Catalytic mechanism and kinetics of malate dehydrogenase

Abstract

Malate dehydrogenase (MDH) is a ubiquitous and central enzyme in cellular metabolism, found in all kingdoms of life, where it plays vital roles in the cytoplasm and various organelles. It catalyzes the reversible NAD+-dependent reduction of L-malate to oxaloacetate. This review describes the reaction mechanism for MDH and the effects of mutations in and around the active site on catalytic activity and substrate specificity, with a particular focus on the loop that encloses the active site after the substrates have bound. While MDH exhibits selectivity for its preferred substrates, mutations can alter the specificity of MDH for each cosubstrate. The kinetic characteristics and similarities of a variety of MDH isozymes are summarized, and they illustrate that the KM values are consistent with the relative concentrations of the substrates in cells. As a result of its existence in different cellular environments, MDH properties vary, making it an attractive model enzyme for studying enzyme activity and structure under different conditions.

Keywords: enzyme kinetics; enzyme specificity; malate dehydrogenase; reaction mechanism.

Figure 1. Overall reaction catalyzed by malate dehydrogenase

The illustrated reaction involves the conversion of L-malate to oxaloacetate, accompanied by the reduction of NAD⁺ to NADH + H⁺. Rib: Ribose, H_S: pro-S hydrogen, H_R: pro-R hydrogen.

Figure 2. Reaction mechanism for malate dehydrogenase

The diagram on the top illustrates the mechanism of oxidation of L-malate, while the diagram on the bottom is the reverse, the reduction of oxaloacetate.

Figure 3. Free energy diagram for MDH

The reaction profile for conversion of malate to oxaloacetate is shown using thermodynamic data from [4,6].

Figure 4. Movement of active site loop upon NAD⁺ and L-malate binding to MDH

(A) Overall loop movement with open in green (4kde) and closed in cyan (6ihe). (B) Close-up of loop movement. L-Malate is shown in sticks with yellow carbons, and NAD⁺ is shown in sticks with gray carbons.