GEL FILTRATION APPLIED TO THE STUDY OF LIPASES AND OTHER ESTERASES

Abstract

Sephadex G-100 and G-200 gel-filtration columns were calibrated for molecular-weight estimation with proteins of known molecular weights, and used to study the composition of several lipase or ester-ase preparations. Enzymes from cow''s milk, rat adipose tissue and pig pancreas were detected in the column effluents by their ability to liberate free acid from emulsified tributyrin at pH 8[center dot]5. Four tributy-rinases were detected in preparations from individual cow''s milks. Molecular weights 62,000, 75,000 and 112,000 were estimated for 3 of them, but although the 4th may be of unusually low molecular weight an estimate was not possible. Extracts of rat adipose tissue apparently contained 6 tributyrinases (molecular weights 39,000, 47,000, 55,000, 68,000, 75,000 and 200,000) but the relative amounts of these enzymes varied widely from rat to rat. Tributyrinase activity in juice expressed from pig pancreatic tissue was due mainly to 1 enzyme (molecular weight 42,000). On the other hand, activity in extracts of acetone-dried pancreas was confined to material of molecular weight > 106, which may be an aggregated form of the lower-molecular-weight enzyme. Activity in fractionated wheat-germ extracts was assayed with emulsified triacetin substrate, and was evidently due to 1 enzyme (molecular weight 51,000). Some problems arising in the application of gel filtration to the study of lipase-esterase systems were indicated.