Identification of the Binding Site for Fibrinogen Recognition Peptide γ383-395 within the αMI-Domain of Integrin α Mβ2

Valentin P. Yakubenko, Dmitry A. Solovjov, Li Zhang, Vivien C. Yee, Edward F. Plow, Tatiana P. Ugarova

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Abstract

The leukocyte integrin αMβ2 (Mac-1, CD11b/CD18) is a cell surface adhesion receptor for fibrinogen. The interaction between fibrinogen and αMβ2 mediates a range of adhesive reactions during the immune-inflammatory response. The sequence γ383TMKIIPFNRLTIG395, P2-C, within the γ-module of the D-domain of fibrinogen, is a recognition site for αMβ2 and αXβ 2. We have now identified the complementary sequences within the αMI-domain of the receptor responsible for recognition of P2-C. The strategy to localize the binding site for P2-C was based on distinct P2-C binding properties of the three structurally similar I-domains of α Mβ2, αXβ2, and αLβ2, i.e. the αMI- and αXI-domains bind P2-C, and the αLI-domain did not bind this ligand. The Lys245-Arg261 sequence, which forms a loop βD-α5 and an adjacent helix α5 in the three-dimensional structure of the αMI-domain, was identified as the binding site for P2-C. This conclusion is supported by the following data: 1) mutant cell lines in which the αMI-domain segments 245KFG and Glu253-Arg261 were switched to the homologous αLI-domain segments failed to support adhesion to P2-C; 2) synthetic peptides duplicating the Lys245-Tyr252 and Glu253-Arg261 sequences directly bound the D fragment and P2-C derivative, γ384-402, and this interaction was blocked efficiently by the P2-C peptide; 3) mutation of three amino acid residues within the Lys245-Arg261 segment, Phe246, Asp254, and Pro257, resulted in the loss of the binding function of the recombinant αMI-domains; and 4) grafting the αM(Lys245-Arg261) segment into the αLI-domain converted it to a P2-C-binding protein. These results demonstrate that the αM(Lys245-Arg 261) segment, a site of the major sequence and structure difference among αMI-, αXI-, αXI-, and αLI-domains, is responsible for recognition of a small segment of fibrinogen, γThr383-Gly395, by serving as ligand binding site.

Original languageEnglish (US)
Pages (from-to)13995-14003
Number of pages9
JournalJournal of Biological Chemistry
Volume276
Issue number17
DOIs
StatePublished - Apr 27 2001
Externally publishedYes

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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