Fig. 3. N-glycosylation of integrin α₅β₁ bridges interface to FN.

(A) Table listing the N-glycosylation sites and the types of glycan trees as determined in our structure. (B) Electrostatic potential map of the FN-integrin interface and the N275-glycan. (C) Two views of the N275-glycan on α₅, which is part of the extensive α₅β₁-FN interface. The N275-glycan contacts with FN9. (D) Comparison of glycan position between integrin α₅β₁ and integrin α_vβ₃ [PDB ID: 3ije (43)] and α_vβ₃ in complex with FN10 [PDB ID: 4mmz (42)]. The N275-glycan (α₅β₁) is a conserved glycosylation site among FN-recognizing integrins such as α_vβ₃. (E) Solid-phase binding assay to measure the binding affinity of integrin α₅(N275A)β₁ ectodomain mutant in comparison to wild-type ectodomain. Both wild-type and N275A ectodomain mutant were unclasped and binding to FN7-10 wild type evaluated in the presence of Mn²⁺. Bars represent means + SD of three measurements. Unspecific binding of integrin to bovine serum albumin (BSA)–coated control wells was subtracted from the total absorbance values. Normalized binding is expressed as the ratio of mutant absorbance relative to wild-type absorbance (ratio = 1 represents wild-type binding).