Immunochemical mapping of the conformation of human fibrinogen. The gamma 95-264 segment in inaccessible to antibody in native fibrinogen but progressively exposed by plasmic cleavage.
The accessibility of the gamma 95-264 sequence to specific antibody probes in the native fibrinogen molecule and its plasmic cleavage fragments have been investigated. The gamma 95-264 segment was generated by cyanogen bromide cleavage of the gamma chain and isolated by gel filtration and ion exchange chromatography. Rabbit antisera to this peptide and to gamma chain recognized at least five antigenic loci uniformly distributed throughout this segment. In primary binding assays, antibodies to gamma 95-264 bound gamma 95-264, free gamma chain, and fibrinogen fragment D, but not native fibrinogen. Also, gamma 95-264 was bound by antibodies to gamma chain and fibrinogen fragment D, but not by antibodies generated to native fibrinogen. Thus, the gamma 95-264 sequence was not accessible to antibody in the native structure. In competitive equilibrium radioimmunoassays, neither native fibrinogen nor highly soluble fibrinogen fraction I-9 inhibited the binding of gamma 95-264 by its antiserum or anti-gamma chain. With plasmic cleavage, however, the gamma 95-264 sequence became accessible to antibody and the series of fragments D greater than Y greater than D:E = X describes the relative reactivity of the gamma chain sequence in fibrinogen degradation products. Differential expression of gamma 95-264 antigenic loci was also observed with D fragments differing in molecular weight. Plasmic cleavage of cross-linked and noncross-linked fibrin generated D fragments which did not express gamma 95-264 as well as fibrinogen D derivatives, indicating that the D domains of fibrinogen and fibrin are immunochemically distinguishable. These findings indicate that the central segment of the gamma chain is inaccessible to antibody in native fibrinogen, but that proper surface orientation is achieved upon plasmic degradation.
D000906:Antibodies; D000936:Antigen-Antibody Complex; D003488:Cyanogen Bromide; D000939:Epitopes; D005337:Fibrin / Q000276:immunology; D005338:Fibrin Fibrinogen Degradation Products / Q000276:immunology; D005340:Fibrinogen / Q000276:immunology; D006801:Humans; D010446:Peptide Fragments / Q000032:analysis; D011863:Radioimmunoassay
D000906:Antibodies; D000936:Antigen-Antibody Complex; D000939:Epitopes; D005338:Fibrin Fibrinogen Degradation Products; D010446:Peptide Fragments; C031564:fibrinogen (gamma 95-264); D005337:Fibrin; D005340:Fibrinogen; D003488:Cyanogen Bromide