Isolation and characterization of a platelet membrane protein related to the vitronectin receptor.
Glycoprotein IIb-IIIa is the most prominent Arg-Gly-Asp (RGD)-binding adhesion receptor on platelets. By affinity chromatography on an immobilized RGD peptide, we have investigated the possible existence of other platelet-associated adhesion receptors that bind RGD peptides. When an octyl glucoside extract of surface-radioiodinated platelets was applied to an affinity matrix of KYGRGDS-coupled Sepharose 4B, a 160-kDa-labeled protein (P160) and GPIIb-IIIa bound and were specifically eluted by soluble GRGDSP peptide, but not by the variant GRGESP peptide. Furthermore, a dodecapeptide corresponding to fibrinogen gamma 400-411 eluted only GPIIb-IIIa but not P160 from the RGD affinity matrix. Characterization of P160 by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and by the O'Farrell gel electrophoresis system indicated that P160 is a component of platelet GPIc. GoH3, a monoclonal antibody recognizing the alpha subunit of the very late antigen-6, failed to immunoprecipitate P160 from the RGD eluate, indicating that it did not contain the very late antigen-6 alpha subunit. In immunoblots, P160 reacted specifically with a polyclonal anti-peptide antibody recognizing the alpha subunit of the vitronectin receptor (VnR), but not with the monoclonal anti-GPIIb antibody PMI-1, suggesting that P160 is the alpha subunit of platelet VnR. This possibility was further substantiated by the complete identity between the determined amino-terminal sequence of P160 and the known sequence of the VnR alpha subunit. Moreover, direct association of P160 with a beta subunit having an apparent molecular weight similar to that of GPIIIa was demonstrated by immunoprecipitation with LM609, an anti-VnR complex monoclonal antibody. These results indicate that the VnR complex is present on platelets and may play a functional role in platelet adhesive reactions.
Lam
S C
SC
Department of Immunology, Research Institute of Scripps Clinic, La Jolla, California 92037.
Plow
E F
EF
D'Souza
S E
SE
Cheresh
D A
DA
Frelinger
A L
AL
Ginsberg
M H
MH
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D001792:Blood Platelets / Q000032:analysis*; D015153:Blotting, Western; D002448:Cell Adhesion; D002846:Chromatography, Affinity; D006023:Glycoproteins / Q000502:physiology*; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010980:Platelet Membrane Glycoproteins / Q000302:isolation & purification* / Q000502:physiology; D011233:Precipitin Tests; D011971:Receptors, Immunologic / Q000302:isolation & purification* / Q000502:physiology; D019030:Receptors, Vitronectin; D019096:Vitronectin
D000911:Antibodies, Monoclonal; D006023:Glycoproteins; D010980:Platelet Membrane Glycoproteins; D011971:Receptors, Immunologic; D019030:Receptors, Vitronectin; D019096:Vitronectin