Roles of Trp31 in high membrane binding and proinflammatory activity of human group V phospholipase A2.
Group V phospholipase A2 is a recently discovered secretory phospholipase A2 (PLA2) that has been shown to be involved in eicosanoid formation in inflammatory cells, such as macrophages and mast cells. We have demonstrated that human group V PLA2 (hsPLA2-V) can bind phosphatidylcholine (PC) membranes and hydrolyze PC substrates much more efficiently than human group IIa PLA2, which makes it better suited for acting on the outer plasma membrane (Han, S.-K., Yoon, E. T., and Cho, W. (1998) Biochem. J. 331, 353-357). In this study, we demonstrate that exogenous hsPLA2-V has much greater activity than does group IIa PLA2 to release fatty acids from various mammalian cells and to elicit leukotriene B4 formation from human neutrophils. To understand the molecular basis of these activities, we mutated two surface tryptophans of hsPLA2-V to alanine (W31A and W79A) and measured the effects of these mutations on the kinetic activity toward various substrates, on the binding affinity for vesicles and phospholipid-coated beads, on the penetration into phospholipid monolayers, and on the activity to release fatty acids and elicit eicosanoid formation from various mammalian cells. These studies show that the relatively high ability of hsPLA2-V to induce cellular eicosanoid formation derives from its high affinity for PC membranes and that Trp31 on its putative interfacial binding surface plays an important role in its binding to PC vesicles and to the outer plasma membrane.
Han
S K
SK
Department of Chemistry, University of Illinois, Chicago, Illinois 60607, USA.
Kim
K P
KP
Koduri
R
R
Bittova
L
L
Munoz
N M
NM
Leff
A R
AR
Wilton
D C
DC
Gelb
M H
MH
Cho
W
W
D002462:Cell Membrane / Q000201:enzymology; D015777:Eicosanoids / Q000378:metabolism; D005227:Fatty Acids / Q000378:metabolism; D006801:Humans; D018836:Inflammation Mediators / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008958:Models, Molecular; D016296:Mutagenesis; D010741:Phospholipases A / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D054467:Phospholipases A2; D013379:Substrate Specificity; D014364:Tryptophan / Q000378:metabolism*
D015777:Eicosanoids; D005227:Fatty Acids; D018836:Inflammation Mediators; D014364:Tryptophan; D010741:Phospholipases A; D054467:Phospholipases A2