Rapid purification and partial characterization of human platelet glycoprotein IIIb. Interaction with thrombospondin and its role in platelet aggregation.
Glycoprotein IIIb (also known as glycoprotein IV) is a major glycoprotein present on the surface of human platelets. Recent studies suggest that glycoprotein IIIb may be a receptor site for thrombospondin. Thrombospondin, a multifunctional adhesive glycoprotein released from stimulated platelets, plays an important role in the stabilization of platelet aggregates. In this study, a new method for the purification of glycoprotein IIIb is described. Glycoprotein IIIb was isolated from Triton X-114 platelet membrane extracts, under nondenaturing conditions, by tandem anion-exchange and size exclusion fast protein liquid chromatography. The purified glycoprotein had the same apparent molecular mass (88 kDa) under nonreducing or reducing conditions. The tryptic peptide map of the purified protein was identical to that of bona fide glycoprotein IIIb as isolated from two-dimensional polyacrylamide gels of platelet membrane proteins. In addition, the purified glycoprotein was recognized by an anti-GPIIIb monoclonal antibody (OKM5). The purified glycoprotein specifically bound to thrombospondin in the presence of calcium. Monospecific anti-GPIIIb antibodies interfered with the expression of endogenous thrombospondin on thrombin-activated platelets and partially inhibited collagen- and thrombin-induced platelet aggregation without a significant effect on platelet secretion. Glycoprotein IIIb, by interacting with thrombospondin on the activated platelet surface, may play an important role in the platelet aggregation process.
McGregor
J L
JL
Université Claude Bernard Lyon I, Faculté de Médecine Alexis Carrel, Institut National de la Santé et de la Recherche Médicale U63-Laboratoire d'Hémobiologie, Lyon, France.
Catimel
B
B
Parmentier
S
S
Clezardin
P
P
Dechavanne
M
M
Leung
L L
LL
D001792:Blood Platelets / Q000502:physiology*; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D006023:Glycoproteins / Q000378:metabolism*; D006801:Humans; D007140:Immunoglobulin Fab Fragments; D007700:Kinetics; D010446:Peptide Fragments / Q000032:analysis; D010974:Platelet Aggregation; D010980:Platelet Membrane Glycoproteins / Q000302:isolation & purification* / Q000502:physiology; D019699:Thrombospondins; D014357:Trypsin
D006023:Glycoproteins; D007140:Immunoglobulin Fab Fragments; D010446:Peptide Fragments; D010980:Platelet Membrane Glycoproteins; D019699:Thrombospondins; D014357:Trypsin