Cloning, characterization, and functional studies of a nonintegrin platelet receptor for type I collagen

Thomas M. Chiang, Augustinus Rinaldy, Andrew Kang

Research output: Contribution to journalArticle

69 Citations (Scopus)

Abstract

A cDNA (1.6 kb) encoding a platelet protein receptor that binds type I collagen has been isolated from a human bone marrow cDNA library by using a degenerate oligonucleotide probe derived from the amine acid sequence of a CNBr fragment of the purified receptor. Computer search revealed that this cDNA represents the coding sequence era unique protein. Using the prokaryotic expression system pKK 223-3-65 cDNA, a 54-kD recombinant protein was obtained and purified to apparent homogeneity. In an eukaryotic expression vector (pcDNA3-65 cDNA), a 65-kD protein was identified that was recognized by monoclonal anti-65 kD antibody (anti-65m). The recombinant protein binds to type I, but not to type III collagen by affinity column chromatography. The binding of the recombinant protein to type I collagen-coated Petri dishes is inhibited by anti-65m in a dose-dependent manner. The pcDNA3-65 cDNA- transfected nonadherent T cells express the protein, allowing them to attach to a type I collagen matrix, and are inhibited by anti-65m in a dose- dependent manner. Like the receptor protein purified from platelet membranes, the recombinant protein inhibits type I collagen-induced platelet aggregation and the adhesion of [14C]serotonin-labeled platelets to type I collagen in a dose-dependent manner. The recombinant protein neither binds to type III collagen-coated Petri dishes nor inhibits type III collagen and ADP-induced platelet aggregation, indicating specificity for type I collagen.

Original languageEnglish (US)
Pages (from-to)514-521
Number of pages8
JournalJournal of Clinical Investigation
Volume100
Issue number3
DOIs
StatePublished - Aug 1 1997

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Collagen Type I
Recombinant Proteins
Organism Cloning
Complementary DNA
Collagen Type III
Blood Platelets
Proteins
Platelet Aggregation
Oligonucleotide Probes
Gene Library
Affinity Chromatography
Adenosine Diphosphate
Amines
Anti-Idiotypic Antibodies
Serotonin
Carrier Proteins
Membrane Proteins
Bone Marrow
type I collagen receptor
T-Lymphocytes

All Science Journal Classification (ASJC) codes

  • Medicine(all)

Cite this

Cloning, characterization, and functional studies of a nonintegrin platelet receptor for type I collagen. / Chiang, Thomas M.; Rinaldy, Augustinus; Kang, Andrew.

In: Journal of Clinical Investigation, Vol. 100, No. 3, 01.08.1997, p. 514-521.

Research output: Contribution to journalArticle

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