title: Interfacial kinetic and binding properties of mammalian group IVB phospholipase A2 (cPLA2beta) and comparison with the other cPLA2 isoforms.
abstract: The cytosolic (group IV) phospholipase A(2) (cPLA(2)s) family contains six members. We have prepared recombinant proteins for human α, mouse β, human γ, human δ, human ε, and mouse ζ cPLA(2)s and have studied their interfacial kinetic and binding properties in vitro. Mouse cPLA(2)β action on phosphatidylcholine vesicles is activated by anionic phosphoinositides and cardiolipin but displays a requirement for Ca(2+) only in the presence of cardiolipin. This activation pattern is explained by the effects of anionic phospholipids and Ca(2+) on the interfacial binding of mouse cPLA(2)β and its C2 domain to vesicles. Ca(2+)-dependent binding of mouse cPLA(2)β to cardiolipin-containing vesicles requires a patch of basic residues near the Ca(2+)-binding surface loops of the C2 domain, but binding to phosphoinositide-containing vesicles does not depend on any specific cluster of basic residues. Human cPLA(2)δ also displays Ca(2+)- and cardiolipin-enhanced interfacial binding and activity. The lysophospholipase, phospholipase A(1), and phospholipase A(2) activities of the full set of mammalian cPLA(2)s were quantified. The relative level of these activities is very different among the isoforms, and human cPLA(2)δ stands out as having relatively high phospholipase A(1) activity. We also tested the susceptibility of all cPLA(2) family members to a panel of previously reported inhibitors of human cPLA(2)α and analogs of these compounds. This led to the discovery of a potent and selective inhibitor of mouse cPLA(2)β. These in vitro studies help determine the regulation and function of the cPLA(2) family members.
authors: Department of Chemistry, University of Washington, Seattle, Washington 98195, USA.
authors: Farideh
authors: F
authors: Ghomashchi
authors: Gajendra S
authors: GS
authors: Naika
authors: James G
authors: JG
authors: Bollinger
authors: Ahmed
authors: A
authors: Aloulou
authors: Matthias
authors: M
authors: Lehr
authors: Christina C
authors: CC
authors: Leslie
authors: Michael H
authors: MH
authors: Gelb
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D055162:Biocatalysis / Q000187:drug effects; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology; D054513:Group IV Phospholipases A2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis / Q000187:drug effects; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009154:Mutation; D054510:Phospholipases A2, Cytosolic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010743:Phospholipids / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D018411:Spodoptera
chemical_list: D004791:Enzyme Inhibitors; D007527:Isoenzymes; D010743:Phospholipids; D011994:Recombinant Proteins; D054513:Group IV Phospholipases A2; D054510:Phospholipases A2, Cytosolic; D002118:Calcium