Role of a membrane-associated serine esterase in the oxidant activation of phospholipase A2 by t-butyl hydroperoxide.
Exposure of bovine pulmonary-arterial endothelial cells to the oxidant lipid t-butyl hydroperoxide (t-Bu-OOH) increases cell-membrane-associated phospholipase A2 (PLA2) activity and stimulates arachidonic acid (AA) release. To test the hypothesis that a membrane-associated serine esterase plays an important role in activating PLA2, the present study was undertaken. In addition to increasing PLA2 activity and AA release, t-Bu-OOH also enhances the activity of a membrane-associated serine esterase that cleaves the synthetic substrate N alpha-p-tosyl-L-arginine methyl ester (TAME). Changes in the activity of this membrane-bound serine esterase correlate directly with changes in the activity of PLA2. Serine esterase inhibitors such as phenylmethanesulphonyl fluoride, di-isopropyl fluorophosphate and alpha 1-proteinase inhibitor, and TAME, a synthetic substrate for serine esterase, prevent the increase in serine esterase activity, PLA2 activity and AA release caused by t-Bu-OOH. Pretreatment of the endothelial cells with the antioxidant vitamin E prevents t-Bu-OOH-induced stimulation of AA release and the cell-membrane-associated serine esterase and PLA2 activities. Adding t-Bu-OOH or the serine esterase trypsin to the endothelial-cell membrane fraction also significantly augments PLA2 activity, implying that these treatments activate latent PLA2. These results suggest that t-Bu-OOH stimulates a membrane-associated serine esterase that plays a crucial role in activating PLA2 and releasing AA.
Department of Biochemistry and Biophysics, University of Kalyani, India.
S
S
Chakraborti
J R
JR
Michael
G H
GH
Gurtner
S S
SS
Ghosh
G
G
Dutta
A
A
Merker
D000595:Amino Acid Sequence; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D002417:Cattle; D002462:Cell Membrane / Q000201:enzymology; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000201:enzymology; D004789:Enzyme Activation; D004950:Esterases / Q000378:metabolism*; D008969:Molecular Sequence Data; D016877:Oxidants / Q000494:pharmacology*; D010545:Peroxides / Q000494:pharmacology*; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D013379:Substrate Specificity; D020122:tert-Butylhydroperoxide
D016877:Oxidants; D010545:Peroxides; D016718:Arachidonic Acid; D020122:tert-Butylhydroperoxide; D004950:Esterases; C022635:serine esterase; D010741:Phospholipases A; D054467:Phospholipases A2