Abstract
The hereditary bone disorder osteogenesis imperfecta is often caused by missense mutations in type I collagen that change one Gly residue to a larger residue and that break the typical (Gly-Xaa-Yaa)(n) sequence pattern. Site-directed mutagenesis in a recombinant bacterial collagen system was used to explore the effects of the Gly mutation position and of the identity of the residue replacing Gly in a homogeneous collagen molecular population. Homotrimeric bacterial collagen proteins with a Gly-to-Arg or Gly-to-Ser replacement formed stable triple-helix molecules with a reproducible 2 °C decrease in stability. All Gly replacements led to a significant delay in triple-helix folding, but a more dramatic delay was observed when the mutation was located near the N terminus of the triple-helix domain. This highly disruptive mutation, close to the globular N-terminal trimerization domain where folding is initiated, is likely to interfere with triple-helix nucleation. A positional effect of mutations was also suggested by trypsin sensitivity for a Gly-to-Arg replacement close to the triple-helix N terminus but not for the same replacement near the center of the molecule. The significant impact of the location of a mutation on triple-helix folding and conformation could relate to the severe consequences of mutations located near the C terminus of type I and type III collagens, where trimerization occurs and triple-helix folding is initiated.
Highlights
Dues, and most of these substituted residues are well represented in the more than 600 missense mutations in type I collagen reported to result in osteogenesis imperfecta (OI) [7]
Recombinant Bacterial Collagens Designed to Include Gly Substitutions—Gly substitutions were introduced into a control protein that consisted of a portion of the collagen-like protein Scl2.28 from S. pyogenes
OI collagens synthesized by fibroblasts always represent a mixture of molecular species, containing zero, one, or two mutant chains, and it is not feasible to purify a single molecular species containing mutations in amounts needed for study
Summary
Dues, and most of these substituted residues are well represented in the more than 600 missense mutations in type I collagen reported to result in OI [7]. The control protein, referred to as VCL, contained an N-terminal globular trimerization domain (V) and a triple-helix domain of sequence (Gly-XaaYaa)79 (Fig. 1).
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.