Abstract
Intermolecular contacts between integrin LFA-1 (αLβ2) and ICAM-1 derive solely from the integrin αL I domain and the first domain (D1) of ICAM-1. This study presents a crystal structure of the engineered complex of the αL I domain and ICAM-1 D1. Previously, we engineered the I domain for high affinity by point mutations that were identified by a directed evolution approach. In order to examine αL I domain allostery between the C-terminal α7-helix (allosteric site) and the metal-ion dependent adhesion site (active site), we have chosen a high affinity variant without mutations directly influencing either the position of the α7-helix or the active sites. In our crystal, the αL I domain was found to have a high affinity conformation to D1 with its α7-helix displaced downward away from the binding interface, recapitulating a current understanding of the allostery in the I domain and its linkage to neighboring domains of integrins in signaling. To enable soluble D1 of ICAM-1 to fold on its own, we also engineered D1 to be functional by mutations, which were found to be those that would convert hydrogen bond networks in the solvent-excluded core into vdW contacts. The backbone structure of the β-sandwich fold and the epitope for I domain binding of the engineered D1 were essentially identical to those of wild-type D1. Most deviations in engineered D1 were found in the loops at the N-terminal region that interacts with human rhinovirus (HRV). Structural deviation found in engineered D1 was overall in agreement with the function of engineered D1 observed previously, i.e., full capacity binding to αL I domain but reduced interaction with HRV.
Highlights
Integrins are noncovalently associated ab heterodimeric cell surface receptors that mediate cell-cell and cell-extracellular matrix adhesions, signaling bidirectionally across the plasma membrane
Compared to the rationally designed activating mutations in the lymphocyte functionassociated antigen (LFA)-1 a I domain, we previously reported an application of directed evolution to select active I domains from a library through a selective pressure for binding to intercellular adhesion molecules (ICAMs)-1 [22]
In the process of integrins engaging with their ligands, the downward ‘pull’ of the a7-helix located at the C-terminal end of the a I domain switches the metal iondependent adhesion site (MIDAS) from low to high affinity conformation
Summary
Integrins are noncovalently associated ab heterodimeric cell surface receptors that mediate cell-cell and cell-extracellular matrix adhesions, signaling bidirectionally across the plasma membrane. Integrins play important roles in development, immune cell trafficking and responses, and homeostasis [1,2,3]. One of the major leukocyte integrins is the lymphocyte functionassociated antigen (LFA)-1, which provides the interactions necessary for immunological synapse formation and adhesion to endothelial cells [4]. As one of the most biologically important ligands for LFA-1, ICAM-1 is expressed at a low constitutive level in diverse types of cells and tissues, while its expression is greatly upregulated in response to inflammation [7] and in some tumors and their stroma [8,9,10,11,12,13,14]. The interaction of LFA-1 and ICAM-1 is contained within the single domains called the a I domain in LFA-
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