L-malate to pyruvate

The practical usefulness of Equations 11.46 through 11.53 has been demonstrated for the malic enzyme catalyzed conversion of L-malate to pyruvate (Equation 11.72). Table 11.1 lists experimentally determined isotope effects for this reaction. Comparison of carbon kinetic isotope effects for protio and deutero-malate substituted at position 2 (the carbon that undergoes sp3 to sp2 transition) rules out the possibility that the hydride transfer and the decarboxylation events are concerted. This conclusion follows from Equation 11.48 which, for a concerted reaction, predicts that 13(V/K) should be smaller than 13(V/K)D, which is opposite to the order observed experimentally. 

Linked oxidation and decarboxylation. Metabolic pathways often make use of oxidation of a (3-hydroxy acid to a (3-oxoacid followed by decarboxylation in the active site of the same enzyme. An example is conversion of L-malate to pyruvate (Eq. 13-45). The Mg2+ or Mn2+-dependent decarboxylating malic dehydrogenase that catalyzes the reaction is usually called the malic enzyme. It is found in most organisms.237-240 While a concerted decarboxylation and dehydrogenation may sometimes occur,241-242 the enzymes of this group appear usually to operate with bound oxoacid intermediates as in Eq. 13-45. 

Malic enzymels), L-malatt-NADP oxidortdm-tase, decarboxylating (EC 1.1.1.40) an important enzyme found in most organisms, which catalyses the decarboxylation of L-malate to pyruvate and CO2, with concomitant reduction of NADP to NADPH (or the synthesis of malate by the reverse reaction) HOOC-CHj-CHOH-COOH + NADF CHj-CO-COOH + CO2 + NADPH + H. M. e. has various metabolic roles 1. Synthesis of malate by the action of M.e. may serve as an Anaplerotic reaction (see) of the TCA-cycle 2. An important route for the total combustion of any TCA-[Pg.380]

In discussion of nomenclature of malic acid decomposing enzymes, mention should be made of malate-lactate transhydrogenase. This enzyme, isolated from Micrococcus lactiyticus (VielloneUa alcalescens), a bacterium found in vertibrates, can catalyze reversibly the conversion of L-malic and pyruvic acids to L-lactic and oxaloacetic acids with NAD as coenzyme (36). 

The problem of stepwise versus concerted oxidative decarboxylation of L-maleate to yield pyruvate CO2 and reduced dinucleotide catalysed by malic enzyme615 has been reinvestigated recently616. The new D and T KIE determinations, using L-malate-2D... 

In the presence of malate deshydrogenase (MDH), oxaloacetate and pyruvate, its decarboxylation derivative, are reduced to L-malate by reduced NADH. The equilibrium of the reaction is forced in the direction of the products by the judicious selection of operating conditions (buffer pH7.8 and an excess of NADH) ... 

In the presence of Sn(OTf)2, (5)-l-pentyl-2-[(piperidin-l-yl)methyl]pyrrolidine, and BusSnF, l-(5)-ethyl-l-trimethylsiloxyethene reacts with methyl pyruvate to give the desired adduct in 92 % ee (Eq. 21). Methyl isopropylglyoxylate and methyl phe-nylglyoxylate also react with l-(5)-ethyl-l-trimethylsiloxyethene to give the corresponding 2-substituted malates in good yields and excellent enantioselectivity [36],... 

Burstein et al. (1986) have shown that in addition to induction and inhibition mechanical and chemical changes of the cell structure effect the selectivity of microbial sensors. Escherichia coli cells were treated by ultrasound, extrusion, or crosslinked with glutaraldehyde before used in sensors for D- and L-lactate, succinate, L-malate, 3-glycero-phosphate, pyruvate, NADH, and NAD PH. The treatment caused different selectivity of the respiratory chain and thus permitted an increase in the specificity of the respective sensors. 

Creighton and Rose (45) have explored the route shown in Scheme 7 to synthesize chiral pyruvate. The method takes advantage of the fact that pyruvate kinase decarboxylates oxalacetate with retention of configuration. The requisite labeled oxalacetates were prepared as shown from the appropriately labeled L-malates, available from equilibration with fumarase in either deuterated or tritiated water. This method is convenient for the preparation of small amounts of chiral acetate but suffers from the low oxalacetate decarboxylase activity of pyruvate kinase. 

The oxidation of L-malate in most living organisms is catalyzed by two distinct types of pyridine nucleotide-dependent enzymes. In one case the principal product is oxaloacetate, while in the other it is pyruvate and CO2. The enzymes of the malate-oxaloacetate class, which utilize NAD+, have been referred to as simple dehydrogenases, while enzymes of the malate-pyruvate type, which, in contrast, use NADP+, have been designated decarboxylating dehydrogenases and are commonly known as malic enzymes (1). 

See also Relationship of Gluconeogenesis to Glycolysis, Gluconeogenesis, Lactate Dehydrogenase, Pyruvate, Lactate, Oxaloacetate, L-Malate. 

Since oxaloacetate is easily decarboxylated into pyruvate and CO2, these two reactions lead to the formation of pyruvate from L-malate. Since the final product of the malolactic transformation in wine is L-lactic acid, MDH or the malic enzyme would be associated to an LDH catalyzing the reduction of pyruvate into L-lactate in this metabolic pathway. At least for wine bacteria, this concept is not acceptable since these bacteria only possess a d-LDH. Malic acid would only lead to the formation of D-lactic acid. 

All reactions mediating fixation of CO by reductive carboxylation are readily reversible. Their equilibrium constant is sUghtly in favor of carboxylation, but at the low tensions of CO prevailing in cells and biological fluids the reverse action, i.e., oxidative decarboxylation, is favored unless appropriate mechanisms are broi t into play to displace the equilibrium position in favor of carboxylation. The two primary processes in each of these reactions, oxidation and decarboxylation or carboxylation and reduction, are intiinately interconnected and appear to be catalyzed by the same enzyme protein. The reactions to be considered in this section are the reductive car-boxylations of pyruvate, a-ketoglutarate, and ribulose 5-phosphate to L-malate, d-isocitrate, and 6-phosphogluconate, respectively. 

Lactate and alanine enter as pyruvate following the activities of lactate dehydrogenase (Figure 11.4) and alanine aminotransferase (Section 16.3). The first of the bypass reactions, the objective of which is to overcome the unfavourable energetics of a reversal of the pyruvate kinase reaction, seems a tortuous route (Figure 11.11). The reaction sequence relies on two important enzymes pyruvate carboxylase and phosphoenolpyruvate carboxykinase. Since pyruvate carboxylase is located exclusively in the mitochondrion, pyruvate must cross the inner mitochondrial membrane (Section 12.2). Oxaloacetate produced by pyruvate carboxylase cannot traverse the inner membrane and is reduced by malate dehydrogenase into l-malate. This step is the reversal of the tricarboxylate cycle reaction (Section 12.4). Malate may, of... 

NADH as an end product. This implicates oxidized malic acid, either pyruvic or oxaloacetic acid, as another end product. By adding commercial preparations of L-lactic dehydrogenase or malic dehydrogenase to the reaction mixture, Morenzoni (90) concluded that the end product was pyruvic acid. Attempts were then made to show whether two enzymes—malate carboxy lyase and the classic malic enzyme, malate oxidoreductase (decarboxylating), were involved or if the two activities were on the same enzyme. The preponderance of evidence indicated that only one enzyme is involved. This evidence came from temperature inactivation studies, heavy-metal inhibition studies, and ratio measurements of the two activities of partially purified preparations of Schiitz and Radlers malo-lactic enzyme (76, 90). This is not the first case of a single enzyme having two different activities (91). 

The ratio [NAD+]/[NADH] appears to be maintained at a relatively constant value and in equilibrium with a series of different reduced and oxidized substrate pairs. Thus, it was observed that in the cytoplasm of rat liver cells, the dehydrogenations catalyzed by lactate dehydrogenase, sn-glycerol 3-phosphate dehydrogenase, and malate dehydrogenase are all at equilibrium with the same ratio of [NAD+]/[NADH].166 In one experiment rat livers were removed and frozen in less than 8 s by "freeze-clamping" (Section L,2) and the concentrations of different components of the cytoplasm determined167 the ratio [NAD+] / [NADH] was found to be 634, while the ratio of [lactate]/[pyruvate] was 14.2. From these values an... 

Answer Because pyruvate carboxylase is a mitochondrial enzyme, the [14C]oxaloacetate (OAA) formed by this reaction mixes with the OAA pool of the citric acid cycle. A mixture of [1-14C] and [4-14C] OAA eventually forms by randomization of the C-l and C-4 positions in the reversible conversions OAA —> malate —> succinate. [1-14C] OAA leads to formation of [3,4-14C]glucose. 

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