The Specificity and Function of the Metal-binding Sites in the Integrin β3 A-domain

Abstract

The A-domains within integrin ␤ subunits contain three metal sites termed the metal ion-dependent adhesion site (MIDAS), site adjacent to the metal ion-dependent adhesion site (ADMIDAS), and ligand-induced metal-binding site (LIMBS), and these sites are involved in ligand engagement. The ␤ 3 integrin subfamily consists of two members, ␣ v ␤ 3 and ␣ llb ␤ 3 . The two receptors share the same ␤ 3 subunit, and the amino acid sequences of their ␣ subunits are 36% identical (1). Both are typical integrin ␣/␤ heterodimers, in which each subunit is composed of a large extracellular region, a transmembrane region, and a short cytoplasmic tail. ␣ llb ␤ 3 expression is restricted primarily to platelets and their megakaryocyte precursors and plays an indispensable role in thrombus formation (2-4). ␣ v ␤ 3 is expressed by a variety of vascular cells, including endothelial cells, where it influences the adhesive and migratory properties of these cells (5-7). Both ␤ 3 integrins bind multiple adhesive ligands, and many of the ligands bind to both receptors, including fibrinogen (8 -10). However, there is a fundamental difference in the specificity involved in fibrinogen recognition by the two integrins; ␣ llb ␤ 3 binds fibrinogen through the sequence at the carboxyl terminus of its ␥ chain 406 KQAGDV 411 (11,12), and ␣ v ␤ 3 binds to one of the two 572 RGD 574 sequences in its A␣ chain (13). In addition, sites within the ␥ chain, distinct from the KQAGDV, have been implicated in recognition of fibrinogen by ␣ v ␤ 3 (14). According to recently published crystal structures of the extracellular regions of ␣ v ␤ 3 (15) and ␣ llb ␤ 3 (16) with bound ligand mimetics, the main area of ligand binding lies between the ␤-propeller of the ␣ subunit and the A-domain in the ␤ 3 subunit. This direct demonstration of ␤ 3 A-domain involvement in ligand engagement was preceded by numerous studies using mutational, immunological, and biochemical approaches (17-21), which indicated that this region played a critical role in ligand binding to the two ␤ 3 integrins.The binding of most ligands to integrins requires metal ions. This is also true for the ␤ 3 integrins and the cellular responses arising from ligand engagement (e.g. platelet aggregation (22-24)). The ␤-propeller domain of the ␣ subunit contains four divalent ion-binding sites within the loops of blades 4 -7 near the bottom of the propeller, but these are not directly involved in ligand engagement (25). The ␤ 3 A-domain contains three metal sites for binding ligands, which are more directly involved in ligand binding. These are referred to as the metal ion-dependent adhesion site (MIDAS), 4 a site adjacent to the MIDAS (ADMIDAS), and a ligand-induced metal-binding site (LIMBS). The ␣ v ␤ 3 and ␣ llb ␤ 3 crystal structures establish that cation bound in the MIDAS is involved directly in ligand engagement; the ADMIDAS cation is not directly involved in contacting ligand but may help to regulate ligand binding; and the LIMBS cation may aid in stabilizing the receptor-ligand complex (26). ...

Expression of Integrin Subunits in the Herniated Intervertebral Disc

“…The receptors of fibronectin are α 5 β 1 , α v β 1 , and α v β 3 [15,20,23,24], and the receptors of collagens are α 1 β 1 , α 2 β 1 , and α v β 1 [20]. And in chondrocyte, fibronectin fragments (Fn-f), which were the breakdown product of fibronectin, stimulated α 5 and β 1 integrin subunits production [25][26][27].…”

Targeted molecular dynamics reveals overall common conformational changes upon hybrid domain swing‐out in β3 integrins

“…The first and third pairs are conserved in αVβ3 (αV D306-β3 K384 and αV K308-β3 E358), whereas in the latter the αIIb R320 residue is replaced by a glycine in αV (G307) 9,56-60. 2) β3 S300 (α5 helix) and β3 R360 (hybrid domain),32 whose Cβ distance increases from ∼9 to ∼34 Å; 3) Residues G349-R352 of the C-terminal region of α7 helix that contacts the hybrid domain;33-36 4) Residues S337-N339 and D126-S130 of the N-terminal portions of the α7 helix61,62 and the α1 helix,36,63-67 respectively, whose interactions are lost when the β6-α7 loop moves away from the ADMIDAS ion. Selective modification of these residues may provide valuable information on the role of these residues in attaining the high affinity ligand binding state.…”

Integrin αvβ3 on human endothelial cells binds von Willebrand factor strings under fluid shear stress

“…However, this apparent discrepancy appears to be explained by differences in experimental conditions. Ligand binding to integrin ␣ v ␤ 3 and P-selectin depends on divalent cations, 21,44,45 and changing the concentrations of Ca 2ϩ and Mg 2ϩ can selectively alter the dependence of VWF or platelet binding on these adhesive proteins. Under physiologic conditions, with Ca 2ϩ and Mg 2ϩ present, the attachment of VWF strings to HUVECs involves integrin ␣ v ␤ 3 ( Figure 6A-D), but without divalent cations the attachment of VWF appears to be independent of integrin ␣ v ␤ 3 ( Figure 6E,F).…”