Interaction of fibrin(ogen) with leukocyte receptor α Mβ2 (Mac-1): Further characterization and identification of a novel binding region within the central domain of the fibrinogen γ-module

Sergiy Yakovlev, Li Zhang, Tatiana Ugarova, Leonid Medved

Research output: Contribution to journalArticle

40 Scopus citations

Abstract

The fibrinogen γ-module sequences, γ190-202 or P1, and γ377-395 or P2, were implicated in interaction with the αMI-domain of the leukocyte receptor αMβ 2. P1 is an integral part of the γ-module central domain, while P2 is inserted into this domain forming an antiparallel β-strand with P1. We hypothesized earlier that separation of P2 from P1 may regulate interaction of fibrin(ogen) with leukocytes during the inflammatory response. To test the relative contributions of these sequences to the interaction and the effect of their separation, we prepared the recombinant γ-module (γ148-411) and its halves, γ148-286 and γ287-411 fragments containing P1 and P2, respectively, and evaluated their affinities for the recombinant αMI-domain. In a solid-phase binding assay, the immobilized γ-module exhibited high affinity for αMI (Kd = 22 nM), while the affinities of the isolated γ148-286 and γ287-411 halves were much lower (Kd's = 521 and 194 nM, respectively), indicating that both halves contribute to the interaction in a synergistic manner. This is consistent with the above hypothesis. Further, we prepared the recombinant γ148-191 and γ192-286 fragments corresponding to the NH2-terminal and central domains, respectively, as well as γ148-226 containing P1, and tested their interaction with αMI. The immobilized γ192-286 fragment bound to αMI with Kd = 559 nM, while both γ148-191 and γ148-226 failed to bind suggesting that P1 does not contribute substantially to the binding and that the binding occurs mainly through the γ227-286 region. To further localize a putative binding sequence, we cleaved γ192-286 and analyzed the resulting peptides. The only αMI-binding activity was associated with the γ228-253 peptide, indicating that this region of the central domain contains a novel αMβ2-binding sequence.

Original languageEnglish (US)
Pages (from-to)617-626
Number of pages10
JournalBiochemistry
Volume44
Issue number2
DOIs
StatePublished - Jan 18 2005

ASJC Scopus subject areas

  • Biochemistry

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