Purification and Some Properties of Intracellolar Esterase from Pseudomonas fluorescens

A novel esterase was found in Pseudomonas fluorescens cells and purified to homo-geneity as determined by polyacrylamide gel electrophoresis. The esterase was extracted from the cells by freeze-thawing and hypotonic treatment. Purification was achieved by ammonium sulfate precipitation, followed by successive chromatog-raphies on DEAE-cellulose and benzylamine-agarose and then electrophoresis. The enzyme catalyzed the hydrolysis of methyl esters, such as methyl butyrate, but its hydrolyzing activity decreased with increase in the chain length of the alcohol moi-ety, and it did not catalyze the hydrolysis of triacylglycerols, such as triacetin. In contrast, the enzyme acted on various acyl residues in a series of methyl esters, such as dimethyl succinate, methyl methacrylate, and dimethyl malate. The optimum pH for activity of this enzyme with methyl butyrate was 7.0-8.5. The enzyme was inhibited by phenylmethylsulfonylfiuoride. Its molecular weight was estimated as 48,000 by molecular sieve electrophoresis and gel filtration on Sephadex G-150. Carboxylesterase [carboxylic-ester hydrolase EC 3.1.1.1] has been purified from several tissues of vertebrates {1-4), but its physiological function is still obscure. We have been studying the relation between carboxylesterase and lipase [triacylglycerol acylhydrolase EC 3.1.1.3]. Various carboxyl-esterases are found in the same tissues as lipase (5-7), and our previous paper suggested that rat liver esterase and lipase may act reciprocally (5, 6). There are many reports on bacterial lipase (8-10). The lipase from Pseudomonas fluorescens has been purified, crystallized, and characterized (11, 12). We found an esterase in intact cells of the same strain from which the lipase was obtained. The present paper describes the purification and some characteristics of this esterase from P. fluo-rescens