Evidence that platelet glycoprotein IIIa has a large disulfide-bonded loop that is susceptible to proteolytic cleavage.
The proteolytic digestion of GPIIIa on intact platelets by chymotrypsin, thrombin, plasmin, trypsin, and staphylococcal V8 protease was monitored in immunoblot studies employing three different antibodies to GPIIIa, one of which was made against a 13-residue synthetic peptide containing the amino terminus of GPIIIa. Chymotrypsin, plasmin, and trypsin gave similar patterns, from which it could be inferred that the major products after extensive digestion were two-chain molecules composed of amino-terminal fragments of Mr approximately 17,000-18,000 disulfide bonded to carboxyl-terminal remnants of Mr approximately 58,000-70,000. These patterns suggest that GPIIIa contains a large disulfide-bonded loop of at least 325 amino acids that is susceptible to proteolytic cleavage, and that the 4 cysteine residues between residues 177 and 273 bond with each other. Such a structure can also account for the presence of the PIA1 epitope, which has recently been localized to a polymorphism at position 33 on these late digestion products. Thrombin did not proteolyze GPIIIa even at 2.5 units/ml. Still to be resolved is whether the minor immunoreactive GPIIIa bands found on normal platelets are related to in vivo or in vitro proteolysis and whether GPIIIa proteolysis plays a role in chymotrypsin-induced exposure of the GPIIb/IIIa receptor.
Beer
J
J
Department of Medicine, State University of New York, Stony Brook 11794-8151.
Coller
B S
BS
D001792:Blood Platelets / Q000378:metabolism*; D002918:Chymotrypsin / Q000378:metabolism; D004220:Disulfides / Q000378:metabolism*; D005341:Fibrinolysin / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D008970:Molecular Weight; D010446:Peptide Fragments / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism*; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D011480:Protease Inhibitors / Q000494:pharmacology; D011487:Protein Conformation; D012697:Serine Endopeptidases / Q000378:metabolism; D013917:Thrombin / Q000378:metabolism; D014357:Trypsin / Q000378:metabolism
D004220:Disulfides; D010446:Peptide Fragments; D010980:Platelet Membrane Glycoproteins; D011480:Protease Inhibitors; D010447:Peptide Hydrolases; D012697:Serine Endopeptidases; D002918:Chymotrypsin; C028344:glutamyl endopeptidase; D014357:Trypsin; D013917:Thrombin; D005341:Fibrinolysin