Summary Quantitative structure activity relationships suggest that the binding of quinoline and phenanthroline analogs to D-lactate dehydrogenase from the barnacle,Balanus nubilus Darwin, does not involve primarily hydrophobic effects. This phenomenon appears to exist also for other lactate dehydrogenases.

G. L. Long; J. R. Cook; W. R. Ellington

1978-01-01

215

[Myoglobinuria due to enzyme abnormalities in glycolytic pathway--especially lactate dehydrogenase M subunit deficiency].  

PubMed

Glycolysis is an important energy productive system. Enzyme abnormalities the in glycolytic pathway, which cause myoglobinuria, are deficiencies of phosphofructokinase, phosphoglycerate kinase, phosphoglycerate mutase, and lactate dehydrogenase (LDH). Common symptoms of these enzyme abnormalities are muscle cramp, muscle pain, and rhabdomyolysis after strenuous exercise. Acute renal failure owing to myoglobinuria is the most noteworthy symptom. In daily life, symptoms are rarely observed and prognosis is usually good. Correct and fast diagnosis of such latent symptomatic disorders is important to prevent a turn for the worse of these symptoms. LDH M subunit deficiency was first discovered by urinary discoloration and a discrepancy of laboratory data. Since then, only four cases have been reported in the Japanese population. The response to ischemic forearm work is characteristic (an increase of venous lactate concentration after ischemic work is not observed and a marked increase of venous pyruvate is found). The increase of pyruvate concentration is specific in LDH-M subunit deficiency, and is not observed in other abnormalities of the glycolytic pathway. Glycolysis was markedly retarded in the patient's muscle in the glyceraldehyde 3-phosphate dehydrogenase (GA3PD) step, possibly due to the impaired reoxidation of NADH produced by GA3PD activity. Then, the excess NADH is reoxidized by alpha-glycerophosphate dehydrogenase and triose phosphates are drained to alpha-glycerophosphate and glycerol. Therefore ATP production is significantly impaired and muscle tissue is damaged. A genetical study revealed a deletion of 20 base-pairs in exon 6 in LDH-M subunit deficiency. This mutation results in a frame-shift translation and premature termination. PMID:1828277

Maekawa, M; Kanno, T; Sudo, K

1991-02-01

216

Biotransformation of R-2-hydroxy-4-phenylbutyric acid by D-lactate dehydrogenase and Candida boidinii cells containing formate dehydrogenase coimmobilized in a fibrous bed bioreactor.  

PubMed

R-2-hydroxy-4-phenylbutyric acid (R-HPBA) is an important intermediate in the manufacture of angiotensin converting enzyme inhibitors. In this work, a recombinant D-lactate dehydrogenase (LDH) was used to transform 2-oxo-4-phenylbutyric acid (OPBA) to R-HPBA, with concomitant oxidation of beta-nicotinamide adenine dinucleotide (NADH) to NAD(+). The cofactor NADH was regenerated by formate dehydrogenase (FDH) present in whole cells of Candida boidinii, which were pre-treated with toluene to make them permeable. The whole cells used in the process were more stable and easier to prepare as compared with the isolated FDH from the cells. Kinetic study showed that the reaction rate was dependent on the concentration of cofactor, NAD(+), and that both R-HPBA and OPBA inhibited the reaction. A novel method for co-immobilization of whole cells and LDH enzyme on cotton cloth was developed using polyethyleneimine (PEI), which induced the formation of PEI-enzyme-cell aggregates and their adsorption onto cotton cloth, leading to multilayer co-immobilization of cells and enzyme with high loading (0.5 g cell and 8 mg LDH per gram of cotton cloth) and activity yield ( > 95%). A fibrous bed bioreactor with co-immobilized cells and enzyme on the cotton cloth was then evaluated for R-HPBA production in fed-batch and repeated batch modes, which gave relatively stable reactor productivity of 9 g/L . h and product yield of 0.95 mol/mol OPBA when the concentrations of OPBA and R-HPBA were less than 10 g/L. PMID:16037987

Bai, Yunling; Yang, Shang-Tian

2005-10-20

217

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.

Abboud, Jean

2013-01-01

218

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

PubMed

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 V max, K m and I 50 values between control and frozen LDH. However, control and dehydrated LDH differed significantly in K m values for pyruvate, lactate, and NAD, I 50 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; Storey, Kenneth B

2013-02-12

219

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...

Zammit, V A; Newsholme, E A

1976-01-01

220

Compartmentation of cytosolic dehydrogenases studied by transfer of tritium from labelled substrates into lactate in rat hepatocytes.  

PubMed

This paper describes the transfer of tritium from [2-(3)H]xylitol or (1R)-[1-(3)H]ethanol into lactate in cells from fed rats either untreated or triiodothyronine-treated. The labelling pattern of lactate during the metabolism of [2-(3)H]xylitol or (1R)-[1-(3)H]ethanol follows the equation L = K(1 - e-t/tau) (mumol tritium/mumol lactate). The yield in lactate together with the minimum value of the total flux of reducing equivalents are used to estimate the specific radioactivity of NADH. We have calculated the lactate dehydrogenase-catalysed oxidation rate of NADH from the experimental values of lactate labelling and the specific radioactivity of NADH. We found the calculated flux of reducing equivalents from NADH to pyruvate to be of the same order of magnitude whether labelled ethanol or labelled xylitol was metabolized. We found the flux to be only a few percent of the maximal activity of lactate dehydrogenase. The results obtained suggest that the cytoplasm can be regarded as one compartment, containing a single pool of NAD(H). PMID:7049253

Vind, C; Grunnet, N

1982-06-01

221

Probing Lactate Dehydrogenase Activity in Tumors by Measuring Hydrogen/Deuterium Exchange in Hyperpolarized l-[1-13C,U-2H]Lactate  

PubMed Central

13C magnetic resonance spectroscopy and spectroscopic imaging measurements of hyperpolarized 13C label exchange between exogenously administered [1-13C]pyruvate and endogenous lactate, catalyzed by lactate dehydrogenase (LDH), has proved to be a powerful approach for probing tissue metabolism in vivo. This experiment has clinical potential, particularly in oncology, where it could be used to assess tumor grade and response to treatment. A limitation of the method is that pyruvate must be administered in vivo at supra-physiological concentrations. This problem can be avoided by using hyperpolarized [1-13C]lactate, which can be used at physiological concentrations. However, sensitivity is limited in this case by the relatively small pyruvate pool size, which would result in only low levels of labeled pyruvate being observed even if there was complete label equilibration between the lactate and pyruvate pools. We demonstrate here a more sensitive method in which a doubly labeled lactate species can be used to measure LDH-catalyzed exchange in vivo. In this experiment exchange of the C2 deuterium label between injected hyperpolarized l-[1-13C,U-2H]lactate and endogenous unlabeled lactate is observed indirectly by monitoring phase modulation of the spin-coupled hyperpolarized 13C signal in a heteronuclear 1H/13C spin–echo experiment.

2012-01-01

222

A Quantitative Assay of Recombinant Malarial Lactate Dehydrogenase as a Platform for Screening Inhibitors from Crude Herbal Extracts  

Microsoft Academic Search

The potential of malaria parasite (Plasmodium falciparum) generating resistance to currently used antimalarial drugs is a major scourge for malaria patients. The multiple targets on which antimalarial drugs act hinder the occurrence of such resistance. As an essential metabolic enzyme involved in energy production in the parasite, the lactate dehydrogenase of P. falciparum (PfLDH) provides an alternative target for parasite

Xiao-Ling XU; Rui-Yi YANG; Xue-Qin YANG; Pei-Quan CHEN; Qing-Ping ZENG

2007-01-01

223

The Detection of Pentachlorophenol by Its Inhibitory Effectiveness on Lactate Dehydrogenase of Rabbit Muscle and Bovine Heart  

Microsoft Academic Search

There is a requirement for on-line monitoring of pentachlorophenol (PCP) in water supplies. One method of PCP detection may be to use enzyme-inhibition biosensors. In the present study, assay conditions were optimised, comparing two isozymes of lactate dehydrogenase (LDH) with regard to inhibition by PCP. Rabbit muscle LDH displayed greater inhibition of enzyme activity when exposed to 70 ?M PCP

Sarah J. Young; Antony A. Dowman; David C. Cowell

1999-01-01

224

An Immunoglobulin A1 that Inhibits Lactate Dehydrogenase Activity, with Reversal of Inhibition by Addition of NADH  

Microsoft Academic Search

We discovered a patient with low serum lactate dehydrogenase (LD) activity and an abnormal LD isozyme pattern. We analyzed the patient's LD inhibitor using electrophoresis, affinity chromatography, and immunochemical technologies. The LD activity of the patient's serum was inhibited more strongly at 4°C than at 37°C. The decrease of LD activity was more marked in a mixture of the patient's

Kiyotaka Fujita; Hirohisa Sato; Fumiko Kameko; Fumiko Terasawa; Nobuo Okumura; Mitsutoshi Sugano; Kazuyoshi Yamauchi; Masato Maekawa; Ikunosuke Sakurabayashi

225

Nuclear magnetic resonance and molecular genetic studies of the membrane-bound D-lactate dehydrogenase of Escherichia coli  

SciTech Connect

In this study the authors demonstrate the potential of combining fluorine-19 nuclear magnetic resonance (NMR) spectroscopy with molecular genetics. They are using the membrane-bound enzyme D-lactate dehydrogenase of Escherichia coli as a model system to characterize interactions between proteins and lipids. They have labeled D-lactate dehydrogenase with 4-, 5-, and 6-fluorotryptophans and obtained high-resolution fluorine-19-NMR spectra showing five resonances, in agreement with the five tryptophan residues expected from the DNA sequence. The five /sup 19/F resonances in the spectra have been assigned to the specific tryptophan residues in the primary sequence of D-lactate dehydrogenase by site-directed oligonucleotide mutagenesis of the cloned gene. They observe large differences in the relative fluorine-19 chemical shifts of each tryptophan residue when labeled by different isomers of fluorotryptophan. On the basis of /sup 18/F NMR spectroscopy of the labeled tryptophan residues, the conformation of D-lactate dehydrogenase is similar in aqueous solution and in the present of a variety of lipids and detergents. This result indicates that the presence of lipids or detergents is not required to maintain the tertiary structure of this membrane-bound enzyme. In contrast, Triton X-100 induces a change to an abnormal conformation of the enzyme as judged from both NMR spectroscopy and the effect of temperature on the maximal velocity of the enzyme in the presence of this detergent.

Rule, G.S.; Pratt, E.A.; Simplaceanu, V.; Ho, C.

1987-01-27

226

The Quaternary Structure of Lactate Dehydrogenase. I. The Subunit Molecular Weight and the Reversible Association at Acid PH.  

National Technical Information Service (NTIS)

The monomer subunit molecular weight of lactate dehydrogenase has been shown to be 18,000 by sedimentation equilibrium studies in 6.2 M guanidine hydrochloride at pH 2. It was demonstrated that 7 M guanidine hydrochloride at pH 7 was not sufficient to com...

D. B. Millar V. Frattali G. E. Willick

1968-01-01

227

Pyruvate Dehydrogenase and Pyruvate Dehydrogenase Kinase Expression in Non Small Cell Lung Cancer and Tumor-Associated Stroma  

Microsoft Academic Search

Pyruvate dehydrogenase (PDH) catalyzes the conver- sion 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 hyp- oxic conditions, conversion of pyruvate to lactate occurs, a reaction catalyzed by lactate dehydrogenase 5 (LDH5). In cancer cells, however, pyruvate is

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

2005-01-01

228

Purification and Properties of Hypoxically Induced Lactate Dehydrogenase from Barley Roots 1  

PubMed Central

Using Affigel Blue and oxamate-agarose affinity chromatography, lactate dehydrogenase (LDH) was purified 2000-fold from hypoxically induced barley roots. Molecular weights of the native and sodium dodecyl sulfate-denatured LDH protein were 157 and 40 kilodaltons, respectively, indicating a tetramer. Purified barley LDH was very similar in size and kinetic properties to potato LDH. However, their amino acid compositions differed substantially and antibodies raised against barley LDH did not cross-react with potato LDH on immunoblots, implying that the barley and potato LDHs are not closely related proteins. In vivo [35S] methionine labeling and immunoprecipitation experiments indicated that hypoxia increased the rate of LDH protein synthesis, and immunoblot analysis showed that LDH protein levels rose during hypoxia. We conclude that increased enzyme synthesis plays a major part in the induction of LDH enzyme activity by low O2 levels in barley roots. Images Fig. 2 Fig. 4 Fig. 5 Fig. 6 Fig. 7

Hoffman, Neil E.; Hanson, Andrew D.

1986-01-01

229

Redox balance via lactate dehydrogenase is important for multiple stress resistance and virulence in Enterococcus faecalis.  

PubMed

Enterococcus faecalis is a highly stress resistant opportunistic pathogen. The intrinsic ruggedness of this bacterium is supposed to be the basis of its capacity to colonize the hostile environments of hospitals and to cause several kinds of infections. We show in this work that general resistance to very different environmental stresses depends on the ability of E. faecalis to maintain redox balance via lactate dehydrogenase (LDH). Furthermore, LDH-deficient mutants are less successful than the wild type at colonizing host organs in a murine model of systemic infection. Taken together, our results, as well as those previously published for Staphylococcus aureus (A. R. Richardson, S. J. Libby, and F. C. Fang, Science 319:1672-1676, 2008), identify LDH as an attractive drug target. These drugs may have additional applications, as in the fight against glycopeptide antibiotic-resistant bacteria and even cancer. PMID:23649090

Rana, Nosheen Fatima; Sauvageot, Nicolas; Laplace, Jean-Marie; Bao, Yinyin; Nes, Ingolf; Rincé, Alain; Posteraro, Brunella; Sanguinetti, Maurizio; Hartke, Axel

2013-05-06

230

Active and inactive state structures of unliganded Lactobacillus casei allosteric L-lactate dehydrogenase.  

PubMed

Lactobacillus casei L-lactate dehydrogenase (LCLDH) is activated through the homotropic and heterotropic activation effects of pyruvate and fructose 1,6-bisphosphate (FBP), respectively, and exhibits unusually high pH-dependence in the allosteric effects of these ligands. The active (R) and inactive (T) state structures of unliganded LCLDH were determined at 2.5 and 2.6 A resolution, respectively. In the catalytic site, the structural rearrangements are concerned mostly in switching of the orientation of Arg171 through the flexible intersubunit contact at the Q-axis subunit interface. The distorted orientation of Arg171 in the T state is stabilized by a unique intra-helix salt bridge between Arg171 and Glu178, which is in striking contrast to the multiple intersubunit salt bridges in Lactobacillus pentosus nonallosteric L-lactate dehydrogenase. In the backbone structure, major structural rearrangements of LCLDH are focused in two mobile regions of the catalytic domain. The two regions form an intersubunit linkage through contact at the P-axis subunit interface involving Arg185, replacement of which with Gln severely decreases the homotropic and hetertropic activation effects on the enzyme. These two regions form another intersubunit linkage in the Q-axis related dimer through the rigid NAD-binding domain, and thus constitute a pivotal frame of the intersubunit linkage for the allosteric motion, which is coupled with the concerted structural change of the four subunits in a tetramer, and of the binding sites for pyruvate and FBP. The unique intersubunit salt bridges, which are observed only in the R state structure, are likely involved in the pH-dependent allosteric equilibrium. PMID:19787773

Arai, Kazuhito; Ishimitsu, Toshihiro; Fushinobu, Shinya; Uchikoba, Hiroyuki; Matsuzawa, Hiroshi; Taguchi, Hayao

2010-02-15

231

Effect of tubulin on the activity of the muscle isoenzyme of lactate dehydrogenase.  

PubMed

The interaction between muscle-type lactate dehydrogenase (LDHm) and tubulin was investigated by monitoring the combined effect of NADH and tubulin on steady-state kinetics and the combined effect of NADH and pH on complex formation between tubulin and the enzyme. Steady-state kinetics showed that LDHm is inhibited by tubulin. Experiments with heart-type lactate dehydrogenase (LDHh) showed that the inhibition is unique to the muscle-type enzyme. The magnitude of the inhibition is dependent upon the concentration of NADH as well as the pH of the buffer medium. The enzyme was less sensitive to inhibition at 50 microM NADH than at 10 microM NADH. Since this effect of NADH is not due to an ionic strength contribution, it is deemed to be specific. In contrast to the absence of tubulin, its presence induced a modification of the kinetic behavior of LDHm; i.e., the velocity dependence on NADH concentration displayed a marked sigmoid response. The inhibition of LDHm by tubulin is more pronounced at lower pH values than at higher pH values. The pH-dependent inhibitory profile is shifted to the left (i.e., pKa is decreased) with increasing concentrations of NADH. This pattern is remarkably similar to that observed for the binding of the enzyme on Sepharose immobilized tubulin and is consistent with the premise that inhibition is a result of interaction between these proteins. NAD+ was much less effective than NADH in dissociating LDHm from immobilized tubulin. Results from these in vitro studies are consistent with similar observations dealing with other glycolytic enzymes and cytoskeleton proteins, which show that enzyme catalytic properties are modified upon binding. PMID:7986093

Marmillot, P; Keith, T; Srivastava, D K; Knull, H R

1994-12-01

232

In situ regeneration of NADH via lipoamide dehydrogenase-catalyzed electron transfer reaction evidenced by spectroelectrochemistry  

PubMed Central

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 ?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

2012-01-01

233

Activity, stability and structural studies of lactate dehydrogenases adapted to extreme thermal environments.  

PubMed

Lactate dehydrogenase (LDH) catalyzes the conversion of pyruvate to lactate with concomitant oxidation of NADH during the last step in anaerobic glycolysis. In the present study, we present a comparative biochemical and structural analysis of various LDHs adapted to function over a large temperature range. The enzymes were from Champsocephalus gunnari (an Antarctic fish), Deinococcus radiodurans (a mesophilic bacterium) and Thermus thermophilus (a hyperthermophilic bacterium). The thermodynamic activation parameters of these LDHs indicated that temperature adaptation from hot to cold conditions was due to a decrease in the activation enthalpy and an increase in activation entropy. The crystal structures of these LDHs have been solved. Pairwise comparisons at the structural level, between hyperthermophilic versus mesophilic LDHs and mesophilic versus psychrophilic LDHs, have revealed that temperature adaptation is due to a few amino acid substitutions that are localized in critical regions of the enzyme. These substitutions, each having accumulating effects, play a role in either the conformational stability or the local flexibility or in both. Going from hot- to cold-adapted LDHs, the various substitutions have decreased the number of ion pairs, reduced the size of ionic networks, created unfavorable interactions involving charged residues and induced strong local disorder. The analysis of the LDHs adapted to extreme temperatures shed light on how evolutionary processes shift the subtle balance between overall stability and flexibility of an enzyme. PMID:17936781

Coquelle, Nicolas; Fioravanti, Emanuela; Weik, Martin; Vellieux, Frédéric; Madern, Dominique

2007-09-22

234

Epitopes of human testis-specific lactate dehydrogenase deduced from a cDNA sequence  

SciTech Connect

The sequence and structure of human testis-specific L-lactate dehydrogenase (LDHC/sub 4/, LDHX; (L)-lactate:NAD/sup +/ oxidoreductase, EC 1.1.1.27) has been derived from analysis of a complementary DNA (cDNA) clone comprising the complete protein coding region of the enzyme. From the deduced amino acid sequence, human LDHC/sub 4/ is as different from rodent LDHC/sub 4/ (73% homology) as it is from human LDHA/sub 4/ (76% homology) and porcine LDHB/sub 4/ (68% homology). Subunit homologies are consistent with the conclusion that the LDHC gene arose by at least two independent duplication events. Furthermore, the lower degree of homology between mouse and human LDHC/sub 4/ and the appearance of this isozyme late in evolution suggests a higher rate of mutation in the mammalian LDHC genes than in the LDHA and -B genes. Comparison of exposed amino acid residues of discrete anti-genic determinants of mouse and human LDHC/sub 4/ reveals significant differences. Knowledge of the human LDHC/sub 4/ sequence will help design human-specific peptides useful in the development of a contraceptive vaccine.

Millan, J.L.; Driscoll, C.E.; LeVan, K.M.; Goldberg, E.

1987-08-01

235

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

236

A molecular design that stabilizes active state in bacterial allosteric L-lactate dehydrogenases.  

PubMed

l-Lactate dehydrogenase (l-LDH) of Lactobacillus casei (LCLDH) is a typical bacterial allosteric l-LDH that requires fructose 1,6-bisphosphate (FBP) for its enzyme activity. A mutant LCLDH was designed to introduce an inter-subunit salt bridge network at the Q-axis subunit interface, mimicking Lactobacillus pentosus non-allosteric l-LDH (LPLDH). The mutant LCLDH exhibited high catalytic activity with hyperbolic pyruvate saturation curves independently of FBP, and virtually the equivalent K(m) and V(m) values at pH 5.0 to those of the fully activated wild-type enzyme with FBP, although the K(m) value was slightly improved with FBP or Mn(2+) at pH 7.0. The mutant enzyme exhibited a markedly higher apparent denaturating temperature (T(1/2)) than the wild-type enzyme in the presence of FBP, but showed an even lower T(1/2) without FBP, where it exhibited higher activation enthalpy of inactivation (?H(‡)). This result is consistent with the fact that the active state is more unstable than the inactive state in allosteric equilibrium of LCLDH. The LPLDH-like network appears to be conserved in many bacterial non-allosteric l-LDHs and dimeric l-malate dehydrogenases, and thus to be a key for the functional divergence of bacterial l-LDHs during evolution. PMID:21828088

Arai, Kazuhito; Ichikawa, Jun; Nonaka, Shinta; Miyanaga, Akimasa; Uchikoba, Hiroyuki; Fushinobu, Shinya; Taguchi, Hayao

2011-08-09

237

Design, synthesis, and biological evaluation of Plasmodium falciparum lactate dehydrogenase inhibitors.  

PubMed

Plasmodium falciparum lactate dehydrogenase (pfLDH) is a key enzyme for energy generation of malarial parasites and is a potential antimalarial chemotherapeutic target. It is known that the oxamate moiety, a pyruvate analog, alone shows higher inhibition against pfLDH than human LDHs, suggesting that it can be used for the development of selective inhibitors. Oxamic acid derivatives were designed and synthesized. Derivatives 5 and 7 demonstrated activities against pfLDH with IC50 values of 3.13 and 1.75 muM, respectively, and have 59- and 7-fold selectivity over mammalian LDH, respectively. They also have micromolar range activities against Plasmodium falciparum malate dehydrogenase (pfMDH), which may fill the role of pfLDH when the activity of pfLDH is reduced. Thus, certain members of these oxamic acid derivatives may have dual inhibitory activities against both pfLDH and pfMDH. It is presumed that dual LDH/MDH inhibitors would have enhanced potential as antimalarial drugs. PMID:17636950

Choi, Seoung-ryoung; Pradhan, Anupam; Hammond, Nicholas L; Chittiboyina, Amar G; Tekwani, Babu L; Avery, Mitchell A

2007-07-18

238

Assessment of Lactate Dehydrogenase, Alkaline Phosphatase and Aspartate Aminotransferase Activities in Cow’s Milk as an Indicator of Subclinical Mastitis  

Microsoft Academic Search

This study examined the activities of lactate dehydrogenase (LDH), alkaline phosphatase (ALP) and aspartate aminotransferase\\u000a (AST) in the milk of lactating Holstein cows in association with subclinical mastitis (SCM). A total of 94 milk samples were\\u000a collected from 58 lactating dairy cows representing stages of lactation from the second to the tenth week after calving. Those\\u000a which were classified as

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

2007-01-01

239

A comparison of biospecific affinity chromatographic methodologies for the purification of NAD +-dependent dehydrogenases: studies with bovine l-lactate dehydrogenase  

Microsoft Academic Search

The general potential of biospecific affinity chromatography in enzyme isolation and purification is well recognised. In the present study, three distinct approaches were assessed for the rapid, one-chromatographic-step affinity purification of l-lactate dehydrogenase (l-LDH, EC 1.1.1.27) from bovine heart: (i) the specific ligand approach using an immobilised oxamate derivative; (ii) the general ligand approach (immobilised NAD+ derivatives) in conjunction with

L Oakey; A Martung; M McMahon; M O’Flaherty; P Mulcahy

1999-01-01

240

Oxidation of D-lactate and L-lactate by Neisseria meningitidis: purification and cloning of meningococcal D-lactate dehydrogenase.  

PubMed Central

Neisseria meningitidis was found to contain at least two lactate-oxidizing enzymes. One of these was purified 460-fold from spheroplast membranes and found to be specific primarily for D-lactate, with low-affinity activity for L-lactate. The gene for this enzyme (dld) was cloned, and a dld mutant was constructed by insertional inactivation of the gene. The mutant was unable to grow on D-lactate but retained the ability to grow on L-lactate, providing evidence for a second lactate-oxidizing enzyme with specificity for L-lactate. High-affinity L-lactate-oxidizing activity was detected in intact bacteria of both the dld+ and dld mutant strains. This L-lactate-oxidizing activity was also seen in sonicated bacteria but was reduced substantially on detergent solubilization or on preparation of spheroplast membranes. Images

Erwin, A L; Gotschlich, E C

1993-01-01

241

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

242

Purification and determination of the binding site of lactate dehydrogenase from chicken breast muscle on immobilized ferric ions.  

PubMed

Lactate dehydrogenase from chicken breast muscle was purified to homogeneity in one step by immobilized metal ion affinity chromatography. The purified enzyme was used to localize the binding site to immobilized Fe(III) ions. After cyanogen bromide degradation and digestion with trypsin, small enzyme fragments capable of binding to immobilized Fe(III) ions were obtained. It is proposed that several histidyl groups are involved in the binding. PMID:1487526

Chaga, G; Andersson, L; Porath, J

1992-12-25

243

Regulation of cyclic GMP, cyclic AMP and lactate dehydrogenase by putative neurotransmitters in the C6 rat glioma cell line  

Microsoft Academic Search

In C6 cells norepinephrine and dopamine caused transient increases in cyclic GMP and cyclic AMP, as well as an induction of lactate dehydrogenase. All of these responses were blocked by l-propranolol, suggesting mediation by a ..beta..-receptor. Phentolamine potentiated the NE-increased cAMP levels by 5-fold when NE was used at suboptimal doses, suggesting the presence of ..cap alpha..-adrenergic receptors in C6

Jane E. Bottenstein; Jean de Vellis

1978-01-01

244

Regulation of cyclic GMP, cyclic amp and lactate dehydrogenase by putative neutrotransmitters in the C6 rat glioma cell line  

Microsoft Academic Search

In C6 cells norepinephrine and dopamine caused transient increases in cyclic GMP and cyclic AMP, as well as an induction of lactate dehydrogenase. All of these responses were blocked by 1-propranolol, suggesting mediation by a ..beta..-receptor. Phentolamine potentiated the NE-increased cAMP levels by 5-fold when NE was used at suboptimal doses, suggesting the presence of ..cap alpha..-adrenergic receptors in C6

J. E. Bottenstein; J. de Vellis

1977-01-01

245

Effects of Urea on M 4Lactate Dehydrogenase from Elasmobranchs and Urea-Accumulating Australian Desert Frogs  

Microsoft Academic Search

We measured the effect of urea on M4-lactate dehydrogenase (M4-LDH) from elasmobranchs and Australian desert frogs (urea accumulators) and from two animals that do not accumulate urea, the axolotl and the rabbit. An analysis of the effect of urea on the Kd(NADH), V, V\\/Km(pyr) and V\\/Km(NADH) shows that in all cases the major effect of urea was on the binding

Caroline J Fuery; Paul V Attwood; Philip C Withers; Paul H Yancey; John Baldwin; Michael Guppy

1997-01-01

246

Characteristic Changes in Platelet-Large Cell Ratio, Lactate Dehydrogenase and C-Reactive Protein in Thrombocytosis-Related Diseases  

Microsoft Academic Search

We examined the clinical usefulness of 3 parameters of routine laboratory tests [platelet-large cell ratio (P-LCR), lactate dehydrogenase (LDH) and C-reactive protein (CRP)] in 84 patients with thrombocytosis-related diseases (reactive thrombocytosis, chronic myeloid leukemia, essential thrombocythemia and polycythemia vera). These thrombocytosis-related diseases were characterized using the 3 parameters P-LCR, LDH and CRP as follows: high P-LCR and high LDH in

Osamu Kabutomori; Yuzuru Kanakura; Yoshinori Iwatani

2007-01-01

247

Differences in some properties of lactate dehydrogenase from muscles of the carp Cyprinus carpio and trout Salmo gairdneri  

Microsoft Academic Search

Some catalytic and kinetic properties of lactate dehydrogenase (LDH) isolated from trout and carp skeletal muscles were compared.\\u000a The specific activity of LDH in the carp muscle was lower by about one third than the activity in the trout muscle. Temperature\\u000a and pH optima for LDH isolated from the carp muscle were higher than those for the trout muscle LDH.

A. Tylicki; D. Masztaleruk; S. Strumilo

2006-01-01

248

Structural relatedness of mouse lactate dehydrogenase isozymes, A 4 (muscle), B 4 (heart), and C 4 (testis)  

Microsoft Academic Search

Three homotetrameric lactate dehydrogenase isozymes, LDH-M(A4), LDH-H(B4), and LDH-X(C4), from DBA\\/2J mice have been purified by affinity chromatography. The amino acid compositions of the subunits A, B, and C, based on a molecular weight of 36,000, have been determined. The compositional relatedness of these isozymes indicates that subunits A (muscle) and B (heart) are more closely related to each other

Shou-Mei T. Chang; Chi-Yu Lee; Steven S.-L. Li

1979-01-01

249

DNA sequence determination and biochemical analysis of the immunogenic protein P36, the lactate dehydrogenase (LDH) of Mycoplasma hyopneumoniae  

Microsoft Academic Search

The DNA sequence of the gene encoding the early and specific immunogenic protein P36 of Mycoplasmu hyopneumoniue has been determined. Comparison of the DNA sequence and the deduced amino acid sequence of P36 with known genes and proteins in data banks indicated that P36 is a L-lactate dehydrogenase (LDH) (EC 1.1.1.27). Biochemical analysis of protein P36 expressed from the cloned

ANDREAS HALDIMANN; JACQUES NICOLET; JOACHIM FREY

1993-01-01

250

Hot Spots in Cold Adaptation: Localized Increases in Conformational Flexibility in Lactate Dehydrogenase A4 Orthologs of Antarctic Notothenioid Fishes  

Microsoft Academic Search

To elucidate mechanisms of enzymatic adaptation to extreme cold, we determined kinetic properties, thermal stabilities, and deduced amino acid sequences of lactate dehydrogenase A4 (A4-LDH) from nine Antarctic (-1.86 to 1 degrees C) and three South American (4 to 10 degrees C) notothenioid teleosts. Higher Michaelis-Menten constants (Km) and catalytic rate constants (kcat) distinguish orthologs of Antarctic from those of

Peter A. Fields; George N. Somero

1998-01-01

251

The cold-induced denaturation of lactate dehydrogenase at sub-zero temperatures in the absence of perturbants  

Microsoft Academic Search

The cold-induced denaturation of lactate dehydrogenase has been determined in an unfrozen, cryoprotectant-free solution at sub-zero temperatures. The cold-induced denaturation temperature (TL) has been found to be -28°C. The results for the first time clearly establish that temperature alone can induce denaturation in a cooled protein solution. The validity of earlier data, obtained in the presence of perturbants (particularly pH

Ross H. M. Hatley; Felix Franks

1989-01-01

252

Novel Bioassay Based on Acetylcholinesterase and Lactate Dehydrogenase Activities to Evaluate the Toxicity of Chemicals to Soil Isopods  

Microsoft Academic Search

This study developed a bioassay with the isopod Porcellio dilatatus based on the activity of the enzymes acetylcholinesterase (AChE) and lactate dehydrogenase (LDH). The in vivo effects of the insecticides parathion-ethyl and endosulfan-sulfate on AChE and LDH activities of P. dilatatus under laboratory conditions were investigated. The route of uptake of the pesticides was through the food (alder leaves). Isopods

Sónia Ribeiro; L. Guilhermino; J. P. Sousa; A. M. V. M. Soares

1999-01-01

253

Native and modified lactate dehydrogenase expression in a fumaric acid producing isolate Rhizopus oryzae 99-880  

Microsoft Academic Search

Rhizopus oryzae is subdivided into two groups based on genetic and phenotypic differences. Type-I isolates accumulate primarily lactic acid\\u000a when grown in the presence of a fermentable carbon source and contain two lactate dehydrogenase genes, ldhA and ldhB. Type-II isolates synthesize predominantly fumaric acid and only have an ldhB gene. In this study, we determined that ldhB transcript is only

Christopher D. Skory; Ashraf S. Ibrahim

2007-01-01

254

A mutation affecting the lactate dehydrogenase locus Ldh-1 in the mouse—I. Genetical and electrophoretical characterization  

Microsoft Academic Search

(101\\/El × C3H\\/El)F1 male mice were injected intraperitoneally with the mutagen procarbazine hydrochloride and immediately caged with untreated test-stock females. Crude liver extracts from the offspring were subjected to polyacrylamide gel isoelectric focusing, and the gels were stained for six enzymes. In the experimental group (mutagen treated spermatogonial germ-cell stage), a dominant inherited banding alteration of the lactate dehydrogenase (LDH)

Daniel J. Charles; Walter Pretsch

1981-01-01

255

A novel hypoxia-response element in the lactate dehydrogenase-B gene of the killifish Fundulus heteroclitus  

Microsoft Academic Search

Previous studies have suggested that the lactate dehydrogenase-B gene (Ldh-B) of the Atlantic killifish, Fundulus heteroclitus, is a hypoxia-responsive gene. Here, we demonstrate that the F. heteroclitus Ldh-B promoter confers hypoxia-dependence upon reporter gene expression in transiently transfected mammalian (Hep3B) and fish (RTG-2 and RTH-149) cells in culture. Mutation and deletion analyses identified a putative hypoxia-response element (HRE) between 109

Bernard B. Rees; Yanira G. Figueroa; Thomas E. Wiese; Barbara S. Beckman; Patricia M. Schulte

2009-01-01

256

Purification and properties of a fructose-1,6-diphosphate activated l-lactate dehydrogenase from Staphylococcus epidermidis  

Microsoft Academic Search

l-(+)-lactate dehydrogenase (LDH) from Staphylococcus epidermidis ATCC 14990 was purified by affinity chromatography. The purified enzyme was specifically activated by fructose-1,6-diphosphate (FDP). The concentration of FDP required for 50% maximal activity was about 0.15 mM. The enzyme activity was inhibited by adenosine diphosphate (ADP) and oxamate. The inhibition by ADP appeared to be competitive with respect to reduced nicotinamide adenine

Friedrich Götz; Karl Heinz Schleifer

1975-01-01

257

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

2013-02-27

258

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

259

Docking studies on the binding of quinoline derivatives and hematin to Plasmodium falciparum lactate dehydrogenase.  

PubMed

The literature has reported that ferriprotoporphyrin IX (hematin) intoxicates the malarial parasite through competition with NADH for the active site of the enzyme lactate dehydrogenase (LDH). In order to avoid this, the parasite polymerizes hematin to hemozoin. The quinoline derivatives are believed to form complexes with dimeric hematin, avoiding the formation of hemozoin and still inhibiting LDH. In order to investigate this hypothesis we calculated the docking energies of NADH and some quinoline derivatives (in the free forms and in complex with dimeric hematin) in the active site of the Plasmodium falciparum LDH (PfLDH). Ours results showed better docking score values to the complexes when compared to the free compounds, pointing them as more efficient inhibitors of Pf_LDH. Further we performed Molecular Dynamics (MD) simulations studies on the best docking conformation of the complex chloroquine-dimeric hematin with PfLDH. Our in silico results corroborate experimental data suggesting a possible action route for the quinoline derivatives in the inhibition of PfLDH. PMID:21696234

Cortopassi, Wilian A; Oliveira, Aline A; Guimarães, Ana P; Rennó, Magdalena N; Krettli, Antoniana U; França, Tanos C C

2011-08-01

260

High energy electron beam inactivation of lactate dehydrogenase suspended in different aqueous media  

NASA Astrophysics Data System (ADS)

The direct and indirect effects of 5 MeV electron beam irradiation in the range (0-400 Gy) at 20°C, 0°C, -3°C and -196°, as well as the influence of the aqueous suspending medium (ultrapure water and heavy water) on the total enzymatic activity of lactate dehydrogenase (LDH) have been studied. Our results showed an exponential decrease on the enzymatic activity of irradiated LDH, at all irradiation temperatures, independently of the direct or indirect action of radiation. The temperature gradient used to lower the temperature of the samples to -196°C drastically influences the results. Freeze-thawing in two steps down to -196°C protects LDH to radiation, in the dose range used. The data obtained here inform on the high energy electrons effects on the enzymatic activity loss during irradiation and during thawing, when the subsequent growth of the water crystals [5] influences the three dimensional structure of the enzyme. A 99.98% concentration of D2O in the suspending medium of the enzyme decreases the global enzymatic activity, but reduces the rate of radiation inactivation of the enzyme. The rate of radiation inactivation of the enzyme suspended in ultrapture water is reduced when compared to the enzyme suspended in bidistilled water, but compared to the D2O suspended enzyme is lightly increased.

Hategan, A.; Popescu, A.; Butan, C.; Oproiu, C.; Hategan, D.; Morariu, V. V.

1999-01-01

261

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

262

Molecular properties of pyruvate bound to lactate dehydrogenase: A Raman spectroscopic study  

SciTech Connect

Lactate dehydrogenase catalyzes the addition of pyruvate to the four position of the nicotinamide ring of bound NAD+; this NAD-pyruvate adduct is bound tightly to the enzyme. We have used the adduct as a model for pyruvate in a competent ternary complex by comparing the Raman spectrum of the bound adduct with that for unliganded pyruvate. To understand the observed normal modes of pyruvate both as the bound adduct and in water, we have taken the Raman spectra of a series of /sup 13/C- and /sup 18/O-labeled pyruvates. We find that the carboxylate COO- moiety of pyruvate remains unprotonated at LDH's active site and forms an ion pair complex. The frequency of pyruvate's carbonyl C = O moiety shifts from 1710 cm-1 in water downward 34 cm-1 when pyruvate binds to LDH. This frequency shift corresponds to a ca. 34% polarization of the carbonyl bond, indicates a substantial interaction between the C = O group and enzyme, and is direct evidence for and is a measure of enzyme-induced electronic perturbation of the substrate needed for catalysis. This bond polarization is likely brought about by electrostatic interactions between the carbonyl moiety and the protonated imidazole group of His-195 and the guanidino group from Arg-109. We discuss how the data bear on the enzymatic chemistry of LDH.

Deng, H.; Zheng, J.; Burgner, J.; Callender, R.

1989-06-01

263

Early diagnosis of radiodermatitis using lactate dehydrogenase isozymes in hairless mice (SKH1-hr)  

PubMed Central

In this study, we evaluate a method for the early diagnosis of radiodermatitis for use in the prevention and therapy of this condition. Hairless mice (SKH1-hr) were used to study the early diagnosis of radiodermatitis. Lactate dehydrogenase (LDH, EC 1.1.1.27) isozymes were analyzed using native-polyacrylamide gel electrophoresis and western blotting of blood serum and tissues collected from SKH1-hr mice. Radiodermatitis developed 24 days after the first X-irradiation. Reduced spleen weight was observed after the last X-irradiation (P<0.05). Thereafter the weight increased until 24 days after the first irradiation, finally reaching levels comparable to those in the sham-irradiated control group. LDH activity was the highest in skeletal muscle and lowest in blood serum. LDH C4, A4, A3B, A2B2, AB3, and B4 isozymes were detected, in the mentioned order, from the cathode. This result was similar in other mouse strains. In the irradiated group, LDH A4 isozyme levels were reduced in the serum until inflammation occurred, whereas those of B4 isozyme were elevated. The subunits A and B followed a similar trend to that of LDH A4 and B4 isozyme, respectively. Importantly, antibodies against LDH B4 isozyme could prove useful in the early diagnosis of radiodermatitis.

Kim, Won-Dong

2012-01-01

264

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

265

Design of novel dihydroxynaphthoic acid inhibitors of Plasmodium falciparum lactate dehydrogenase.  

PubMed

We have studied inhibition of Plasmodium falciparum lactate dehydrogenase (pfLDH) by dihydroxynaphthoic acid (DHNA) analogues derivatives of hemigossypol-sesquiterpene found in cottonseed known to exhibit antimalarial activity. Molecular models of pfLDH-DHNA complexes were prepared from high-resolution crystal structures containing DHNA and azole inhibitors and binding affinities of the inhibitors were computed by molecular mechanics - polarizable continuum model of solvation (MM-PCM) approach. The 3D structures of the pfLDH-DHNA complexes were validated by a QSAR model, which confirmed consistency between the computed binding affinities and experimental inhibition constants for a training set and validation set of twelve DHNA inhibitors obtained from literature. Novel more potent DHNA analogs were identified by structure-based molecular design and predicted to inhibit pfLDH in the low nanomolar concentration range. In addition, the designed DHNA analogs displayed favorable predicted ADME-related profiles and an elevated selectivity for the pfLDH over the human isoform. PMID:22741776

Megnassan, Eugene; Keita, Melalie; Bieri, Cecile; Esmel, Akori; Frecer, Vladimir; Miertus, Stanislav

2012-09-01

266

Selective active site inhibitors of human lactate dehydrogenases A 4, B 4, and C 4 1 1 Abbreviations: LDH, lactate dehydrogenase; and LDH-A 4, B 4, and -C 4, human lactate dehydrogenases A 4, B 4, and C 4  

Microsoft Academic Search

Human lactate dehydrogenases (LDH-A4, -B4, and -C4) are highly homologous with 84–89% sequence similarities and 69–75% amino acid identities. Active site residues are especially conserved. Gossypol, a natural product from cotton seed, is a non-selective competitive inhibitor of NADH binding to LDH, with Ki values of 1.9, 1.4, and 4.2 ?M for LDH-A4, -B4, and -C4, respectively. However, derivatives of

Yue Yu; Jason A. Deck; Lucy A. Hunsaker; Lorraine M. Deck; Robert E. Royer; Erwin Goldberg; David L. Vander Jagt

2001-01-01

267

Kinetic study of heavy metal salt effects on the activity of L-lactate dehydrogenase in solution or immobilized on an oxygen electrode  

Microsoft Academic Search

A sensitive and convenient biosensor for detection of heavy metal salts has been developed. The method is based on the effects of heavy metal salts on the catalytic activity of L-lactate dehydrogenase (LDH) in solution or coimmobilized with L-lactate oxidase (LOD) on an oxygen electrode. At metal concentrations below 100 ?M, the kinetic behavior, with the LDH substrate NADH, showed

S. Fennouh; V. Casimiri; A. Geloso-Meyer; C. Burstein

1998-01-01

268

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

269

Lactate dehydrogenase undergoes a substantial structural change to bind its substrate.  

PubMed

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 here. The on-rate of the formation of the encounter complex from LDH/NADH with oxamate (a substrate mimic) is determined as a function of temperature and in the presence of small concentrations of a protein destabilizer (urea) and protein stabilizer (TMAO). It shows a strong temperature dependence with inverse Arrhenius behavior and a temperature-dependent enthalpy (heat capacity of 610 +/- 84 cal/Mol K), is slowed in the presence of TMAO and speeded up in the presence of urea. These results suggest that LDH/NADH occupies a range of conformations, some competent to bind substrate (open structure; a minority population) and others noncompetent (closed), in fast equilibrium with each other in accord with a select fit model of binding. From the thermodynamic results, the two species differ in the rearrangement of low energy hydrogen bonds as would arise from changes in internal hydrogen bonding and/or increases in the solvation of the protein structure. The binding-competent species can bind ligand at or very near diffusion-limited speeds, suggesting that the binding pocket is substantially exposed to solvent in these species. This would be in contrast to the putative closed structure where the binding pocket resides deep within the protein interior. PMID:17483169

Qiu, Linlin; Gulotta, Miriam; Callender, Robert

2007-05-04

270

CcpA-independent glucose regulation of lactate dehydrogenase 1 in Staphylococcus aureus.  

PubMed

Lactate Dehydrogenase 1 (Ldh1) is a key enzyme involved in Staphylococcus aureus NO·-resistance. Full ldh1-induction requires the presence of glucose, and mutants lacking the Carbon-Catabolite Protein (CcpA) exhibit decreased ldh1 transcription and diminished Ldh1 activity. The redox-regulator Rex represses ldh1 directly by binding to Rex-sites within the ldh1 promoter (P(ldh1)). In the absence of Rex, neither glucose nor CcpA affect ldh1 expression implying that glucose/CcpA-mediated activation requires Rex activity. Rex-mediated repression of ldh1 depends on cellular redox status and is maximal when NADH levels are low. However, compared to WT cells, the ?ccpA mutant exhibited impaired redox balance with relatively high NADH levels, yet ldh1 was still poorly expressed. Furthermore, CcpA did not drastically alter Rex transcript levels, nor did glucose or CcpA affect the expression of other Rex-regulated genes indicating that the glucose/CcpA effect is specific for P(ldh1). A putative catabolite response element (CRE) is located ?30 bp upstream of the promoter-distal Rex-binding site in P(ldh1). However, CcpA had no affinity for P(ldh1) in vitro and a genomic mutation of CRE upstream of P(ldh1) in S. aureus had no affect on Ldh1 expression in vivo. In contrast to WT, ?ccpA S. aureus preferentially consumes non-glycolytic carbon sources. However when grown in defined medium with glucose as the primary carbon source, ?ccpA mutants express high levels of Ldh1 compared to growth in media devoid of glucose. Thus, the actual consumption of glucose stimulates Ldh1 expression rather than direct CcpA interaction at P(ldh1). PMID:23342123

Crooke, Adrianne K; Fuller, James R; Obrist, Markus W; Tomkovich, Sarah E; Vitko, Nicholas P; Richardson, Anthony R

2013-01-14

271

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

2012-06-23

272

Regulation of the activity of lactate dehydrogenases from four lactic acid bacteria.  

PubMed

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

2013-05-17

273

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.

2011-01-01

274

Down-regulation of lactate dehydrogenase-A by siRNAs for reduced lactic acid formation of Chinese hamster ovary cells producing thrombopoietin  

Microsoft Academic Search

Lactate, one of the major waste products in mammalian cell culture, can inhibit cell growth and affect cellular metabolism\\u000a at high concentrations. To reduce lactate formation, lactate dehydrogenase-A (LDH-A), an enzyme catalyzing the conversion\\u000a of glucose-derived pyruvate to lactate, was down-regulated by an expression vector of small interfering RNAs (siRNA) in recombinant\\u000a Chinese hamster ovary (rCHO) cells producing human thrombopoietin

Sung Hyun Kim; Gyun Min Lee

2007-01-01

275

The separate function of d -lactate-, octopine, and alanopine dehydrogenases in the foot muscle of the jumping cockle Cardium tuberculatum during anaerobiosis  

Microsoft Academic Search

Summary  \\u000a1. \\u000aLactate dehydrogenase (LDH), octopine dehydrogenase (ODH), and alanopine dehydrogenase (AlDH) from muscle tissues of the cockleCardium tuberculatum have been separated. Purification (ca. 100-fold) was carried out by ammonium sulfate fractionation, ion exchange chromatography, gel chromatography, and affinity chromatography.\\u000a2. \\u000aIon exchange chromatography and electrophoresis separated four isoenzymes of AlDH comigrating with strombine dehydrogenase (StDH) activity (Mr ca. 44

Georg Meinardus-Hager; Gerd Gäde

1986-01-01

276

Effects of nitrate respiration on expression of the Arc-controlled operons encoding succinate dehydrogenase and flavin-linked L-lactate dehydrogenase.  

PubMed Central

Expression of sdhCDAB (encoding succinate dehydrogenase) and lctD (encoding the flavin-linked L-lactate dehydrogenase) is elevated aerobically and repressed anaerobically in Escherichia coli. The repression is initiated by autophosphorylation of the sensor protein ArcB, followed by phosphoryl group transfer to the regulator ArcA. ArcA-P, a global transcriptional regulator, then prevents sdh and lct expression. The stimulus for ArcB is not O2 deficiency per se. In vitro experiments showed that ArcB phosphorylation is enhanced by pyruvate, D-lactate, acetate, and NADH, the concentrations of which are likely to increase with the lack of an effective exogenous electron sink. In addition to their aerobic function, the two primary dehydrogenases also have roles in anaerobic nitrate respiration. Results presented here indicate that the increase of sdh and lct expression by nitrate depended on its chemical reduction, which in turn diminished the ArcA-P pool. Unexpectedly, a mutation in the fnr gene (encoding a global regulator involved in anaerobic metabolism) also alleviated the anaerobic repressions. Mutations in arcB or arcA were epistatic over that of fnr. Moreover, since this relief was counteracted by pyruvate in the growth medium, Fnr appears to affect formation of stimuli for ArcB. It is possible that Fnr also indirectly affects some of the other members of the arcA modulon, e.g., cyoABCDE (encoding the cytochrome o complex), cydAB (encoding the cytochrome d complex), and sodA (encoding the manganese-dependent superoxide dismutase).

Iuchi, S; Aristarkhov, A; Dong, J M; Taylor, J S; Lin, E C

1994-01-01

277

Electrochemical characteristics of a carbon-based thick-film l-lactate biosensor using l-lactate dehydrogenase  

Microsoft Academic Search

Carbon-based thick-film electrodes employing nicotinamide adenine dinucleotide (NAD+)-dependent dehydrogenases were constructed by screen printing. Cyclic voltammetric and amperometric investigations of the thick-film electrode showed significantly different electrocatalytic properties toward NADH compared to conventional polished electrodes. Electrochemical oxidation of NADH was attained at the low potential of +350 mV vs. AgAgCl at the thick-film carbon electrode, and this can be attributed

Hyun C Yoon; Hak-Sung Kim

1996-01-01

278

Importance of lactate dehydrogenase for the regulation of glycolytic flux and insulin secretion in insulin-producing cells.  

PubMed Central

The role of lactate dehydrogenase (LDH) in the generation of the metabolic signal for insulin secretion was studied after stable overexpression in INS-1 and RINm5F insulin-producing cells. INS-1 cells with a 25-fold overexpression of LDH-A, the highest level achieved, showed a 20-30% decrease in the glucose oxidation rate at glucose concentrations above 5 mM when compared with control cells, whereas values were unchanged at lower glucose concentrations. Lactate release increased in parallel with a decrease in the glucose oxidation rate. However, the INS-1 cell glucose-induced insulin secretory response, together with the rate of glucose utilization, were not significantly affected by LDH-A overexpression. Despite 3-fold overexpression of LDH-A in glucose-unresponsive RINm5F cells, there was no change in insulin secretion, glucose metabolism or lactate production in these cells. Exogenously added pyruvate and lactate potentiated glucose-stimulated insulin secretion in INS-1 cells, an effect that was abolished after LDH-A overexpression. Both compounds significantly decreased glucose oxidation rates in control cells. After overexpression of LDH-A in INS-1 cells, the effects of pyruvate and lactate on glucose oxidation were diminished. On the other hand, after LDH-A overexpression, both glycolytic metabolites decreased the glucose utilization rate at 5 mM glucose. The present data suggest that the level of LDH expression in insulin-secreting cells is critical for correct channelling of pyruvate towards mitochondrial metabolism. Interestingly, glucokinase-mediated glycolytic flux was decreased after LDH-A overexpression. Thus preferential channelling of glucose towards aerobic metabolism by glucokinase may be determined, at least in part, by the low level of constitutive expression of LDH-A in pancreatic beta-cells. In conclusion, the level of LDH expression in insulin-secreting cells is an important determinant of the physiological insulin-secretory capacity, and also determines how pyruvate and lactate affect insulin secretion.

Alcazar, O; Tiedge, M; Lenzen, S

2000-01-01

279

Oxidation of NADH produced by a lactate dehydrogenase immobilised on poly(aniline)–poly(anion) composite films  

Microsoft Academic Search

The immobilisation of enzymes is important for applications in bioelectrochemistry such as biosensors or biofuel cells. In this paper we report the immobilisation of lactate dehydrogenase (LDH) on poly(aniline)–poly(acrylate) [PANi–PAA] and poly(aniline)–poly(vinylsulfonate) [PANi–PVS] composite films. Two genetically engineered forms of LDH (E.C.1.1.1.27) from Bacillus stearothermophilus, one with a poly(histidine) tag on the C-Terminus (LDH-CHis) the other with a poly(histidine) tag

Evelyne Simon; Catherine M Halliwell; Chee Seng Toh; Anthony E. G Cass; Philip N Bartlett

2002-01-01

280

Identification of substituted 2-thio-6-oxo-1,6-dihydropyrimidines as inhibitors of human lactate dehydrogenase.  

PubMed

A novel 2-thio-6-oxo-1,6-dihydropyrimidine-containing inhibitor of human lactate dehydrogenase (LDH) was identified by high-throughput screening (IC50=8.1 ?M). Biochemical, surface plasmon resonance, and saturation transfer difference NMR experiments indicated that the compound specifically associated with human LDHA in a manner that required simultaneous binding of the NADH co-factor. Structural variation of the screening hit resulted in significant improvements in LDHA biochemical inhibition activity (best IC50=0.48 ?M). A crystal structure of an optimized compound bound to human LDHA was obtained and explained many of the observed structure-activity relationships. PMID:23628333

Dragovich, Peter S; Fauber, Benjamin P; Corson, Laura B; Ding, Charles Z; Eigenbrot, Charles; Ge, HongXiu; Giannetti, Anthony M; Hunsaker, Thomas; Labadie, Sharada; Liu, Yichin; Malek, Shiva; Pan, Borlan; Peterson, David; Pitts, Keith; Purkey, Hans E; Sideris, Steve; Ultsch, Mark; VanderPorten, Erica; Wei, BinQing; Xu, Qing; Yen, Ivana; Yue, Qin; Zhang, Huihui; Zhang, Xuying

2013-04-10

281

[Antigenic activity of the N-terminal peptide of porcine muscle lactate dehydrogenase, obtained upon cyanogen bromide cleavage].  

PubMed

N-terminal peptide was isolated from BrCN hydrolysate of pig muscle lactate dehydrogenase (LDH), its antigenic properties were studied using solid-phase immunoenzyme assay. N-terminal fragment containing 1-32 amino acids of LDH exhibited the antigenic activity towards antiserum and specific antibodies to native LDH5 and inhibited formation of the antigen-antibody complex by 25-35%, thus suggesting the presence of at least one antigenic determinant in the N-terminal fragment. The N-terminal peptide of LDH5 conjugated with bovine albumin maintained the antigenic activity towards specific antibodies against native LDH5 (inhibition by 20-25%). PMID:2815682

Alekseeva, A E; Grebenshchikova, O G; Potapkina, T A; Prozorovski?, V N

282

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.

Barrett, M C; Dawson, A P

1975-01-01

283

In Vivo Regulation of Alcohol Dehydrogenase and Lactate Dehydrogenase in Rhizopus Oryzae to Improve l Lactic Acid Fermentation  

Microsoft Academic Search

Rhizopus oryzae is becoming more important due to its ability to produce an optically pure l-lactic acid. However, fermentation by Rhizopus usually suffers from low yield because of production of ethanol as a byproduct. Limiting ethanol production in living immobilized\\u000a R. oryzae by inhibition of alcohol dehydrogenase (ADH) was observed in shake flask fermentation. The effects of ADH inhibitors added

Sitanan Thitiprasert; Sarintip Sooksai; Nuttha Thongchul

2011-01-01

284

The ldhA Gene, Encoding Fermentative L-Lactate Dehydrogenase of Corynebacterium glutamicum, Is under the Control of Positive Feedback Regulation Mediated by LldR  

Microsoft Academic Search

Corynebacterium glutamicum ldhA encodes L-lactate dehydrogenase, a key enzyme that couples L-lactate production to reoxidation of NADH formed during glycolysis. We previously showed that in the absence of sugar, SugR binds to the ldhA promoter region, thereby repressing ldhA expression. In this study we show that LldR is another protein that binds to the ldhA promoter region, thus regulating ldhA

Koichi Toyoda; Haruhiko Teramoto; Masayuki Inui; Hideaki Yukawa

2009-01-01

285

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

Microsoft Academic Search

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,

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

2003-01-01

286

Molecular mechanism of SugR-mediated sugar-dependent expression of the ldhA gene encoding l -lactate dehydrogenase in Corynebacterium glutamicum  

Microsoft Academic Search

This paper reports on the transcriptional regulation mechanism of the Corynebacterium glutamicum ldhA gene encoding l-lactate dehydrogenase responsible for production of l-lactate. DNA affinity purification allowed us to identify SugR, a global repressor of genes involved in sugar uptake and\\u000a glycolysis, as a protein binding to the ldhA promoter region. Whereas ldhA gene expression and ldhA promoter activity were completely

Koichi Toyoda; Haruhiko Teramoto; Masayuki Inui; Hideaki Yukawa

2009-01-01

287

Maternal effect and embryonic gene expression for lactate dehydrogenase B (LDH-B), glucose-6-phosphate dehydrogenase (G6PDH), and peptidase (PEP1) during the development of Pleurodeles waltl (urodele amphibian)  

Microsoft Academic Search

The polymorphism of three enzymes [lactate dehydrogenase B (LDH-B), glucose-6-phosphate dehydrogenase, (G6PDH), and peptidase-1 (PEP-1)] in Pleurodeles waltl has allowed the expression of the corresponding loci to be followed during the development of spawnings arising from various crosses. A maternal effect lasting up to the late tail-bud stage (approx. stage 28) was shown for PEP-1. A similar situation was observed

F. Gasser; V. Ferrier

1984-01-01

288

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.

1989-01-01

289

Overproduction of a 37-kilodalton cytoplasmic protein homologous to NAD+-linked D-lactate dehydrogenase associated with vancomycin resistance in Staphylococcus aureus.  

PubMed Central

We previously reported the isolation of a laboratory-derived Staphylococcus aureus mutant, 523k, that has constitutive low-level resistance to vancomycin (MIC = 5 micrograms/ml) and teicoplanin (MIC = 5 micrograms/ml) and elaborates a ca. 39-kDa cytoplasmic protein that was not detected in the parent strain 523 (MIC = 1 micrograms/ml). We have now detected the protein in strain 523 by immunoblotting with antiserum raised against the protein. Consistent with our initial observations, densitometric analysis of the immunoblots revealed an increased production of the protein in 523k compared with that of the susceptible parent 523. The 5' region of the gene encoding the protein of interest was identified by nucleotide sequencing a PCR product amplified from the genome of 523k with degenerate primers designed to encode the amino acid sequence of proteolytic peptides obtained from the protein. The remainder of the gene was identified by library screening, PCR, and nucleotide sequencing. The gene encodes a 36.7-kDa protein with homology to a family of bacterial NAD+-dependent, D-specific 2-hydroxyacid dehydrogenases which includes both D-lactate dehydrogenase and the enterococcal vancomycin resistance protein VanH and is therefore designated ddh. Increased production of the product of ddh, Ddh, was associated with increased D-lactate dehydrogenase activity in 523k, a finding which suggested that Ddh is likely to be the D-lactate dehydrogenase previously identified in S. aureus. The increased D-lactate dehydrogenase activity in strain 523k and the structural similarities among Ddh, D-lactate dehydrogenase, and VanH suggest that overproduction of Ddh might play a role in vancomycin resistance in this strain.

Milewski, W M; Boyle-Vavra, S; Moreira, B; Ebert, C C; Daum, R S

1996-01-01

290

Crystal structure of the MJ0490 gene product of the hyperthermophilic archaebacterium Methanococcus jannaschii, a novel member of the Lactate\\/Malate family of dehydrogenases 1 1 Edited by R. Huber  

Microsoft Academic Search

The MJ0490 gene, one of the only two genes of Methanococcus jannaschii showing sequence similarity to the lactate\\/malate family of dehydrogenases, was classified initially as coding for a putative l-lactate dehydrogenase (LDH). It has been re-classified as a malate dehydrogenase (MDH) gene, because it shows significant sequence similarity to MT0188, MDH II from Methanobacterium thermoautotrophicum strain ?H. The three-dimensional structure

Byung Il Lee; Changsoo Chang; Seung-Je Cho; Soo Hyun Eom; Kyeong Kyu Kim; Yeon Gyu Yu; Se Won Suh

2001-01-01

291

Quantification of Viability in Organotypic Multicellular Spheroids of Human Malignant Glioma using Lactate Dehydrogenase Activity: A Rapid and Reliable Automated Assay  

Microsoft Academic Search

Organotypic spheroids from malignant glioma resemble the biological complexity of the original tumor and are therefore appealing to study anticancer drug responses. Accurate and reproducible quantification of response effect has been lacking to determine drug responses in this three-dimensional tumor model. Lactate dehydrogenase (LDH) activity was demonstrated in cryostat sections of spheroids using the tetrazolium salt method. Calibrated digital image

Witt Hamer Philip C. De; Ard Jonker; Sieger Leenstra; Jan M. Ruijter; Cornelis J. F. Van Noorden

2005-01-01

292

Studies on the antioxidant effect and interaction of diphenyl diselenide and dicholesteroyl diselenide with hepatic ?-aminolevulinic acid dehydratase and isoforms of lactate dehydrogenase  

Microsoft Academic Search

Studies on the interaction of dicholesteroyl diselenide (DCDS) and diphenyl diselenide (DPDS) with hepatic ?-aminolevulinic acid dehydratase (ALA-D) and different isoforms of lactate dehydrogenase (LDH) from different tissues were investigated. In addition, their antioxidant effects were tested in vitro by measuring the ability of the compounds to inhibit the formation of hepatic thiobarbituric acid reactive species (TBARS) induced by both

I. J. Kade; M. W. Paixão; O. E. D. Rodrigues; E. O. Ibukun; A. L. Braga; G. Zeni; C. W. Nogueira; J. B. T. Rocha

2009-01-01

293

Cadmium-induced production of superoxide anion and nitric oxide, DNA single strand breaks and lactate dehydrogenase leakage in J774A.1 cell cultures  

Microsoft Academic Search

The involvement of reactive oxygen species in the toxicity of cadmium (Cd) has been proposed. We have, therefore, examined the effects of this cation on the production of Superoxide anion and nitric oxide and DNA single strand breaks in J774A.1 macrophage cells in culture as well as the effects on lactate dehydrogenase (LDH) leakage and cell viability. Following a 48-h

Ezdihar A. Hassoun; Sidney J. Stohs

1996-01-01

294

[Activity of lactate dehydrogenase in the brain cortex and hippocampus of Mongolian gerbils after global ischemia and reperfusion injuries].  

PubMed

Cerebral ischemia results in severe derangements of energy metabolism in the nervous tissue including activation of glycolytic pathway. Activity of cytosolic lactate dehydrogenase (LDH) in the specific brain structures remains unclear. The recent study was aimed at investigation into the LDH activity in the cytoplasm of both hippocampal and cortical neurons in Mongolian gerbils (Meriones unguiculatus) at different durations of reperfusion after global ischemia. Analysis showed that the activity of LDH in pyramidal neurons of various hippocampal areas and neurons of II, III and V cortical layers after 7-minute forebrain ischemia depended on both localization of the neurons and duration ofreperfusion. In general, the changes in postischemic cytosolic LDH activity include significant decrease in the LDH activity 2 days after reperfusion with varying degree of recovery on day 7 of reperfusion. PMID:22650061

Shcherbak, N S; Galagudza, M M; Ovchinnikov, D A; Kuz'menkov, A N; Iukina, G Iu; Barantsevich, E R; Tomson, V V; Shliakhto, E V

2012-02-01

295

Effect of neem limonoids on lactate dehydrogenase (LDH) of the rice leaffolder, Cnaphalocrocis medinalis (Guenée) (Insecta: Lepidoptera: Pyralidae).  

PubMed

Neem is derived from the neem tree Azadirachta indica A. Juss. (Meliaceae), and its primary insecticidal component is the tetranortriterpenoid azadirachtin and other limonoids. The effect of neem limonoids azadirachtin, salannin, deacetylgedunin, gedunin, 17-hydroxyazadiradione and deacetylnimbin on enzyme lactate dehydrogenase (LDH) activity of the rice leaffolder (RLF) Cnaphalocrocis medinalis (Lepidoptera: Pyralidae) larvae was investigated. There was a decrease in enzyme activity relative to the control at all concentrations tested. When fed a diet of rice leaves treated with neem limonoids in bioassays, gut tissue enzyme, LDH levels in rice leaffolder larvae are affected. These results indicate neem limonoids affect LDH activity. These effects are most pronounced in early instar larvae. Azadirachtin was the most potent in of all the limonoids in all experiments indicating strong enzyme inhibition. Clear dose-response relationships were established with respect to LDH activity. PMID:16154614

Senthil Nathan, Sengottayan; Kalaivani, Kandaswamy; Chung, Paul Gene; Murugan, Kadarkarai

2005-09-12

296

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

PubMed

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

Ilmén, Marja; Koivuranta, Kari; Ruohonen, Laura; Rajgarhia, Vineet; Suominen, Pirkko; Penttilä, Merja

2013-05-25

297

The reaction of trimethylamine dehydrogenase with electron transferring flavoprotein.  

PubMed

The kinetics of electron transfer between trimethylamine dehydrogenase (TMADH) and its physiological acceptor, electron transferring flavoprotein (ETF), has been studied by static and stopped-flow absorbance measurements. The results demonstrate that reducing equivalents are transferred from TMADH to ETF solely through the 4Fe/4S center of the former. The intrinsic limiting rate constant (klim) and dissociation constant (Kd) for electron transfer from the reduced 4Fe/4S center of TMADH to ETF are about 172 s-1 and 10 microM, respectively. The reoxidation of fully reduced TMADH with an excess of ETF is markedly biphasic, indicating that partial oxidation of the iron-sulfur center in 1-electron reduced enzyme significantly reduces the rate of electron transfer out of the enzyme in these forms. The interaction of the two unpaired electron spins of flavin semiquinone and reduced 4Fe/4S center in 2-electron reduced TMADH, on the other hand, does not significantly slow down the electron transfer from the 4Fe/4S center to ETF. From a comparison of the limiting rate constants for the oxidative and reductive half-reactions, we conclude that electron transfer from TMADH to ETF is not rate-limiting during steady-state turnover. The overall kinetics of the oxidative half-reaction are not significantly affected by high salt concentrations, indicating that electrostatic forces are not involved in the formation and decay of reduced TMADH-oxidized ETF complex. PMID:7592591

Huang, L; Rohlfs, R J; Hille, R

1995-10-13

298

IMMUNOCAPTURE DIAGNOSTIC ASSAYS FOR MALARIA USING PLASMODIUM LACTATE DEHYDROGENASE (pLDH)  

Microsoft Academic Search

We have developed two diagnostic assays based on the specific detection of Plasmodium lactate de- hydrogenase (pLDH) activity. These assays exploit a panel of monoclonal antibodies that capture the parasite enzyme and allow for the quantitation and speciation of human malaria infections. An immunocapture pLDH activity assay (ICpLDH) allows for the rapid purification and measurement of pLDH from infected blood

ROBERT PIPER; JACQUES LEBRAS; LAURA WENTWORTH; ANGELA HUNT-COOKE; SANDRA HOUZE; PETER CHIODINI; MICHAEL MAKLER

299

Interaction of lactate dehydrogenase with anthraquinone dyes: characterization of ligands for dye–ligand chromatography  

Microsoft Academic Search

Anthraquinone dyes (ADs), originally developed for the textile industry, are useful nucleotide-specific ligands for the purification of proteins by affinity techniques. Their specific feature is to mimic the adenine nucleotides ATP, ADP, NAD, NADH, which enables them to interact with the nucleotide-binding sites of enzymes such as dehydrogenases, kinases and ATPases. In the present study, the interactions and\\/or inhibitory effects

V. Bohá?ová; P. Do?olomanský; A. Breier; P. Gemeiner; A. Ziegelhöffer

1998-01-01

300

Ab initio modeling of ethylbenzene dehydrogenase reaction mechanism.  

PubMed

Density functional theory calculations were performed to study the mechanism of ethylbenzene oxidation by ethylbenzene dehydrogenase (EBDH). EBDH is a bacterial molybdopterin enzyme capable of stereospecific anaerobic hydroxylation of alkylaromatic compounds to secondary alcohols. It is a key biocatalyst in the metabolism of ethylbenzene-degrading bacteria such as Aromatoleum aromaticum , which converts ethylbenzene to (S)-1-phenylethanol. The recently determined EBDH structure enabled the theoretical description of the ethylbenzene oxidation mechanism. In this work, theoretical calculations and kinetic isotopic experiments were conducted and combined in order to elucidate the reaction mechanism. We considered three aspects: (i) Does the reaction concur with one two-electron or two one-electron transfers? (ii) Is the active site His192 important for the reaction and what is its protonation state? (iii) What catalytic consequences have different possible arrangements of the molybdopterin ligand? The most important outcome of the calculations is that mechanisms involving two one-electron transfers and a radical-type intermediate have lower energy barriers than the corresponding two-electron transfer mechanisms and are, therefore, more plausible. The mechanism involves two transition states: radical-type TS1 associated with the C-H bond cleavage, and carbocation-type TS2 associated with the transfer of the second electron and OH rebound. Using models with protonated and nonprotonated His 192, we conclude that this amino acid takes part in the mechanism. However, as both models yielded plausible reaction pathways, its protonation state cannot be easily predicted. Qualitative agreement was reached between the calculated kinetic isotope effects (KIE) obtained for radical TS1 and the KIE measured experimentally at optimum pH, but we observed a very strong pH dependence of KIE throughout the investigated pH range (3.1 for pH 6, 5.9 for pH 7, up to 10.5 at pH 8.). This may be explained by assuming a gradual shift of the rate-determining step from TS1 associated with high KIE to TS2 associated with low KIE with lowered pH and an increasing contribution of proton/deuteron tunneling associated with high pH. Finally, models were calculated with different signs of the conformational twist of the pterin ligands, yielding only slightly different energy profiles of the reaction pathways. PMID:20387836

Szaleniec, Maciej; Borowski, Tomasz; Schühle, Karola; Witko, Malgorzata; Heider, Johann

2010-05-01

301

Interdependence of coenzyme-induced conformational work and binding potential in yeast alcohol and porcine heart lactate dehydrogenases: a hydrogen-deuterium exchange study  

SciTech Connect

Binding of NAD coenzymes to yeast alcohol dehydrogenase (YADH) and porcine heart lactate dehydrogenase (PHLDH) was studied by hydrogen-deuterium exchange with the infrared technique. Conformational changes in the enzymes specific to the coenzymes and their fragments were observed, and the pH dependence of the exchange reaction shows that it conforms to the EX-2 scheme. In both YADH and PHLDH the magnitude of the conformational change as measured by exchange retardation is considerably larger for the NAD/sup +/ than for NADH. Studies with coenzyme fragments like ADP-ribose, ADP, and AMP also highlight the lack of rigorous correlation between structural features such as charge and size and their influence on exchange behavior. Ternary complexes such as YADH-NAD/sup +/-pyrazole, PHLDH-NAD/sup +/-oxalate, and PHLDH-NADH-oxamate, which mimic the transition state, have a significantly more pronounced effect on exchange rates than the corresponding binary complexes. The outstanding feature of this study is the demonstration that in the binary enzyme-coenzyme complexes the more loosely bound NAD/sup +/ is more effective in retarding exchange than the more firmly bound NADH. These differences are attributed to the unequal structural constraints exerted by the two coenzymes upon the enzymes, which translate to unequal expenditure of transconformational work in the formation of the two complexes. The opposing variation in the free energy of binding and the transconformational work expended can be viewed as an unequal partitioning of the net free energy gain resulting from the protein-ligand interaction into a binding term and that required for conformational change.

De Weck, Z.; Pande, J.; Kaegi, J.H.R.

1987-07-28

302

Metal Cu(II) and Zn(II) bipyridyls as inhibitors of lactate dehydrogenase  

Microsoft Academic Search

Metal complex–protein interaction is an evolving concept for determining cellular targets of metallodrugs. Lacatate dehydrogenase\\u000a (LDH) is critically implicated in tumor growth and therefore, considered to be an important target protein for anti-tumor\\u000a metal complexes. Due to efficient biocompatibility of copper (Cu2+) and zinc (Zn2+), we synthesized CubpyAc2 · H2O (Cu-bpy) and ZnbpyAc2 · H2O (Zn-bpy; where bpy = 2,2? bipyridine, Ac = CH3COO?) complexes and evaluated their

Raj Kumar Koiri; Surendra Kumar Trigun; Santosh Kumar Dubey; Santosh Singh; Lallan Mishra

2008-01-01

303

Synthesis and application of a photoaffinity analog of nicotinamide adenosine dinucleotide: Identification of the active sites of glutamate and lactate dehydrogenases  

Microsoft Academic Search

A photoaffinity analog of NAD{sup +} has been synthesized by chemically coupling (³²P)2-azido-AMP and NMN{sup +} to produce (³²P)nicotinamide 2-azidoadenosine dinucleotide (2-azido-NAD{sup +}). The utility of 2-azido-NAD{sup +} as an effective active-site-directed photoprobe was demonstrated using bovine liver glutamate dehydrogenase and porcine muscle lactate dehydrogenase as model enzymes. In the absence of ultraviolet light 2-azido-NAD{sup +} is a substrate for

1990-01-01

304

The role of an NAD-independent lactate dehydrogenase and acetate in the utilization of lactate by Clostridium acetobutylicum strain P262  

Microsoft Academic Search

Clostridium acetobutylicum strain P262 utilized lactate at a rapid rate [600 nmol min?1 (mg protein)?1], but lactate could not serve as the sole energy source. When acetate was provided as a co-substrate, the growth rate was\\u000a 0.05h?1. Butyrate, carbon dioxide and hydrogen were the end products of lactate and acetate utilization, and the stoichiometry was\\u000a 1 lactate + 0.4 acetate

Francisco Diez-Gonzalez; James B. Russell; Jean B. Hunter

1995-01-01

305

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.

2012-01-01

306

Lactate Dehydrogenase 5 Expression in Non-Hodgkin Lymphoma Is Associated with the Induced Hypoxia Regulated Protein and Poor Prognosis  

PubMed Central

Lactate dehydrogenase 5 (LDH-5) is one of the major isoenzymes catalyzing the biochemical process of pyruvate to lactate. The purpose of this study was to investigate the expression of serum LDH-5 and test whether this enzyme is regulated by tumor hypoxia and represents a prognostic marker in patients with Non-Hodgkin’s lymphoma (NHL). In this study, LDH-5 levels were detected using agarose gel electrophoresis in NHL patients (n?=?266) and non-NHL controls including benign lymphadenectasis (n?=?30) and healthy cohorts (n?=?233). We also explored the expression of LDH-5 and hypoxia-inducible factor (HIF) 1? in NHL and benign controls by immunohistochemistry and immunofluorescence staining, respectively. Moreover, the role of LDH-5 in the progression of NHL was assessed by multivariate Cox analyses and Kaplan-Meier survival estimates. Serum concentrations of LDH-5 were significantly higher in NHL patients (9.3%) than in benign patients and healthy controls (7.5% and 7.2%, respectively, P<0.01). Application of LDH-5 detection increased the sensitivity of NHL detection, identifying 53.4% of NHL patients as positive, compared with the measurement of total LDH levels (36.5% sensitivity). LDH-5 concentrations increased with clinical stage, extra-nodal site involvement, and WHO performance status of patients with NHL. Exposure to a hypoxic environment induced the expression of LDH-5 and its overexpression correlated with HIF1? cytoplasmic accumulation in NHL cells. In multivariate analyses, LDH-5 was an independent marker for progression-free survival in patients with NHL (P<0.001). Overall, the expression of LDH-5 was elevated in NHL, showing an association with tumor hypoxia and unfavorable prognosis. Thus, LDH-5 emerges as a promising prognostic predictor for NHL patients.

Chen, Zhujun; Xu, Xiaofeng; Hu, Hongfeng; Zhao, Xinmin; Gao, Xiang; Guo, Lin

2013-01-01

307

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.

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

2013-01-01

308

Coulometric bioelectrocatalytic reactions based on NAD-dependent dehydrogenases in tricarboxylic acid cycle  

Microsoft Academic Search

This paper describes the characterization of mediated electro-enzymatic electrolysis systems based on NAD-dependent dehydrogenase reactions in the tricarboxylic acid (TCA) cycle. A micro-bulk electrolysis system with a carbon felt anode immersed in an electrolysis solution with a value of about 10?L was constructed for coulometric analysis of the substrate oxidation. Diaphorase (DI) was used to couple the NAD-dependent dehydrogenase reaction

Jun Fukuda; Seiya Tsujimura; Kenji Kano

2008-01-01

309

Evaluation of cerebrospinal fluid adenosine deaminase activity in HIV-seropositive subjects and its association with lactate dehydrogenase and protein levels.  

PubMed

The purpose of this study was to investigate the role of ADA as additional marker of HIV infection as well as its association with other biochemical markers. This study included 55 patients, 26 being diagnosed as HIV positive and 29 patients diagnosed as HIV negative. Glucose, total protein, lactate dehydrogenase, and adenosine deaminase (ADA) activity were measured on cerebrospinal fluid (CSF). ADA activity on CSF was statistically different in HIV-seropositive subjects compared with HIV-negative subjects. The sensitivity and specificity of ADA activity on CSF was 50 and 82.76%, respectively. ADA activity was positively correlated with lactate dehydrogenase and protein in patients with HIV positive and it was negatively correlated with glucose levels. ADA determination in CSF could add information about inflammatory processes in patients with HIV infection. PMID:20347569

Pinheiro, F V; Pimentel, V C; Moresco, R N; Moretto, M B

2009-10-23

310

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

311

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

PubMed Central

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 in terms of redox balance.

Gupta, S; Clark, D P

1989-01-01

312

A chemically modified carbon paste electrode with d-lactate dehydrogenase and alanine aminotranferase enzyme sequences for d-lactic acid analysis  

Microsoft Academic Search

An amperometric biosensor was constructed for the analysis of d-lactic acid based on immobilizing d-lactate dehydrogenase(d-LDH), alanine aminotransferase (ALT), NAD+, a redox polymer and polyethylenimine in carbon paste. The effect of addition of ALT in the paste, using enzyme sequences of ALT\\/d-LDH, was insignificant for d-lactic acid analysis. The responses of d-lactic acid in ALT\\/d-LDH paste electrode are the same

Hun-Chi Shu; Ning-ping Wu

2001-01-01

313

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

314

Cibacron Blue F3GA anchored monolayers with biospecific affinity for NAD(H)-dependent lactate dehydrogenase: characterization by FTIR-spectroscopy and atomic force microscopy  

Microsoft Academic Search

Fourier transform infrared reflection absorption spectroscopy (FT-IRAS) and atomic force microscopy have been used to analyze several alkanethiol self-assembled monolayers derivatized with the triazine dye Cibacron Blue F3G-A. This compound, when in solution, works as a competitive inhibitor for NAD(H)-dependent lactate dehydrogenase in building a complex by a site-specific affinity binding at the enzyme's NAD+-binding pocket. Therefore, the chemisorption of

Lars Bertilsson; Hans-Jürgen Butt; Gabriele Nelles; Daniela D. Schlereth

1997-01-01

315

Crystal structure of Plasmodium berghei lactate dehydrogenase indicates the unique structural differences of these enzymes are shared across the Plasmodium genus  

Microsoft Academic Search

As Plasmodium rely extensively on homolactic fermentation for energy production, Plasmodium falciparum lactate dehydrogenase (PfLDH)—the key enzyme in this process—has previously been suggested as a novel target for antimalarials. This enzyme has distinctive kinetic and structural properties that distinguish it from its human homologues. In this study, we now describe the expression, kinetic characterisation and crystal structure determination of the

V. J. Winter; A. Cameron; R. Tranter; R. B. Sessions; R. L. Brady

2003-01-01

316

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

Microsoft Academic Search

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) con- generic perciformes (Lates calcarifer and Lates niloticus) we characterized both coding and non-coding regions of lactate dehydrogenase-B (ldh-b), a locus which exhibits tempera- ture-adaptive differences among temperate and sub-tropical populations of the

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

317

Surface-modified gold electrodes with biospecific affinity for lactate dehydrogenase based on Cibacron Blue 3FGA self-assembled monolayers  

Microsoft Academic Search

Surface-modified gold electrodes with biospecific affinity for NAD(H)-dependent lactate dehydrogenase (LDH) have been prepared from mixed self-assembled monolayers bearing the coenzyme-analogous, Cibacron Blue 3FG-A, as anchored pendant group. Enzymatic monolayers are formed by formation of a complex upon the site-specific binding of Cibacron Blue to the enzyme through its NAD+-binding pocket.In such monolayers the enzyme may be expected to lie

Daniela D. Schlereth

1997-01-01

318

Arginine to tryptophan substitution in the active site of a human lactate dehydrogenase variant — LDHB ?GUA1: postulated effects on subunit structure and catalysis  

Microsoft Academic Search

A variant of lactate dehydrogenase (LDHB?GUA1) was previously identified among the Guaymi Indians of Panama and Costa Rica. The LDHB?GUA1 variant is enzymatically inactive; however, the variant subunits alter the electrophoresic mobility of the tetramers that include active LDHA and LDHB subunits. The kinetic properties of the tetramerc enzyme, comprised of inactive B plus active A subunits, are similar to

Gisela C. Shonnard; Nicholas V. Hud; H. W. Mohrenweiser

1996-01-01

319

The kinetic properties and sensitivities to inhibitors of lactate dehydrogenases (LDH1 and LDH2) from Toxoplasma gondii: comparisons with pLDH from Plasmodium falciparum  

Microsoft Academic Search

Toxoplasma gondii differentially expresses two forms of lactate dehydrogenase in tachyzoites and bradyzoites, respectively, designated LDH1 and LDH2. Previously it was demonstrated that LDH1 and LDH2 share a unique structural feature with LDH from the malarial parasite Plasmodium falciparum (pLDH), namely, the addition of a five-amino acid insert into the substrate specificity loops. pLDH exhibits a number of kinetic properties

Caroline Dando; Eric R Schroeder; Lucy A Hunsaker; Lorraine M Deck; Robert E Royer; Xiulan Zhou; Stephen F Parmley; David L Vander Jagt

2001-01-01

320

D175 Discriminates Between NADH and NADPH in the Coenzyme Binding Site of Lactobacillus delbrueckii subsp. Bulgaricus D-Lactate Dehydrogenase  

Microsoft Academic Search

The NAD-dependent D-(-)-lactate dehydrogenase (D-LDH) from Lactobacillus delbrueckii subsp. bulgaricus (in short, L. bulgaricus) has been modified at position 175 by site-directed mutagenesis, changing a conserved aspartate residue into an alanine. The D175A mutant enzyme displays a 40-fold shift in coenzyme preference from NADH to NADPH. This demonstrates that D175 truly belongs to the amino acid consensus GXGXXGX(17)D (where X

N. Bernard; K. Johnsen; J. J. Holbrook; J. Delcour

1995-01-01

321

A novel in-frame deletion mutation in a case of lactate dehydrogenase (LD) H subunit deficiency showing an atypical LD isoenzyme pattern in serum and erythrocytes  

Microsoft Academic Search

Objective: We report a case showing an atypical lactate dehydrogenase (LD) isoenzyme pattern involving deficiency only of LD-1 and LD-2 in serum and erythrocytes. LD activity in serum from this patient was extremely low, similar to complete LD-H deficiency, and also that in erythrocytes was low.Design: The DNA fragment containing exon 1 through 7 of the LD-H gene were amplified

Kayoko Sudo; Masato Maekawa; Nobuyuki Houki; Takanari Okuda; Setsuko Akizuki; Tadao Magara; Kazuhiro Kawano

1999-01-01

322

Trypan blue dye uptake and lactate dehydrogenase in adult rat hepatocytes—Freshly isolated cells, cell suspensions, and primary monolayer cultures  

Microsoft Academic Search

Summary  Leakage of lactate dehydrogenase and staining by the vital dye trypan blue were investigated in adult rat hepatocytes at the\\u000a time of isolation, in suspensions up to 3 h and in primary monolayer cultures up to 3 d. These two parameters of plasma membrane\\u000a integrity were found to correlate closely in hepatocyte suspensions, but to a lesser degree in monolayer

Hugo O. Jauregui; Nancy T. Hayner; James L. Driscoll; Rhonda Williams-Holland; Milton H. Lipsky; Pierre M. Galletti

1981-01-01

323

Downstream processing of diagnostic enzymes: Optimised protocols for the simultaneous separation and purification of lactate dehydrogenase and pyruvate kinase from rabbit muscle  

Microsoft Academic Search

Cibacron Blue 3GA-Sepharose CL6B was used to design two optimised, inexpensive and easy to scale-up processes for the simultaneous separation and purification of l-lactate dehydrogenase (LDH) and pyruvate kinase (PK) from rabbit muscles. The tissue was homogenised, filtered, and the liquid treated by DEAE-cellulose and Sephadex-G25 gel to obtain the “pre-treated” extract which was used in the dye-column. The first

G. Tsamadis; N. Papageorgakopoulou; Y. Clonis

1992-01-01

324

Affinity chromatography of d -lactate dehydrogenases from Limulus polyphemus (horseshoe crab) and Haliotus sp. (abalone) voluntary muscle on 8-(6-aminohexyl)-amino-adenine nucleotide-sepharose  

Microsoft Academic Search

An improved method of purification employing sequential isoenzyme elution on DEAE-cellulose at pH 6.5 and biologically specific\\u000a elution with the reduced NAD-pyruvate adduct from 8-(6-aminohexyl)-amino-NAD?-Sepharose is described forLimulus (horseshoe crab) muscle D- lactate dehydrogenase. The protein is judged as being at least 98%pure by its constant specific activity in the terminal purification steps, a molar extinction coefficient (280nm) identical with

George L. Long; Elizabeth L. Moore

1976-01-01

325

Purification and Properties of the Threespine Stickleback ( Gasterosteus aculeatus) Lactate Dehydrogenase LDH-B 4 and LDH-C 4 Isoenzymes  

Microsoft Academic Search

In the threespine stickleback (Gasterosteus aculeatus) lactate dehydrogenase (LDH, EC 1.1.1.27) is encoded by three loci, Ldh-A, Ldh-B, and Ldh-C. LDH-B4 isoenzyme restricted its function to eye and brain, while LDH-C4 isoenzyme functions in the eye. In the Dead Vistula stickleback population, none of LDH loci is polymorphic. The LDH-B4 and LDH-C4 isoenzymes from the eye were purified to homogeneity

Marek S. Zitara; Jadwiga Gronczewska; Edward F. Skorkowski

1997-01-01

326

Bone-specific alkaline phosphatase protein, total alkaline phosphatase activity and lactate dehydrogenase in sera of patients with sickle cell disease.  

PubMed

Serum bone-specific alkaline phosphatase protein (bAP) was evaluated as indicator of bone turnover by immunoradiometric assay (IRMA) in twenty patients with sickle cell disease and in twenty healthy control subjects. Serum bAP was also compared with serum total alkaline phosphatase activity and serum lactate dehydrogenase in the same group. The concentrations of serum bAP and serum lactate dehydrogenase were significantly higher in the study group than in the control group (p < 0.05, p < 0.01, respectively). The serum total alkaline phosphatase activity showed no significant difference with the control healthy subjects. There was no correlation between serum bAP and total alkaline phosphatase or lactate dehydrogenase levels in the patient group. In conclusion, serum bAP protein measured by IRMA can be considered a sensitive marker of bone turnover and could be especially useful as valuable non-invasive biochemical marker for identifying sickle cell patients with skeletal complications. PMID:11345404

Bolarin, D M

2001-01-01

327

Monoclonal suppressor factor specific for lactate dehydrogenase B. I. Mechanism of interaction between the factor and its target cells  

PubMed Central

Hybridomas secreting a monoclonal T suppressor-effector factor (TseF) were produced by fusion of a lactate dehydrogenase B (LDHB)-specific long-term T suppressor-effector (Tse) cell line with the BW5147 thymoma. A short exposure (4 h) to TseF completely suppresses the antigen-specific and A-restricted proliferation of LDHB-primed Lyt-1+2- [possibly helper (Th)] cells. The action of TseF on Th cells, as that of the Tse cells themselves, is antigen-specific and A-restricted. The interaction of TseF with Th cells involves two binding events, of which one occurs via antigen bridge, and the other represents the recognition of a factor-derived Ak-like moiety by the anti-Ak receptor of Th cells. The Ak-like moiety of the TseF carries the determinants that serve as restriction elements for antigen recognition by Th cells, and additional determinants demonstrable by T cell-specific monoclonal "anti-Ak" antibodies, however, it lacks serologically detectable determinants of the B cell-derived A alpha A beta class II Mhc molecules.

1983-01-01

328

Comparison of parasite lactate dehydrogenase based immunochromatographic antigen detection assay (optimal) with microscopy for detection of malaria parasites.  

PubMed

This study was done to compare the ability of a newly developed rapid malaria test OPtiMAL, an immunochromatographic antigen detection assay for the diagnosis of malaria using parasite lactate dehydrogenase, against standard microscopy. Blood samples were obtained from 232 patients suspected of having malaria. A total of 122 samples (52.5%) were positive by blood films while 118 (50.8%) were positive by OPtiMAL test. The blood film indicated that 21.4% (26 of 122) of the patients were positive for P. falciparum and 78.6% (96 of 122) were infected with P. vivax. OPtiMAL test showed that 21.2% (25 of 118) were positive for P. falciparum and 78.8% (93 of 118) were infected with P. vivax. This assay had sensitivities of 88.4% and 96.8% compared to traditional blood films for detection of P. falciparum and P. vivax malaria respectively. Thus OPtiMAL test can be used with or without traditional blood film examination for detection of both P. vivax and P. falciparum malaria and can be effectively used for the rapid diagnosis of malaria. PMID:17642705

Chayani, N; Das, B; Sur, M; Bajoria, S

329

Lactate dehydrogenase, catalase, and superoxide dismutase in tumor tissue of breast cancer patients in respect to mammographic findings.  

PubMed

Lactate dehydrogenase (LDH), marker of anaerobic metabolism, is associated with highly invasive and metastatic breast cancer. Novel studies show that increased anaerobic metabolism (LDH), as well as activity of antioxidative enzymes (superoxide dismutase (SOD) and catalase (CAT)), is correlated with higher mammographic density, as known predictor of breast cancer risk. In this study, we measured LDH, MDH, and SOD activity in tumor and adjacent tissues of breast cancer patients by spectrophotometric assay. Mammograms were evaluated according to the American College of Radiology Breast Imaging Reporting and Data system. Mammographically dense breast tissue is associated with higher activity of LDH in tumor tissue of breast cancer patients. Moreover, patients with masses have significantly higher activity of LDH compared to patients with focal asymmetries or architectural distortion. Patients with spiculated mass margin had higher activity of LDH compared to patients with focal asymmetries or architectural distortion. Activity of LDH in patients significantly increases, while activity of CAT significantly decreases with the increase of BIRADS category. These results suggest that the association of activity of LDH and CAT in tumor tissue with mammographic characteristics could help in defining aggressive breast cancers. PMID:23197387

Radenkovic, Sandra; Milosevic, Zorica; Konjevic, Gordana; Karadzic, Katarina; Rovcanin, Branislav; Buta, Marko; Gopcevic, Kristina; Jurisic, Vladimir

2013-06-01

330

Transcriptome, proteome, and metabolite analyses of a lactate dehydrogenase-negative mutant of Enterococcus faecalis V583.  

PubMed

A constructed lactate dehydrogenase (LDH)-negative mutant of Enterococcus faecalis V583 grows at the same rate as the wild type but ferments glucose to ethanol, formate, and acetoin. Microarray analysis showed that LDH deficiency had profound transcriptional effects: 43 genes in the mutant were found to be upregulated, and 45 were found to be downregulated. Most of the upregulated genes encode enzymes of energy metabolism or transport. By two-dimensional (2D) gel analysis, 45 differentially expressed proteins were identified. A comparison of transcriptomic and proteomic data suggested that for several proteins the level of expression is regulated beyond the level of transcription. Pyruvate catabolic genes, including the truncated ldh gene, showed highly increased transcription in the mutant. These genes, along with a number of other differentially expressed genes, are preceded by sequences with homology to binding sites for the global redox-sensing repressor, Rex, of Staphylococcus aureus. The data indicate that the genes are transcriptionally regulated by the NADH/NAD ratio and that this ratio plays an important role in the regulatory network controlling energy metabolism in E. faecalis. PMID:21296946

Mehmeti, Ibrahim; Jönsson, Maria; Fergestad, Ellen M; Mathiesen, Geir; Nes, Ingolf F; Holo, Helge

2011-02-04

331

Trimethylamine-N-oxide counteracts urea effects on rabbit muscle lactate dehydrogenase function: a test of the counteraction hypothesis.  

PubMed Central

Trimethylamine-N-oxide (TMAO) in the cells of sharks and rays is believed to counteract the deleterious effects of the high intracellular concentrations of urea in these animals. It has been hypothesized that TMAO has the generic ability to counteract the effects of urea on protein structure and function, regardless of whether that protein actually evolved in the presence of these two solutes. Rabbit muscle lactate dehydrogenase (LDH) did not evolve in the presence of either solute, and it is used here to test the validity of the counteraction hypothesis. With pyruvate as substrate, results show that its Km and the combined Km of pyruvate and NADH are increased by urea, decreased by TMAO, and in 1:1 and 2:1 mixtures of urea:TMAO the Km values are essentially equivalent to the Km values obtained in the absence of the two solutes. In contrast, values of k(cat) and the Km for NADH as a substrate are unperturbed by urea, TMAO, or urea:TMAO mixtures. All of these effects are consistent with TMAO counteraction of the effects of urea on LDH kinetic parameters, supporting the premise that counteraction is a property of the solvent system and is independent of the evolutionary history of the protein.

Baskakov, I; Wang, A; Bolen, D W

1998-01-01

332

Serum markers lactate dehydrogenase and S100B predict independently disease outcome in melanoma patients with distant metastasis  

PubMed Central

Background: Established prognostic factors are of limited value to predict long-term survival and benefit from metastasectomy in advanced melanoma. This study aimed to identify prognostic factors in patients with distant metastasis. Methods: We analysed overall survival of 855 institutional melanoma patients with distant metastasis by bivariate Kaplan–Meier survival probabilities and multivariate Cox hazard regression analysis. Results: Serum lactate dehydrogenases (LDH), S100B, the interval between initial diagnosis and occurrence of distant metastasis, the site of distant metastases, and the number of involved distant sites were significant independent prognostic factors in both bivariate and multivariate analyses. Visceral metastases other than lung (hazard ratio (HR) 1.8), elevated S100B (HR 1.7) and elevated LDH (HR 1.6) had the highest negative impact on survival. Complete metastasectomy was likewise an independent prognostic factor in multivariate analysis. This treatment was associated with favourable survival for patients with normal LDH and S100B values (5-year survival, 37.2%). Conclusion: The serum markers LDH and S100B were both found to be prognostic factors in melanoma patients with distant metastasis. Furthermore, complete metastasectomy had an independent favourable prognostic impact in particular for the patient subgroup with normal LDH and S100B values.

Weide, B; Elsasser, M; Buttner, P; Pflugfelder, A; Leiter, U; Eigentler, T K; Bauer, J; Witte, M; Meier, F; Garbe, C

2012-01-01

333

Biochemical, biophysical, and molecular genetic studies on the membrane-bound D-lactate dehydrogenase from Escherichia coli  

SciTech Connect

The membrane-bound D-lactate dehydrogenase (D-LDH) from E. coli has been used as a model system in order to study structure-function relationships in membrane proteins. This enzyme is activated by various lipids and detergents in vitro, and appears to provide energy for the active transport of amino acids and sugars in E. coli membrane vesicles. In order to obtain sufficient amounts of enzyme for /sup 19/F nuclear magnetic resonance (NMR) and circular dichroism (CD) experiments, the cloned dld gene was used to construct a plasmid in which the expression of D-LDH is induced by a temperature shift. Upon temperature induction, the presence of this plasmid results in levels of D-LDH in the cell which are 300-fold higher than wild type levels. The cloned dld gene was also sequenced and the primary structure of D-LDH, as deduced from the DNA sequence, was verified by determination of the amino-terminal sequence and the amino acid composition of D-LDH. The dld gene codes for a protein which does not contain a signal sequence and is 571 residues long.

Rule, G.S.

1986-01-01

334

[Characterization of aldehyde dehydrogenase gene fragment from mung bean Vigna radiata using the polymerase chain reaction].  

PubMed

Two degenerate oligonucleotide sequence primers and polymerase chain reactions on total DNA have been utilized to clone on 651--bp gene fragment coding the central part of amino acid sequence of an earlier unknown aldehyde dehydrogenase (ALDH) from mung bean. The deduced partial amino acid sequence for this aldehyde dehydrogenase shows about 65% sequence identity to ALDHs of Vibrio cholerae Rhodococcus sp., Alcaligenes eutrophus and about 45% sequence identity to mammalian ALDHs 1 and 2, ALDHs of Aspergillus niger and A, nidulans, the betain aldehyde dehydrogenase from spinach. Alignment of the mung bean aldehyde dehydrogenase partial amino acid sequence with the sequence of 16 NAD(P)(+)-dependent aldehyde dehydrogenases has demonstrated that all strictly conserved amino acid residues and all three conservative regions are identical. PMID:9778740

Ponomarev, A G; Bubiakina, V V; Tatarinova, T D; Zelenin, S M

1998-01-01

335

Cloning of a Neisseria meningitidis gene for L-lactate dehydrogenase (L-LDH): evidence for a second meningococcal L-LDH with different regulation.  

PubMed Central

We report the cloning of lldA, a Neisseria meningitidis gene for L-lactate dehydrogenase (L-LDH). Escherichia coli contains a single L-LDH gene (lldD) in the lld operon (previously lct). E. coli grown in complex media does not have L-LDH activity, but the activity is induced by growth in defined medium with L-lactate as the carbon source. In contrast, meningococci contain at least one L-LDH in addition to the lldA gene product. These enzymes are active in meningococci grown in complex media and are not dependent on growth in L-lactate. The predicted amino acid sequence of lldA is homologous to that of E. coli lldD and of other prokaryotic and eukaryotic flavin mononucleotide-containing enzymes that catalyze the oxidation of L-lactate and other small alpha-hydroxy acids. A mutant with a deletion in lldA was found to have reduced L-LDH activity. However, this mutant was able to grow on L-lactate, indicating that a second L-LDH must exist. Activity of the lldA enzyme was affected by growth conditions, being increased by growth on a defined medium with either L-lactate or pyruvate as the carbon source. For meningococci grown on a complex medium, activity of the lldA enzyme was increased by growth on plates or in well-aerated broth. A second L-lactate-oxidizing activity was seen in bacteria grown in poorly aerated broth. Neisseria gonorrhoeae contains a homolog of lldA. As for meningococci, mutation of the gonococcal lldA reduced L-LDH activity but did not affect growth on L-lactate.

Erwin, A L; Gotschlich, E C

1996-01-01

336

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

PubMed

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%) K m for L-lactate and a higher V max 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 K m of L-lactate of control LDH to anoxic levels, whereas (b) stimulation of kinases increased the K m 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

2013-02-21

337

Changes of Reaction Time and Blood Lactate Concentration of Elite Volleyball Players During a Game  

PubMed Central

The purpose of this study was to investigate changes in reaction time of elite volleyball players during a game. Fourteen volleyball players participated in the study. Reaction time was measured using the Optojump system. In addition, blood lactate concentration was assessed to monitor physiological load during the game. All measurements were performed during a pre-game test and during sets 1, 2, 3 and 4. Reaction time during set 1 decreased significantly by 13,3 % compared with pre-game values, from 600 ms during the pre-game test to 520 ms during set 1 (p<0,05). Blood lactate concentration increased significantly during set 1, 2, 3 and 4 compared with pre-game conditions (p<0,05). Reaction time stays in the first phase of its changes pattern and elite volleyball players do not reach psychomotor fatigue threshold throughout the game.

Mroczek, Dariusz; Kawczynski, Adam; Chmura, Jan

2011-01-01

338

Hot spots in cold adaptation: Localized increases in conformational flexibility in lactate dehydrogenase A4 orthologs of Antarctic notothenioid fishes  

PubMed Central

To elucidate mechanisms of enzymatic adaptation to extreme cold, we determined kinetic properties, thermal stabilities, and deduced amino acid sequences of lactate dehydrogenase A4 (A4-LDH) from nine Antarctic (?1.86 to 1°C) and three South American (4 to 10°C) notothenioid teleosts. Higher Michaelis–Menten constants (Km) and catalytic rate constants (kcat) distinguish orthologs of Antarctic from those of South American species, but no relationship exists between adaptation temperature and the rate at which activity is lost because of heat denaturation. In all species, active site residues are conserved fully, and differences in kcat and Km are caused by substitutions elsewhere in the molecule. Within geographic groups, identical kinetic properties are generated by different substitutions. By combining our data with A4-LDH sequences for other vertebrates and information on roles played by localized conformational changes in setting kcat, we conclude that notothenioid A4-LDHs have adapted to cold temperatures by increases in flexibility in small areas of the molecule that affect the mobility of adjacent active-site structures. Using these findings, we propose a model that explains linked temperature-adaptive variation in Km and kcat. Changes in sequence that increase flexibility of regions of the enzyme involved in catalytic conformational changes may reduce energy (enthalpy) barriers to these rate-governing shifts in conformation and, thereby, increase kcat. However, at a common temperature of measurement, the higher configurational entropy of a cold-adapted enzyme may foster conformations that bind ligands poorly, leading to high Km values relative to warm-adapted orthologs.

Fields, Peter A.; Somero, George N.

1998-01-01

339

Alternative splicing generates two lactate dehydrogenase subunits differentially expressed during hypoxia via HIF-1 in the shrimp Litopenaeus vannamei.  

PubMed

Metabolic adjustment to low oxygen exposure (hypoxia) in the white shrimp Litopenaeus vannamei implies a shift to anaerobic metabolism. Lactate dehydrogenase (LDH) is a key enzyme of the anaerobic metabolism described in most organisms. The structure and expression of the LDH gene, as well as the LDH isoenzymes in marine crustacean are not well defined. In the present study we characterized a gene that codes for two LDH subunits, measured their expression and detected the isoenzymes in tissues from white shrimp. We also silenced the transcriptional activator hypoxia inducible factor 1 (HIF-1) to elucidate the regulation of LDH in tissues from white shrimp in response to hypoxia. The complete LDH gene coding sequence is 7571 bp (LvanLDH) and encodes two different LDH subunits (LDHvan-1 and LDHvan-2) generated by alternative splicing and composed of 332 amino acids with conserved domains important for the function and regulation. Phylogenetic analysis shows that LvanLDH -1 and -2 are closer to the invertebrate counterparts. The LDHvan-1 transcript increased 2.5-fold after hypoxia in gills but not in hepatopancreas, while the LDHvan-2 transcript decreased 14-fold in muscle but not in gills and hepatopancreas. Three bands with LDH activity of ?60–90 kDa were detected in hepatopancreas, while one band of ?140 kDa was detected in gills and muscle. The silencing of HIF-1 blocked the increase of LDH mRNA and activity produced by hypoxia in gills. These results demonstrate a single gene for LDH (LvanLDH) that by alternative splicing generates two different LDH subunits (LDHvan-1 and LDHvan-2) that are expressed in a tissue-specific manner during hypoxia via the HIF-1 pathway. PMID:22586706

Soñanez-Organis, José Guadalupe; Rodriguez-Armenta, Mariana; Leal-Rubio, Bertha; Peregrino-Uriarte, Alma Beatriz; Gómez-Jiménez, Silvia; Yepiz-Plascencia, Gloria

2012-05-01

340

Prognostic value of ferritin, neuron-specific enolase, lactate dehydrogenase, and urinary and plasmatic catecholamine metabolites in children with neuroblastoma  

PubMed Central

Different plasma and urinary parameters have been tested as valuable prognostic markers for children with neuroblastoma (NB), but conclusive results from multivariate analyses are still lacking. Samples collected at diagnosis from 505 patients diagnosed in Italy between June 1994 and November 2010 were analyzed at the Italian reference laboratory according to standard methodologies. Patient clinical data were retrieved from the Italian NB Registry. For statistical analysis, patients were grouped according to stage, age, MYCN status, and outcome. Cumulative survival was calculated by the Kaplan–Meier procedure using the first quartile of the marker distribution as a cut-off value to stratify the patients. Multivariate analysis was performed by the Cox regression model by considering only the significant variables. When the entire cohort of patients was considered, none of the different parameters had an independent prognostic value. However, in patients with localized disease without MYCN amplification the significant positive associations between urinary and plasmatic vanillylmandelic acid (VMA)/homovanillic acid (HVA) ratio and a better prognosis remained significant (P < 0.05 and P < 0.01, respectively), as well as, the positive association between high lactate dehydrogenase (LDH) values and a worse prognosis (P < 0.001). Moreover, in stage 4 patients without MYCN amplification, neuron-specific enolase levels above 200 ng/mL and LDH levels above 2500 IU/mL maintained their significant association with a worse outcome (P = 0.01 and P = 0.0001, respectively). In conclusion, LDH had an independent prognostic value in patients of all stages without MYCN amplification. Moreover, the urinary and plasmatic VMA/HVA ratio was an independent predictor of prognosis in patients with localized disease without MYCN amplification. Since LDH and catecholamine metabolites are measured in all patients at diagnosis, these findings may be helpful for an easy, cost-effective, patient risk stratification.

Cangemi, Giuliana; Reggiardo, Giorgio; Barco, Sebastiano; Barbagallo, Laura; Conte, Massimo; D'Angelo, Paolo; Bianchi, Maurizio; Favre, Claudio; Galleni, Barbara; Melioli, Giovanni; Haupt, Riccardo; Garaventa, Alberto; Corrias, Maria V

2012-01-01

341

Prognostic value of ferritin, neuron-specific enolase, lactate dehydrogenase, and urinary and plasmatic catecholamine metabolites in children with neuroblastoma.  

PubMed

Different plasma and urinary parameters have been tested as valuable prognostic markers for children with neuroblastoma (NB), but conclusive results from multivariate analyses are still lacking. Samples collected at diagnosis from 505 patients diagnosed in Italy between June 1994 and November 2010 were analyzed at the Italian reference laboratory according to standard methodologies. Patient clinical data were retrieved from the Italian NB Registry. For statistical analysis, patients were grouped according to stage, age, MYCN status, and outcome. Cumulative survival was calculated by the Kaplan-Meier procedure using the first quartile of the marker distribution as a cut-off value to stratify the patients. Multivariate analysis was performed by the Cox regression model by considering only the significant variables. When the entire cohort of patients was considered, none of the different parameters had an independent prognostic value. However, in patients with localized disease without MYCN amplification the significant positive associations between urinary and plasmatic vanillylmandelic acid (VMA)/homovanillic acid (HVA) ratio and a better prognosis remained significant (P < 0.05 and P < 0.01, respectively), as well as, the positive association between high lactate dehydrogenase (LDH) values and a worse prognosis (P < 0.001). Moreover, in stage 4 patients without MYCN amplification, neuron-specific enolase levels above 200 ng/mL and LDH levels above 2500 IU/mL maintained their significant association with a worse outcome (P = 0.01 and P = 0.0001, respectively). In conclusion, LDH had an independent prognostic value in patients of all stages without MYCN amplification. Moreover, the urinary and plasmatic VMA/HVA ratio was an independent predictor of prognosis in patients with localized disease without MYCN amplification. Since LDH and catecholamine metabolites are measured in all patients at diagnosis, these findings may be helpful for an easy, cost-effective, patient risk stratification. PMID:23226699

Cangemi, Giuliana; Reggiardo, Giorgio; Barco, Sebastiano; Barbagallo, Laura; Conte, Massimo; D'Angelo, Paolo; Bianchi, Maurizio; Favre, Claudio; Galleni, Barbara; Melioli, Giovanni; Haupt, Riccardo; Garaventa, Alberto; Corrias, Maria V

2012-11-30

342

A novel hypoxia-response element in the lactate dehydrogenase-B gene of the killifish Fundulus heteroclitus.  

PubMed

Previous studies have suggested that the lactate dehydrogenase-B gene (Ldh-B) of the Atlantic killifish, Fundulus heteroclitus, is a hypoxia-responsive gene. Here, we demonstrate that the F. heteroclitus Ldh-B promoter confers hypoxia-dependence upon reporter gene expression in transiently transfected mammalian (Hep3B) and fish (RTG-2 and RTH-149) cells in culture. Mutation and deletion analyses identified a putative hypoxia-response element (HRE) between 109 and 90 nucleotides upstream of the major start site. This HRE is characterized by the sequence 5'-GATGTG-3' spaced by 8 nucleotides from a perfect inverted repeat, and both sites are necessary for hypoxic induction of reporter gene expression in mammalian and fish cells. This HRE differs from the canonical sequence at one nucleotide position that is invariant among HREs from a wide range of hypoxia-sensitive genes. In fish cells, maximal induction of reporter gene expression driven by this HRE occurred at the lowest oxygen level tested (0.5%), took 48 h to 96 h, and was independent of glucose concentration (between 5.6 and 25 mM). Under all conditions tested, hypoxic induction of gene expression was lower in RTH-149 cells than in RTG-2, suggesting a potential defect in hypoxia signaling in RTH-149 cells. These results demonstrate that the F. heteroclitus Ldh-B promoter contains a novel HRE that is capable of driving reporter gene expression in a sequence-specific and oxygen-, time-, and cell line-dependent manner. PMID:19439190

Rees, Bernard B; Figueroa, Yanira G; Wiese, Thomas E; Beckman, Barbara S; Schulte, Patricia M

2009-05-08

343

Structure and sequence conservation of a putative hypoxia response element in the lactate dehydrogenase-B gene of Fundulus.  

PubMed

Many aquatic habitats are characterized by periodic or sustained episodes of low oxygen concentration, or hypoxia, and organisms that survive in these habitats do so by utilizing a suite of behavioral, physiological and biochemical adjustments to low oxygen (1-3). In the killifish Fundulus heteroclitus, one response to prolonged exposure to hypoxia is an increase in the activity of lactate dehydrogenase-B (LDH-B), the terminal enzyme of anaerobic glycolysis, in liver tissue (4). An increase in glycolytic enzyme activity also occurs in mammalian cells during hypoxia, a process due, in part, to increased rates of gene transcription mediated by the hypoxia-inducible transcription factor, HIF-1 (5). Given that a homolog of HIF-1 has been identified in fish (6), we hypothesized that HIF might be involved in the observed up-regulation of LDH-B in F. heteroclitus. Herein, we describe the presence of DNA elements in intron 2 of the Ldh-B gene from F. heteroclitus that resemble hypoxia response elements (HRE) describedfor mammalian genes (7-10). Specifically, over a region of approximately 50 base pairs we identified two consensus HIF-1 binding sites, as well as DNA elements that may bind other transcription factors (e.g., cyclic AMP response elements; CRE). We found that these sites were perfectly conserved among geographically diverse populations of F. heteroclitus, as well as being highly conserved among multiple species in the genus Fundulus. The spacing, orientation, and sequence conservation of these putative regulatory elements suggest that they may be functionally involved in the hypoxic regulation of Ldh-B in these fish. PMID:11441966

Rees, B B; Bowman, J A; Schulte, P M

2001-06-01

344

Molecular genetic characterization of the L-lactate dehydrogenase gene (ldhL) of Lactobacillus helveticus and biochemical characterization of the enzyme.  

PubMed Central

The Lactobacillus helveticus L-(+)-lactate dehydrogenase (L-LDH) gene (ldhL) was isolated from a lambda library. The nucleotide sequence of the ldhL gene was determined and shown to have the capacity to encode a protein of 323 amino acids (35.3 kDa). The deduced sequence of the 35-kDa protein revealed a relatively high degree of identity with other lactobacillar L-LDHs. The highest identity (80.2%) was observed with the Lactobacillus casei L-LDH. The sizes and 5' end analyses of ldhL transcripts showed that the ldhL gene is a monocistronic transcriptional unit. The expression of ldhL, studied as a function of growth, revealed a high expression level at the logarithmic phase of growth. The ldhL gene is preceded by two putative -10 regions, but no corresponding -35 regions could be identified. By primer extension analysis, the ldhL transcripts were confirmed to be derived from the -10 region closest to the initiation codon. However, upstream of these regions additional putative -10/-35 regions could be found. The L-LDH was overexpressed in Escherichia coli and purified to homogeneity by two chromatographic steps. The purified L-LDH was shown to be a nonaliosteric enzyme, and amino acid residues involved in allosteric regulation were not conserved in L. helveticus L-LDH. However, a slight enhancement of enzyme activity was observed in the presence of fructose 1,6-diphosphate, particularly at neutral pH. A detailed enzymatic characterization of L-LDH was performed. The optimal reaction velocity was at pH 5.0, where the kinetic parameters K(m), and Kcat for pyruvate were 0.25 mM and 643 S-1, respectively.

Savijoki, K; Palva, A

1997-01-01

345

Differences and similarities in binding of pyruvate and l-lactate in the active site of M 4 and H 4 isoforms of human lactate dehydrogenase  

Microsoft Academic Search

We present QM\\/MM calculations that show differences in geometries of active sites of M4 and H4 isoforms of human LDH ligated with oxamate, pyruvate or l-lactate. As the consequence of these differences, binding isotope effects of the methyl hydrogen atoms of pyruvate and l-lactate may be used to experimentally distinguish these isoforms. Based on the FEP calculations we argue that

Katarzyna ?widerek; Piotr Paneth

2011-01-01

346

Secondary sup 15 N isotope effects on the reactions catalyzed by alcohol and formate dehydrogenases  

SciTech Connect

Secondary {sup 15}N isotope effects at the N-1 position of 3-acetylpyridine adenine dinucleotide have been determined, by using the internal competition technique, for horse liver alcohol dehydrogenase (LADH) with cyclohexanol as a substrate and yeast formate dehydrogenase (FDH) with formate as a substrate. On the basis of less precise previous measurements of these {sup 15}N isotope effects, the nicotinamide ring of NAD has been suggested to adopt a boat conformation with carbonium ion character at C-4 during hydride transfer. If this mechanism were valid, as N-1 becomes pyramidal an {sup 15}N isotope effect for the reaction catalyzed by LADH was measured. These values suggest that a significant {sup 15}N kinetic isotope effect is not associated with hydride transfer for LADH and FDH. Thus, in contrast with the deformation mechanism previously postulated, the pyridine ring of the nucleotide apparently remains planar during these dehydrogenase reactions.

Rotberg, N.S.; Cleland, W.W. (Univ. of Wisconsin, Madison (USA))

1991-04-23

347

Contribution of a buried aspartate residue towards the catalytic efficiency and structural stability of Bacillus stearothermophilus lactate dehydrogenase.  

PubMed Central

The X-ray structure of lactate dehydrogenase (LDH) shows the side-chain carboxylate group of Asp-143 to be buried in the hydrophobic interior of the enzyme, where it makes hydrogen-bonding interactions with both the side-chain hydroxyl group of Ser-273 and the main-chain amide group of His-195. This is an unusual environment for a carboxylate side-chain as hydrogen bonding normally occurs with water molecules at the surface of the protein. A charged hydrogen-bonding interaction in the interior of a protein would be expected to be much stronger than a similar interaction on the solvent-exposed exterior. In this respect the side-chain carboxylate group of Asp-143 appears to be important for maintaining tertiary structure by providing a common linkage point between three discontinuous elements of the secondary structure, alpha 1F, beta K and the beta-turn joining beta G and beta H. The contribution of the Asp-143 side-chain to the structure and function of Bacillus stearothermophilus LDH was assessed by creating a mutant enzyme containing Asn-143. The decreased thermal stability of both unactivated and fructose-1,6-diphosphate (Fru-1,6-P2)-activated forms of the mutant enzyme support a structural role for Asp-143. Furthermore, the difference in stability of the wild-type and mutant enzymes in guanidinium chloride suggested that the carboxylate group of Asp-143 contributes at least 22 kJ/mol to the conformational stability of the wild-type enzyme. However, there was no alteration in the amount of accessible tryptophan fluorescence in the mutant enzyme, indicating that the mutation caused a structural weakness rather than a gross conformational change. Comparison of the wild-type and mutant enzyme steady-state parameters for various 2-keto acid substrates showed the mutation to have a general effect on catalysis, with an average difference in binding energy of 11 kJ/mol for the transition-state complexes. The different effects of pH and Fru-1,6-P2 on the wild-type and mutant enzymes also confirmed a perturbation of the catalytic centre in the mutant enzyme. As the side-chain of Asp-143 is not sufficiently close to the active site to be directly involved in catalysis or substrate binding it is proposed that the effects on catalysis shown by the mutant enzyme are induced either by a structural change or by charge imbalance at the active site.(ABSTRACT TRUNCATED AT 400 WORDS) Images Figure 1

Nobbs, T J; Cortes, A; Gelpi, J L; Holbrook, J J; Atkinson, T; Scawen, M D; Nicholls, D J

1994-01-01

348

Differences and similarities in binding of pyruvate and L-lactate in the active site of M4 and H4 isoforms of human lactate dehydrogenase.  

PubMed

We present QM/MM calculations that show differences in geometries of active sites of M(4) and H(4) isoforms of human LDH ligated with oxamate, pyruvate or L-lactate. As the consequence of these differences, binding isotope effects of the methyl hydrogen atoms of pyruvate and l-lactate may be used to experimentally distinguish these isoforms. Based on the FEP calculations we argue that L-lactate is a better candidate for the experimental studies. Our calculations of energies of interactions of ligands with the active site residues provide explanation for the observed experimentally sensitivity to inhibition of the M(4) isoenzyme isoform and pinpoint the differences to interactions of the ligand with the histidine residue. We conclude that pyruvate interacts much stronger in the active site of H(4) than M(4) isoform and that the latter interactions are weaker than with water molecules in the aqueous solution. PMID:20951115

Swiderek, Katarzyna; Paneth, Piotr

2010-10-14

349

Lactate dehydrogenase test  

MedlinePLUS

... Normal value ranges may vary slightly among different laboratories. Some labs use different measurements or test different samples. Talk to your doctor about the meaning of your specific test results.

350

The ldhA gene, encoding fermentative L-lactate dehydrogenase of Corynebacterium glutamicum, is under the control of positive feedback regulation mediated by LldR.  

PubMed

Corynebacterium glutamicum ldhA encodes L-lactate dehydrogenase, a key enzyme that couples L-lactate production to reoxidation of NADH formed during glycolysis. We previously showed that in the absence of sugar, SugR binds to the ldhA promoter region, thereby repressing ldhA expression. In this study we show that LldR is another protein that binds to the ldhA promoter region, thus regulating ldhA expression. LldR has hitherto been characterized as an L-lactate-responsive transcriptional repressor of L-lactate utilization genes. Transposon mutagenesis of a reporter strain carrying a chromosomal ldhA promoter-lacZ fusion (PldhA-lacZ) revealed that ldhA disruption drastically decreased expression of PldhA-lacZ. PldhA-lacZ expression in the ldhA mutant was restored by deletion of lldR, suggesting that LldR acts as a repressor of ldhA in the absence of L-lactate and the LldR-mediated repression is not relieved in the ldhA mutant due to its inability to produce L-lactate. lldR deletion did not affect PldhA-lacZ expression in the wild-type background during growth on either glucose, acetate, or L-lactate. However, it upregulated PldhA-lacZ expression in the sugR mutant background during growth on acetate. The binding sites of LldR and SugR are located around the -35 and -10 regions of the ldhA promoter, respectively. C. glutamicum ldhA expression is therefore primarily repressed by SugR in the absence of sugar. In the presence of sugar, SugR-mediated repression of ldhA is alleviated, and ldhA expression is additionally enhanced by LldR inactivation in response to L-lactate produced by LdhA. PMID:19429617

Toyoda, Koichi; Teramoto, Haruhiko; Inui, Masayuki; Yukawa, Hideaki

2009-05-08

351

Impact of critical process and formulation parameters affecting in-process stability of lactate dehydrogenase during the secondary drying stage of lyophilization: a mini freeze dryer study.  

PubMed

The stresses during the secondary-drying stage of lyophilization were investigated using a controlled humidity mini-freeze-dryer [Luthra S, Obert J-P, Kalonia DS, Pikal MJ. 2007. Investigation of drying stresses on proteins during lyophilization: Differentiation between primary and secondary-drying stresses on lactate dehydrogenase using a humidity controlled mini freeze-dryer. J Pharm Sci 96: 61-70.]. Lactate dehydrogenase (LDH), was formulated in: (1) Tween 80, (2) citrate buffer, and (3) both Tween 80 and citrate buffer. Protein activity recovery was measured as a function of relative humidity (RH), product temperature, and drying duration. Studies were also conducted with different concentrations of sucrose, sorbitol, and poly (vinyl pyrrolidone) (PVP). LDH stability was affected to a small extent by RH and significantly by drying temperature and duration. Complete stabilization of LDH was observed when lyophilized with sucrose and PVP but only a partial stabilization was observed with sorbitol. The mini-freeze-dryer enabled studying the process parameters independently, unlike a conventional study where these effects are generally convoluted. The results suggest that the stability of the protein is a function of the dynamics of the system during lyophilization. The origin of the stabilization effect of sucrose, which could, in principle, be attributed both to direct interaction with the protein or vitrification of the protein was elucidated using lyoprotectants that can either hydrogen bond well with the protein (sorbitol) or form a good glass (PVP). It appears both effects are required for complete stabilization of the protein. PMID:17621675

Luthra, Sumit; Obert, Jean-Philippe; Kalonia, Devendra S; Pikal, Michael J

2007-09-01

352

Cloning and expression of the gene encoding the fructose-1,6-diphosphate-dependent L-(+)-lactate dehydrogenase of Streptococcus mutans.  

PubMed Central

The fructose-1,6-diphosphate-dependent lactate dehydrogenase from Streptococcus mutans JH1000 was purified by a modification of published methods. The sequence of 27 amino-terminal amino acids was determined, which allowed us to construct a 17-base DNA probe that had 32-fold degeneracy. The probe was used to screen a genomic library in pBR322. Of 18 reactive clones, 1 was found that expressed lactate dehydrogenase (LDH) activity identical to that of S. mutans with regard to dependence on fructose-1,6-diphosphate, thermal inactivation profile, and inhibition by sodium oxamate. Extracts of this clone possessed a protein band that comigrated in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with purified LDH from JH1000. Compared with controls, the clone was shown to produce elevated amounts of L-(+)-lactic acid during growth in the presence of glucose, thereby indicating that the activity was expressed in vivo. This result was substantiated by demonstrating that the activity could complement a mutation in the fermentative D-(-)-LDH of Escherichia coli. Subcloning showed that the S. mutans LDH subunit is encoded by a 1.2-kilobase gene. Our ability to clone this gene is expected to have great practical significance in the construction of an effector strain for use in the replacement therapy of dental caries. Images

Hillman, J D; Duncan, M J; Stashenko, K P

1990-01-01

353

Low potential detection of NADH based on Fe?O? nanoparticles/multiwalled carbon nanotubes composite: fabrication of integrated dehydrogenase-based lactate biosensor.  

PubMed

Fe(3)O(4) magnetic nanoparticles were in situ loaded on the surface of multiwalled carbon nanotubes (MWCNTs) by a simple coprecipitation procedure. The resulting Fe(3)O(4)/MWCNTs nanocomposite brings new capabilities for electrochemical sensing by combining the advantages of Fe(3)O(4) magnetic nanoparticles and MWCNTs. It was found that Fe(3)O(4) has redox properties similar to those of frequently used mediators used for electron transfer between NADH and electrode. The cyclic voltammetric results indicated the ability of Fe(3)O(4)/MWCNTs modified GC electrode to catalyze the oxidation of NADH at a very low potential (0.0 mV vs. Ag/AgCl) and subsequently, a substantial decrease in the overpotential by about 650 mV compared with the bare GC electrode. The catalytic oxidation current allows the stable and selective amperometric detection of NADH at an applied potential of 0.0 mV (Ag/AgCl) with a detection limit of 0.3 ?M and linear response up to 300 ?M. This modified electrode can be used as an efficient transducer in the design of biosensors based on coupled dehydrogenase enzymes. Lactate dehydrogenase (LDH) and NAD(+) were subsequently immobilized onto the Fe(3)O(4)/MWCNTs nanocomposite film by covalent bond formation between the amine groups of enzyme or NAD(+) and the carboxylic acid groups of the Fe(3)O(4)/MWCNT film. Differential pulse voltammetric detection of lactate on Fe(3)O(4)/MWCNT/LDH/NAD(+) modified GC electrode gives linear responses over the concentration range of 50-500 ?M with the detection limit of 5 ?M and sensitivity of 7.67 ?A mM(-1). Furthermore, the applicability of the sensor for the analysis of lactate concentration in human serum samples has been successfully demonstrated. PMID:22230696

Teymourian, Hazhir; Salimi, Abdollah; Hallaj, Rahman

2011-12-26

354

Mitochondrial Lactate Dehydrogenase Is Involved in Oxidative-Energy Metabolism in Human Astrocytoma Cells (CCF-STTG1)  

Microsoft Academic Search

Lactate has long been regarded as an end product of anaerobic energy production and its fate in cerebral metabolism has not been precisely delineated. In this report, we demonstrate, for the first time, the ability of a human astrocytic cell line (CCF-STTG1) to consume lactate and to generate ATP via oxidative phosphorylation. 13C-NMR and HPLC analyses aided in the identification

Joseph Lemire; Ryan J. Mailloux; Vasu D. Appanna

2008-01-01

355

Homo-d-Lactic Acid Fermentation from Arabinose by Redirection of the Phosphoketolase Pathway to the Pentose Phosphate Pathway in l-Lactate Dehydrogenase Gene-Deficient Lactobacillus plantarum?  

PubMed Central

Optically pure d-lactic acid fermentation from arabinose was achieved by using the Lactobacillus plantarum NCIMB 8826 strain whose l-lactate dehydrogenase gene was deficient and whose phosphoketolase gene was substituted with a heterologous transketolase gene. After 27 h of fermentation, 38.6 g/liter of d-lactic acid was produced from 50 g/liter of arabinose.

Okano, Kenji; Yoshida, Shogo; Tanaka, Tsutomu; Ogino, Chiaki; Fukuda, Hideki; Kondo, Akihiko

2009-01-01

356

Immobilized metal ion affinity chromatography on Co2+-carboxymethylaspartate-agarose Superflow, as demonstrated by one-step purification of lactate dehydrogenase from chicken breast muscle.  

PubMed

A rapid method for the purification of lactate dehydrogenase from whole chicken muscle extract in one chromatographic step is reported. The purification procedure can be accomplished in less than 1 h. A new type of immobilized metal ion affinity chromatography adsorbent is used that can be utilized at linear flow rates higher than 5 cm/min. The final preparation of the enzyme was with purity higher than 95% as ascertained by SDS-PAGE. Three immobilized metal ions (Ni2+, Zn2+ and Co2+) were compared for their binding properties towards the purified enzyme. The binding site of the enzyme for immobilized intermediate metal ions was determined after cleavage with CNBr and binding studies of the derivative peptides on immobilized Co2+. A peptide located on the N-terminus of the enzyme, implicated in the binding, has great potential as a purification tag in fusion proteins. PMID:9889081

Chaga, G; Hopp, J; Nelson, P

1999-02-01

357

Supplementation of medium with diammonium hydrogen phosphate enhanced the D-lactate dehydrogenase levels leading to increased D-lactic acid productivity.  

PubMed

The production of D-lactic acid by Lactobacillus lactis RM2-24 was investigated using modified media to increase the efficiency of the fermentation process. The results indicated that the addition of 5 g/l peptone and 1 g/l (NH4)2HPO4 enhanced D-lactic acid production by 32%, as compared to that obtained from non supplemented media, with a productivity of 3.0 g/l/h. Lactate dehydrogenase (LDH) expression profile in these different media was studied which resulted in appearance of additional LDH isoform produced by cells when they were grown in HSYE supplemented with (NH4)2HPO4. The additional LDH appears to be L-LDH contributing to production of L-lactic acid in the fermented broth. This is totally new information in the lactic acid fermentation and could be very useful to industries engaged in D-lactic acid production. PMID:23932744

Singhvi, Mamata; Jadhav, Akanksha; Gokhale, Digambar

2013-07-20

358

Elevated pretreatment serum levels of soluble vascular cell adhesion molecule 1 and lactate dehydrogenase as predictors of survival in cutaneous metastatic malignant melanoma.  

PubMed Central

Very rapid progression of disease with a median survival of 6-9 months is a common feature of metastatic cutaneous malignant melanoma. Nevertheless, substantial variability of survival suggests that metastatic cutaneous malignant melanoma can be divided into several biological subgroups. Pretreatment serum levels of soluble adhesion molecules and various clinical parameters in cutaneous metastatic malignant melanoma were evaluated to determine their prognostic value. In this study pretreatment serum levels of soluble vascular cell adhesion molecule 1 (sVCAM-1), soluble intercellular cell adhesion molecule 1 (sICAM-1), soluble endothelial leukocyte adhesion molecule 1 (sE-selectin) and multiple clinical factors were assessed in relation to overall survival of 97 consecutive patients with metastatic cutaneous malignant melanoma seen at our institution between May 1990 and April 1996. For statistical analysis, both univariate and multivariate Cox proportional-hazards models were used. Elevated pretreatment serum levels of sVCAM-1 (P < 0.005) and of lactate dehydrogenase (P < 0.002) were rendered statistically independent and were significantly associated with unfavourable outcome. Patients were assigned to one of three risk categories (low, intermediate and high) according to a cumulative risk score defined as the function of the sum of these two variables. There were significant differences in overall survival (P < 0.0001) between low- (n = 53, 5-year survival probability of 23.3%), intermediate- (n = 29, 5-year survival probability of 9.9%) and high-risk (n = 15) patients. Elevated pretreatment serum levels of sVCAM-1 and of lactate dehydrogenase correlate with poor outcome in metastatic cutaneous malignant melanoma. These data support risk stratification for future therapeutic trials and identify factors that need to be validated in prospective studies and may potentially influence decision-making in palliative management of patients with disseminated cutaneous malignant melanoma.

Franzke, A.; Probst-Kepper, M.; Buer, J.; Duensing, S.; Hoffmann, R.; Wittke, F.; Volkenandt, M.; Ganser, A.; Atzpodien, J.

1998-01-01

359

Cytosolic ratio of malate dehyrogenase\\/lactate dehydrogenase activity in peripheral leukocytes of race horses with training  

Microsoft Academic Search

The activities of the enzymes involved in the malate–aspartate shuttle and m RNA expression of malate dehydrogenase (MDH), a crucial enzyme for the NADH shuttle that produces ATP in glucose metabolism in the peripheral leukocytes of horses, were measured to investigate the change in metabolic states with training. There were no significant differences in plasma glucose and immunoreactive insulin concentrations

T. Arai; M. Hosoya; M. Nakamura; E. Magoori; Y. Uematsu; T. Sako

2002-01-01

360

Efficient Production of l-Lactic Acid by Metabolically Engineered Saccharomyces cerevisiae with a Genome-Integrated l-Lactate Dehydrogenase Gene  

PubMed Central

We developed a metabolically engineered yeast which produces lactic acid efficiently. In this recombinant strain, the coding region for pyruvate decarboxylase 1 (PDC1) on chromosome XII is substituted for that of the l-lactate dehydrogenase gene (LDH) through homologous recombination. The expression of mRNA for the genome-integrated LDH is regulated under the control of the native PDC1 promoter, while PDC1 is completely disrupted. Using this method, we constructed a diploid yeast transformant, with each haploid genome having a single insertion of bovine LDH. Yeast cells expressing LDH were observed to convert glucose to both lactate (55.6 g/liter) and ethanol (16.9 g/liter), with up to 62.2% of the glucose being transformed into lactic acid under neutralizing conditions. This transgenic strain, which expresses bovine LDH under the control of the PDC1 promoter, also showed high lactic acid production (50.2 g/liter) under nonneutralizing conditions. The differences in lactic acid production were compared among four different recombinants expressing a heterologous LDH gene (i.e., either the bovine LDH gene or the Bifidobacterium longum LDH gene): two transgenic strains with 2?m plasmid-based vectors and two genome-integrated strains.

Ishida, Nobuhiro; Saitoh, Satoshi; Tokuhiro, Kenro; Nagamori, Eiji; Matsuyama, Takashi; Kitamoto, Katsuhiko; Takahashi, Haruo

2005-01-01

361

Hypoxia/anoxia as signaling for increased alcohol dehydrogenase activity in saffron (Crocus sativus L.) corm.  

PubMed

Alcohol dehydrogenase, NAD-dependent lactate dehydrogenase, and NAD-independent lactate dehydrogenase activities were investigated in corms cultivated in normoxic and hypoxic/anoxic conditions. Depending on the developmental stage, hypoxia/anoxia was a signal for increase in either alcohol dehydrogenase or NAD-dependent lactate dehydrogenase. NAD-independent lactate dehydrogenase contributed to the recycling of lactate, thus preventing acidosis. PMID:15659829

Keyhani, Ezzatollah; Keyhani, Jacqueline

2004-12-01

362

Role of pyruvate kinase, phosphoenolpyruvate carboxykinase, malic enzyme and lactate dehydrogenase in anaerobic energy metabolism of Tubifex spec  

Microsoft Academic Search

1.During anaerobic exposure ofTubifex, lactate and alanine increase only within the first 24 h, while concentrations of succinate, propionate and also acetate continually increase under prevailing anaerobic conditions. Enzymes involved in anaerobic energy metabolism were isolated, and the effects of various metabolites and inorganic compounds on their catalytic properties studied.2.The specific activities of the cytosolic enzymes LDH, PK, MDH, and

K. H. Hoffmann; T. Mustafa; J. B. Jørgensen

1979-01-01

363

Mechanism of the dehydrogenase reaction of DmpFG and analysis of inter-subunit channeling efficiency and thermodynamic parameters in the overall reaction.  

PubMed

The bifunctional, microbial enzyme DmpFG is comprised of two subunits: the aldolase, DmpG, and the dehydrogenase, DmpF. DmpFG is of interest due to its ability to channel substrates between the two spatially distinct active sites. While the aldolase is well studied, significantly less is known about the dehydrogenase. Steady-state kinetic measurements of the reverse reaction of DmpF confirmed that the dehydrogenase uses a ping-pong mechanism, with substrate inhibition by acetyl CoA indicating that NAD(+)/NADH and CoA/acetyl CoA bind to the same site in DmpF. The Km of DmpF for exogenous acetaldehyde as a substrate was 23.7 mM, demonstrating the necessity for the channel to deliver acetaldehyde directly from the aldolase to the dehydrogenase active site. A channeling assay on the bifunctional enzyme gave an efficiency of 93% indicating that less than 10% of the toxic acetaldehyde leaks out of the channel into the bulk media, prior to reaching the dehydrogenase active site. The thermodynamic activation parameters of the reactions catalyzed by the aldolase, the dehydrogenase and the DmpFG complex were determined. The Gibb's free energy of activation for the dehydrogenase reaction was lower than that obtained for the full DmpFG reaction, in agreement with the high kcat obtained for the dehydrogenase reaction in isolation. Furthermore, although both the DmpF and DmpG reactions occur with small, favorable entropies of activation, the full DmpFG reaction occurs with a negative entropy of activation. This supports the concept of allosteric structural communication between the two enzymes to coordinate their activities. PMID:23742989

Smith, Natalie E; Tie, Wan Jun; Flematti, Gavin R; Stubbs, Keith A; Corry, Ben; Attwood, Paul V; Vrielink, Alice

2013-06-03

364

Meso-alpha,epsilon-diaminopimelate D-dehydrogenase: distribution and the reaction product.  

PubMed Central

A high activity of meso-alpha-epsilon-diaminopimelate dehydrogenase was found in extracts of Bacillus sphaericus, Brevibacterium sp., Corynebacterium glutamicum, and Proteus vulgaris among bacteria tested. B. sphaericus IFO 3525, in which the enzyme is most abundant, was chosen to study the enzyme reaction. The enzyme was not induced by the addition of meso-alpha-epsilon-diaminopimelate to the growth medium. The reaction product was isolated and identified as alpha-amino-epsilon-ketopimelate by a comparison of the properties of its 2,4-dinitrophenylhydrazone with those of an authentic sample in silica gel thin-layer chromatography, absorption, infrared and proton nuclear magnetic resonance spectrometry, and elemental analyses. The alpha-amino-epsilon-ketopimelate formed enzymatically was decarboxylated by H2O2 to yield L-alpha-aminoadipate. This suggests that the amino group with D-configuration in the substrate is oxidatively deaminated; the enzyme is a D-amino acid dehydrogenase. L-alpha-Amino-epsilon-ketopimelate undergoes spontaneous dehydration to the cyclic delta1-piperideine-2,6-dicarboxylate. The enzyme reaction is reversible, and meso-alpha-epsilon-diaminopimelate was formed in the reductive amination of L-alpha-epsilon-ketopimelate.

Misono, H; Togawa, H; Yamamoto, T; Soda, K

1979-01-01

365

Direct production of allitol from D-fructose by a coupling reaction using D-tagatose 3-epimerase, ribitol dehydrogenase and formate dehydrogenase.  

PubMed

Allitol was produced from D-fructose via a new NADH-regenerating enzymatic reaction system using D-tagatose 3-epimerase (D-TE), ribitol dehydrogenase (RDH), and formate dehydrogenase (FDH). D-fructose was epimerized to D-psicose by the D-TE of Pseudomonas cichorii ST-24 and the D-psicose was subsequently reduced to allitol by the RDH of an RDH-constitutive mutant, X-22, derived from Klebsiella pneumoniae IFO 3321. NADH regeneration for the reduction of D-psicose by the RDH was achieved by the irreversible formate dehydrogenase reaction, which allowed the D-psicose produced from d-fructose to be successively transformed to allitol with a production yield from D-fructose of almost 100%. The reactions progressed without any by-product formation. After separation of the product from the reaction mixture by a simple procedure, a crystal of allitol was obtained in a yield exceeding 90%. This crystal was characterized and determined to be allitol by HPLC analysis, its IR and NMR spectra, its melting point, and optical rotation measurement. PMID:16232907

Takeshita, K; Ishida, Y; Takada, G; Izumori, K

2000-01-01

366

Overexpression of pyruvate dehydrogenase kinase 1 and lactate dehydrogenase A in nerve cells confers resistance to amyloid ? and other toxins by decreasing mitochondrial respiration and reactive oxygen species production.  

PubMed

We previously demonstrated that nerve cell lines selected for resistance to amyloid ? (A?) peptide exhibit elevated aerobic glycolysis in part due to increased expression of pyruvate dehydrogenase kinase 1 (PDK1) and lactate dehydrogenase A (LDHA). Here, we show that overexpression of either PDK1 or LDHA in a rat CNS cell line (B12) confers resistance to A? and other neurotoxins. Treatment of A?-sensitive cells with various toxins resulted in mitochondrial hyperpolarization, immediately followed by rapid depolarization and cell death, events accompanied by increased production of cellular reactive oxygen species (ROS). In contrast, cells expressing either PDK1 or LDHA maintained a lower mitochondrial membrane potential and decreased ROS production with or without exposure to toxins. Additionally, PDK1- and LDHA-overexpressing cells exhibited decreased oxygen consumption but maintained levels of ATP under both normal culture conditions and following A? treatment. Interestingly, immunoblot analysis of wild type mouse primary cortical neurons treated with A? or cortical tissue extracts from 12-month-old APPswe/PS1dE9 transgenic mice showed decreased expression of LDHA and PDK1 when compared with controls. Additionally, post-mortem brain extracts from patients with Alzheimer disease exhibited a decrease in PDK1 expression compared with nondemented patients. Collectively, these findings indicate that key Warburg effect enzymes play a central role in mediating neuronal resistance to ?? or other neurotoxins by decreasing mitochondrial activity and subsequent ROS production. Maintenance of PDK1 or LDHA expression in certain regions of the brain may explain why some individuals tolerate high levels of A? deposition without developing Alzheimer disease. PMID:22948140

Newington, Jordan T; Rappon, Tim; Albers, Shawn; Wong, Daisy Y; Rylett, R Jane; Cumming, Robert C

2012-09-04

367

Kinetic study of heavy metal salt effects on the activity of L-lactate dehydrogenase in solution or immobilized on an oxygen electrode.  

PubMed

A sensitive and convenient biosensor for detection of heavy metal salts has been developed. The method is based on the effects of heavy metal salts on the catalytic activity of L-lactate dehydrogenase (LDH) in solution or coimmobilized with L-lactate oxidase (LOD) on an oxygen electrode. At metal concentrations below 100 microM, the kinetic behavior, with the LDH substrate NADH, showed a competitive inhibition with high affinity during the first 10 s. With increased incubation time, irreversible first order inactivation with respect to enzyme concentration was observed. This irreversible inactivation of LDH in solution was dose dependent. The efficiencies obtained for the different heavy metal salts were: HgGl2 > AgNO3 > Pb(COOCH3)2 > CuSO4 > ZnCl2. HgCl2 and AgNO3 were effective in the nanomolar range while the other metal salts acted at the micromolar level. LDH is protected by saturating amounts of substrate NADH against the effects of the heavy metal salts studied. The pKs for LDH catalytic activity and inactivation by heavy metal salts were similar. The results suggest binding of the heavy metal salts to the enzyme active site. Except for lead acetate, all heavy metal detection was in the range of European norms. For AgNO3, CuSO4 and HgCl2, the sensor limit of detection reached the European norm values whereas with ZnCl2 it was well below. The immobilization of LDH considerably decreased the amount of enzyme consumed by permitting repetitive assays. The efficiency of inactivation by the heavy metal salts was reduced in comparison with LDH in solution. Restoration of activity of the inactivated immobilized enzyme was obtained with DTT, EDTA, KCN and NADH treatment. This opens up possibilities for detection of toxic compounds using simple procedures suitable for assays in a variety of monitoring conditions in environmental and food pollution control. PMID:9828387

Fennouh, S; Casimiri, V; Geloso-Meyer, A; Burstein, C

1998-10-01

368

Disequilibrium in the malate dehydrogenase reaction in rat liver mitochondria in vivo  

PubMed Central

1. When [2-14C]pyruvate is injected into rats the C3-position of liver glutamate becomes more heavily labelled than the C2-position, thus establishing that oxaloacetate and fumarate are not in equilibrium in rat liver mitochondria in vivo. The amount of disequilibrium was shown to be simply related to the value that the C3-label/C2-label ratio would have were no label recycled. This ratio, z, was calculated for post-absorptive rats in environmental temperatures of 20° and 30°C from determinations of the distribution of label within glutamate 1, 3 and 10min after intravenous injection of [2-14C]pyruvate. The values of z (best estimate and range) were 1.65 (1.60–1.69) in rats at 20°C and 2.43 (2.23–2.63) in rats at 30°C. These values of z imply the following rates of interconversion in mitochondria of fumarate and oxaloacetate (in terms of the oxaloacetate?citrate flux, R) in rats at 20°C: [Formula: see text] and in rats at 30°C: [Formula: see text] 2. The kinetic parameters of malate dehydrogenase and fumarate hydratase and the intramitochondrial concentrations of NAD+ and NADH under (as far as could be judged) conditions in vivo were collated. From them and the best estimates of R now available were calculated the rates of interconversion of fumarate, malate and oxaloacetate required to give the found values of z. These rates showed that the fumarate hydratase reaction was nearly in equilibrium, but that the malate dehydrogenase reaction was considerably out of equilibrium. The calculations also led to the following conclusions. 3. In livers of rats at 20° and 30°C mitochondrial malate concentrations were respectively about 5 and 1.5 times mean cellular concentrations. 4. Mitochondrial oxaloacetate concentrations were less than 0.2 of the mean cellular concentrations. They were also only 0.65 and 0.55 of the equilibrium concentrations for the malate dehydrogenase reaction in rats at 20° and 30°C respectively. 5. Malate dehydrogenase activity was low because of the very low oxaloacetate concentrations in the mitochondria and the very small fraction of the enzyme complexed with NAD+, i.e. in each direction one substrate concentration was very sub-optimal.

Heath, D. F.; Phillips, J. C.

1972-01-01

369

Interaction of cytoplasmic dehydrogenases: quantitation of pathways of ethanol metabolism.  

PubMed

The interaction between xylitol, alcohol and lactate dehydrogenase has been studied in hepatocytes from rats by applying specifically tritiated substrates. A simple model, describing the metabolic fate of tritium from [2-3H] xylitol and (1R) [1-3H]ethanol is presented. The model allows calculation of the specific radioactivity of free, cytosolic NADH, based on transfer of tritium to lactate, glucose and water. From the initial labelling rate of lactate and the specific radioactivity of cytosolic NADH, we have determined the reversible flow through the lactate dehydrogenase catalyzed reaction to 1-5 mumol/min . g wet wt. The results suggest that xylitol, alcohol and lactate dehydrogenase share the same pool of NAD(H) in the cytoplasma. This finding allows estimation of the ethanol oxidation rate by the non-alcohol dehydrogenase pathways from the relative yield of tritium in water and glucose. The calculations are based on a comparison of the fate of the 1-pro-R hydrogen of ethanol and the hydrogen bound to carbon 2 of xylitol or carbon 2 of lactate under identical conditions. PMID:6356159

Vind, C; Grunnet, N

1983-01-01

370

Homo-D-lactic acid fermentation from arabinose by redirection of the phosphoketolase pathway to the pentose phosphate pathway in L-lactate dehydrogenase gene-deficient Lactobacillus plantarum.  

PubMed

Optically pure d-lactic acid fermentation from arabinose was achieved by using the Lactobacillus plantarum NCIMB 8826 strain whose l-lactate dehydrogenase gene was deficient and whose phosphoketolase gene was substituted with a heterologous transketolase gene. After 27 h of fermentation, 38.6 g/liter of d-lactic acid was produced from 50 g/liter of arabinose. PMID:19502433

Okano, Kenji; Yoshida, Shogo; Tanaka, Tsutomu; Ogino, Chiaki; Fukuda, Hideki; Kondo, Akihiko

2009-06-05

371

Improved Production of Homo-d-Lactic Acid via Xylose Fermentation by Introduction of Xylose Assimilation Genes and Redirection of the Phosphoketolase Pathway to the Pentose Phosphate Pathway in l-Lactate Dehydrogenase Gene-Deficient Lactobacillus plantarum?  

PubMed Central

The production of optically pure d-lactic acid via xylose fermentation was achieved by using a Lactobacillus plantarum NCIMB 8826 strain whose l-lactate dehydrogenase gene was deficient and whose phosphoketolase genes were replaced with a heterologous transketolase gene. After 60 h of fermentation, 41.2 g/liter of d-lactic acid was produced from 50 g/liter of xylose.

Okano, Kenji; Yoshida, Shogo; Yamada, Ryosuke; Tanaka, Tsutomu; Ogino, Chiaki; Fukuda, Hideki; Kondo, Akihiko

2009-01-01

372

Improved production of homo-D-lactic acid via xylose fermentation by introduction of xylose assimilation genes and redirection of the phosphoketolase pathway to the pentose phosphate pathway in L-Lactate dehydrogenase gene-deficient Lactobacillus plantarum.  

PubMed

The production of optically pure d-lactic acid via xylose fermentation was achieved by using a Lactobacillus plantarum NCIMB 8826 strain whose l-lactate dehydrogenase gene was deficient and whose phosphoketolase genes were replaced with a heterologous transketolase gene. After 60 h of fermentation, 41.2 g/liter of d-lactic acid was produced from 50 g/liter of xylose. PMID:19820147

Okano, Kenji; Yoshida, Shogo; Yamada, Ryosuke; Tanaka, Tsutomu; Ogino, Chiaki; Fukuda, Hideki; Kondo, Akihiko

2009-10-09

373

Site-specific incorporation of 5-fluorotryptophan as a probe of the structure and function of the membrane-bound D-lactate dehydrogenase of Escherichia coli: A sup 19 F nuclear magnetic resonance study  

Microsoft Academic Search

The structure and function of the membrane-bound D-lactate dehydrogenase of Escherichia coli have been investigated by fluorine-19 nuclear magnetic resonance spectroscopy of 5-fluorotryptophan-labeled enzyme in conjunction with oligonucleotide-directed, site-specific mutagenesis. 5-Fluorotryptophan has been substituted for nine phenylalanine, tyrosine, and leucine residues in the enzyme molecule without loss of activity. The ¹⁹F signals from these additional tryptophan residues have been used

Olve B. Peersen; E. A. Pratt; H. T. N. Truong; Chien Ho; Gordon S. Rule

1990-01-01

374

Ultracentrifugation studies of the location of the site involved in the interaction of pig heart lactate dehydrogenase with acidic phospholipids at low pH. A comparison with the muscle form of the enzyme  

Microsoft Academic Search

Lactate dehydrogenase (LDH) from the pig heart interacts with liposomes made of acidic phospholipids most effectively at low\\u000a pH, close to the isoelectric point of the protein (pH = 5.5). This binding is not observed at neutral pH or high ionic strength.\\u000a LDH-liposome complex formation requires an absence of nicotinamide adenine dinucleotides and adenine nucleotides in the interaction\\u000a environment. Their

Grzegorz Terlecki; El?bieta Czapi?ska; Katarzyna Hotowy

2007-01-01

375

An analysis of reaction pathways for proton tunnelling in methylamine dehydrogenase  

PubMed Central

Computational methods have now become a valuable tool to understand the way in which enzymes catalyse chemical reactions and to aid the interpretation of a diverse set of experimental data. This study focuses on the influence of the condensed-phase environment structure on proton transfer mechanisms, with an aim to understand how C–H bond cleavage is mediated in enzymatic reactions. We shall use a combination of molecular simulation, ab initio or semi-empirical quantum chemistry and semi-classical multidimensional tunnelling methods to consider the primary kinetic isotope effects of the enzyme methylamine dehydrogenase (MADH), with reference to an analogous application to triosephosphate isomerase. Analysis of potentially reactive conformations of the system, and correlation with experimental isotope effects, have highlighted that a quantum tunnelling mechanism in MADH may be modulated by specific amino acid residues, such as Asp428, Thr474 and Asp384.

Nunez, Sara; Tresadern, Gary; Hillier, Ian H; Burton, Neil A

2006-01-01

376

Activity of lactate dehydrogenase but not its concentration of messenger RNA increases with body size in barred sand bass, Paralabrax nebulifer (Teleostei).  

PubMed

In white skeletal muscle of conspecific pelagic fishes, the activity of enzymes associated with anaerobic glycolysis, e.g., lactate dehydrogenase (LDH), usually scale positively with increasing body size; this pattern is opposite to that found for enzymes of aerobic metabolism, which decrease in mass-specific activity with size (1-3). The higher mass-specific capacities for anaerobic ATP generation in larger conspecifics are thought to facilitate conservation of high-speed ("burst") swimming ability in fishes of different sizes (1). To investigate the mechanisms responsible for scaling of LDH activity, total RNA, and the specific mRNA for LDH-A (the skeletal muscle isoform of the enzyme) in white muscle of paralabrax nebulifer, the barred sand bass. We also measured total protein concentration and the concentration of actin, the major protein of thin filaments, and its specific mRNA. Although LDH activity scaled significantly with body size as predicted (1-4), no other biochemical trait measured showed a significant size-dependent concentration. We conclude that the regulation of LDH activity in white muscle of this species is not governed by LDH-A mRNA concentrations, but rather by one or more other mechanisms, for example rate of translation of LDH message or a reduced rate of degradation of LDH-A in larger fish. PMID:8916541

Yang, T H; Somero, G N

1996-10-01

377

[Dynamics of changes in the activity of plasma lactate dehydrogenase, leucylarylamidase and GOT and GPT aminotransferases in poultry following damage to the organism caused by cadmium and tetrachloromethane].  

PubMed

In the experiments performed with broilers, after a per os application of tetrachloromethane and cadmium (Cd+2), the activity of lactate dehydrogenase (LDH), leucylarylamidase (LA), and the GOT and GPT amino transferase was determined in the blood plasm. By means of an electrophoretic division of the blood plasm of clinically healthy broilers one enzymatically active fraction of LA and LDH was ascertained. The injury caused to the organism of poultry by carbon tetrachloride and cadmium did not affect the heterogeneity of these two enzymes. After an application of tetrachloromethane (CCl4) in the dose of 8 and 10 ml kg-1 the total activity of LDH and LA changed. The influence of cadmium on the organism in the total dose of 200 mg kg-1 resulted in the changed activity of LDH, GOT, and LA. From the results obtained it has been assumed that the examined activity of LDH, GOT, and LA can be utilized for the diagnosing of non-specific injuries of the organism of poultry. PMID:180643

Kacmár, P; Samo, A

1976-01-01

378

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

379

Dynamics of a Lactate Dehydrogenase Polymorphism in the Wood Louse PORCELLIO SCABER Latr.: Evidence for Partial Assortative Mating and Heterosis in Natural Populations  

PubMed Central

Electrophoretic separation of lactate dehydrogenase (LDH) of Porcellio scaber from 14 natural populations in California, and one each in Oregon, Delaware and Massachusetts, indicates a biallelic polymorphism. Phenotypes are recovered from laboratory matings of virgin females in frequencies agreeing with simple Mendelian inheritance, and the frequency distributions of phenotypes in natural populations are typically in agreement with the appropriate Hardy-Weinberg distributions for these same populations. The same allele predominates in all natural populations examined. Temporal stability within populations suggests that the polymorphism is at, or near, equilibrium. The spatial distribution of allele frequencies, however, is apparently mosaic. Abrupt discontinuities in gene frequency over short distances (50 m to 1 km) suggest that interpopulation migration is insufficient to swamp local differences in gene frequency. Analysis of the transmission dynamics of the polymorphism in natural populations using mother-offspring genotype comparisons suggests that the allelic frequencies of transmitted male gametes are not independent of female genotype. Specifically, the observed mating scheme in natural populations appears to be partially assortative. Comparisons of progeny genotype distributions with yearling (or adult) genotype distributions from the same populations indicate a superior post-partum viability of heterozygous individuals relative to homozygotes. The distortion of progeny genotypic distributions created by assortment is thus apparently counteracted by subsequent heterosis.

Sassaman, Clay

1978-01-01

380

Rationally re-designed mutation of NAD-independent l-lactate dehydrogenase: high optical resolution of racemic mandelic acid by the engineered Escherichia coli  

PubMed Central

Background NAD-independent l-lactate dehydrogenase (l-iLDH) from Pseudomonas stutzeri SDM can potentially be used for the kinetic resolution of small aliphatic 2-hydroxycarboxylic acids. However, this enzyme showed rather low activity towards aromatic 2-hydroxycarboxylic acids. Results Val-108 of l-iLDH was changed to Ala by rationally site-directed mutagenesis. The l-iLDH mutant exhibited much higher activity than wide-type l-iLDH towards l-mandelate, an aromatic 2-hydroxycarboxylic acid. Using the engineered Escherichia coli expressing the mutant l-iLDH as a biocatalyst, 40 g·L-1 of dl-mandelic acid was converted to 20.1 g·L-1 of d-mandelic acid (enantiomeric purity higher than 99.5%) and 19.3 g·L-1 of benzoylformic acid. Conclusions A new biocatalyst with high catalytic efficiency toward an unnatural substrate was constructed by rationally re-design mutagenesis. Two building block intermediates (optically pure d-mandelic acid and benzoylformic acid) were efficiently produced by the one-pot biotransformation system.

2012-01-01

381

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

PubMed Central

The first structure of an NAD-dependent tartrate dehydrogenase (TDH) has been solved to 2?Å resolution by single anomalous diffraction (SAD) phasing as a complex with the intermediate analog oxalate, Mg2+ 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.

2010-01-01

382

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

383

Some effects of polyvinyl alcohol and polyvinyl pyrrolidone on the activity of lactate dehydrogenase and its isoenzymes  

Microsoft Academic Search

The activity of Sigma type II LDH (containing all LDH isoenzymes), type VII (LDH 1) and type V (LDH 5) has been evaluated in 0.1 M phosphate buffer and 0.4 M tris-HCl buffer, both at pH 7.4, before and after addition of PVP and PVA. A slight reduction of reaction velocity occurred when PVP and PVA were added, probably because

Hans A. Dahl

1971-01-01

384

Initial reaction kinetics of succinate dehydrogenase in mouse liver studied with a real-time image analyser system  

Microsoft Academic Search

The initial reaction kinetics of succinate dehydrogenase in situ were investigated in sections of mouse unfixed liver using an ARGUS-100 image analyser system. The sections were incubated on substrate-containing agarose gel films. Images of a section, illuminated with monochromatic light (584 nm), were captured with the image analyser in real time at intervals of 10 s during the incubation. The

Y. Nakae; P. J. Stoward

1992-01-01

385

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.

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

2009-01-01

386

Elevated serum lactate dehydrogenase isoenzymes and aspartate transaminase distinguish Albers-Schönberg disease (Chloride Channel 7 Deficiency Osteopetrosis) among the sclerosing bone disorders.  

PubMed

Osteopetrosis (OPT) refers to the consequences of generalized failure of skeletal resorption during growth. Most cases are explained by loss-of-function mutation within the genes that encode either chloride channel 7 (CLCN7) or a vacuolar proton pump subunit (TCIRG1), each compromising acid secretion by osteoclasts. Patients suffer fractures and sometimes cranial nerve entrapment and insufficient medullary space for hematopoiesis. In 1996, we reported that a high serum level of the brain isoenzyme of creatine kinase (BB-CK), the CK of osteoclasts, characterizes OPT dueamong the sclerosing bone disorders (J Clin Endocrinol Metab. 1996;11:1438). Now, we show that elevation in serum of multiple lactate dehydrogenase (LDH) isoenzymes with aspartate transaminase (AST) distinguishes autosomal dominant OPT due to loss-of-function mutation in CLCN7 [Albers-Schönberg disease (A-SD)] among these conditions. Serum total LDH and AST levels as high as 3× and 2×, respectively, the upper limits of normal for age-appropriate controls, were persistent and essentially concordant in A-SD. Serum LDH was elevated in 7 of 9 children and in the 2 adults studied with A-SD. LDH isoenzyme quantitation showed excesses of LDH-2, -3, and -4. Neither total LDH nor AST increases were found in other forms of OPT, including bisphosphonate-induced OPT, or in 41 children and 6 adults representing 20 additional sclerosing bone disorders. Serum TRACP-5b and BB-CK also were markedly elevated in A-SD. Hence, high serum levels of several enzymes characterize A-SD. Elevated serum LDH isoenzymes and AST indicate a disturbance (of uncertain clinical significance) within multiple extraosseous tissues when there is CLCN7 deficiency. PMID:20499337

Whyte, Michael P; Kempa, Lydia G; McAlister, William H; Zhang, Fan; Mumm, Steven; Wenkert, Deborah

2010-11-01

387

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

PubMed

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 degrees C over the next century. PMID:19787021

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

2009-09-01

388

The role of lactate dehydrogenase, alkaline phosphatase and aspartate aminotransferase in the diagnosis of subclinical intramammary infections in dairy sheep and goats.  

PubMed

The objective was to investigate the changes occurring in the activities of the enzymes lactate dehydrogenase (LDH), alkaline phosphatase (ALP) and aspartate aminotransferase (AST) in sheep and goat milk as a result of subclinical intramammary infections (IMI) and to evaluate the use of these enzymes for the diagnosis of subclinical IMI in dairy sheep and goats. A total of 206 samples of sheep milk and 162 samples of goat milk, obtained from equal udder halves, were used in the study. For each species they were divided into two groups: a no-infection group and a subclinical infection group. Activities of LDH, ALP and AST were significantly higher in the subclinical infection group than in the no-infection group (P<0.05) in both sheep (LDH: 350.42+/-11.25 v. 120.91+/-4.41; ALP: 2773.43+/-105.18 v. 2189+/-94.24; AST: 29.57+/-0.74 v. 17.32+/-0.46) and goats (LDH: 354.07+/-13.33 v. 103.79+/-3.75; ALP: 311.13+/-25.74 v. 137.24+/-19.62; AST: 27.59+/-6.42 v. 15.87+/-0.45). The activity of LDH was identified as indicator for subclinical IMI in both sheep and goats. The optimum cut-off values for LDH activity, offering the highest diagnostic sensitivity (DSn) and diagnostic specificity (DSp), determined by receiver operating characteristic (ROC) analysis, were at 197 U/l, 185 U/l and 197 U/l for sheep, goats and both species, respectively. DSn for sheep, goats and both species at these cut-off values was 92.8%, 98.2% and 94.0%, whereas DSp was 95.4%, 96.3% and 96.3%, respectively. It was concluded that the determination of LDH activity in milk serum is a sensitive and reliable method for the detection of subclinical IMI in dairy sheep and goats. PMID:19919725

Katsoulos, Panagiotis D; Christodoulopoulos, Georgios; Minas, Anastasios; Karatzia, Maria A; Pourliotis, Konstantinos; Kritas, Spyridon K

2009-11-18

389

ERYTHROCYTE ALDEHYDE DEHYDROGENASE ACTIVITY: LACK OF ASSOCIATION WITH ALCOHOL USE AND DEPENDENCE OR ALCOHOL REACTIONS IN AUSTRALIAN TWINS  

Microsoft Academic Search

Aim: Aldehyde dehydrogenase 1 (ALDH1) has been advocated as a marker of alcohol intake. The absence or low levels of ALDH1 may be associated with alcohol-induced flushing or other reactions to alcohol in Europeans and therefore, with reduced alcohol use. This study tested whether variation in erythrocyte ALDH1 activity was associated with alcohol use, alcohol dependence or reactions to alcohol

NARELLE K. HANSELL; DONA PANG; ANDREW C. HEATH; NICHOLAS G. MARTIN; JOHN B. WHITFIELD

2005-01-01

390

Catching catalysis in the act: using single crystal kinetics to trap methylamine dehydrogenase reaction intermediates.  

PubMed

Methylamine dehydrogenase (MADH) is produced by a range of gram-negative methylotrophic and autotrophic bacteria, and allows the organisms to utilise methylamine as the sole source of carbon. The enzyme catalyses the oxidation of methylamine to formaldehyde and ammonia, leaving it in a two-electron reduced state. To complete the catalytic cycle, MADH is reoxidised via an electron transfer (ET) chain. The redox center in the enzyme is the organic cofactor tryptophan tryptophylquinone (TTQ) derived from the posttranslational modification of two Trp residues in the protein. This cofactor has spectral features in the visible region, which change during catalytic turnover, defining spectrally distinct reaction intermediates that reflect the electronic state of the TTQ. In the case of the Paracoccus denitrificans enzyme the physiologic ET chain involves the protein redox partner amicyanin (a blue copper protein). A stable binary (MADH/amicyanin) complex can be formed, and its crystal structure has been solved to 2.5 A resolution by Chen et al. [Biochemistry 21 (1992) 4959]. These crystals were shown to be competent for catalysis and ET by Merli et al. [J. Biol. Chem. 271 (1996) 9177] using single crystal polarised absorption spectroscopy. Through a novel combination of single crystal visible microspectrophotometry, X-ray crystallography and freeze-trapping, we have trapped reaction intermediates of the enzyme in complex with its physiological redox partner amicyanin in the crystalline state. We will present data confirming that catalysis and ET in the binary complex crystals can be tracked by single crystal visible microspectrophotometry. We will also show that the reaction pathway is unperturbed by the presence of cryoprotectant solution, enabling direct freeze-trapping of reaction intermediates within the crystal. We will present new data demonstrating that the binary complex crystals are also capable of exhibiting UV light-dependent oxidase activity, as observed in solution [Biochim. Biophys. Acta 1364 (1998) 297]. PMID:12686162

Pearson, Arwen R; Wilmot, Carrie M

2003-04-11

391

Regression of Dalton’s lymphoma in vivo via decline in lactate dehydrogenase and induction of apoptosis by a ruthenium(II)-complex containing 4-carboxy N -ethylbenzamide as ligand  

Microsoft Academic Search

Summary  A novel ruthenium(II)-complex containing 4-carboxy N-ethylbenzamide (Ru(II)-CNEB) was found to interact with and inhibit M4-lactate dehydrogenase (M4-LDH), a tumor growth supportive\\u000a enzyme, at the tissue level. The present article describes modulation of M4-LDH by this compound in a T-cell lymphoma (Dalton’s\\u000a Lymphoma: DL) vis a vis regression of the tumor in vivo. The compound showed a dose dependent cytotoxicity to

Raj K. Koiri; Surendra K. Trigun; Lallan Mishra; Kiran Pandey; Deobrat Dixit; Santosh K. Dubey

2009-01-01

392

Base compositions of genes encoding alpha-actin and lactate dehydrogenase-A from differently adapted vertebrates show no temperature-adaptive variation in G + C content.  

PubMed

There is a long-standing debate in molecular evolution concerning the putative importance of GC content in adapting the thermal stabilities of DNA and RNA. Most studies of this relationship have examined broad-scale compositional patterns, for example, total GC percentages in genomes and occurrence of GC-rich isochores. Few studies have systematically examined the GC contents of individual orthologous genes from differently thermally adapted species. When this has been done, the emphasis has been on comparing large numbers of genes in only a few species. We have approached the GC-adaptation temperature hypothesis in a different manner by examining patterns of base composition of genes encoding lactate dehydrogenase-A (ldh-a) and alpha-actin (alpha-actin) from 51 species of vertebrates whose adaptation temperatures ranged from -1.86 degrees C (Antarctic fishes) to approximately 45 degrees C (desert reptile). No significant positive correlation was found between any index of GC content (GC content of the entire sequence, GC content of the third codon position [GC(3)], and GC content at fourfold degenerate sites [GC(4)]) and any index of adaptation temperature (maximal, mean, or minimal body temperature). For alpha-actin, slopes of regression lines for all comparisons did not differ significantly from zero. For ldh-a, negative correlations between adaptation temperature and total GC content, GC(3), and GC(4) were observed but were shown to be due entirely to phylogenetic influences (as revealed by independent contrast analyses). This comparison of GC content across a wide range of ectothermic ("cold-blooded") and endothermic ("warm-blooded") vertebrates revealed that frogs of the genus Xenopus, which have commonly been used as a representative cold-blooded species, in fact are outliers among ectotherms for the alpha-actin analyses, raising concern about the appropriateness of choosing these amphibians as representative of ectothermic vertebrates in general. Our study indicates that, whereas GC contents of isochores may show variation among different classes of vertebrates, there is no consistent relationship between adaptation temperature and the percentage of thermal stability-enhancing G + C base pairs in protein-coding genes. PMID:12519912

Ream, Rachael A; Johns, Glenn C; Somero, George N

2003-01-01

393

Decreases in activation energy and substrate affinity in cold-adapted A4-lactate dehydrogenase: evidence from the Antarctic notothenioid fish Chaenocephalus aceratus.  

PubMed

Enzyme function is strongly affected by temperature, and orthologs from species adapted to different thermal environments often show temperature compensation in kinetic properties. Antarctic notothenioid fishes live in a habitat of constant, extreme cold (-1.86 +/- 2 degrees C), and orthologs of the enzyme A4-lactate dehydrogenase (A4-LDH) in these species have adapted to this environment through higher catalytic rates, lower Arrhenius activation energies (Ea), and increases in the apparent Michaelis constant for the substrate pyruvate (Km(PYR)). Here, site-directed mutagenesis was used to determine which amino acid substitutions found in A4-LDH of the notothenioid Chaenocephalus aceratus, with respect to orthologs from warm-adapted teleosts, are responsible for these adaptive changes in enzyme function. Km(PYR) was measured in eight single and two double mutants, and Ea was tested in five single and two double mutants in the temperature range 0 degrees C-20 degrees C. Of the four mutants that had an effect on these parameters, two increased Ea but did not affect Km(PYR) (Gly224Ser, Ala310Pro), and two increased both Ea and Km(PYR) (Glu233Met, Gln317Val). The double mutants Glu233Met/Ala310Pro and Glu233Met/Gln317Val increased Km(PYR) and Ea to levels not significantly different from the A4-LDH of a warm temperate fish (Gillichthys mirabilis, habitat temperature 10 degrees C-35 degrees C). The four single mutants are associated with two alpha-helices that move during the catalytic cycle; those that affect Ea but not Km(PYR) are further from the active site than those that affect both parameters. These results provide evidence that (1) cold adaptation in A4-LDH involves changes in mobility of catalytically important molecular structures; (2) these changes may alter activation energy alone or activation energy and substrate affinity together; and (3) the extent to which these parameters are affected may depend on the location of the substitutions within the mobile alpha-helices, perhaps due to differences in proximity to the active site. PMID:15317880

Fields, Peter A; Houseman, Daniel E

2004-08-18

394

A lactate dehydrogenase ELISA-based assay for the in vitro determination of Plasmodium berghei sensitivity to anti-malarial drugs  

PubMed Central

Background Plasmodium berghei rodent malaria is a well-known model for the investigation of anti-malarial drug efficacy in vivo. However, the availability of drug in vitro assays in P. berghei is reduced when compared with the spectrum of techniques existing for Plasmodium falciparum. New alternatives to the current manual or automated methods described for P. berghei are attractive. The present study reports a new ELISA drug in vitro assay for P. berghei using two monoclonal antibodies against the parasite lactate dehydrogenase (pLDH). Methods This procedure includes a short-in vitro culture, the purification of schizonts and the further generation of synchronized mice infections. Early stages of the parasite are then incubated against different concentrations of anti-malarial drugs using micro-plates. The novelty of this procedure in P. berghei relies on the quantification of the drug activity derived from the amount of pLDH estimated by an ELISA assay using two monoclonal antibodies: 14C1 and 19G7. The IC50s obtained through the ELISA assay were compared with those from the micro-test. Results The initial parameters of the synchronized samples used in the in vitro assays were a parasitaemia of 0.5% and haematocrit of 1%, with an incubation period of 22 hours at 36.5°C. pLDH detection using a 14C1 coating at 10 ?g/ml and 19G7 at 2.5 × 10-3 ?g/ml provided good readouts of optical densities with low background in negative controls and specific detection levels for all parasite stages. IC50s values derived from the ELISA assay for artesunate, chloroquine, amodiaquine and quinine were: 15, 7, 2, and 144 nM, respectively. When artesunate and chloroquine IC50s were evaluated using the micro-test similar values were obtained. Conclusion This ELISA-based in vitro drug assay is easy to implement, fast, and avoids the use radioisotopes or expensive equipment. The utility of this simple assay for screening anti-malarial drug activity against P. berghei in vitro is demonstrated.

2012-01-01

395

Stringency of SubstrateSpecificity of Escherichia coli Malate-Dehydrogenase  

Microsoft Academic Search

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

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

1995-01-01

396

Comparison of benzyl alcohol dehydrogenases and benzaldehyde dehydrogenases from the benzyl alcohol and mandelate pathways in Acinetobacter calcoaceticus and from the TOL-plasmid-encoded toluene pathway in Pseudomonas putida. N-terminal amino acid sequences, amino acid compositions and immunological cross-reactions.  

PubMed Central

1. N-Terminal sequences were determined for benzyl alcohol dehydrogenase, benzaldehyde dehydrogenase I and benzaldehyde dehydrogenase II from Acinetobacter calcoaceticus N.C.I.B. 8250, benzyl alcohol dehydrogenase and benzaldehyde dehydrogenase encoded by the TOL plasmid pWW53 in Pseudomonas putida MT53 and yeast K(+)-activated aldehyde dehydrogenase. Comprehensive details of the sequence determinations have been deposited as Supplementary Publication SUP 50161 (5 pages) at the British Library Document Supply Centre, Boston Spa. Wetherby. West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1991) 273. 5. The extent of sequence similarity suggests that the benzyl alcohol dehydrogenases are related to each other and also to established members of the family of long-chain Zn2(+)-dependent alcohol dehydrogenases. Benzaldehyde dehydrogenase II from Acinetobacter appears to be related to the Pseudomonas TOL-plasmid-encoded benzaldehyde dehydrogenase. The yeast K(+)-activated aldehyde dehydrogenase has similarity of sequence with the mammalian liver cytoplasmic class of aldehyde dehydrogenases but not with any of the Acinetobacter or Pseudomonas enzymes. 2. Antisera were raised in rabbits against the three Acinetobacter enzymes and both of the Pseudomonas enzymes, and the extents of the cross-reactions were determined by immunoprecipitation assays with native antigens and by immunoblotting with SDS-denatured antigens. Cross-reactions were detected between the alcohol dehydrogenases and also among the aldehyde dehydrogenases. This confirms the interpretation of the N-terminal sequence comparisons and also indicates that benzaldehyde dehydrogenase I from Acinetobacter may be related to the other two benzaldehyde dehydrogenases. 3. The amino acid compositions of the Acinetobacter and the Pseudomonas enzymes were determined and the numbers of amino acid residues per subunit were calculated to be: benzyl alcohol dehydrogenase and TOL-plasmid-encoded benzyl alcohol dehydrogenase, 381; benzaldehyde dehydrogenase I and benzaldehyde dehydrogenase II, 525; TOL-plasmid-encoded benzaldehyde dehydrogenase, 538.

Chalmers, R M; Keen, J N; Fewson, C A

1991-01-01

397

A Kinetic Study of the Lipoamide Dehydrogenase-Nadh-Dye Reaction.  

National Technical Information Service (NTIS)

Because of the inhibition of the enzyme lipoamide dehydrogenase, diaphorase, by sulfhydryl binding compounds, interest developed in the mechanism by which this enzyme acts on various substrates. The application of fluorometric and spectrophotometric techn...

G. G. Guilbault P. Goldberg D. N. Kramer

1965-01-01

398

Site-specific incorporation of 5-fluorotryptophan as a probe of the structure and function of the membrane-bound D-lactate dehydrogenase of Escherichia coli: A sup 19 F nuclear magnetic resonance study  

SciTech Connect

The structure and function of the membrane-bound D-lactate dehydrogenase of Escherichia coli have been investigated by fluorine-19 nuclear magnetic resonance spectroscopy of 5-fluorotryptophan-labeled enzyme in conjunction with oligonucleotide-directed, site-specific mutagenesis. 5-Fluorotryptophan has been substituted for nine phenylalanine, tyrosine, and leucine residues in the enzyme molecule without loss of activity. The {sup 19}F signals from these additional tryptophan residues have been used as markers for sensitivity to substrate, exposure to aqueous solvent, and proximity to a lipid-bound spin-label. The nuclear magnetic resonance data show that two mutational sites, at amino acid residues 340 and 361, are near the lipid environment used to stabilize the enzyme. There are a number of amino acid residues on the carboxyl side of this region that are strongly sensitive to the aqueous solvent. The environment of the wide-type tryptophan residue at position 469 changes as a result of two of the substitution mutations, suggesting some amino acid residue-residue interactions. Secondary structure prediction methods indicate a possible binding site for the flavin adenine dinucleotide cofactor in the carboxyl end of the enzyme molecule. These results suggest that the membrane-bound D-lactate dehydrogenase may have the two-domain structure of many cytoplasmic dehydrogenases but with the addition of a membrane-binding domain between the catalytic and cofactor-binding domains. This type of three-domain structure may be of general significance for understanding the structure of membrane-bound proteins which do not traverse the lipid bilayer of membranes.

Peersen, O.B.; Pratt, E.A.; Truong, H.T. N.; Ho, C. (Carnegie Mellon Univ., Pittsburgh, PA (USA)); Rule, G.S. (Univ. of Virginia, Charlottesville (USA))

1990-04-03

399

Regulation of lactate production and glucose transport as well as of glucose transporter 1 and lactate dehydrogenase A mRNA levels by basic fibroblast growth factor in rat Sertoli cells  

Microsoft Academic Search

By using cultured rat Sertoli cells as a model, both the action of basic fibroblast growth factor (bFGF) on lactate production and the site of this action were studied. bFGF stimulated Sertoli cell lactate production in a dose- dependent manner (basal: 7·30·5; 0·1 ng\\/ml bFGF: 7·50·5; 1 ng\\/ml bFGF: 7·50·6; 10 ng\\/ml bFGF: 10·31·0; 30 ng\\/ml bFGF: 15·21·5; 50 ng\\/ml

M F Riera; S B Meroni; H F Schteingart; E H Pellizzari; S B Cigorraga

2002-01-01

400

Inhibition of Cellular Immune Reactions in Mice infected with Lactic Dehydrogenase Virus  

Microsoft Academic Search

CERTAIN viral infections affect the immune response of the host to heterogenous antigens. Murine leukaemia viruses, such as Moloney, Friend, Rauscher and Gross passage A virus, inhibit antibody formation1-4. In addition, the passage A virus depresses cellular immunity5. On the other hand, lactic dehydrogenase virus (LDV) enhances humoral antibody formation in mice and prevents the development of immunological tolerance to

R. J. Howard; A. L. Notkins; S. E. Mergenhagen

1969-01-01

401

Azotobacter vinelandii glucose 6-phosphate dehydrogenase properties of NAD and NADP-linked reactions  

Microsoft Academic Search

Glucose 6-phosphate oxidation, catalyzed by purified Azotobacter vinelandii glucose 6-phosphate dehydrogenase, was studied with respect to the selective utilization of NAD, NADP, thionicotinamide adenine dinucleotide or thionicotinamide adenine dinucleotide phosphate as coenzyme. A sigmoidal relationship was observed for the effect of substrate concentration on initial velocities when either NAD, NADP or thionicotinamide adenine dinucleotide was used as coenzyme, with N

Bruce M Anderson; Darla J Wise; Constance D Anderson

1997-01-01

402

Deuterium isotope effects on initial rates of the liver alcohol dehydrogenase reaction  

Microsoft Academic Search

Initial steady-state velocities using highly purified deuterated ; substrates and coenzymes as well as an isoenzymatically homogeneous enzyme ; preparation were measured for liver alcohol dehydrogenase at 25 deg in 20 mm ; phosphate (pH 7.0) or 20 mm pyrophosphate (pH 8.8). Significant isotope effects ; of v\\/sub 0,H\\/\\/v\\/sub 0,D\\/ were observed. In the reduction of acetaldehyde by A-; NADD

Karen Bush; Vernon J. Shiner; H. R. Mahler

1973-01-01

403

Computational study of the mechanism of half-reactions in class 1A dihydroorotate dehydrogenase from Trypanosoma cruzi.  

PubMed

Chagas' disease is considered to be a health problem affecting millions of people in Latin America. This disease is caused by the parasite Trypanosoma cruzi. Recently dihydroorotate dehydrogenase class 1A from Trypanosoma cruzi (TcDHODA) was shown to be essential for the survival and growth of T. cruzi and proposed as a drug target against Chagas' disease. This enzyme catalyzes the oxidation of (S)-dihydroorotate to orotate, with a proposed catalytic cycle consisting of two half-reactions. In the first half-reaction dihydroorotate is oxidized to orotate, with the consequent reduction of the flavin mononucleotide cofactor. In the second half-reaction fumarate is reduced to succinate. The first oxidation half-reaction may occur via a concerted or a stepwise mechanism. Herein, the catalytic mechanism of TcDHODA has been studied using hybrid Quantum Mechanical/Molecular Mechanical (QM/MM) Molecular Dynamics (MD) simulations. The free energy profiles derived from the bidimensional potential of mean force reveal more details for two half-reaction processes. PMID:24084894

de Farias Silva, Natália; Lameira, Jerônimo; Alves, Cláudio Nahum; Martí, Sergio

2013-10-16

404

Primary deuterium and tritium isotope effects upon V/K in the liver alcohol dehydrogenase reaction with ethanol.  

PubMed

The primary isotope effect upon V/K when ethanol stereospecifically labeled with deuterium or tritium is oxidized by liver alcohol dehydrogenase has been measured between pH 6 and 9. The deuterium isotope effect was obtained with high reproducibility by the use of two different radioactive tracers, viz. 14C and 3H, to follow the rate of acetaldehyde formation from deuterium-labeled ethanol and normal ethanol, respectively. Synthesis of the necessary labeled compounds is described in this and earlier work referred to. V/K isotope effects for both tritium and deuterium have been measured with three different coenzymes, NAD+, thio-NAD+, and acetyl-NAD+. With NAD+ at pH 7, D(V/K) was 3.0 and T(V/K) was 6.5. With increasing pH, these values decreased to 1.5 and 2.5 at pH 9. The intrinsic isotope effect evaluated by the method of Northrop [Northrop, D.B. (1977) in Isotope Effects on Enzyme-Catalyzed Reactions (Cleland, W. W., O'Leary, M, H., & Northrop, D. B., Eds.) pp 112-152, University Park Press, Baltimore] varies little with pH. It amounts to about 10 with NAD+ and about 5 with the coenzyme analogues. Commitment functions and their dependence upon pH calculated in this connection appear to be in agreement with known kinetic parameters of liver alcohol dehydrogenase. This assay method was also applied in vivo in the rat. Being a noninvasive method because only trace amounts of isotopes are needed, it may yield information about alternative routes of ethanol oxidation in vivo. In naive rats at low concentrations of ethanol, it confirms the discrete role of the non alcohol dehydrogenase systems. PMID:7028109

Damgaard, S E

1981-09-29

405

Genetic polymorphism of blood proteins in the troops of Japanese macaques, Macaca fuscata : II. Erythrocyte lactate dehydrogenase polymorphism in Macaca fuscata  

Microsoft Academic Search

In investigating the genetic marker for population genetics of Japanese macaques by electrophoresis, the author found the erythrocyte lacate dehydrogenase (LDH) polymorphism existing in some troops. There were four kinds of variations which seemed to be controlled by two loci, controlling A and B subunits of this enzyme. The variant phenotypes were named LDH-Amac2-1 LDH-Bmac1-1, LDH-Amac3-1 LDH-Bmac1-1, LDH-Amac 1-1 LDH-Bmac2-1,

Takayoshi Shotake

1974-01-01

406

Molecular Characterization of the Soybean Alcohol Dehydrogenase Gene Family Amplified in Vitro by the Polymerase Chain Reaction 1  

PubMed Central

Soybean (Glycine max) alcohol dehydrogenase (ADH) cDNAs were amplified in vitro from total RNA by the polymerase chain reaction (PCR). The amplification strategy involved first strand cDNA synthesis from anaerobic cotyledon total RNA using an 18-thymidine primer. The second strand cDNA primer was a conserved sequence near the 5? end of known plant ADH transcripts. The PCR products were ligated into a plasmid vector and unique clones were isolated on the basis of size and restriction pattern. Sequence analysis revealed three distinct classes of soybean ADH cDNAs, all of which showed high homology to Adh genes from maize and peas. RNA blot hybridization analyses showed differential expression patterns for these genes. One gene, expressed constitutively in all seedling organs, was inducible by anaerobiosis, one gene was expressed only in anaerobic organs, and the third gene was expressed predominantly in anaerobic roots. Images Figure 2 Figure 3 Figure 6

Newman, Kurt D.; VanToai, Tara T.

1992-01-01

407

The Reaction of the Molybdenum- and Copper-Containing Carbon Monoxide Dehydrogenase from Oligotropha carboxydovorans with Quinones†  

PubMed Central

Carbon monoxide dehydrogenase (CODH) from Oligotropha carboxydovorans catalyzes the oxidation of carbon monoxide to carbon dioxide, providing the organism both a carbon source and energy for growth. In the oxidative half of the catalytic cycle, electrons gained from CO are ultimately passed to the electron transport chain of the Gram-negative organism, but the proximal acceptor of reducing equivalents from the enzyme has not been established. Here we investigate the reaction of reduced enzyme with various quinones, and find them to be catalytically competent. Benzoquinone has a kox of 125.1 s-1 and Kd of 48 ?M; ubiquinone-1 has a kox/Kd value of 2.88 × 105 M-1s-1; 1,4-napthoquinone has a kox of 38 s-1 and Kd of 140 ?M; and 1,2-napthoquinone-4-sulfonic acid a kox/Kd of 1.31 × 105 M-1s-1. An extensive effort to identify a cytochrome that was reducible by CO/CODH was unsuccessful. Steady-state studies with benzoquinone indicate that the rate-limiting step is in the reductive half of the reaction (that is, the reaction of oxidized enzyme with CO). On the basis of the inhibition of CODH by diphenyliodonium chloride we conclude that quinone substrates interact with CODH at the enzyme’s flavin site. Our results strongly suggest that CODH donates reducing equivalents directly to the quinone pool without using a cytochrome as an intermediary.

Wilcoxen, Jarett; Zhang, Bo; Hille, Russ

2011-01-01

408

Kinetics of the Reaction of Baker?s Yeast Glucose6-Phosphate-Dehydrogenase with 5,5?-Dithiobis(2-Nitrobenzoic Acid)  

Microsoft Academic Search

The kinetics of the reaction of baker?s yeast glucose-6-phosphate dehydrogenase with excess 5,5?-dithiobis(2-nitrobenzoic acid) (DTNB) were studied at pH 8.5 and 30°C and at constant ionic strength of 0.01 and in the absence and in the presence of NADP+ or glucose 6-phosphate. The reaction follows pseudo-first-order kinetics irrespective of whether these substrates are absent or present. The observed pseudo-first-order rate

S. A. Adediran; M. R. Gbadegesin

1995-01-01

409

Competence in Haemophilus Influenzae. A Role for Inosine and Lactate in the Primary Cell-Deoxyribonucleic Acid Attachment Reaction.  

National Technical Information Service (NTIS)

Cells of Haemophilus influenzae grown with or without competence promoting factors, inosine and lactate, were compared in their ability to bind deoxyribonucleic acid (DNA) and to undergo a genetic transformation. It was found that cells grown with the abo...

J. M. Ranhand

1969-01-01

410

Efficient Production of Optically Pure d-Lactic Acid from Raw Corn Starch by Using a Genetically Modified l-Lactate Dehydrogenase Gene-Deficient and ?-Amylase-Secreting Lactobacillus plantarum Strain?  

PubMed Central

In order to achieve direct and efficient fermentation of optically pure d-lactic acid from raw corn starch, we constructed l-lactate dehydrogenase gene (ldhL1)-deficient Lactobacillus plantarum and introduced a plasmid encoding Streptococcus bovis 148 ?-amylase (AmyA). The resulting strain produced only d-lactic acid from glucose and successfully expressed amyA. With the aid of secreting AmyA, direct d-lactic acid fermentation from raw corn starch was accomplished. After 48 h of fermentation, 73.2 g/liter of lactic acid was produced with a high yield (0.85 g per g of consumed sugar) and an optical purity of 99.6%. Moreover, a strain replacing the ldhL1 gene with an amyA-secreting expression cassette was constructed. Using this strain, direct d-lactic acid fermentation from raw corn starch was accomplished in the absence of selective pressure by antibiotics. This is the first report of direct d-lactic acid fermentation from raw starch.

Okano, Kenji; Zhang, Qiao; Shinkawa, Satoru; Yoshida, Shogo; Tanaka, Tsutomu; Fukuda, Hideki; Kondo, Akihiko

2009-01-01

411

Efficient production of optically pure D-lactic acid from raw corn starch by using a genetically modified L-lactate dehydrogenase gene-deficient and alpha-amylase-secreting Lactobacillus plantarum strain.  

PubMed

In order to achieve direct and efficient fermentation of optically pure D-lactic acid from raw corn starch, we constructed L-lactate dehydrogenase gene (ldhL1)-deficient Lactobacillus plantarum and introduced a plasmid encoding Streptococcus bovis 148 alpha-amylase (AmyA). The resulting strain produced only D-lactic acid from glucose and successfully expressed amyA. With the aid of secreting AmyA, direct D-lactic acid fermentation from raw corn starch was accomplished. After 48 h of fermentation, 73.2 g/liter of lactic acid was produced with a high yield (0.85 g per g of consumed sugar) and an optical purity of 99.6%. Moreover, a strain replacing the ldhL1 gene with an amyA-secreting expression cassette was constructed. Using this strain, direct D-lactic acid fermentation from raw corn starch was accomplished in the absence of selective pressure by antibiotics. This is the first report of direct D-lactic acid fermentation from raw starch. PMID:19011066

Okano, Kenji; Zhang, Qiao; Shinkawa, Satoru; Yoshida, Shogo; Tanaka, Tsutomu; Fukuda, Hideki; Kondo, Akihiko

2008-11-14

412

A quantitative study of the biospecific desorption of rat liver (M4) lactate dehydrogenase from 10-carboxydecylamino-Sepharose. Determination of the number of ligand-binding sites blocked on adsorption.  

PubMed Central

1. The theory of Nichol, Ogston, Winzor & Sawyer [(1974) Biochem. J. 143, 435-443] for quantitative affinity chromatography, when adapted for use with a non-specific column from which a multi-site protein can be specifically desorbed by its free ligand, permits determination of the concentration of adsorption sites on the column, their adsorptive affinity (as an association constant) and either the intrinsic (site) constant for ligand-binding to the protein or an 'occlusion coefficient' (defined as the number of ligand-binding sites blocked on adsorption), one of which must be known. 2. The theory has been applied to the NADH-specific desorption of rat liver M4 lactate dehydrogenase from 10-carboxydecylamino-Sepharose. It suggests that most of the enzyme molecules are adsorbed with at least two NADH-binding sites blocked, indicating an extensive adsorption interface in relation to the protein surface. Other chromatographic parameters were also determined for the system. 3. Among topics discussed are (a) factors affecting the experimentally determined value for the number of blocked sites, (b) the nature of the adsorption sites on the column and (c) the similarity of the analysis to that for determining Hill coefficients, and other possible applications.

Kyprianou, P; Yon, R J

1982-01-01

413

Contribution of K99 and D319 to Substrate Binding and Catalysis in the Saccharopine Dehydrogenase Reaction  

PubMed Central

Saccharopine dehydrogenase catalyzes the NAD-dependent oxidative deamination of saccharopine to L-lysine and ?-ketoglutarate. Lysine 99 is within hydrogen-bond distance to the ?-carboxylate of the lysine substrate and D319 is in the vicinity of the carboxamide side chain of NADH. Both are conserved and may be important to the overall reaction. Replacing K99 with M gives decreases of 110-, 80- and 20-fold in the V2/Km values for lysine, ?-ketoglutarate and NADH, respectively. Deuterium isotope effects on V and V/KLys increase, while the solvent deuterium isotope effects decrease compared to the C205S mutant enzyme. Data for K99M suggest a decreased affinity for all reactants and a change in the partition ratio of the imine intermediate to favor hydrolysis. A change in the bound conformation of the imine and/or the distance of the imine carbon to C4 of the nicotinamide ring of NADH is also suggested. Changing D319 to A decreases V2/KNADH by 33-fold. Primary deuterium and solvent deuterium isotope effects decrease compared to C205S suggesting a non-isotope sensitive step has become slower. NADH binds to enzyme first, and sets the site for binding of lysine and ?-ketoglutarate. The slower step is likely the conformational change generated upon binding of NADH.

Ekanayake, Devi K.; West, Ann H.; Cook, Paul. F.

2011-01-01

414

On the rate of proton exchange with solvent of the catalytic histidine in flavocytochrome b2 (yeast L-lactate dehydrogenase).  

PubMed Central

The family of FMN-dependent, alpha-hydroxy acid-oxidizing enzymes catalyzes substrate dehydrogenation by a mechanism the first step of which is abstraction of the substrate alpha-proton (so-called carbanion mechanism). For flavocytochrome b2 and lactate oxidase, it was shown that once on the enzyme this proton is lost only slowly to the solvent (Lederer F, 1984, In: Bray RC, Engel PC, Mayhew SG, eds, Flavins & flavoproteins, Berlin: Walter de Gruyter & Co., pp 513-526; Urban P, Lederer F, 1985, J Biol Chem 260:11115-11122). This suggested the occurrence of a pKa increase of the catalytic histidine upon enzyme reduction by substrate. For flavocytochrome b2, the crystal structure indicated 2 possible origins for the stabilization of the imidazolium form of His 373: either a network of hydrogen bonds involving His 373, Tyr 254, flavin N5 and O4, a heme propionate, and solvent molecules, and/or electrostatic interactions with Asp 282 and with the reduced cofactor N1 anion. In this work, we probe the effect of the hydrogen bond network at the active site by studying proton exchange with solvent for 2 mutants: Y254F and the recombinant flavodehydrogenase domain, in which this network should be disrupted. The rate of proton exchange, as determined by intermolecular hydrogen transfer experiments, appears identical in the flavodehydrogenase domain and the wild-type enzyme, whereas it is about 3-fold faster in the Y254F mutant. It thus appears that specific hydrogen bonds to the solvent do not play a major role in stabilizing the acid form of His 373 in reduced flavocytochrome b2. Removal of the Y254 phenol group induces a pKa drop of about half a pH unit for His 373 in the reduced enzyme. Even then, the rate of exchange of the imidazolium proton with solvent is still lower by several orders of magnitude than that of a normally ionizing histidine. Other factors must then also contribute to the pKa increase, such as the electrostatic interactions with D282 and the anionic reduced cofactor, as suggested by the crystal structure.

Balme, A.; Lederer, F.

1994-01-01

415

cis-Chlorobenzene Dihydrodiol Dehydrogenase (TcbB) from Pseudomonas sp. Strain P51, Expressed in Escherichia coli DH5?(pTCB149), Catalyzes Enantioselective Dehydrogenase Reactions  

PubMed Central

cis-Chlorobenzene dihydrodiol dehydrogenase (CDD) from Pseudomonas sp. strain P51, cloned into Escherichia coli DH5?(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) enantiomer 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, Henning; Fleischmann, Thomas; Van Der Meer, Jan Roelof; Kohler, Hans-Peter E.

1999-01-01

416

Regulation of glycolysis in the erythrocyte: role of the lactate/pyruvate and NAD/NADH ratios.  

PubMed

Mature erythrocytes, when removed from the circulation, exhibit severe disturbances of glycolytic flow, with accumulation not only of lactate, the ultimate product of glycolysis, but also of several upstream metabolic intermediates, primarily fructose-1,6-diphosphate, glyceraldehyde-3-phosphate, and dihydroxyacetone phosphate. This accumulation may be prevented (and also reverted) by allowing the diffusible end products lactate and pyruvate to leave the cell by equilibrating with a much larger extracellular compartment. The disturbance of erythrocyte glycolysis does not result from direct inhibition by lactate itself but from the interplay between the lactate dehydrogenase and glyceraldehyde-3-phosphate dehydrogenase (3-PGAD) reactions. The accumulation of intermediates reflects the increased lactate-to-pyruvate ratio; this leads to a secondary imbalance of the nicotinamide adenine dinucleotide-to-reduced nicotinamide adenine dinucleotide (NAD-to-NADH) ratio, which in turn slows down glycolysis at the 3-PGAD step, whose upstream metabolites then pile up. No accumulation, however, takes place if the lactate-to-pyruvate ratio is maintained constant in the extracellular compartment, regardless of concentrations. These studies demonstrate that orderly glycolysis in the erythrocyte is regulated by the NAD-to-NADH ratio and also provide a method that makes possible the in vitro study of erythrocyte glycolysis. PMID:1856577

Tilton, W M; Seaman, C; Carriero, D; Piomelli, S

1991-08-01

417

Lactate, not pyruvate, is neuronal aerobic glycolysis end product: An in vitro electrophysiological study  

Microsoft Academic Search

For over 60 years, a distinction has been made between aerobic and anaerobic glycolysis based on their respective end products: pyruvate of the former, lactate of the latter. Recently we hypothesized that, in the brain, both aerobic and anaerobic glycolysis terminate with the formation of lactate from pyruvate by the enzyme lactate dehydrogenase (LDH). If this hypothesis is correct, lactate

A. Schurr; R. S. Payne

2007-01-01

418

Onchocerca fasciata : Enzyme histochemistry and tissue distribution of various dehydrogenases in the adult female worm  

Microsoft Academic Search

Histochemistry studies of key dehydrogenases in the glycolytic pathway and related enzymes and the tricarboxylic acid (TCA)-cycle\\u000a enzymes were carried out on adult female Onchocerca fasciata. The distribution pattern and enzymatic activities of 6-phosphogluconate dehydrogenase (6-GPDH), lactate dehydrogenase (LDH),\\u000a mitochondrial glycerol-3-phosphate dehydrogenase (GPDH), nicotinamide adenine dinucleotide (phosphate) [NAD+(P)]-linked isocitrate\\u000a dehydrogenase (ICDH), and NAD+(P)-linked malate dehydrogenase (MDH) in various tissues of

M. S. Omar; A. M. S. Raoof; O. M. Al-Amari

1996-01-01

419

Steady-state kinetic mechanism of the NADP+- and NAD+-dependent reactions catalysed by betaine aldehyde dehydrogenase from Pseudomonas aeruginosa.  

PubMed Central

Betaine aldehyde dehydrogenase (BADH) catalyses the irreversible oxidation of betaine aldehyde to glycine betaine with the concomitant reduction of NAD(P)(+) to NADP(H). In Pseudomonas aeruginosa this reaction is a compulsory step in the assimilation of carbon and nitrogen when bacteria are growing in choline or choline precursors. The kinetic mechanisms of the NAD(+)- and NADP(+)-dependent reactions were examined by steady-state kinetic methods and by dinucleotide binding experiments. The double-reciprocal patterns obtained for initial velocity with NAD(P)(+) and for product and dead-end inhibition establish that both mechanisms are steady-state random. However, quantitative analysis of the inhibitions, and comparison with binding data, suggest a preferred route of addition of substrates and release of products in which NAD(P)(+) binds first and NAD(P)H leaves last, particularly in the NADP(+)-dependent reaction. Abortive binding of the dinucleotides, or their analogue ADP, in the betaine aldehyde site was inferred from total substrate inhibition by the dinucleotides, and parabolic inhibition by NADH and ADP. A weak partial uncompetitive substrate inhibition by the aldehyde was observed only in the NADP(+)-dependent reaction. The kinetics of P. aeruginosa BADH is very similar to that of glucose-6-phosphate dehydrogenase, suggesting that both enzymes fulfil a similar amphibolic metabolic role when the bacteria grow in choline and when they grow in glucose.

Velasco-Garcia, R; Gonzalez-Segura, L; Munoz-Clares, R A

2000-01-01

420

Infection of central nervous system cells by ecotropic murine leukemia virus in C58 and AKR mice and in in utero-infected CE/J mice predisposes mice to paralytic infection by lactate dehydrogenase-elevating virus.  

PubMed Central

Certain mouse strains, such as AKR and C58, which possess N-tropic, ecotropic murine leukemia virus (MuLV) proviruses and are homozygous at the Fv-1n locus are specifically susceptible to paralytic infection (age-dependent poliomyelitis [ADPM]) by lactate dehydrogenase-elevating virus (LDV). Our results provide an explanation for this genetic linkage and directly prove that ecotropic MuLV infection of spinal cord cells is responsible for rendering anterior horn neurons susceptible to cytocidal LDV infection, which is the cause of the paralytic disease. Northern (RNA) blot hybridization of total tissue RNA and in situ hybridization of tissue sections demonstrated that only mice harboring central nervous system (CNS) cells that expressed ecotropic MuLV were susceptible to ADPM. Our evidence indicates that the ecotropic MuLV RNA is transcribed in CNS cells from ecotropic MuLV proviruses that have been acquired by infection with exogenous ecotropic MuLV, probably during embryogenesis, the time when germ line proviruses in AKR and C58 mice first become activated. In young mice, MuLV RNA-containing cells were found exclusively in white-matter tracts and therefore were glial cells. An increase in the ADPM susceptibility of the mice with advancing age correlated with the presence of an increased number of ecotropic MuLV RNA-containing cells in the spinal cords which, in turn, correlated with an increase in the number of unmethylated proviruses in the DNA extracted from spinal cords. Studies with AKXD recombinant inbred strains showed that possession of a single replication-competent ecotropic MuLV provirus (emv-11) by Fv-1n/n mice was sufficient to result in ecotropic MuLV infection of CNS cells and ADPM susceptibility. In contrast, no ecotropic MuLV RNA-positive cells were present in the CNSs of mice carrying defective ecotropic MuLV proviruses (emv-3 or emv-13) or in which ecotropic MuLV replication was blocked by the Fv-1n/b or Fv-1b/b phenotype. Such mice were resistant to paralytic LDV infection. In utero infection of CE/J mice, which are devoid of any endogenous ecotropic MuLVs, with the infectious clone of emv-11 (AKR-623) resulted in the infection of CNS cells, and the mice became ADPM susceptible, whereas littermates that had not become infected with ecotropic MuLV remained ADPM resistant.

Anderson, G W; Palmer, G A; Rowland, R R; Even, C; Plagemann, P G

1995-01-01

421

Drosophila lactate dehydrogenase: Molecular and genetic aspects  

Microsoft Academic Search

(LDH) obtained from larvae of Drosophila melanogaster was purified to homogeneity by affinity chromatography on oxamate-Sepharose. The purification procedure is simple to operate and gives a homogeneous preparation in a good yield (34.86%) after only two steps. Utilizing the homogeneous LDH preparation, an attempt was made to characterize the LDH molecule. Thus, it was found that the N-terminal amino acid

A. Onoufriou; S. N. Alahiotis

1982-01-01

422

Differential effects of trilostane and cyanoketone on the 3?-hydroxysteroid dehydrogenase-isomerase reactions in androgen and 16-androstene biosynthetic pathways in the pig testis  

Microsoft Academic Search

3?-Hydroxysteroid dehydrogenase-isomerase (3?-HSD-I) activity in the pig testis is responsible for the conversion of dehydroepiandrosterone (DHA) to 4-androstenedione and also for the conversion of 5,16-androstadien-3?-o1 (andien-?) to 4,16-androstadien-3-one (dienone). Therefore, 3?-HSD-I plays an essential role in the biosynthesis of hormonally and pheromonally active steroids. Previous studies from this laboratory have suggested that the 3?-HSD-I reactions in the androgen and 16-androstene

Gerard M. Cooke

1996-01-01

423

Methionyl-tRNA synthetase shows the nucleotide binding fold observed in dehydrogenases  

Microsoft Academic Search

A striking common structural feature has emerged from the comparison of the X-ray crystallographic studies of several dehydrogenases. In lactate dehydrogenase1, soluble malate dehydrogenase2, alcohol dehydrogenase3 and glyceraldehyde-3-phosphate dehydrogenase4 similar foldings have been described in the region which binds the coenzyme NAD, whereas no significant similarities were observed in the chemical sequences. The occurrence of a characteristic `nucleotide binding fold'

J. L. Risler; C. Zelwer; S. Brunie

1981-01-01

424

D-lactate metabolism in the alga, Chlamydomonas Reinhardtii  

SciTech Connect

(/sup 14/C)D-lactate rapidly accumulates in Chlamydomonas cells under anaerobic conditions from the sugar-phosphate pools which are labeled during photosynthesis with /sup 14/CO/sub 2/. A soluble D-lactate dehydrogenase (30 ..mu..mol NADH oxidized/h/mg Chl), which functions only in the direction of pyruvate reduction, has been partially purified and characterized. The D-lactate is reoxidized in Chlamydomonas by a mitochondrial membrane-bound dehydrogenase. This enzyme is known in the plant literature as glycolate dehydrogenase, an enzyme of the oxidative photosynthetic carbon (C/sub 2/) cycle. This dehydrogenase may be linked to the mitochondrial electron transport chain, although the direct electron acceptor is unknown. Therefore, D-lactate accumulation may be, in part, due to the shut down of electron transport during anaerobiosis. In vivo chase experiments have shown that the D-lactate turns over rapidly when algal cells, which have been grown with air levels of CO/sub 2/ (0.04%), are returned to aerobic conditions in the light. Such turnover is not observed in cells which had been grown with 1 to 5% CO/sub 2/. Cells grown with high CO/sub 2/ have lower levels of glycolate dehydrogenase activity. They are currently using mutants of Chlamydomonas deficient in mitochondrial respiration to study the role of D-lactate oxidation in these algae.

Husic, D.W.; Tolbert, N.E.

1986-05-01

425

Multiple isotope effects as a probe of proton and hydride transfer in the 6-phosphogluconate dehydrogenase reaction.  

PubMed

Primary solvent deuterium, primary substrate deuterium, multiple solvent deuterium/substrate deuterium, and multiple solvent deuterium/13C isotope effects on V/K6PG have been measured for the Candida utilis and sheep liver 6-phosphogluconate dehydrogenases (6PGDH). Proton inventory data suggest the presence of a significant medium effect in a step preceding hydride transfer and the presence of a kinetic solvent deuterium isotope effect on hydride transfer. Multiple isotope effect data confirm the presence of multiple solvent deuterium sensitive steps, likely including a conformational change preceding hydride transfer, hydride transfer, and decarboxylation. PMID:9843374

Hwang, C C; Cook, P F

1998-11-10

426

Tunneling and classical paths for proton transfer in an enzyme reaction dominated by tunneling: oxidation of tryptamine by aromatic amine dehydrogenase.  

PubMed

Proton tunneling dominates the oxidative deamination of tryptamine catalyzed by the enzyme aromatic amine dehydrogenase. For reaction with the fast substrate tryptamine, a H/D kinetic isotope effect (KIE) of 55 +/- 6 has been reported-one of the largest observed in an enzyme reaction. We present here a computational analysis of this proton-transfer reaction, applying combined quantum mechanics/molecular mechanics (QM/MM) methods (PM3-SRP//PM3/CHARMM22). In particular, we extend our previous computational study (Masgrau et al. Science 2006, 312, 237) by using improved energy corrections, high-level QM/MM methods, and an ensemble of paths to estimate the tunneling contributions. We have carried out QM/MM molecular dynamics simulations and variational transition state theory calculations with small-curvature tunneling corrections. The results provide detailed insight into the processes involved in the reaction. Transfer to the O2 oxygen of the catalytic base, Asp128beta, is found to be the favored reaction both thermodynamically and kinetically, even though O1 is closer in the reactant complex. Comparison of quantum and classical models of proton transfer allows estimation of the contribution of hydrogen tunneling in lowering the barrier to reaction in the enzyme. A reduction of the activation free energy due to tunneling of 3.1 kcal mol-1 is found, which represents a rate enhancement due to tunneling by 2 orders of magnitude. The calculated KIE of 30 is significantly elevated over the semiclassical limit, in agreement with the experimental observations; a semiclassical value of 6 is obtained when tunneling is omitted. A polarization of the C-H bond to be broken is observed due to the close proximity of the catalytic aspartate and the (formally) positively charged imine nitrogen. A comparison is also made with the related quinoprotein methylamine dehydrogenase (MADH)-the much lower KIE of 11 that we obtain for the MADH/methylamine system is found to arise from a more endothermic potential energy surface for the MADH reaction. PMID:17388439

Masgrau, Laura; Ranaghan, Kara E; Scrutton, Nigel S; Mulholland, Adrian J; Sutcliffe, Michael J

2007-03-01

427

Glucose and lactate metabolism by Actinomyces naeslundii.  

PubMed

Actinomyces are among the predominant bacteria in the oral microflora. This review discusses the glucose and lactate metabolism of Actinomyces naeslundii and its ecological significance in dental plaque. This bacterium has the Embden-Meyerhof-Parnas (EMP) pathway as the main route to degrade glucose. The EMP pathway-derived metabolic intermediates, phosphoenolpyruvate (PEP) and pyruvate, are further converted into different end-products, depending on the environment. Under anaerobic conditions in the absence of bicarbonate, the pyruvate is converted into lactate by a lactate dehydrogenase. In the presence of bicarbonate, the PEP is combined with bicarbonate and then converted into succinate through the succinate pathway, while the pyruvate is converted into formate and acetate through the pyruvate formate-lyase pathway. Under aerobic conditions, the pyruvate liberates acetate and CO2 through a pathway initiated by a pyruvate dehydrogenase. A. naeslundii strains also degrade lactate, aerobically, to acetate and CO2 through the conversion of lactate into pyruvate by a NAD-independent lactate dehydrogenase. These strains also synthesize glycogen from a glycolytic intermediate, glucose 6-phosphate. Besides atmospheric conditions and bicarbonate, the intracellular reduction-oxidation potential, carbohydrate concentration, and environmental pH also modulate the metabolism of A. naeslundii. Some of the phosphorylating enzymes involved in A. naeslundii metabolism--e.g., GTP/polyphosphate (PPn)-dependent glucokinase, pyrophosphate (PPi)-dependent phosphofructokinase, UDP-glucose pyrophosphorylase, and GDP/IDP-dependent PEP carboxykinase--are unique to A. naeslundii and have not been found in other oral bacteria. The utilization of PPn and PPi as phosphoryl donors, together with glycogen synthesis and lactate utilization, could contribute to the efficient energy metabolism found in A. naeslundii. Through this flexible and efficient metabolic capacity, A. naeslundii can adapt to fluctuating environments and compete with other bacteria in dental plaque. Further, this bacterium may modify the dental plaque environment and promote the microbial population shifts in dental plaque. PMID:10634585

Takahashi, N; Yamada, T

1999-01-01

428

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.

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

2012-01-01

429

Pyruvate into lactate and back: From the Warburg effect to symbiotic energy fuel exchange in cancer cells  

Microsoft Academic Search

Tumor cells fuel their metabolism with glucose and glutamine to meet the bioenergetic and biosynthetic demands of proliferation. Hypoxia and oncogenic mutations drive glycolysis, with the pyruvate to lactate conversion being promoted by increased expression of lactate dehydrogenase A and inactivation of pyruvate dehydrogenase. The NAD+ pool is consecutively regenerated and supports the high glycolytic flux required to produce anabolic

Olivier Feron

2009-01-01

430

Mitochondrial lactate oxidation complex and an adaptive role for lactate production.  

PubMed

The intracellular lactate shuttle (ILS) hypothesis holds that lactate produced as the result of glycolysis and glycogenolysis in the cytosol is balanced by oxidative removal in mitochondria of the same cell. Also, the ILS is a necessary component of the previously described cell-cell lactate shuttle (CCLS), because lactate supplied from the interstitium and vasculature can be taken up and used in highly oxidative cells (red skeletal and cardiac myocytes, hepatocytes, and neurons). This ILS emphasizes the role of mitochondrial redox in creating the proton and lactate anion concentration gradients necessary for the oxidative disposal of lactate in the mitochondrial reticulum during exercise and other conditions. The hypothesis was initially supported by direct measurement of lactate oxidation in isolated mitochondria as well as findings of the existence of mitochondrial monocarboxylate transporters (mMCT) and lactate dehydrogenase (mLDH). Subsequently, the presence of a mitochondrial lactate oxidation complex (composed of mMCT1, CD147 (basigin), mLDH, and cytochrome oxidase (COX)) was discovered, which lends support to the presence of the ILS. Most recently, efforts have been made to evaluate the role of lactate as a cell-signaling molecule (i.e., a "lactormone") that is involved in the adaptive response to exercise. Lactate is capable of upregulating MCT1 and COX gene and protein expression. Current findings allow us to understand how lactate production during exercise represents a physiological signal for the activation of a vast transcription network affecting MCT1 protein expression and mitochondrial biogenesis, thereby explaining how training increases the capacity for lactate clearance via oxidation. PMID:18379211

Hashimoto, Takeshi; Brooks, George A

2008-03-01

431

Malate dehydrogenase from the green gliding bacterium Chloroflexus aurantiacus is phylogenetically related to lactic dehydrogenases  

Microsoft Academic Search

The gene encoding malate dehydrogenase (MDH) from Chloroflexus aurantiacus was cloned, sequenced, and analyzed. The mdh gene corresponded to a polypeptide of 309 amino acids with a molecular mass of 32,717 Da. The primary structure and the coenzyme-binding\\u000a domain showed a high degree of similarity to lactate dehydrogenase (LDH), whereas the conserved amino acids that participate\\u000a in substrate binding were

Bjørnar Synstad; Oddmund Emmerhoff; Reidun Sirevåg

1996-01-01

432

Micro-coulometric study of bioelectrochemical reaction coupled with TCA cycle.  

PubMed

The mediated electro-enzymatic electrolysis systems based on the tricarboxylic acid (TCA) cycle reaction were examined on a micro-bulk electrolytic system. A series of the enzyme-catalyzed reactions in the TCA cycle was coupled with electrode reaction. Electrochemical oxidation of NADH was catalyzed by diaphorase with an aid of a redox mediator with a formal potential of -0.15 V vs. Ag|AgCl. The mediator was also able to shuttle electrons between succinate dehydrogenase and electrode. The charge during the electrolysis increased on each addition of dehydrogenase reaction in a cascade of the TCA cycle. However, the electrolysis efficiencies were close to or less than 90% because of the product inhibition. Lactate oxidation to acetyl-CoA catalyzed by two NAD-dependent dehydrogenases was coupled with the bioelectrochemical TCA cycle reaction to achieve the 12-electron oxidation of lactate to CO(2). The charge passed in the bioelectrocatalytic oxidation of 5 nmol of lactate was 4 mC, which corresponds to 70% of the electrolysis efficiency. PMID:22391482

Tsujimura, Seiya; Fukuda, Jun; Shirai, Osamu; Kano, Kenji; Sakai, Hideki; Tokita, Yuichi; Hatazawa, Tsuyonobu

2012-02-17

433

Potential of mean force calculation for the proton and hydride transfer reactions catalyzed by medium-chain acyl-CoA dehydrogenase: effect of mutations on enzyme catalysis.  

PubMed

Potential of mean force calculations have been performed on the wild-type medium-chain acyl-CoA dehydrogenase (MCAD) and two of its mutant forms. Initial simulation and analysis of the active site of the enzyme reveal that an arginine residue (Arg256), conserved in the substrate-binding domain of this group of enzymes, exists in two alternate conformations, only one of which makes the enzyme active. This active conformation was used in subsequent computations of the enzymatic reactions. It is known that the catalytic alpha,beta-dehydrogenation of fatty acyl-CoAs consists of two C-H bond dissociation processes: a proton abstraction and a hydride transfer. Energy profiles of the two reaction steps in the wild-type MCAD demonstrate that the reaction proceeds by a stepwise mechanism with a transient species. The activation barriers of the two steps differ by only approximately 2 kcal/mol, indicating that both may contribute to the rate-limiting process. Thus this may be a stepwise dissociation mechanism whose relative barriers can be tuned by suitable alterations of the substrate and/or enzyme. Analysis of the structures along the reaction path reveals that Arg256 plays a key role in maintaining the reaction center hydrogen-bonding network involving the thioester carbonyl group, which stabilizes transition states as well as the intervening transient species. Mutation of this arginine residue to glutamine increases the activation barrier of the hydride transfer reaction by approximately 5 kcal/mol, and the present simulations predict a substantial loss of catalytic activity for this mutant. Structural analysis of this mutant reveals that the orientation of the thioester moiety of the substrate has been changed significantly as compared to that in the wild-type enzyme. In contrast, simulation of the active site of the Thr168Ala mutant shows no significant change in the relative orientation of the substrate and the cofactor in the active site; as a result, this mutation has very little effect on the overall reaction barrier, and this is consistent with the experimental data. This study demonstrates that significant insights into the catalytic mechanism can be obtained from simulation studies, and the results can be used to design novel mechanistic probes for the enzyme. PMID:16342946

Bhattacharyya, Sudeep; Ma, Shuhua; Stankovich, Marian T; Truhlar, Donald G; Gao, Jiali

2005-12-20

434

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

435

The role of pyruvate dehydrogenase, phosphofructo-1-kinase and acetylCoA carboxylase in the regulation of fatty acid synthesis in the lactating rat mammary gland during the starved to re-fed transition  

Microsoft Academic Search

Re-feeding 24-h-starved lactating rats resulted in a rapid (within 0.5 h) restoration of glucose uptake by the mammary gland and a slower (within 3 h) restoration of fatty acid synthesis. The rapid reactivation of glucose uptake (82% of fed value within 0.5 h of re-feeding) correlated with a rapid reactivation of 6-phosphofructo-1-kinase (6-PF-1-K) and glycolysis (as determined by a 97%

Kevork Hagopian; Michael R Munday

1997-01-01

436

Lactate conversion to acetate, CO 2 and H 2 in cell suspensions of Desulfovibrio vulgaris (Marburg): indications for the involvement of an energy driven reaction  

Microsoft Academic Search

Cell suspensions of Desulfovibrio vulgaris were found to catalyze, in the absence of sulfate, the complete conversion of 1 lactate to 1 acetate, 1 CO2, and 2 H2 (?G'0=-8.8 kJ\\/mol) and of 1 pyruvate to 1 acetate, 1 CO2, and 1 H2 (?G'0=-52 kJ\\/mol). Protonophores, the proton translocating ATPase inhibitor N,N'-dicyclohexylcarbodiimide, and arsenate specifically inhibited H2 formation from lactate but

Ishverlal P. Pankhania; Alfred M. Spormann; W. Allan Hamilton; Rudolf K. Thauer

1988-01-01

437

Reversible Inactivation of Dehydrogenases.  

National Technical Information Service (NTIS)

The reversible dissociation of lactic and malic dehydrogenases has been studied in detail and compared with the dissociation properties of triosephosphate dehydrogenase, alpha-glycerophosphate dehydrogenase, and liver alcohol dehydrogenase. Evidence is pr...

O. P. Chilson G. B. Kitto J. Pudles N. O. Kaplan

1965-01-01