Structural and functional studies with antibodies to the integrin β2 subunit -: A model for the I-like domain

To establish a structure and function map of the beta 2 integrin subunit, we mapped the epitopes of a panel of beta 2 monoclonal antibodies including function-blocking, nonblocking, and activating antibodies using human/mouse beta 2 subunit chimeras. Activating antibodies recognize the C-terminal half of the cysteine-rich region, residues 522-612. Antibodies that do not affect ligand binding map to residues 1-98 and residues 344-521. Monoclonal antibodies to epitopes within a predicted I-like domain (residues 104-341) strongly inhibit LFA-1-dependent adhesion. These function-blocking monoclonal antibodies were mapped to specific residues with human --> mouse knockout or mouse --> human knock-in mutations. Combinatorial epitopes involving residues distant in the sequence provide support for a specific alignment between the beta-subunit and I domains that was used to construct a three-dimensional model. Antigenic residues 133, 332, and 339 are on the first and last predicted alpha-helices of the I-like domain, which are adjacent on its front, Other antigenic residues in beta 2 and in other integrin beta subunits are present on the front. No antigenic residues are present on the back of the domain, which is predicted to be in an interface with other domains, such as the alpha subunit beta-propeller domain. Most mutations in the beta 2 subunit in leukocyte adhesion deficiency are predicted to be buried in the beta 2 subunit I-like domain. Two long insertions are present relative to alpha-subunit I-domains. One is tied down to the back of the I-like domain by a disulfide bond. The other corresponds to the specificity-determining loop defined in beta 1 and beta 3 integrins and contains the antigenic residue Glu(175) in a disulfide-bonded loop located near the top of the domain.