Computational design of an integrin I domain stabilized in the open high affinity conformation

We have taken a computational approach to design mutations that stabilize a large protein domain of similar to 200 residues in two alternative conform ations. Mutations in the hydrophobic core of the alpha M beta 2 integrin I domain were designed to stabilize the crystallographically defined open or closed conformers. When expressed on the cell surface as part of the intact heterodimeric receptor, binding of the designed open and closed I domains to the ligand iC3b, a form of the complement component C3, was either incre ased or decreased, respectively compared to wild type. Moreover, when expre ssed in isolation from other integrin domains using an artificial transmemb rane domain, designed open I domains were active in ligand binding, whereas designed closed and wild type I domains were inactive, Comparison to a hum an expert designed open mutant showed that the computationally designed mut ants are far more active, Thus, computational design can be used to stabili ze a molecule in a desired conformation, and conformational change in the I domain is physiologically relevant to regulation of ligand binding.