Inhibition of the self-assembly of collagen I into fibrils with synthetic peptides. Demonstration that assembly is driven by specific binding sites on the monomers.
A series of experiments were carried out to test the hypothesis that the self-assembly of collagen I monomers into fibrils depends on the interactions of specific binding sites in different regions of the monomer. Six synthetic peptides were prepared with sequences found either in the collagen triple helix or in the N- or C-telopeptides of collagen I. The four peptides with sequences found in the telopeptides were found to inhibit self-assembly of collagen I in a purified in vitro system. At concentrations of 2.5 mM, peptides with sequences in the C-telopeptides of the alpha1(I) and alpha2(I) chain inhibited assembly at about 95%. The addition of the peptide with the alpha2-telopeptide sequence was effective in inhibiting assembly if added during the lag phase and early propagation phase but not later in the assembly process. Experiments with biotinylated peptides indicated that both the N- and C-telopeptides bound to a region between amino acid 776 and 822 of the alpha(I) chain. A fragment of nine amino acids with sequences in the alpha2-telopeptide was effective in inhibiting fibril assembly. Mutating two aspartates in the 9-mer peptide to serine had no effect on inhibition of fibril assembly, but mutating two tyrosine residues and one phenylalanine residue abolished the inhibitory action. Molecular modeling of the binding sites demonstrated favorable hydrophobic and electrostatic interactions between the alpha2telopeptide and residues 781-794 of the alpha(I) chain.
Prockop
D J
DJ
Center for Gene Therapy, Allegheny University of the Health Sciences, Philadelphia, Pennsylvania 19102, USA.
Fertala
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D003094:Collagen / Q000378:metabolism*; D024042:Collagen Type I; D017364:Collagenases / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis / Q000378:metabolism*; D017433:Protein Structure, Secondary
D024042:Collagen Type I; D010455:Peptides; C052929:collagen type I trimeric cross-linked peptide; D003094:Collagen; D017364:Collagenases