Pigeon liver malic enzyme

Hsu, Robert Y.

doi:10.1007/bf00229535

Cited by 99 publications

(96 citation statements)

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“…Data from several different analyses, including affinity labeling with bromopyruvate which alkylates sulfhydryls, indicate that a sulfhydryl does occur in the active site of the animal enzyme, but it is thought to bind the metal and not be directly involved in catalysis (7,10). Wedding's group (15) supports this idea, and they contend that the pK of this sulfhydryl can be detected in the response of Vma to pH for NAD-ME.…”

Section: Resultsmentioning

confidence: 99%

“…up to pH 8.0 (7,10), while the C3 plant enzyme appears to have an optimum activity around pH 6.4 to 7.3 (4,14). The plant NAD-ME, located in mitochondria, is similar to the C3 NADP-ME with respect to the pH at which it exhibits maximum activity (15).…”

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confidence: 94%

“…Extensive mechanistic information is available for the pigeon liver NADP-ME (7,10) and is becoming available for plant NAD-ME (15), but such information for the forms of plant NADP-ME is minimal. In light of the differences in the pH optima for the various MEs, we have used the effect of pH on the kinetic parameters with respect to malate for the F. trinervia enzyme as a means of comparing the pK values of functional groups of this putative special form of ME with those published for the pigeon liver enzyme and the plant NAD-ME.…”

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Effect of pH on the Kinetic Parameters of NADP-Malic Enzyme from a C₄Flaveria (Asteraceae) Species

Holaday

Lowder²

1989

Plant Physiol.

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We have used the pH variation in the kinetic parameters with respect to malate of NADP-malic enzyme purified from the C4 species, Flaveria trinervia, to compare the pK values of its functional groups with those for the pigeon liver NADP-malic enzyme (Ml Schimerlik, WW Cleland [1977] Biochemistry 16: 576-583) and the plant NAD-malic enzyme (KO Willeford, RT Wedding ] Plant Physiol 84: 1084-1087. Like the other enzymes, the C4 enzyme has a group with a pK of about 6.0 (6.6 for the C4 enzyme), as indicated from plots of the log V,mx/Km (Vmax = maximum rate of catalysis) versus pH, which must lose a proton for malate binding and subsequent catalysis. The optimum ionization for the C4 enzyme-NADP-Mg2+ complex occurs at pH 7.1 to 7.5. From pH 7.5 to 8.4, the Km increases, but V,,., remains constant. The log VmaxIKm plot in this pH range indicates a group with a pK of about 7.7. The other malic enzymes exhibit a similar pK. Above pH 8.4, deprotonation leads to a marked increase in Km and a decrease in V,,, for the C4 enzyme. As in the case of the animal enzyme, the log Vm,x/Km plot for the C4 enzyme appears to approach a slope of two. The curve suggests an average pK of 8.4 for the groups involved, while the animal enzyme exhibits an average pK of 9.0. The NAD-malic enzyme does not exhibit any pK values at these high pK values. We hypothesize that the putative groups with the high pK values may be at least partially responsible for the ability of the C4 NADP-malic enzyme to maintain high activity at pH 8.0 in illuminated chloroplasts.Vma. up to pH 8.0 (7, 10), while the C3 plant enzyme appears to have an optimum activity around pH 6.4 to 7.3 (4, 14). The plant NAD-ME, located in mitochondria, is similar to the C3 NADP-ME with respect to the pH at which it exhibits maximum activity (15). Of course, several factors determine the optimum pH to use for in vitro assays, but a superficial look at all MEs suggests that the C4 enzyme is a special form that has evolved for decarboxylation at high rates in illuminated choroplasts during C4 photosynthesis.Extensive mechanistic information is available for the pigeon liver NADP-ME (7, 10) and is becoming available for plant NAD-ME (15), but such information for the forms of plant NADP-ME is minimal. In light of the differences in the pH optima for the various MEs, we have used the effect of pH on the kinetic parameters with respect to malate for the F. trinervia enzyme as a means of comparing the pK values of functional groups of this putative special form of ME with those published for the pigeon liver enzyme and the plant NAD-ME. MATERIALS AND METHODS Plant MaterialPlants of Flaveria trinervia (Spreng.) C. Mohr were grown in a greenhouse under a 14 h photoperiod during the summer months. The seed was collected locally, and the plants were fertilized with a Hoagland solution three times a week.

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Section: Resultsmentioning

confidence: 99%

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confidence: 94%

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confidence: 99%

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Effect of pH on the Kinetic Parameters of NADP-Malic Enzyme from a C₄Flaveria (Asteraceae) Species

Holaday

Lowder²

1989

Plant Physiol.

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“…A stepwise oxidative decarboxylation of malate by the NADPmalic enzyme was proposed early in the investigation of its mechanism (Viega Salles & Ochoa, 1950;Hsu, 1970;Tang & Hsu, 1973) with hydride transfer from malate to NADP preceding decarboxylation of the resulting oxalacetate intermediate. The proposed mechanism was based on the observation that the NADP-enzyme will catalyze the reduction of oxalacetate to malate and the decarboxylation of oxalacetate to pyruvate and CO, in the presence of NADPH.…”

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Role of the divalent metal ion in the NAD:malic enzyme reaction: An ESEEM determination of the ground state conformation of malate in the E:Mn: Malate complex

et al. 1996

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The conformation of L-malate bound at the active site of Ascaris m u m malic enzyme has been investigated by electron spin echo envelope modulation spectroscopy. Dipolar interactions between Mn2+ bound to the enzyme active site and deuterium specifically placed at the 2-position, the 3R-position, and the 3s-position of L-malate were observed. The intensities of these interactions are related to the distance between each deuterium and Mn2+. Several models of possible Mn-malate complexes were constructed using molecular graphics techniques, and conformational searches were conducted to identify conformers of malate that meet the distance criteria defined by the spectroscopic measurements. These searches suggest that L-malate binds to the enzyme active site in the trans conformation, which would be expected to be the most stable conformer in solution, not in the gauche conformer, which would be more similar to the conformation required for oxidative decarboxylation of oxalacetate formed from L-malate at the active site of the enzyme.Keywords: decarboxylation; EPR spectroscopy; malic enzyme; substrate conformationThe mitochondrial NAD-malic enzyme (EC 1.1.1.39) from Ascaris suum catalyzes the divalent metal ion-dependent oxidative decarboxylation of L-malate utilizing NAD to yield pyruvate, CO,, and NADH. Several different divalent metal ions including Mg2+, Mn2+, Co2+, Ni2+, Cd2+, and Zn2+ (Schimerlik et at.,

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“…The importance of sulfhydryl residues in pigeon liver malic enzyme has been investigated extensively (10). Studies of the inactivation of this enzyme with reagents selective for sulfhydryl groups, e.g.…”

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Evidence for the Existence of Two Essential and Proximal Cysteinyl Residues in NADP-Malic Enzyme from Maize Leaves

Drincovich

Spampinato²,

Andreo³

1992

Plant Physiol.

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Incubation of maize (Zea mays) leaf NADP-malic enzyme with monofunctional and bifunctional N-substituted maleimides results in an irreversible inactivation of the enzyme. Inactivation by the monofunctional reagents, N-ethylmaleimide (NEM) and N-phenylmaleimide, followed pseudo-first-order kinetics. The maximum inactivation rate constant for phenylmaleimide was 10-fold higher than that for NEM, suggesting a possible hydrophobic microenvironment of the residue(s) involved in the modification of the enzyme. In contrast, the inactivation kinetics with the bifunctional maleimides, ortho-, meta-, and para-phenylenebismaleimide, were biphasic, probably due to different reactivities of the groups reacting with the two heads of these bifunctional reagents, with a possible cross-linking of two sulfhydryl groups. plants such as maize. The enzyme plays a key role in C4 photosynthetic metabolism because it generates reducing power and CO2 in the bundle sheath chloroplasts where Rubisco and the Calvin cycle operate (7,8).Thiol groups play a key role in the activity of the enzyme obtained from different sources. The importance of sulfhydryl residues in pigeon liver malic enzyme has been investigated extensively (10). Studies of the inactivation of this enzyme with reagents selective for sulfhydryl groups, e.g. 5,5'-dithiobis(2-nitrobenzoic acid) or NEM2, confirmed the presence of one sulfhydryl group near each substrate site on the enzyme tetramer (24). In contrast, evidence for the presence of two essential sulfhydryl residues per monomer of the enzyme from maize leaves has been recently reported (4).To obtain further information concerning the function and microenvironment of the active site thiol group(s), the reaction of NADP-malic enzyme from maize leaves with different N-substituted maleimides was studied. Bifunctional maleimides were used to obtain further evidence about the existence of two proximal sulfhydryl groups near the NADP-binding site of the C4 enzyme (4). Several isomers of phenylenebismaleimide with different estimated cross-linking distances, as well as maleimides that differ in hydrophobic properties, were all found to modify the enzyme in an irreversible fashion.NADP-dependent malic enzyme (L-malate: NADP oxidoreductase/decarboxylase; EC 1.1.1.40) acts in a wide range of metabolic pathways in both animals and plants. The enzyme catalyzes the reversible oxidative decarboxylation of L-malate in the presence of NADP and a bivalent metal ion to produce pyruvate, NADPH, and CO2. In animals, cytosolic malic enzyme generates reducing power for the biosynthesis of fatty acids (9, 28). In plants, two forms of this enzyme are known to occur and have important metabolic roles. The cytosolic form is thought to participate in the regulation of intracellular pH (3,20) and/or in the provision of reducing power that can be used in processes requiring NADPH (8).The chloroplast stromal form is found specifically in the bundle sheath chloroplasts of NADP-malic enzyme type C4

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Pigeon liver malic enzyme

Cited by 99 publications

References 50 publications

Effect of pH on the Kinetic Parameters of NADP-Malic Enzyme from a C₄Flaveria (Asteraceae) Species

Effect of pH on the Kinetic Parameters of NADP-Malic Enzyme from a C₄Flaveria (Asteraceae) Species

Role of the divalent metal ion in the NAD:malic enzyme reaction: An ESEEM determination of the ground state conformation of malate in the E:Mn: Malate complex

Evidence for the Existence of Two Essential and Proximal Cysteinyl Residues in NADP-Malic Enzyme from Maize Leaves

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Pigeon liver malic enzyme

Cited by 99 publications

References 50 publications

Effect of pH on the Kinetic Parameters of NADP-Malic Enzyme from a C4Flaveria (Asteraceae) Species

Effect of pH on the Kinetic Parameters of NADP-Malic Enzyme from a C4Flaveria (Asteraceae) Species

Role of the divalent metal ion in the NAD:malic enzyme reaction: An ESEEM determination of the ground state conformation of malate in the E:Mn: Malate complex

Evidence for the Existence of Two Essential and Proximal Cysteinyl Residues in NADP-Malic Enzyme from Maize Leaves

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Effect of pH on the Kinetic Parameters of NADP-Malic Enzyme from a C₄Flaveria (Asteraceae) Species

Effect of pH on the Kinetic Parameters of NADP-Malic Enzyme from a C₄Flaveria (Asteraceae) Species