title: Mapping the Effect of Gly Mutations in Collagen on α2β1 Integrin Binding.
abstract: The replacement of one Gly in the essential repeating tripeptide sequence of the type I collagen triple helix results in the dominant hereditary bone disorder osteogenesis imperfecta. The mechanism leading to pathology likely involves misfolding and autophagy, although it has been hypothesized that some mutations interfere with known collagen interactions. Here, the effect of Gly replacements within and nearby the integrin binding GFPGER sequence was investigated using a recombinant bacterial collagen system. When a six-triplet human type I collagen sequence containing GFPGER was introduced into a bacterial collagen-like protein, this chimeric protein bound to integrin. Constructs with Gly to Ser substitutions within and nearby the inserted human sequence still formed a trypsin-resistant triple helix, suggesting a small local conformational perturbation. Gly to Ser mutations within the two Gly residues in the essential GFPGER sequence prevented integrin binding and cell attachment as predicted from molecular dynamics studies of the complex. Replacement of Gly residues C-terminal to GFPGER did not affect integrin binding. In contrast, Gly replacements N-terminal to the GFPGER sequence, up to four triplets away, decreased integrin binding and cell adhesion. This pattern suggests either an involvement of the triplets N-terminal to GFPGER in initial binding or a propagation of the perturbation of the triple helix C-terminal to a mutation site. The asymmetry in biological consequences relative to the mutation site may relate to the observed pattern of osteogenesis imperfecta mutations near the integrin binding site.
authors.affiliation: From the Departments of Biomedical Engineering and Chemistry, Tufts University, Medford, Massachusetts 02155 and.
authors.forename: Sezin
authors.initials: S
authors.lastname: Yigit
authors.affiliation: From the Departments of Biomedical Engineering and Chemistry, Tufts University, Medford, Massachusetts 02155 and.
authors.forename: Hongtao
authors.initials: H
authors.lastname: Yu
authors.affiliation: From the Departments of Biomedical Engineering and.
authors.forename: Bo
authors.initials: B
authors.lastname: An
authors.affiliation: Department of Biochemistry, University of Cambridge, Cambridge CB2 1QW, United Kingdom.
authors.forename: Samir
authors.initials: S
authors.lastname: Hamaia
authors.affiliation: Department of Biochemistry, University of Cambridge, Cambridge CB2 1QW, United Kingdom.
authors.forename: Richard W
authors.initials: RW
authors.lastname: Farndale
authors.affiliation: From the Departments of Biomedical Engineering and.
authors.forename: David L
authors.initials: DL
authors.lastname: Kaplan
authors.affiliation: Chemistry, Tufts University, Medford, Massachusetts 02155 and Yu-Shan.Lin@tufts.edu.
authors.forename: Yu-Shan
authors.initials: YS
authors.lastname: Lin
authors.affiliation: From the Departments of Biomedical Engineering and Barbara.Brodsky@tufts.edu.
authors.forename: Barbara
authors.initials: B
authors.lastname: Brodsky
mesh_terms: D019943:Amino Acid Substitution; D024042:Collagen Type I / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D038982:Integrin alpha2beta1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020125:Mutation, Missense; D011485:Protein Binding; D017433:Protein Structure, Secondary
chemical_list: D024042:Collagen Type I; D038982:Integrin alpha2beta1
keywords: binding; collagen; extracellular matrix; integrin; missense mutations; molecular dynamics; osteogenesis imperfecta; recombinant protein expression; triple helix