High Concentrations Of Pyruvate Research Articles

Inhibition of Human Erythrocyte Lactate Dehydrogenase by High Concentrations of Pyruvate. Evidence for the Competitive Substrate Inhibition

The mechanism of the inhibitory effect of high concentrations of pyruvate on human erythrocyte lactate dehydrogenase has been studied by the use of a new parameter, delta, defined as the difference between the reciprocals of initial reaction rates obtained from experimental measurements and hypothetical linear Lineweaver-Burk plots. This parameter served as a method for differentiating between the competitive and umcompetitive substrate inhibition. Results of this study indicate that pyruvate is a competitive substrate inhibitor. It is suggested that the inhibitory effect of pyruvate is due to its competition with NADH for binding to the free enzyme and formation of an inactive enzyme-pyruvate binary complex. The competitive nature of pyruvate inhibition is further supported by the results of a kinetic study with NADH as the variable substrate. The dissociation constnat of the inactive enzyme-pyruvate binary complex was determined to be 101 micrometer. The physiological significance of the inhibitory effect could be to preserve a level of NADH concentration necessary for other vital enzymic reactions of living cells despite the presence of a high concentration of pyruvate.

Pyruvate carboxylase: Mechanism of the second partial reaction

The stoichiometry of the products of the pyruvate carboxylase reaction has been investigated and shown to vary as the concentration of pyruvate was altered. At high concentrations of pyruvate, the ratio of orthophosphate liberated to oxaloacetate produced approached one, but as the pyruvate concentration was decreased, the ratio increased. On the basis of this evidence, a model for the reaction mechanism was proposed in which the ▪ complex could react either with pyruvate to form oxaloacetate, or water to produce ▪ and HCO 3 −. The nonproductive breakdown of the enzyme-substrate complex and the resulting lack of stoichiometry provides an explanation for the nonlinear double reciprocal plots obtained for both the overall reaction and the pyruvate:oxaloacetate exchange reaction. Since neither the rate of breakdown of the isolated ▪ complex nor the rate of decarboxylation of oxaloacetate in the absence of pyruvate could account for the difference in the amount of the two products formed during the overall reaction, it was postulated that the presence of pyruvate was necessary for hydrolysis to occur. Rate equations were derived describing the dependence of the initial velocity of the release of oxaloacetate in the overall reaction and the rate of the pyruvate:oxaloacetate exchange reaction, on pyruvate concentration. By assigning appropriate values to the various rate constants, theoretical curves were generated and fitted to the experimental data.

Inhibition of gluconeogenesis and lactate formation from pyruvate by N6, O2'-dibutyryl adenosine 3':5'-monophosphate.

N6,O2-Dibutyryl adenosine 3':5'-monophosphate (Bt2cAMP) inhibits gluconeogenesis and lactate formation but increases ketogenesis by isolated liver cells incubated with high concentrations of pyruvate. The inhibitory effects can not be explained on the basis of an inhibition of the pyruvate dehydrogenase complex nor by a change in the NAD+ oxidation-reduction potential of the mitochondrial compartment. Both oleate and 3-hydroxybutyrate substantially increase the rates of gluconeogenesis and lactate formation from pyruvate but do not overcome the inhibition caused by Bt2cAMP. A decreased effectiveness of pyruvate kinase is proposed to account for the inhibition of both gluconeogenesis and lactate formation by Bt2cAMP. This enzyme catalyzes a step required in the transfer of reducing equivalents from the mitochondrial compartment to the cytoplasm and participates in the formation of glucose and lactate from pyruvate by the overall reaction: 2 pyruvate- + 2 NADHmito + 4 ATP4- + 4 H2O leads to 1/2 glucose + lactate- + 2 NAD+ mito + 4 ADP3- + 4 HPO4(2)- + H+. Inhibition of pyruvate kinase promotes gluconeogenesis with most substrates but inhibits gluconeogenesis from pyruvate for want of cytoplasmic reducing equivalents.

Transport of pyruvate nad lactate into human erythrocytes. Evidence for the involvement of the chloride carrier and a chloride-independent carrier.

The kinetics and activation energy of entry of pyruvate and lactate into the erythrocyte were studied at concentrations below 4 and 15mM respectively. The Km and Vmax. values for both substrates are reported, and it is shown that pyruvate inhibits competitively with respect to lactate and vice versa. In both cases the Km for the carboxylate as a substrate was the same as its Ki as an inhibitor. Alpha-Cyano-4-hydroxycinnamate and its analogues inhibited the uptake of both lactate and pyruvate competitively. Inhibition was also produced by treatment of cells with fluorodinitrobenzene but not with the thiol reagents or Pronase. At high concentrations of pyruvate or lactate (20mM), uptake of the carboxylate was accompanied by an efflux of Cl-ions. This efflux of Cl- was inhibited by alpha-cyano-4-hydroxycinnamate and picrate and could be totally abolished by very low (less than 10 muM) concentrations of the inhibitor of Cl- transport, 4,4'-di-isothiocyanostilbene-2,2'-disulphonic acid. This inhibitor titrated out the chlordie efflux induced by pyruvate, bicarbonate, formate and fluoride, in each case total inhibition becoming apparent when approximately 1.2x10(6) molecules of inhibitor were present per erythrocyte, that is, about one inhibitor molecule per molecule of the Cl- carrier. Evan when Cl- efflux was totally blocked pyruvate and lactate uptake occurred. Kinetic evidence is presented which suggests that the Cl- carrier can transport pyruvate and lactate with a high Km and high Vmax., but that an additional carrier with a low Km and a low Vmax. also exists. This carrier catalyses the exchange of small carboxylate anions with intracellular lactate, is competitively inhibited by alpha-cyano-4-hydroxycinnamate and non-competitively inhibited by picrate. The Cl- carrier shows a reverse pattern of inhibition. It is concluded that net efflux of lactic acid from the cell must occur on the Cl- carrier and involve exchange with HCO3 - followed by loss of CO2. The low Km carrier might be used in pyruvate/lactate or acetoacetate/beta-hydroxybutyrate exchanges involved in transferring reducing power across the cell membrane. The possibility that the Cl- carrier exists in cells other than the erythrocyte is discussed. It is concluded that its presence in other cell membranes together with a low intracellular Cl- concentration would explain why the pH in the cytoplasm is lower than that of the blood, and why permeable carboxylate anions do not accumulate within the cell when added from outside.

Pyruvate flux into resealed ghosts from human erythrocytes

The kinetics of pyruvate transport across the isolated red blood cell membrane were studied by a simple and precise spectrophotometric method: following the oxidation of NADH via lactate dehydrogenase trapped within resealed ghosts. The initial rate of pyruvate entry was linear. Influx was limited by saturation at high pyruvate concentration. Pyruvate influx was greatly stimulated by increasing ionic strength in the outer but not the inner aqueous compartment. The K m ranged from 15.0 mM at μ = 0.05 to 3.7 mM at μ = 0.01, while the V went from 0.611 · 10 -15 to 0.137 · 10 -15 mol · min -1 · ghost -1 . Ionic strength was shown to affect the translocation step and not pyruvate binding. The energy of activation of pyruvate flux into resealed ghosts was 25 kcal/mol, similar to that found in intact red blood cells. Inhibitors of pyruvate influx included such anions as thiocyanate, chloride, bicarbonate, α-cyanocinnamate, salicylate and ketomalonate (but not acetate); noncompetitive inhibitors were phloretin, 1-fluoro-2,4-dinitrobenzene, 4-acetamido-4′-isothiocyanate-stilbene-2,2′-disulfonic acid and o-phenanthroline/CuSO 4 mixtures. The last reagent, known to induce disulfide links in certain membrane proteins, blocked the ionic strength stimulation of pyruvate influx in this study.

A fructose 1,6-diphosphate-independent l-lactate dehydrogenase in a strain of Streptococcus mutans

NAD-linked l-lactate dehydrogenase (LDH) activity of Streptococcus mutans strain FIL was not activated by fructose 1,6-diphosphate even at high concentrations of pyruvate and at every pH tested (4.6–7.6) in contrast to other streptococcal NAD-linked l-LDHs. This LDH activity was not affected by the addition of phosphate, ATP nor Mn 2+.

Regulation of lactate dehydrogenase and change of fermentation products in streptococci.

Streptococcus mutans JC 2 produced mainly lactate as a fermentation product when grown in nitrogen-limited continuous culture in the presence of an excess of glucose and produced formate, acetate, and ethanol, but no lactate, under glucose-limited conditions. The levels of lactate dehydrogenase (LDH) in these cultures were of the same order of magnitude, and the activity of LDH was completely dependent on fructose-1,6-diphosphate (FDP). The intracellular level of FDP was high and the level of phosphoenolpyruvate (PEP) was low under the glucose-excess conditions. In the glucose-limited cultures, all glycolytic intermediates studied, except PEP, were low. S. mutans FIL, which had an FDP-independent LDH and similar levels of glycolytic intermediates as S. mutans JC2, produced mainly lactate under glucose-excess or under glucose-limited conditions. LDH of Streptococcus bovis ATCC 9809 was dependent on FDP for activity at a low concentration of pyruvate but had a significant activity without FDP at a high concentration of pyruvate. This strain also produced mainly lactate both under glucose-excess and glucose-limited conditions. The levels and characteristics of these LDHs were not changed by the culture conditions. These results indicate that changes in the intracellular level of FDP regulate LDH activity, which in turn influences the type of fermentation products produced by streptococci. PEP, adenosine 5'-monophosphate, adenosine 5'-diphosphate, and inorganic phosphate significantly inhibited LDH activity from S. mutans JC 2 and may also participate in the regulation of LDH activity in other streptococci.

New Instrument for Rapid Determination of Activities of Lactate Dehydrogenase Isoenzymes

Lactate dehydrogenase isoenzymes can be distinguished kinetically by the fact that isoenzyme H is strongly inhibited a few seconds after the reaction is started if high concentrations of pyruvate are present, in contrast to the M isoenzyme. A new instrument that exploits this fact can measure both the total activity and the proportion of H isoenzyme in serum or plasma in 8 to 10 s. The instrument consists of a simplified stopped-flow apparatus in which the plasma is assayed for lactate dehydrogenase activity, and an electronic device that measures the rate of the reaction at two pre-set time intervals. The first rate is taken between 0.2 and 0.4 s after the reaction is started, a time at which both isoenzymes are fully active, and at which the rate obtained thus reflects total lactate dehydrogenase activity in the plasma sample. The second rate is measured 4 to 6 s after the start of the reaction, at which time the H isoenzyme has become inhibited and the observed rate compared to the initial rate is therefore proportional to the percentage of H isoenzyme activity in the serum. These two rates are electronically displayed on two three-digit voltmeters, the first display being the total activity, the second a number proportional to the inhibited slope. The percentage of M isoenzyme can then be calculated from the initial and final rate. A total of five to six repeat assays may be done within a minute on 1 ml of plasma or serum. This instrument may be of significant value in following the progress of myocardial infarctions and other diseases.

Cyclic AMP induced inhibition of pyruvate kinase flux in the intact liver cell

The rate of pyruvate kinase flux in the intact cell is estimated by a new procedure, involving trapping of 14C from NaH 14CO 3 in a large pyruvate + lactate pool, and calculation of the specific activity of phosphoenol pyruvate. With high concentrations of pyruvate as substrate for isolated rat liver cells, cyclic AMP (0.1 mM) depresses pyruvate kinase flux by about 45%, in addition to inhibiting both glucose and lactate formation. The inhibition of pyruvate kinase may cause an inhibition of hydrogen translocation from the mitochondria to the cytosol.

Malic Enzymes of Rabbit Heart Mitochondria: SEPARATION AND COMPARISON OF SOME CHARACTERISTICS OF A NICOTINAMIDE ADENINE DINUCLEOTIDE-PREFERRING AND A NICOTINAMIDE ADENINE DINUCLEOTIDE PHOSPHATE-SPECIFIC ENZYME

Abstract Two distinct malic enzymes have been separated from rabbit heart mitochondria. Both enzymes require Mn2+ or Mg2+ for activity. One of these enzymes (Enzyme 1) catalyzes the oxidative decarboxylation of malate in the presence of either NAD+ or NADP+, the former being much the more effective. After purification 100- to 200-fold by Sephadex and DEAE-cellulose chromatography and removal of contaminating malic dehydrogenase, some kinetic and physical characteristics of this enzyme were evaluated. The pH optimum of activity is near 7.0, and its isoelectric pH is 5.4. The enzyme appears not to be reversible and does not catalyze the decarboxylation of oxalacetate at pH 4.5 or 7.4. This enzyme is inhibited by ATP competitively with respect to malate, and noncompetitively by oxalate. A similar or identical enzyme is also present in mitochondria from guinea pig and pigeon heart and from pigeon breast muscle. The second enzyme (Enzyme 2), which is readily separated from Enzyme 1 by ammonium sulfate precipitation, is specific for NADP+. This enzyme, which is similar to the enzyme isolated from bovine heart mitochondria (Frenkel, R. (1971) J. Biol. Chem. 246, 3069–3074) also requires low concentrations of Mn2+ or Mg2+ for activity. Enzyme 2 carries out the reverse reaction (malate synthesis) in the presence of high concentrations of CO2 and pyruvate at about 6% of the rate of the forward reaction and catalyzes the decarboxylation of oxalacetate at pH 4.5 at about one-half the rate of forward reaction. The pH optimum (malate oxidation) for Enzyme 2 is between 7.0 and 8.0, and its isoelectric pH is 7.26. In contrast to the bovine heart mitochondrial enzyme, Enzyme 2 showed classical saturation kinetics and was not stimulated by succinate or fumarate at low concentration of malate. Estimates of the molecular weights using Sephadex G-200 indicated values of approximately 180,000 (range in two experiments, 165,000 to 195,000) and 270,000 (range, 260,000 to 280,000) for Enzyme 2 and Enzyme 1, respectively.

Effect of temperature upon inhibition by substrate of lactate dehydrogenase isoenzymes from a poikilotherm

The effect of temperature upon the inhibition by substrate (pyruvate and lactate) and the values of K 1 for NAD + and pyruvate have been studied comparatively on lactate dehydrogenase ( l-lactate : NAD + oxidoreductase, EC 1.1.1.27) isoenzymes purified from tissues of a poikilotherm (the snake Bothrops neuwiedii) and of a homeotherm (Ox). Inhibition of the ophidian isoenzyme B 4 by high concentrations of pyruvate is suppressed at the upper limits of the temperature of the habitat. The data presented suggest that the temperature increment affects only the forces binding the pyruvate in the abortive ternary complex Enzyme-NAD +-pyruvate.

Regulation of glucose synthesis in hormone-sensitive isolated rat hepatocytes.

A simplified procedure was developed for isolation of intact, hormone-sensitive liver cells in a high and reproducible yield. These cells produce glucose from various precursors at rates comparable to those achieved in isolated perfused liver. Glucagon enhanced glucose synthesis from pyruvate, dihydroxyacetone, fructose, or xylitol more effectively at low than at high substrate concentration. At high pyruvate concentrations (>2 mM), glucagon or adenosine 3':5'-cyclic monophosphate (0.1 mM) exerts a curious inhibition of gluconeogenesis that can be reverted to stimulation on addition of ethanol. It is suggested that glucagon and cyclic AMP inhibit pyruvate dehydrogenase and thus limit the supply of reducing equivalents needed for glucose formation. Supporting evidence for hormonal control of pyruvate dehydrogenase in isolated liver cells is provided by the fact that glucagon decreases and insulin increases decarboxylation of [1-(14)C]pyruvate. Calcium salts (1.3 mM) enhance glucose formation from pyruvate but greatly enhance the inhibition exerted by the divalent cationophore, A23187. Inhibition by glucagon of glucose synthesis from pyruvate is additive with the effects of A23187 + Ca(++). However, with dihydroxyacetone as substrate, glucagon partially reverses the inhibition exerted by A23187 + Ca(++). The results are consistent with glucagon effecting an inhibition of pyruvate dehydrogenase and a stimulation of hexosediphosphatase activities.

Identification of lactate dehydrogenase isoenzymes by rapid kinetics.

Stopped-flow kinetics indicate that human and chicken heart-type(4) lactate dehydrogenases (LDH) become inhibited as DPNH is oxidized in the presence of high concentrations of pyruvate. This inhibition is much less marked with the human and chicken muscle-type(4) enzymes. The initial rates and the difference in inhibition between the two types of enzyme have made it possible to determine the amount of, as well as the ratio between, the two types of LDH that are present in a given sample by a single kinetic assay. The stopped-flow kinetic method has been used to analyze amounts of LDH isoenzyme in different tissues, as well as in serum.

Reversibility of the tryptophanase reaction: synthesis of tryptophan from indole, pyruvate, and ammonia.

Degradation of tryptophan to indole, pyruvate, and ammonia by tryptophanase (EC 4....) from Escherichia coli, previously thought to be an irreversible reaction, is readily reversible at high concentrations of pyruvate and ammonia. Tryptophan and certain of its analogues, e.g., 5-hydroxytryptophan, can be synthesized by this reaction from pyruvate, ammonia, and indole or an appropriate derivative at maximum velocities approaching those of the degradative reactions. Concentrations of ammonia required for the synthetic reactions produce specific changes in the spectrum of tryptophanase that differ from those produced by K(+) and indicate that ammonia interacts with bound pyridoxal 5'-phosphate to form an imine. Kinetic results indicate that pyruvate is the second substrate bound, hence indole must be the third. These results favor a modified mechanism for the multitude of tryptophanase-catalyzed reactions in which alpha-aminoacrylate, which functions as a common enzyme-bound intermediate in both synthetic and degradative reactions, is not released into the medium during the latter reactions, but is degraded to pyruvate and ammonia by sequential reversible steps via enzyme-bound intermediates.

A Lactate Dehydrogenase Specific to the Liver of Gadoid Fish

Comparative studies on tissue extracts of haddock, cod, and related species in the suborder Gadoidei indicated the presence of a lactate dehydrogenase specific to liver tissue. Genetic and evolutionary variation of the lactate dehydrogenase isozymes in these fishes indicate that the liver isozyme is encoded at a structural gene locus genetically distinct from the loci encoding the heart and muscle isozymes. The properties of purified liver, heart, and muscle lactate dehydrogenases from haddock were compared to investigate their relationship. The haddock liver enzyme is like other lactate dehydrogenases in having a molecular weight of 140,000 with a subunit size of 35,000. The liver enzyme is distinguished from the heart and muscle isozymes by its greater resistance to heat inactivation. The gadoid heart and muscle isozymes do not hybridize with each other in vivo or in vitro, but the liver enzyme can hybridize with both in vitro, although not in a random fashion. The catalytic properties of the haddock liver and heart isozymes are very similar; they are distinct from the muscle isozyme in having a greater affinity for both lactate and pyruvate and in being inhibited at high pyruvate concentrations. The liver enzyme showed no unusual substrate specificities. Ironically, the liver enzyme can be mistaken for the muscle enzyme since both have nearly identical electrophoretic mobilities. The catalytic properties of the haddock heart and liver enzymes suggest that these two may be homologous and that both might be homologous to the heart-type lactate dehydrogenase of the higher vertebrates. The immunological crossreactivity of these two with each other and with chicken heart lactate dehydrogenase provides confirmation for this hypothesis. The immunological data suggest a considerable degree of sequence homology between the heart and liver lactate dehydrogenases of gadoid fishes. From evolutionary considerations, it appears that the divergence of the heart and liver loci from an ancestor heart-type locus is a relatively recent event, perhaps arising concurrently with the evolution of the gadoid fish into a distinct lineage.

Functional aspects of lactate dehydrogenase isoenzymes of atlantic salmon ( Salmo salar L.)

1. 1. Lactate dehydrogenase (LDH) in heart, skeletal muscle, liver, and blood-plasma of Atlantic salmon ( Salrno solar L.) were investigated with respect to distribution and kinetic properties. 2. 2. Skeletal muscle and blood-plasma contain one group of five electrophoretically slow-migrating isoenzymes of LDH. Heart and liver contain a second group of five fastmigrating isocnzymes. 3. 3. These two groups show differences in inhibition at high concentrations of pyruvate as well as of lactate, in such a way that the fast-migrating forms are more inhibited than the slow-migrating ones. 4. 4. The influence of temperature and pH on the differences in substrate inhibition was studied. 5. 5. Differences in apparent K m values were also obtained. 6. 6. No differences were detected between the activities of the two groups in their dependence on concentrations of NAD or NADH. 7. 7. The electrophoretically extreme LDH forms of each isoenzyme group were isolated. These extreme forms showed the same apparent K m pyruvate as all the isoenzymes of their respective group.

Regulation of gluconeogenesis in the guinea pig liver.

Because of differences in the pattern of enzyme activities involved in glycolysis and gluconeo‐genesis between rat and guinea pig liver, the regulation of gluconeogenesis in guinea pig liver was studied.The following results were obtained: Starvation increases the capacity for gluconeogenesis from pyruvate in vivo. Gluconeogenesis from l‐lactate and from pyruvate in isolated perfused livers is greater in the guinea pig than in the rat, whereas gluconeogenesis from other precursors is similar in both species. Long chain and medium chain fatty acids inhibit gluconeogenesis from l‐lactate in isolated perfused rat liver. In the guinea pig liver the difference does not result from a lower rate of fatty acid oxidation but from a greater reduction in the cytoplasmic NAD+/NADH system.During enhanced fatty acid oxidation in guinea pig liver and in rat liver, similar changes in the concentrations of both acetyl‐S‐CoA and CoA‐SH are observed.With high concentrations of pyruvate as the gluconeogenic precursor, both fatty acids and ethanol lead to a slight stimulation of glucose formation by supplying hydrogen equivalents to the cytosolic NAD+/NADH system. Both glucagon and dibutyryl‐Ado‐3′:5′‐P stimulate ureogenesis and ketogenesis in isolated perfused guinea pig liver. However, the stimulation of gluconeogenesis by glucagon and Ado‐3′:5′‐P normally observed in isolated perfused rat livers was not found in isolated perfused guinea pig livers. Partially purified pyruvate carboxylases from rat and guinea pig liver do not differ with respect to the apparent Km values for pyruvate and ATP, and also the apparent Ka values for acetyl‐S‐CoA at different pH values. At constant concentrations of MgATP2− or MnATP2− the pH‐optima for the guinea pig enzyme are significantly lower. 5‐Methoxyindole‐2‐carbonic acid inhibits gluconeogenesis in guinea pig liver in the same way as in rat liver. Quinolinate, on the other hand is far less inhibitory in guinea pig liver and does not inhibit at all in isolated perfused pigeon livers. Major species differences are found when the activities of the rate limiting enzymes involved in gluconeogenesis are measured under Vmax conditions. Pyruvate carboxylase activity is in the same range in rat and guinea pig liver; pyruvate kinase activity is considerably lower, phospho‐enolpyruvate carboxykinase activity is considerably higher in guinea pig liver. According to our results gluconeogenesis in guinea pig liver from lactate or pyruvate is mainly regulated by the concentration of pyruvate.The difference in the regulation of gluconeogenesis between rat and guinea pig liver probably results from a difference in the compartmentalization of phosphoenopyruvate carboxykinase. A higher rate of futile cycling between phosphoenolpyruvate and pyruvate in rat liver, due to the higher activity of pyruvate kinase in relation to the activities of phosphoenolpyruvate carboxykinase and pyruvate carboxylase, may also contribute to the observed differences.

Thiamine-dependent Neonatal Lactic Acidosis with Hyperalaninemia

The ‘vitamin dependencies’ comprise a rapidly developing group of metabolic diseases. We describe a recurring disorder of pyruvate metabolism responsive to thiamine supplements. Severe neonatal metabolic acidosis was observed in a child who subsequently developed psychomotorretardation and infantile spasms. The acidosis was accompanied by increased concentrations of lactate, pyruvate and -alanine in plasma and urine. Spontaneous acidosis has been intermittent, three episodes being recorded since birth. In later life, thiamine (25 mg i.v.) given after correction of acidosis with bicarbonate, provoked a severe metabolic alkalosis. The last episode could be induced by high carbohydrate diet, corrected in 4 days by thiamine (5 mg i.m.), and controlled by a low carbohydrate, high protein diet. Nutritional deficiency of thiaming has been ruled out. Since -ketoglutarate levels were not increased, a specific thiamine-dependent defect in pyruvate decarboxylase was considered. Assay of this enzyme in fresh peripheral leucocytes, or after phytohemagglutination stimulation, at low and high concentrations of pyruvate, with and without thiamine added, revealed no abnormality. Negative results wre also obtained with cultured skin fibroblasts. A phenotypic variant of the traits described by BLASS and TADA, but limited to liver in this case, is proposed. (Research supported by Grants MRC 1085 and NHW 604-7-643.)

Physiological Concentrations of Lactate Dehydrogenases and Substrate Inhibition

Lactate dehydrogenases at physiological concentrations are inhibited by high concentrations of pyruvate when the enzyme and the pyruvate are incubated in the presence of oxidized nicotinamide-adenine dinucleotide before assay. The inhibition is much more pronounced with the H-type than with the M-type lactate dehydrogenase. These results suggest that substrate inhibition may be operative in vivo.

Inhibition of lactate dehydrogenase by high concentrations of pyruvate: The nature and removal of the inhibitor

Inhibition of lactate dehydrogenase by high concentrations of pyruvate: The nature and removal of the inhibitor