title: Folding delay and structural perturbations caused by type IV collagen natural interruptions and nearby Gly missense mutations.
abstract: The standard collagen triple helix requires Gly as every third residue in the amino acid sequence, yet all nonfibrillar collagens contain sites where this repeating pattern is interrupted. To explore the effects of such natural interruptions on the triple helix, a 4- or 15-residue sequence from human basement membrane type IV collagen was introduced between (Gly-Xaa-Yaa)(n) domains within a recombinant bacterial collagen. The interruptions had little effect on melting temperature, consistent with the high thermal stability reported for nonfibrillar collagens. Although the 4-residue interruption cannot be accommodated within a standard triple helix, trypsin and thermolysin resistance indicated a tightly packed structure. Central residues of the 15-residue interruption were protease-susceptible, whereas residues near the (Gly-Xaa-Yaa)(n) boundary were resistant, supporting a transition from an alternate conformation to a well packed triple helix. Both interruptions led to a delay in triple-helix folding, with the 15-residue interruption causing slower folding than the 4-residue interruption. These results suggest that propagation through interruptions represents a slow folding step. To clarify the relation between natural interruptions and pathological mutations, a Gly to Ser missense mutation was placed three triplets away from the 4-residue interruption. As a result of this mutation, the 4-residue interruption and nearby triple helix became susceptible to protease digestion, and an additional folding delay was observed. Because Gly missense mutations that cause disease are often located near natural interruptions, structural and folding perturbations arising from such proximity could be a factor in collagen genetic diseases.
authors.affiliation: Department of Biochemistry, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
authors.forename: Eileen S
authors.initials: ES
authors.lastname: Hwang
authors.forename: Barbara
authors.initials: B
authors.lastname: Brodsky
mesh_terms: D019943:Amino Acid Substitution; D003095:Collagen Diseases / Q000235:genetics / Q000378:metabolism; D024141:Collagen Type IV / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005998:Glycine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D020125:Mutation, Missense; D017510:Protein Folding; D017433:Protein Structure, Secondary
chemical_list: D024141:Collagen Type IV; D005998:Glycine