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Complex structure of engineered modular domains defining molecular interaction between ICAM-1 and integrin LFA-1.

ABSTRACT: Intermolecular contacts between integrin LFA-1 (?(L)?(2)) and ICAM-1 derive solely from the integrin ?(L) I domain and the first domain (D1) of ICAM-1. This study presents a crystal structure of the engineered complex of the ?(L) 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 ?(L) I domain allostery between the C-terminal ?7-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 ?7-helix or the active sites. In our crystal, the ?(L) I domain was found to have a high affinity conformation to D1 with its ?7-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 ?-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 ?(L) I domain but reduced interaction with HRV.

SUBMITTER: Kang S

PROVIDER: S-EPMC3431320 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Complex structure of engineered modular domains defining molecular interaction between ICAM-1 and integrin LFA-1.

Kang Sungkwon S Kim Chae Un CU Gu Xiaoling X Owens Roisin M RM van Rijn Sarah J SJ Boonyaleepun Vanissra V Mao Yuxin Y Springer Timothy A TA Jin Moonsoo M MM

PloS one 20120830 8

Intermolecular contacts between integrin LFA-1 (α(L)β(2)) and ICAM-1 derive solely from the integrin α(L) I domain and the first domain (D1) of ICAM-1. This study presents a crystal structure of the engineered complex of the α(L) 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 α(L) I domain allostery between the C-terminal α7-helix (allosteric site) and the metal-ion dep ...[more]

PMID: 22956999

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Project description:Using single-molecule force spectroscopy to probe ICAM-1 interactions with recombinant alphaLbeta2 immobilized on microspheres and beta2 integrin on neutrophils, we quantified an impressive hierarchy of long-lived, high-strength states of the integrin bond, which start from basal levels with activation in solutions of divalent cations and shift dramatically upward to hyperactivated states with cell signaling. Taking advantage of very rare events, we used repeated measurements of bond lifetimes under steady ramps of force to achieve a direct assay for the off-rates of ICAM-1 from beta2 integrin throughout the course of each experiment. In our companion article I, we demonstrate the assay using results from tests of a monovalent ICAM-1 probe against recombinant alphaLbeta2 on microspheres in millimolar solutions of divalent cations (Ca2+, Mg2+, Mn2+). In this article, we examine the impact of inside-out and outside-in signaling in neutrophils on the lifetimes and mechanical strengths of ICAM-1 bonds to beta2 integrin on the cell surface. Even though ICAM-1 bonds to recombinant alphaLbeta2 on microspheres in Mg2+ or Mn2+ can live for long periods of time under slow pulling, here we show that stimulation of neutrophils in Mg2+ plus the chemokine IL-8 (i.e., inside-out signaling) induces several-hundred-fold longer lifetimes for ICAM-1 attachments to LFA-1, creating strong bonds at very slow pulling speeds where none are perceived in Mg2+ or Mn2+ alone. Similar changes are observed with outside-in signaling, i.e., long lifetimes and increased bond strength also occur when neutrophils are bound with the activating (anti-CD18) monoclonal 240Q. Limiting our investigation to rare events using very dilute ICAM-1 probes, we show that although the prolonged lifetimes of cell surface attachments for both inside-out and outside-in signaling exhibit single-bond-like statistics for dissociation under force, they are consistent with a tightly coupled dimeric ICAM-1 interaction with a pair of LFA-1 heterodimers.

Dataset Information

Complex structure of engineered modular domains defining molecular interaction between ICAM-1 and integrin LFA-1.

Publications

Complex structure of engineered modular domains defining molecular interaction between ICAM-1 and integrin LFA-1.

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