Metal ion and ligand binding of integrin α ₅ β ₁

Abstract

Integrin α 5 β 1 binds to an Arg-Gly-Asp (RGD) motif in its ligand fibronectin. We report high-resolution crystal structures of a fourdomain α 5 β 1 headpiece fragment, alone or with RGD peptides soaked into crystals, and RGD peptide affinity measurements. The headpiece crystallizes in a closed conformation essentially identical to that seen previously for α 5 β 1 complexed with a Fab that allosterically inhibits ligand binding by stabilizing the closed conformation. Soaking experiments show that binding of cyclic RGD peptide with 20-fold higher affinity than a linear RGD peptide induces conformational change in the β 1 -subunit βI domain to a state that is intermediate between closed (low affinity) and open (high affinity). In contrast, binding of a linear RGD peptide induces no shape shifting. However, linear peptide binding induces shape shifting when Ca 2+ is depleted during soaking. Ca 2+ bound to the adjacent to metal iondependent adhesion site (ADMIDAS), at the locus of shape shifting, moves and decreases in occupancy, correlating with an increase in affinity for RGD measured when Ca 2+ is depleted. The results directly demonstrate that Ca 2+ binding to the ADMIDAS stabilizes integrins in the low-affinity, closed conformation. Comparisons in affinity between four-domain and six-domain headpiece constructs suggest that flexible integrin leg domains contribute to conformational equilibria. High-resolution views of the hybrid domain interface with the plexin-semaphorin-integrin (PSI) domain in different orientations show a ball-and-socket joint with a hybrid domain Arg side chain that rocks in a PSI domain socket lined with carbonyl oxygens.

Leukocyte integrin α L β 2 headpiece structures: The αI domain, the pocket for the internal ligand, and concerted movements of its loops

“…1C). The LFA-1 βI domain β1-α1 loop, α1-helix, β6-α7 loop, and α7-helix have conformations essentially identical to those in closed β1, β3, β6, and β7 integrin structures (19)(20)(21)(22)(23).…”

Regulation of Plasma Membrane Localization of the Na+-Taurocholate Cotransporting Polypeptide (Ntcp) by Hyperosmolarity and Tauroursodeoxycholate

“…5). Because ␤ 1 -integrins were identified as sensors for TUDC in rat liver (19,48), the integrin-inhibitory peptide GRGDSP, which binds to ␣ 5 ␤ 1 -integrin with a K D of 396 nM (62), was used to study the involvement of ␤ 1 -integrins in TUDC-induced Ntcp insertion into the plasma membrane. GRGDSP (10 mol/liter) inhibited the counteracting effect of TUDC on hyperosmotic Ntcp retrieval, although the inactive control peptide GRADSP (10 mol/liter) was ineffective (Fig.…”

Relating conformation to function in integrin α 5 β 1

“…SG/19 recognizes a single species-specific residue, Thr-82, in a hybrid domain loop that only becomes ordered when bound to SG/19 ( Fig. 8A) (25,36). SG/19 simultaneously recognizes human-mouse invariant residues at the end of the α1-helix in the βI domain, and thus recognizes an orientation between the hybrid and βI domains present only in the closed conformation.…”