Dihydroxyacetone Reductase from Mucor javanicus. 2. Identification of the Physiological Substrate and Reactivity towards Related Compounds

Evidence is presented that the oxidoreductase isolated from Mucor javanicus on the basis of its high activity for alicyclic substrates, particularly for trans-1,4-decalindione, is a dihydroxyacetone reductase. This identification rests mainly on the following experimental evidence. (1) From extracts of the mycelium a compound could be isolated, which was reduced at high rate in the presence of the Mucor enzyme and was shown to be dihydroxyacetone. (2) Glycerol oxidation catalyzed by the Mucor enzyme gave exclusively dihydroxyacetone. (3) Comparison of specificity data obtained with the Mucor enzyme and the known glycerol-dihydroxyacetone oxidoreductase from Aerobacter aerogenes revealed a close functional relationship between the two enzymes. (4) The specificity and stereospecificity observed with a large number of substrates was found to be in agreement with an active-site topography of an enzyme specific for dihydroxyacetone. The specificity comparisons with the Aerobacter enzyme disclosed interesting differences: the Mucor enzyme showed high reactivity towards hydrophobic substrates such as cyclohexanone, whereas the Aerobacter enzyme was completely inactive with these substrates. This difference is attributed to the fact that the two enzymes are involved in different metabolic processes: in Mucor javanicus reduction of dihydroxyacetone and in Aerobacter aerogenes oxidation of glycerol. Taking advantage of the broad specificity and the high stereospecificity the usefulness of the Mucor enzyme in preparing compounds of high enantiomeric purity was demonstrated. Since oxidation of glycerol leads to dihydroxyacetone as the sole product, it was possible to determine the equilibrium constant of the dihydroxyacetone/glycerol system.