Orientation of the Carboxy-terminal Regions of Fibrin γ Chain Dimers Determined from the Crosslinked Products Formed in Mixtures of Fibrin, Fragment D, and Factor XIIIa

Kevin R Siebenlist; David A Meh; Joseph S Wall; James F Hainfeld; Michael W Mosesson

doi:10.1055/s-0038-1649890

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 1995; 74(04): 1113-1119
DOI: 10.1055/s-0038-1649890

Original Article

Coagulation

Schattauer GmbH Stuttgart

Orientation of the Carboxy-terminal Regions of Fibrin γ Chain Dimers Determined from the Crosslinked Products Formed in Mixtures of Fibrin, Fragment D, and Factor XIIIa

Authors

Kevin R Siebenlist

¹The University of Wisconsin Medical School Sinai Samaritan Medical Center, Milwaukee, WI, USA

²The Department of Basic Health Sciences, School of Dentistry, Marquette University, Milwaukee, WI, USA
David A Meh

¹The University of Wisconsin Medical School Sinai Samaritan Medical Center, Milwaukee, WI, USA
Joseph S Wall

³The Brookhaven National Laboratory, Biology Department, Upton, NY, USA
James F Hainfeld

³The Brookhaven National Laboratory, Biology Department, Upton, NY, USA
Michael W Mosesson

¹The University of Wisconsin Medical School Sinai Samaritan Medical Center, Milwaukee, WI, USA

Abstract

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Summary

There are two schools of thought regarding the orientation of the intermolecular ∈-amino-(γ-glutamyl) lysine isopeptide bonds formed between γ chains in the D domains of assembled fibrin fibers. Some investigators believe that these bonds are oriented parallel to the direction of fiber growth (longitudinally) at the contacting ends of fibrin D domains (‘DD-long’), whereas others believe that these bonds are oriented across the two-stranded fibril, between D domains in opposing strands (‘DD-transverse’). To distinguish between these two possibilities, the structure of crosslinked products formed in mixtures of fibrin, plasmic fragment D, and factor XIIIa were analyzed, based upon this rationale: Complex formation between D fragments and a fibrin template depends upon the non-covalent ‘D:E’ interaction between each fibrin E domain and two D fragments (‘D:fibrin:D’). If carboxy-terminal γ chains in the D:fibrin:D complex become aligned in a DD-long configuration, only crosslinked fragment D dimers (‘D-D’) will result and the fibrin ‘template’ will not become crosslinked to the associated D fragments. If instead, γ chain crosslinks form transversely between the D fragments and fibrin, covalently linked D-fibrin complexes will result.

SDS-PAGE of factor XIIIa crosslinked mixtures of fibrin and fragment D demonstrated products of a size and subunit composition indicating D-fibrin and D-fibrin-D formation. Small amounts of D dimers were also formed at the same levels as were formed in mixtures of fragment D and factor XIIIa alone. Electron microscopic images of D-fibrin-D complexes prepared under physiological buffer conditions demonstrated that the D fragments were associated with the central E domain of the fibrin molecule, but that they could be dissociated from this non-covalent association in 2% acetic acid. These findings indicate that γ chain crosslinks occur transversely in D:fibrin:D complexes and permit the extrapolated conclusion that γ chain crosslinks are also positioned transversely in an assembled fibrin polymer.

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References

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