Further Studies on Elastase-Like Esterases in Human Leukocyte Granules 1

Abstract

Summary

1. Human peripheral blood leukocyte granules contain a neutral esterase which hydrolyzes t-BOC-l-alanine p-nitrophenol, a synthetic substrate of elastase. The alanine p-nitrophenyl esterase was identified following isoelectric focusing of granule proteins in acrylamide gel and sucrose density-gradient systems. The enzyme proved highly basic, focusing at pH 9.4 in acrylamide gel and at pH 10.8 in the sucrose gradient.

2. At pH 8.0, preparations of human leukocyte granules also actively hydrolyzed N-acetyl-l-alanyl-l-alanyl-l-alanine methyl ester, a second highly specific elastase substrate, releasing 1 mole of acid/mole of substrate. In this reaction, the activity of 10 μg of crude granule extract protein corresponded to that of 1 μg of crystalline pancreatic elastase.

3. Products of hydrolysis of both esters; namely, t-BOC-l-alanine and N-acetyl-l-alanyl-l-alanyl-l-alanine, were shown to inhibit the digestion of denatured hemoglobin at neutral pH by granule extract suggesting that part of the neutral protease activity of human leukocyte granules may be attributed to the elastase-like esterase(s).