Lactic dehydrogenase and cytochrome b2 of baker's yeast. Enzymic and chemical properties of the crystalline enzyme

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Abstract
Chemical and enzymic studies with crystalline yeast lactic dehydrogenase are described. The absorption spectrum of the lactate-reduced enzyme shows maxima at 556.5, 528, 424, 330 and 265 m[mu]; [epsilon] at 556.5 m[mu] is 38.8 X 103. There is 1 mole each of protohaem and of riboflavin phosphate present as prosthetic groups/80 000 g of enzyme. No significant amount of non-haem iron, copper, cobalt, molybdenum or other related metal is present. There is 1 g atom of mg/80,000 g of enzyme; this is probably associated with the deoxyribopolynucleotide component of the crystalline material. Oxidation by hydrogen peroxide, and by added Cu2+ ions under aerobic conditions, causes dissociation of the flavin prosthetic group and loss of dehydrogenase activity. Ethylenediaminetetraacetate protects against metal-catalyzed inactivation. The mechanism of the inactivation process is discussed. The term cytochrome b2 applies to the native, enzymically active flavohaemoprotein. A haemoprotein product containing no riboflavin phosphate and devoid of enzymic activity may be obtained from cytochrome b2. The enzyme catalyses the oxidation of glycollate, L-lactate and [alpha]-hydroxybutyrate only. Lactate is oxidized most rapidly. The activation energies for reduction of heart-muscle cytochrome c, potassium ferricyanide and methylene blue under specific conditions are 7[center dot]2, 7[center dot]6 and 8[center dot]7 kcal./mole, respectively. Under specified conditions, reduction of heart-muscle cytochrome c proceeds by a 1st-order reaction, and of ferricyanide by a zero-order reaction. The enzymic and chemical properties of the crystalline cytochrome b2 are compared with those of non-crystalline preparations obtained by other workers.