Author:
HOUGH SHELLEY R.,PARKS JOHN E.
Abstract
ABSTRACT: Platelet‐activating factor (PAF) is a potent lipid mediator that is inactivated by platelet‐activating factor acetylhydrolase (PAF‐AH). Platelet‐activating factor bioactivity has been detected in bovine sperm phospholipids and PAF‐AH activity is extraordinarily high in bovine seminal plasma. The purpose of this study was to purify and characterize partially the PAF‐AH in bovine seminal plasma. Platelet‐activating factor acetylhydrolase was partially purified from bovine seminal plasma using gelatin—agarose and ion‐exchange chromatography and nondenaturing polyacrylamide gel electrophoresis (PAGE). Enzyme activity was increased 11‐fold over seminal plasma with a yield of 11%. Platelet‐activating factor acetylhydrolase activity was eluted from a single band with a Rf of 0.258 from a nondenaturing preparative PAGE gel along with several other proteins of varying molecular weights. Following separation by sodium dodecyl sulfate (SDS)‐PAGE under reducing conditions, PAF‐AH was identified as a ∼60‐kD band by western blotting using antiserum directed against human blood PAF‐AH. N‐terminal sequencing of the ∼60 kD band, followed by amino acid‐sequence similarity searching, demonstrated a single‐sequence match with PAF‐AH from bovine blood. Based on western blotting, a ∼60‐kD band corresponding to PAF‐AH was detected in seminal vesicle fluid but not in samples of washed, sonicated sperm or sperm plasma membranes where activity was low (<5% and <0.3%, respectively, of that in seminal plasma), suggesting that seminal plasma PAF‐AH does not bind tightly to sperm. Specific PAF‐AH activity measured in seminal vesicle fluid was in the lower range of that in seminal plasma. These results demonstrate that PAF‐AH activity in bovine seminal plasma is due to PAF‐AH secreted by the seminal vesicles with sequence homology to the enzyme in human blood.
Subject
Urology,Endocrinology,Reproductive Medicine,Endocrinology, Diabetes and Metabolism
Cited by
1 articles.
订阅此论文施引文献
订阅此论文施引文献,注册后可以免费订阅5篇论文的施引文献,订阅后可以查看论文全部施引文献