The three overall integrin conformational states. The bent conformation has a closed headpiece and is low affinity. Extension at the α- and β-knees releases an interface between the headpiece and lower legs and yields an extended-closed conformation also with low affinity. Swing-out of the hybrid domain at its interface with the βI domain is connected through the βI α7-helix to rearrangements at the βI interface with the β-propeller domain that greatly (~1,000-fold) increase affinity for ligand in the extended-open conformation. Similar interdomain rearrangements in αI integrins result in activating a binding site for an internal ligand, Glu310 in α_L, which pulls down the αI α7-helix to activate a similar increase in affinity (~1,000 to 10,000-fold) of the α_L I domain for the ligand ICAM-1. Although the integrin headpiece has highly preferred closed and open conformations, the lower β-legs are highly flexible, and thus we speak of “overall” conformational states. This is symbolized by the dashed lower β-leg. Therefore, only very large separations between α and β TMD, such as induced by lateral motion of β when its cytoplasmic domain is associated with the actin cytoskeleton, can be transmitted through the floppy β-leg to stabilize the high-affinity, open headpiece conformation.

The closed and open integrin headpiece conformations. A 2.3 Å movement of the βI-αI loop at the ligand binding site that raises affinity for ligand 1,000 to 10,000-fold is amplified to 4 Å displacement of the ADMIDAS Ca²⁺, 5 Å sideways displacement of the αI-helix as it straightens, 7 Å reshaping of the β6-α7 loop, 5.5 Å connecting rod-like displacement of the α-7 helix, and leverages hybrid domain swing-out to a 75 Å separation at the integrin knees. Structures are of the α_IIbβ₃ headpiece crystallized in absence [12] (a) or presence [11] (b) of RGD peptide with missing portions supplemented by superposition of the α_IIbβ₃ ectodomain [3].

Imaging single ICAM-1 molecules in the synapse. Single Cy5-ICAM-1 imaged in an immune synapse (outline) with ~0.1% labeling. Kymographs for single particles show time vs. x-component of random x-y movement and periods of binding (vertical segments). Courtesy of R. Varma.

Structure of integrins from electron microscopy, electron tomography, and neutron or X-ray scattering in solution reveal three conformational states. Schematics are shown to right. All scale bars = 10 nm. With permission from cited references. Panels show representative class averages of negatively-stained integrins unless otherwise noted. (a) α_Vβ₃ ectodomain with a C-terminal coiled-coil clasp (1) or unclasped with Mn²⁺ (2) or RGD (3) [13]. (b) α_IIbβ₃ ectodomain, clasped (4) or unclasped (5-7) [3]. (c) α_IIbβ₃ purified from platelets in detergent (8, 10, 12) [23], (13) [16] and embedded in lipoprotein nanodiscs (9, 11) [27]. With no additions (8-9), Mn²⁺ (10), talin head domain (11), and RGD peptide or RGD mimetics (12-13). Negative stain electron tomography class average (13). (d) Three-dimensional molecular envelopes of purified, detergent-soluble native α_IIbβ₃ in solution determined by small-angle neutron scattering (14) [21] or X-ray scattering (15-17) [23] with no additions (14-15), Mn²⁺ (16), or Mn²⁺ and RGD mimetic (17). (e) α₅β₁ headpiece [14,15] alone (18), + allosteric inhibitory SG/19 Fab (19), + RGD (20), or + fibronectin domain 7-10 fragment (21). (f) LFA-1 ectodomain [19] clasped (22) or unclasped (23-25). (g) α_Xβ₂ [17,19] clasped from different publications (26-27) and unclasped (28-29). (h) α_Xβ₂ [17,19] clasped with extension and activation-promoting CBR LFA-1/2 Fab (30, 31, 35); and with CBR LFA-1/2 Fab and allosteric inhibitory Fab 7E4 (32-33) and TS1/18 (34) or allosteric activating Fab MEM148 (36) or m24 (37).

Dynamics of LFA-1-ICAM-1 interactions. Naïve T cells adhered to supported planar bilayers with 200 molecules/μm² Cy5-ICAM-1 and 20 molecules/μm² agonist MHC-peptide complexes. A. At T = 0 the ICAM-1 fluorescence was bleached and the contact area was imaged at +5 and +20 seconds. B. Control without bleaching. Courtesy of T. N. Sims.