title: Disulfide bond reduction in fibrinogen: calcium protection and effect on clottability.
abstract: Fibrinogen contains 29 disulfide bonds. Limited reduction in buffers containing calcium led to cleavage of three of them: the two A alpha 442Cys-A alpha 472Cys intrapeptide disulfide bonds and the symmetrical A alpha 28Cys-A alpha 28Cys bond. The limited reduction did not affect clotting by thrombin. However, a prolongation of the thrombin clotting time occurred when the limited reduction took place in the absence of calcium. The bonds reduced under this condition included the three already mentioned and also the two gamma 326Cys-gamma 339Cys intrapeptide disulfide bonds located in the C-terminal ends of the gamma-chain. N-Terminal analysis of thrombin-treated samples showed that thrombin cleavage occurred at the normal A alpha 16-A alpha 17 site in fibrinogen that was partially reduced in the presence of calcium. By contrast, thrombin cleaved at the A alpha 19-A alpha 20 site in fibrinogen that was partially reduced in the absence of calcium, rendering the protein unclottable by removing the A alpha 17Gly-18Pro-19Arg peptide. The loss of thrombin clottability may have also come from gamma 326Cys-gamma 339Cys disulfide bond reduction since the structure supported by this bond may be important for the function of the C-terminal polymerization site. In samples of the partially reduced fibrinogen lacking the A alpha 17-19 residues, gel formation occurred through an oligomerization mechanism catalyzed by factor XIII.
authors.affiliation: Blood Coagulation Biochemistry, New York Blood Center, New York 10021.
authors.forename: R
authors.initials: R
authors.lastname: Procyk
authors.forename: B
authors.initials: B
authors.lastname: Blombäck
mesh_terms: D000478:Alkylation; D000595:Amino Acid Sequence; D002118:Calcium / Q000494:pharmacology*; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002851:Chromatography, High Pressure Liquid; D004220:Disulfides / Q000378:metabolism*; D005340:Fibrinogen / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D013917:Thrombin / Q000378:metabolism
chemical_list: D004220:Disulfides; D005340:Fibrinogen; D013917:Thrombin; D002118:Calcium