pH dependence of the kinetic parameters for oxalacetate decarboxylation and pyruvate reduction reactions catalyzed by malic enzyme

Park, Sang Hoon; Harris, Ben G.; Cook, Paul F.

doi:10.1021/bi00361a004

Cited by 36 publications

(37 citation statements)

References 34 publications

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“…The oxaloacetate decarboxylase activity of other enzymes has been reported. Examples are pyruvate kinase (7) and some MEs (22) that are found to be encoded on the chromosome of L. lactis IL1403 (5).…”

Section: Discussionmentioning

confidence: 99%

Mechanism of Citrate Metabolism by an Oxaloacetate Decarboxylase-Deficient Mutant of Lactococcus lactis IL1403

Pudlik

Lolkema

2011

J Bacteriol

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Citrate metabolism plays an important role in many food fermentation processes involving lactic acid bacteria (LAB) (11). Citrate is the precursor of carbon dioxide and the flavor compounds diacetyl and acetoin that contribute to the organoleptic properties of fermented foods. The flavor compounds are formed from the central metabolite pyruvate in the cytoplasm. Citrate metabolism feeds directly into the pyruvate pool (Fig. 1). Following uptake by the secondary citrate transporter CitP, citrate lyase (CL) converts citrate to acetate and oxaloacetate. Acetate is not further metabolized and leaves the cells, while oxaloacetate is decarboxylated to pyruvate by a soluble oxaloacetate decarboxylase encoded by the mae gene and termed CitM (25). In a recent study it was demonstrated that in the lactic acid bacterium Lactococcus lactis IL1403(pFL3), part of the pyruvate formed from citrate was converted into acetoin and part was converted into acetate in parallel pathways (23). In resting cells, the distribution over the two end products strongly depended on the conditions. A high turnover rate through the pathway favored the formation of acetoin over acetate.The citrate transporter CitP plays a pivotal role in the kinetics of the pathway. Under physiological conditions, when citrate is cometabolized with a carbohydrate, CitP operates in the fast citrate/L-lactate exchange mode; citrate is taken up and at the same time that L-lactate, the product of glycolysis, is excreted (precursor/product exchange) (2,17,19,23). In the absence of L-lactate, CitP operates in the much slower unidirectional H ϩ -symport mode (19), which makes uptake the rate-determining step for the flux through the pathway (23). It was shown that, in addition to L-lactate, CitP has an affinity for intermediates/products of the pathway, which results in the typical biphasic consumption of citrate in the absence of L-lactate. During the first phase, citrate is taken up slowly in symport with an H ϩ , but as intermediates/products accumulate in the cytoplasm, the transporter switches to the fast exchange mode, in which citrate is taken up in exchange with pyruvate, ␣-acetolactate, and/or acetate, resulting in much higher consumption rates in the second phase. Therefore, the kinetics and product profile of citrate metabolism are determined largely by the availability of exchangeable substrates for the transporter CitP.The physiological function of citrate metabolism in LAB is in metabolic energy generation and acid stress resistance (8,18,20,28). The citrate metabolic pathway generates proton motive force (PMF) (3,19,20) by an indirect proton pumping mechanism, by which the two components of the PMF, membrane potential (⌬) and pH gradient (⌬pH), are generated in separate steps (13,14). The exchange of divalent citrate and monovalent lactate catalyzed by CitP generates membrane po-

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Section: Discussionmentioning

confidence: 99%

Mechanism of Citrate Metabolism by an Oxaloacetate Decarboxylase-Deficient Mutant of Lactococcus lactis IL1403

Pudlik

Lolkema

2011

J Bacteriol

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“…Implications to Catalytic Mechanism-A three-step acidbase catalytic mechanism is proposed for ascarid malic enzyme based on pH and isotope partitioning studies (25)(26)(27). The assignment of the general acid, general base, and binding groups shown in Scheme 1 is based on the x-ray structure of the ME⅐NAD complex (11) and site-directed mutagenesis studies (19).…”

Section: Figmentioning

confidence: 99%

Crystallographic Studies on Ascaris suum NAD-Malic Enzyme Bound to Reduced Cofactor and Identification of an Effector Site

Rao

Coleman

Karsten

et al. 2003

Journal of Biological Chemistry

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The crystal structure of the mitochondrial NAD-malic enzyme from Ascaris suum, in a quaternary complex with NADH, tartronate, and magnesium has been determined to 2.0-Å resolution. The structure closely resembles the previously determined structure of the same enzyme in binary complex with NAD. However, a significant difference is observed within the coenzyme-binding pocket of the active site with the nicotinamide ring of NADH molecule rotating by 198°over the C-1-N-1 bond into the active site without causing significant movement of the other catalytic residues. The implications of this conformational change in the nicotinamide ring to the catalytic mechanism are discussed. The structure also reveals a binding pocket for the divalent metal ion in the active site and a binding site for tartronate located in a highly positively charged environment within the subunit interface that is distinct from the active site. The tartronate binding site, presumably an allosteric site for the activator fumarate, shows striking similarities and differences with the activator site of the human NAD-malic enzyme that has been reported recently. Thus, the structure provides additional insights into the catalytic as well as the allosteric mechanisms of the enzyme. Malic enzyme (ME)1 is an oxidative decarboxylase that catalyzes the conversion of L-malate to pyruvate and carbon dioxide, using a divalent metal ion (Mg 2ϩ or Mn 2ϩ ) and NAD ϩ or NAD(P) ϩ as cofactors (1-3). The enzyme is found in prokaryotes and eukaryotes and participates in diverse metabolic pathways such as photosynthesis, lipogenesis, and energy metabolism. Mitochondrial and cytosolic isoforms of the enzyme have been identified (1, 4). A sequence comparison of malic enzymes from different sources shows significant homology within the family but no homology to other proteins with the exception of the dinucleotide binding signature motif (5). Because of its functional importance, the enzyme has been isolated and characterized from several sources (3, 6). The mitochondrial NAD-malic enzyme (m-NAD⅐ME) from the parasitic nematode, Ascaris suum, plays a pivotal role in carbohydrate metabolism in parasitic worms (7). In the anaerobic metabolism of A. suum, malate, an intermediate in the worm's glycolytic pathway, is transported into the mitochondria where it undergoes a dismutation and is converted to pyruvate and NADH via the malic enzyme reaction and to fumarate via the fumarase reaction. Fumarate is then converted to short, branched-chain fatty acids via succinate mediated by the NADH produced in the malic enzyme reaction. The succinate dehydrogenase reaction is also involved in a site 1 oxidative phosphorylation, the main source of mitochondrial ATP (8). Since malic enzyme generates reducing equivalents (NADH) for the conversion of fumarate to succinate, it is not surprising that fumarate regulates its own utilization by activating the malic enzyme reaction (9, 10).The ascarid malic enzyme has been extensively studied in our laboratories from the standpoint of its kine...

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“…Dittrich (7), surveying the occurrence of the NAD and NADP malic enzymes, found that the NADP enzyme appeared in excess of the NAD enzyme in many plants, but it should be noted that these assays were of crude extracts and that the NAD malic enzyme was assayed by allowing the MDH activity to come to equilibrium with NAD, then adding Mn2+. The NADH produced in this procedure is a potent inhibitor of the activity of the NAD malic enzyme (22) and in addition, the assay with NADP would include about 60% of the NAD enzyme activity (10). The relative activity ofthe enzyme varies considerably depending on the tissue and the species.…”

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confidence: 99%

Malic Enzymes of Higher Plants

Wedding¹

1989

Plant Physiol.

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The characteristics and distribution of the malic enzyme in plants is discussed as well as those features which appear to be limited to the plant NAD malic enzyme. Regulation of the malic enzyme as it relates to the physiological roles of this enzyme is also discussed. CHARACTERISTICS Enzyme FormsThe malic enzyme is a MDH' that also decarboxylates malate using NAD(P) yielding pyruvate, CO2, and NAD(P)H. A divalent metal ion is required for the reaction. As presently classified by the International Union of Biochemistry, the malic enzyme exists in three different forms. One, MDH

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pH dependence of the kinetic parameters for oxalacetate decarboxylation and pyruvate reduction reactions catalyzed by malic enzyme

Cited by 36 publications

References 34 publications

Mechanism of Citrate Metabolism by an Oxaloacetate Decarboxylase-Deficient Mutant of Lactococcus lactis IL1403

Mechanism of Citrate Metabolism by an Oxaloacetate Decarboxylase-Deficient Mutant of Lactococcus lactis IL1403

Crystallographic Studies on Ascaris suum NAD-Malic Enzyme Bound to Reduced Cofactor and Identification of an Effector Site

Malic Enzymes of Higher Plants

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