Purification and Some Properties of Soluble Phospholipase B from Baker’s Yeast (Saccharomyces cerevisiae)

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Abstract
Phospholipase B from baker’s yeast (Saccharomyces cerevisiae) was purified by acid treatment of the crude extract, ammonium sulfate fractionation, and column chromatographies on DEAE-Sepharose CL-6B, Sepharose 4B, and Bio-Gel HTP. The purified preparation had lysophospholipase activity and phospholipase B activity in a ratio of 16:1. The optimum pH of both activities was 3.5 ~ 4.0. The enzyme was a glycoprotein and its molecular size was somewhat heterogeneous, ranged from about 280,000 to 420,000 by gel filtration. Phospholipase B activity was strongly stimulated by 0.1 % DOC, but lysophospholipase activity was completely inhibited by the detergent. Neither activity was stimulated by Ca2+ and both were inhibited by SDS, Triton X-100, and Fe3+. The enzyme hydrolyzed the acyl ester bonds of phosphatidylcholine sequentially, first the 2-acyl and then the 1-acyl groups. The Km values for phosphatidylcholine and lysophosphatidylcholine were 0.63 mm and 0.05 mm, respectively.