Purification and properties of rats stomach kallikrein

Abstract

Kallikrein (EC 3.4.21.8) was purified from rat stomach by column chromatography on p- aminobenzamidine-Sepharose , DEAE-Sephadex A-50 and Sephadex G-150 and by isoelectric focusing, measuring its activities to hydrolyse l-prolyl- l-phenylalanyl- l-arginine-4-methyl-coumaryl-7-amide and to release kinin from heat-treated rat plasma. The purified stomach kallikrein showed a single band on polyacrylamide gel electrophoresis at pH 7.0. Its molecular weight was calculated to be 29 000 by gel-filtration on a column of Sephadex G-50. The kallikrein was stable between pH 6–11 and hydrolyzed l-prolyl- l-phenylalanyl- l-arginine-4-methyl-coumaryl-7-amide optimally at pH 11.0. The l-prolyl- l-phenylalanyl- l-arginine-4-methyl-coumaryl-7-amide hydrolyzing activity of rat stomach kallikrein was inhibited by diisopropyl fluorophosphate and Trasylol, but not by trypsin inhibitors from soybean, lima bean and ovomucoid. These properties of rat stomach kallikrein are different from those of partially purified rat plasma kallikrein, but similar to those of grandular kallikreins from other species. From these results, it was concluded that kallikrein is present in rat stomach and that it can be classified as a glandular kallikrein.