A β ₃ Integrin Mutation Abolishes Ligand Binding and Alters Divalent Cation-Dependent Conformation

Abstract

The ligand-binding function of integrin adhesion receptors depends on divalent cations. A mutant alpha IIb beta 3 integrin (platelet gpIIb/IIIa) that lacks ligand recognition shows immunologic evidence of a perturbed interaction with divalent cations. This was found to be caused by a G----T mutation that resulted in an Asp119----Tyr119 substitution in the beta 3 subunit. This residue is proximal to bound ligand and is in a conserved region among integrins that are enriched in oxygenated residues. The spacing of these residues aligns with the calcium-binding residues in EF hand proteins, suggesting interaction with receptor-bound divalent cation as a mechanism of ligand binding common to all integrins.

Probing Conformational Changes in the I-like Domain and the Cysteine-rich Repeat of Human β3 Integrins following Disulfide Bond Disruption by Cysteine Mutations

“…In contrast, only the C177A mutation affected AP2, 7E3, and LM609 binding, while the C273A mutation had no inhibitory effect. (45,46), D119Y (47,48), and S162L (49). Our results suggest that the structural integrity of the two insertions of the I-like domain, in close contact with the ␣ subunit ␤ propeller domain, is important for stable ␣␤ heterodimerization of ␣ IIb ␤ 3 .…”

The Role of α and β Chains in Ligand Recognition by β7 Integrins

“…It is known that the MIDAS motif of the ␤ chain is involved in ligand binding in the context of the A-domain-containing integrins ␣ L ␤ 2 and ␣ M ␤ 2 (43)(44)(45) and in ligand binding in the context of ␣ chains that lack A-domains such as ␣ 4 ␤ 1 , ␣ 5 ␤ 1 , ␣ IIb ␤ 3 , ␣ V ␤ 5 , and ␣ V ␤ 6 (35,(37)(38)(39)(40)(41)(42). We have shown for the first time that the MIDAS motif of a single ␤ chain, ␤ 7 , is critical whether it is paired with an A-domain-containing (␣ E ) or A-domain-lacking (␣ 4 ) ␣ chain.…”

“…Indeed, all ␤ chains possess a relatively conserved region of ϳ250 amino acids that contains a DXSXS sequence, which, together with conserved downstream oxygenated residues, could form a complete MIDAS site. In fact, mutation of such residues in ␤ 1 , ␤ 2 , ␤ 3 , ␤ 5 , and ␤ 6 integrins abolishes ligand binding activity (35,(37)(38)(39)(40)(41)(42)(43)(44)(45). However, it remains likely that non-A-domain-containing integrin ␣ chains also contribute directly to ligand binding.…”

Modeling the αIIbβ3 integrin solution conformation

“…This was done in order to allow greater flexibility in modeling the putative interaction sites within each subunit and between the subunits in the complex (Calvete et al, 1992a,b). At the NH2-terminal tryptic fragment'; RGD cross-link site2 Glycosylation site3; chymotryptic cleavage' RGD cross-link ~i t e~.~; inhibitory point mutation5 Fibrinogen-binding peptide6; tryptic cleavage' Glycosylation site3; RGD, KQAGDV cross-link site2, V8 cleavage' Tryptic cleavage' Tryptic Cysteine-rich region 17*9; glycosylation site3 Cysteine-rich region 27*9 Cysteine-rich region 37*9 glycosylation site3 Cysteine-rich region 4' s9 GIycosyIation site3 a References: 'Calvete et al, 1992a;'Calvete et al, 1992b;'Eirin et al, 1986;4D'Souza et al, 1988;'Loftus et al, 1990;6Charo et al, 1991;7Calvete et al, 1991~;'Ramsamooj et al, 1991;'Fitzgerald et al, 1987;''Poncz et al, 1987;"Calvete et al, 1989a;''Calvete et al, 1989b; '3D'Souza et a]., 1991; I4Nermut et al, 1988;"Chou and Fasman, 1978. end of the assignment procedure, 36 beads were defined: 12, 18, and 3 for the extracellular parts of the P3 and of the heavy and light chains of the q [ b subunit, respectively; 1 for the octyl glucoside micelle containing the transmembrane parts of both subunits, and 2 for the cytoplasmic regions, as reported in Table 3.…”