Historically, most integrin-ligand pairs have been identified either by affinity chromatography or through the ability of subunit-specific monoclonal antibodies to block cell adhesion to specific ligands. In some cases, direct protein-protein binding assays have been used to support biochemical or cell biological data. Despite their wide variety, it is possible to cluster integrin-ligand combinations into four main classes, reflecting the structural basis of the molecular interaction. These classes do not necessarily reflect evolutionary relationships.
(a) RGD-binding integrins
All five αV integrins, two β1 integrins (α5, α8) and αIIbβ3 share the ability to recognise ligands containing an RGD tripeptide active site. Crystal structures of αVβ3 and αIIbβ3 complexed with RGD ligands have revealed an identical atomic basis for this interaction (Xiong et al., 2002
; Xiao et al., 2004
). RGD binds at an interface between the α and β subunits, with the R residue fitting into a cleft in a β-propeller module in the α subunit, and the D coordinating a cation bound in a von Willebrand factor A-domain in the β subunit. The RGD-binding integrins are among the most promiscuous in the receptor family, with β3 integrins in particular binding to a large number of extracellular matrix and soluble vascular ligands. Although many ligands are shared by this subset of integrins, the rank order of ligand affinity varies, presumably reflecting the preciseness of the fit of the ligand RGD conformation with the specific α,β active site pockets.
(b) LDV-binding integrins
α4β1, α4β7, α9β1, the four members of the β2 sub-family and αEβ7 recognise related sequences in their ligands. α4β1, α4β7 and α9β1 bind to an acidic motif, termed ‘LDV’, that is functionally related to RGD. Fibronectin contains the prototype LDV ligand in its type III connecting segment region, but other ligands (such as VCAM-1 and MAdCAM-1) employ related sequences. Although definitive structural information is lacking, it is highly likely that LDV peptides bind similarly to RGD at the junction between the α and β subunits. Osteopontin also interacts with α4β1, α4β7 and α9β1, but this is apparently via a different peptide motif, SVVYGLR, and the location of the ligand-binding site has not been identified.
The β2 family employ a different mode of ligand binding, with the major interaction taking place via an inserted A-domain in the α subunit (see Shimaoka et al., 2003
for the structure of a complex between the αL A-domain and ICAM-1). However, despite this fundamental mechanistic difference, the characterised sites within ligands that bind β2 integrins are structurally homologous to the LDV motif. The major difference is that β1/β7 ligands employ an aspartate residue for cation coordination, while β2 integrins use glutamate. Collectively, therefore, the LDV motif is described by the consensus sequence L/I-D/E-V/S/T-P/S.
(c) A-domain β1 integrins
Four α subunits containing an αA-domain (α1, α2, α10 and α11) combine with β1, and form a distinct laminin/collagen-binding subfamily. Few other validated ligands have been identified for these integrins. A crystal structure of a complex between the α2 A-domain and a triple-helical collagenous peptide has revealed the structural basis of the interaction, with a critical glutamate within a collagenous GFOGER motif providing the key cation-coordinating residue (Emsley et al., 2000
). Currently, the mechanism of laminin binding is unknown.
(d) Non αA-domain-containing laminin-binding integrins
Three β1 integrins (α3, α6 and α7), plus α6β4, are highly selective laminin receptors. Analysis of laminin fragments indicates that these receptors and the A-domain-containing β1 integrins bind to different regions of the ligands. In neither case has the active site been narrowed down to a particular sequence or residue.