Lactic dehydrogenase and cytochrome b2 of baker's yeast. Purification and crystallization

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Abstract
Lactic dehydrogenase was partially purified from extracts of baker''s yeast by selective heat-denaturation of proteins, salting-out and adsorption and elution from calcium phosphate gel. A close relationship between the enzymic activity and the concentration of cytochrome b2 (Bach et al. 1946) was observed. The cytochrome b2 and lactic dehydrogenase activity were purified together by treatment of air-dried yeast with butan-1-ol, fractionation of butanol-saturated extracts of yeast in sodium lactate with acetone at low temperature and crystallization by anaerobic dialysis at 0 against 0.05[image]-sodium lactate-0.01 m[image]-ethylenediamine-tetra-acetate, at pH 6.8. The coral-pink crystals of the flavohaemoprotein (Appleby and Morton, 1954) probably belong to a tetragonal system. The lactic dehydrogenase activities of the crystals, expressed as [mu]moles of hydrogen acceptor reduced/hour/ mg of protein at 20[degree] were as follows: with 0.5 m[image]-potassium ferricyanide, 7750; with 0.05 m[image]-heart-muscle cytochrome c, 5870; with 0.08 m[image]-methylene blue, 2025. These activities are substantially greater than those described for yeast lactic dehydrogenase as purified by other workers.