American Chemical Society
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There are conflicting ideas regarding the location of the carboxyl-terminal regions of cross-linked γ-chain dimers in double-stranded fibrin fibrils. Some investigators believe that the chains are always oriented longitudinally along each fibril strand and traverse the contacting ends of abutting fibrin D domains (“DD-long” cross-linking). Other investigations have indicated instead that the chains are situated transversely between adjacent D domains in opposing fibril strands (transverse cross-linking). To distinguish between these two possibilities, the γ dimer composition of factor XIIIa-cross-linked fibrin/fibrinogen complexes that had been formed through noncovalent D/E interactions between fibrinogen D domains and fibrin E domains was examined. Two factor XIIIa-mediated cross-linking conditions were employed. In the first, fibrin/fibrinogen complexes were formed between 125I-labeled fibrinogen 2 (“peak 2” fibrinogen), each heterodimeric molecule containing one γA and one larger γ‘ chain, and nonlabeled fibrin 1 molecules (“peak 1” fibrin), each containing two γA chains. If DD-long cross-linking occurred, 125I-labeled γA−γA, γA−γ‘, and γ‘−γ‘dimers in a 1:2:1 ratio would result. Transverse cross-linking would yield a 1:1 mixture of 125I-labeled γA−γA and γA−γ‘ dimers, without any γ‘−γ‘ dimers. Autoradiographic analyses of reduced SDS−PAGE gels from protocol 1 revealed 125I-labeled γA−γA and γA−γ‘ dimers at a ratio of ∼1:1. No labeled γ‘−γ‘ dimers were detected. Protocol 2 used a converse mixture, 125I-fibrin 2 and nonlabeled fibrinogen 1. DD-long cross-linking of this mixture would yield only nonradioactive γA−γA dimers, whereas transverse cross-linking would yield a 1:1 mixture of 125I-labeled γA−γA and γA−γ‘ dimers. Autoradiographic analyses of this mixture yielded 125I-labeled γA−γA and γA−γ‘ dimers in a 1:1 ratio. These findings provide no evidence that longitudinal (DD-long) γ chain positioning occurs in cross-linked fibrin and indicate instead that most, if not all, γ-chain positioning in an assembled fibrin polymer is transverse.
Siebenlist, Kevin R.; Meh, David A.; and Mosesson, Michael W., "Position of γ-Chain Carboxy-Terminal Regions in Fibrinogen/Fibrin Cross-Linking Mixtures" (2000). Biomedical Sciences Faculty Research and Publications. 241.
ADA Accessible Version
Accepted version. Biochemistry, Vol. 39, No. 46 (November 1, 2000): 14171-14175. DOI. © American Chemical Society. Used with permission.