Biochemical characterization of copine: a ubiquitous Ca2+-dependent, phospholipid-binding protein.
The copines are a novel group of Ca(2+)-dependent, phospholipid-binding proteins first isolated from Paramecium tetraurelia [Creutz, C. E., et al. (1998) J. Biol. Chem. 273, 1393-1402] and found in a wide range of organisms, from plants to humans. They have a Ca(2+) and phospholipid-binding domain consisting of two C2 domains and a core domain in the C-terminal portion that is homologous to the A domain found in certain integrins. We provide here the first description of the properties and distribution of a native mammalian copine, copine I. This protein is expressed in all major adult rat organs as demonstrated by probing Western blots of rat organ homogenates with anticopine antibodies. The highest levels of copine are found in the spleen. A protocol for purifying copine to homogeneity from bovine spleen is described. Purified native copine is a 58 kDa monomer that exhibits Ca(2+) self-association to form higher-order multimers, and Ca(2+)-dependent, phospholipid binding activity with preference for negatively charged phospholipids over neutral phospholipids and selectivity for Ca(2+) over Mg(2+). Half-maximal association with vesicles enriched in phosphatidylserine occurs at Ca(2+) concentrations between 1 and 10 microM. Copine I exhibits Mn(2+) binding activity that is strongly competed by Mg(2+) and partially competed by Ca(2+), suggesting that the copine I A domain may be a functional MIDAS metal binding site similar to that found in integrins [Lee, J. O., et al. (1995) Cell 80, 631-638]. Roles for copine in binding membranes and target proteins or small molecules are discussed.
Tomsig
J L
JL
Department of Pharmacology, University of Virginia, Charlottesville, Virginia 22908, USA.
Creutz
C E
CE
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000378:metabolism; D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D002413:Cations, Divalent; D002417:Cattle; D005260:Female; D006801:Humans; D016023:Integrins / Q000737:chemistry / Q000378:metabolism; D008081:Liposomes / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008297:Male; D008670:Metals / Q000378:metabolism; D008969:Molecular Sequence Data; D009928:Organ Specificity; D010743:Phospholipids / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D017208:Rats, Wistar; D013154:Spleen / Q000378:metabolism
D002352:Carrier Proteins; D002413:Cations, Divalent; D016023:Integrins; D008081:Liposomes; D008670:Metals; D010743:Phospholipids; C110203:copine; D000255:Adenosine Triphosphate; D002118:Calcium