Muscles malate dehydrogenase

Mitochondrial L-malate dehydrogenase (bovine heart muscle)1501... 

Many of the biochemical processes that generate chemical energy for the cell take place in the mitochondria. These organelles contain the biochemical equipment necessary for fatty acid oxidation, di- and tricarboxylic acid oxidation, amino acid oxidation, electron transport, and ATP generation. In this experiment, a mitochondrial fraction will be isolated from beef heart muscle. The mitochondria will be analyzed for protein content and fractionated into submitochondrial particles. Each fraction will be analyzed for malate dehydrogenase and monoamine oxidase activities. 

Fig. 42.9. Metabolism of the carbon skeletons of BCAA in skeletal muscle. 1. The first step in the metabolism of BCAA is transamination (TA). 2. Carbon from valine and isoleucine enters the TCA cycle as succinyl CoA and is converted to pyruvate by decarboxylating malate dehydrogenase (malic enzyme). 3. The oxidative pathways generate NADH and FAD(2H) even before the carbon skeleton enters the TCA cycle. The rate-limiting enzyme in the oxidative pathways is the a-keto acid dehydrogenase complex. The carbon skeleton also can be converted to glutamate and alanine, shown in blue.

This enzyme is a pyridoxal protein which is present in excessive amounts in blood serum during diseases of the liver or cardiac muscle. It may be assayed by an ultra-violet method using malate dehydrogenase in an indicator reaction [274]. Alternately, the oxalacetate formed is decomposed to pyruvate which is treated with DNP to form pyruvate-dinitrophenylhydrazone. In the presence of sodium hydroxide, an intense brown colour is produced with an absorption maximum at 505 nm [275]. 

All the polymeric NAD(P)+-derivatives have been checked for their cofactor activity and compatibility with enzymatic biocatalytic processes. The polymer-linked NAD(P)" -derivatives were associated with NAD(P) -dependent enzymes such as AlDH [281, 282, 291, 292], lactate dehydrogenase [286, 292], malate dehydrogenase [288, 292] and aldehyde dehydrogenase [287]. It was found that different NAD(P)+-polymers are active as cofactors towards different enzymes. For example, polyethyleneimine and polylysine bound NAD+-derivative revealed 60% and 25% activity, respectively, as compared with the native NAD+ in the presence of rabbit muscle lactate dehydrogenase, but only minute activity (ca. 2-7%) in the presence of alanine dehydrogenase from Bacillus suhtilis [281]. A comparative study of the cofactor activity with different enzymes is a subject of great interest. Even though several studies [279] attempt to predict the structural/functional relationship for the polymer-bound NAD(P)+-derivatives,... 

The malate—aspartate shuttle is the mechanism by which electrons from NADH produced in the cytosol are transported into mitochondria, as the inner membrane is impermeable to NADH itself. Oxaloacetate is reduced to malate in the cytosol by malate dehydrogenase, in the process oxidizing NADH to replenish cytosolic NAD. The malate-aspartate shuttle is found mainly in cardiac muscle and liver cells, while the glycerol 3-phosphate shuttle operates mainly in brain and skeletal muscle cells. Once malate has entered the mitochondria it is oxidized to oxaloacetate, generating NADH within the mitochondrial matrix. Oxaloacetate is then converted to aspartate, which is transported out of the mitochondria in exchange for glutamate. 

Mitochondria from body wall muscle and probably the pharynx lack a functional TCA cycle and their novel anaerobic pathways rely on reduced organic acids as terminal electron acceptors, instead of oxygen (Saz, 1971 Ma et al, 1993 Duran et al, 1998). Malate and pyruvate are oxidized intramitochondrially by malic enzyme and the pyruvate dehydrogenase complex, respectively, and excess reducing power in the form of NADH drives Complex II and [3-oxidation in the direction opposite to that observed in aerobic organelles (Kita, 1992 Duran et al, 1993 Ma et al,... 

Both are abundant in skeletal muscle, myocardium, liver, and erythrocytes, so that hemolysis must be avoided, and in serum they may be assayed spectrophotometrically by their conversion of phosphate-buffered pyruvate to lactate (R6, W16) or oxalacetate to malate (S25) at the expense of added NADH2, when the rate of decrease of optical density at 340 m x thus measmes the serum activities of the respective enzymes. Recently, however, the reverse reaction has been found best for serum lactic dehydrogenase assay (A2a). In conventional spectrophotometric units the normal ranges are 100-600 units per ml for lactic dehydrogenase (W16) and 42-195 xmits per ml for malic dehydrogenase (S25) as before, one conventional spectrophotometric unit per ml = 0.48 pmoles/ minute/liter (W13). 

FIG. 4.2 Malate metabolism in mitochondria from body wall muscle of adult Ascaris smm. (1) Fumarase (2) malic enzyme (3) pyruvate dehydrogenase complex (4) complex I (5) succinate-coenzyme Q reductase (complex II, fumarate reductase) (6) acyl CoA transferase (7) methylmalonyl CoA mutase (8) methyl-malonyl CoA decarboxylase (9) propionyl CoA condensing enzyme (10) 2-methyl acetoacetyl CoA reductase (11) 2-methyl-3-oxo-acyl CoA hydratase (12) electron-transfer flavoprotein (13) 2-methyl branched-chain enoyl CoA reductase (14) acyl CoA transferase. 

Go-dehydrogenase I, co-enzyme I or co-zymase, obtainable from extracts of yeast, muscle, liver and kidney, is the co-enzyme of sugar fermentation, and acts with the dehydrogenases of hexose diphosphate, malate, and alcohol, and also with the lactate dehydrogenase of muscle. 

E.g.—(a) Co-dehydrogenase I systems dehydrogenases of malate, fumarate, glucose (liver), and lactate (muscle). (6) Co-dehydrogenase n systems dehydrogenase of hexose phosphate (yeast and red blood cells). 

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