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Disrupting integrin transmembrane domain heterodimerization increases ligand binding affinity, not valency or clustering.

Abstract：

:Residues important in the interaction between the 23-residue transmembrane (TM) domains of the integrin alpha(IIb)- and beta(3)-subunits were identified by mutating each non-Leu residue to Leu. Leu substitutions of alpha(IIb) at G972, G976, and T981, and of beta(3) at I693 and G708, increased ligand binding. Substitutions with other amino acids at alpha(IIb)G972 and beta(3)G708 could also increase ligand binding. The results are consistent with and extend the helical interface between the integrin alpha- and beta-subunit TM domains previously defined by cysteine scanning and disulfide bond formation. We differentiated between affinity- and valency-based modes of activation by TM domain mutations. The mutant alpha(IIb) W967C forms disulfide-linked alpha(IIb)-subunits within an (alpha(IIb)beta(3))(2) tetramer. This tetramer behaved as an ideal model for the valency mode of regulation, because it exhibited significantly increased binding to multivalent but not monovalent ligands and basally retained the bent conformation. By contrast, the activating Leu mutants showed increased binding to the monovalent, ligand-mimetic PAC-1 Fab and increased exposure of ligand-induced binding site (LIBS) epitopes, suggesting that they partially adopt an extended conformation. Furthermore, the previously described beta(3)G708N mutation in Chinese hamster ovary cells enhanced ligand binding affinity, not valency, and did not alter cell-surface clustering as defined by confocal microscopy. Our studies provide evidence that disrupting the integrin heterodimeric TM helix-helix interface activates ligand binding mainly by increasing the monomeric affinity for ligand, but not the receptor valency, i.e., clustering.

journal_name

Proc Natl Acad Sci U S A

journal_title

Proceedings of the National Academy of Sciences of the United States of America

authors

Luo BH,Carman CV,Takagi J,Springer TA

doi

10.1073/pnas.0409440102

keywords:

subject

Has Abstract

pub_date

2005-03-08 00:00:00

pages

3679-84

issue

eissn

0027-8424

issn

1091-6490

pii

0409440102

journal_volume

102

pub_type

杂志文章

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Disrupting integrin transmembrane domain heterodimerization increases ligand binding affinity, not valency or clustering.