Lactate dehydrogenases keto acids

Adam, W., Lazarus, M., Saha-Moller, C.R. and Schreier, P. (1998) Quantitative transformation of racemic 2-hydroxy acids into (R)-2-hydroxy acids by enantioselective oxidation with glycolate oxidase and subsequent reduction of 2-keto acids with D-lactate dehydrogenase. Tetrahedron Asymmetry, 9 (2), 351-355. 

Dehydrogenases often act primarily to reduce a carbonyl compound rather than to dehydrogenate an alcohol. These enzymes may still be called dehydrogenases. For example, in the lactic acid fermentation lactate is formed by reduction of pyruvate but we still call the enzyme lactate dehydrogenase. In our bodies this enzyme functions in both directions. However, some enzymes that act mainly in the direction of reduction are called reductases. An example is aldose reductase, a member of a family of aldo-keto reductases71 73 which have (a / P)8-barrel structures.74 76... 

R)-2-Hydroxy-4-phenylbutyric acid was produced continuously in an enzyme membrane reactor by enzymatic reductive animation of the a-keto acid with d-lactate dehydrogenase coupled with formate dehydrogenase (FDH) for regeneration of NADH. Reactor performance data matched a kinetic reactor model (Schmidt, 1992). 

Enzyme-catalyzed syntheses of enantiomeric pure hydroxy acids use lactate dehydrogenases (LDH E.C. 1.1.1.27) or hydroxyisocaproate dehydrogenases (HicDH). Both kinds of enzymes are available as D- or L-specific catalysts. L-specific [5, 6] LDHs as well as D-specific [7-10] LDHs favorably catalyze the reduction of pyruvate, HicDHs (and mandelate dehydrogenase, too) convert keto acids with longer aliphatic or aromatic side chains. These enzymes can be isolated from Lactobacillus strains [11-14]. 

Fig. 8 Synthesis of amino acids by a multienzyme system consisting of leucine dehydrogenase (LeuDH) catalyzing the reductive amination of the corresponding keto acid, L-lactate dehydrogenase (l-LDH), and lactate for the regeneration of NADH and urease for the in situ generation of ammonia. The coenzyme NAD+ was covalently bond to dextran, enzymes and dextran-coupled NAD+ were...

L-Lactate dehydrogenase (l-LDH, EC 1.1.1.27) catalyzes the reduction of pyruvate to (S)-lactate with a simultaneous oxidation of NADH. l-LDH is found in all higher organisms. There are two kinds of l-LDHs enzymes from one group are activated by fructose 1,6-diphosphate while the other group stays independent [71]. l-LDH is highly selective for pyruvate, short-chain 2-keto acids and phenylpyruvic acid [80]. All bacterial NAD+-dependent LDHs form lactate from pyruvate in vivo, and there is no evidence at all that they catalyze the other direction as well. The equilibrium constant lies far on the direction of lactate formation, and thus the reaction catalyzed by bacterial LDHs can be considered almost irreversible. LDHs from some lacto-bacilli like Lactobacillus fermentum or L. cellobiosus show no or just poor reaction with lactate [71], whereas mammalian LDHs can be considered as reversible [71]. Well characterized l-LDHs are summarized in Table 2. 

The two-enzyme system was also used to convert L-lactate into D-lactate with a yield better than 97%. L-Lactate is oxidized by L-lactate dehydrogenase to give pymvate. The keto acid is reduced in an electrochemical system at the cathode to racemic lactate and NADH is oxidized to NAD at the anode. The continuous... 

Stereospecific reductions of a-keto acids are well documented. l- and D-lactate dehydrogenases from common mammalian or bacterial sources are available for this purpose. By using a preselected l- or d-LDH, the preparation of a (2S)- or (2/ )-hydroxy acid of >99% ee can be virtually assured. The structural range of a-keto acids (29) that has been subjected to preparative-scale reductions to hydroxy acids... 

Reductions catalyzed by lactate dehydrogenase formation of chiral a-hydroxy acids from ck--keto acids. 

Resistance to vancomycin is via a sensor histidine kinase (VanS) and a response regulator (VanR). VanH encodes a D-lactate dehydrogenase/a-keto acid reductase and generates D-lactate, which is the substrate for VanA, a D-Ala-D-Lac ligase. The result is cell wall precursors terminating in D-Ala-D-... 

It was discovered in 1958 that anaerobically grown yeast contains a form of lactate dehydrogenase which is different from the d- and L-lac-tate cytochrome c reductases of aerobic yeast (306, 319). The enzyme has been partially purified (320, 321), and shown to contain flavin (320-322). Gel filtration studies have suggested a molecular weight of about 100,000 (320, 321). Preparations of the enzyme oxidize several d-2-hydroxyacids to the respective keto acids in a reversible manner (320). For the forward reaction, ferricyanide, 2,6-dichloroindophenol, menadione, and methylene blue have been used as electron acceptors, and for the reverse reaction leucomethyl viologen and FMNHa are effective electron donors (320). A number of L-2-hydroxyacids and 2-keto acids have been shown to be competitive inhibitors. Oxalate, cyanide, o-phenanthro-line, and EDTA are also potent inhibitors (320, 321, 323, 324). The inhibition by metal chelators develops slowly and is reversed by addition of Zn, Co, Mn +, or Fe + (320, 323-326). Substrates prevent the inhibition by chelators at concentrations considerably lower than their respective Km values (327). It has been suggested that EDTA inactivation involves the removal of a metal, most probably Zn +, from the substrate binding site of the enzyme (325, 326, 328, 329). However, others have... 

Lactate dehydrogenase (LDH) catalyzes the interconversion of the hydroxy-acid lactate and the keto-acid pyruvate with the coenzyme nicotinamide adenine dinucleotide [48]. This enzyme plays a fundamental role in respiration, and multiple isozymes have evolved to enable eflRcient production of substrate appropriate for the microenvironment [49]. Two main subunits, referred to as heart and muscle (skeletal), are combined in the functional enzyme as a tetramer to accommodate aero-... 

The usefulness of a mutant dehydrogenase was demonstrated in a practical synthesis of 4-amino-2-hydroxy acids, which themselves are valuable as y-turn mimics for investigations into the secondary structure of peptides[146]. Chemoenzymatic synthesis of these compounds were achieved by lipase catalyzed hydrolysis of a a-keto esters to the corresponding a-keto acids followed by reduction employing a lactate dehydrogenase in one pot. Wild type lactate dehydrogenase from either Bacillus... 

Kinetic resolutions have a maximum yield of only 50%. Therefore, a second enzymatic process was added after completion of the glycolate oxidase-catalyzed kinetic resolution[131). By addition of D-lactate dehydrogenase (E.C. 1.1.1.28) together with formate dehydrogenase for NADH regeneration, enantiospecific reduction of the 2-keto acid was achieved. Overall, a quantitative transformation (deracemization) of the racemic 2-hydroxy acid into the corresponding (R)-2-hydroxy acid was achieved (Fig. 16.2-29). 

Lactate dehydrogenases use several 2-hydroxy and 2 keto acids and other analogs as substrates 75-79). With the rabbit muscle enzyme (77),... 

Lactate dehydrogenase reduces an a-keto acid to a hydroxy acid with NADH. The enzyme will, however, also oxidize glyoxalate (XXIII) to... 

The simplest way of regenerating NAD is simply to transfer the electrons to the keto group of pyruvate, yielding lactate, in the reaction catalyzed by lactate dehydrogenase. This reaction takes place in animal cells, especially muscle cells, and is carried out by lactic acid bacteria in the fermentation of milk to yogurt. 

Williams, J.C. McDermott, A.E. Dynamics of the flexible loop of triose-phosphate isomerase—The loop motion is not ligand-gated. Biochemistry 1995, 34. 8309-8319. Holbrook, J.J. Liljas, A. Steindel, S.J.. et al. Lactate dehpdrogenase. Enzymes 1975,11, 191-292. 3rd ed. Wilks, H.M. Halsall. D.J. Atkinson, T., et al. Designs for a broad substrate-specificity keto acid dehydrogenase. Biochemistry 1990, 29. 8587-8591. 

With the successful development of an isobutanol production pathway in E. coli, the a-keto-acid isobutanol production pathway was transferred to the traditional amino acid producer Corynebacterium glutamicum [89]. The native ilvCD and adhA genes were overexpressed, which led to 2.6 g 1 of isobutanol production with other alcohols as byproducts. By deleting the competing pathways, namely pyruvate carboxylase and lactate dehydrogenase, isobutanol production was increased to 4.9 g 1 . The work showed the potential of both the universality of the a-keto-acid isobutanol production pathway and the potential of utilization of C. glutamicum for higher chain alcohol production [89]. 

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