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 mere carried out to test the hypothesis that t he self-assembly of collagen I monomers into fibrils depends on the in teractions of specific binding sites in different regions of the monom er. Six synthetic peptides were prepared with sequences found either i n the collagen triple helix or in the N- or C-telopeptides of collagen I. The four peptides with sequences found in the telopeptides were fo und to inhibit self-assembly of collagen I in a purified in vitro syst em. At concentrations of 2.5 mM, peptides with sequences in the C-telo peptides of the alpha 1(I) and alpha 2(I) chain inhibited assembly at about 95%. The addition of the peptide with the alpha 2-telopeptide se quence was effective in inhibiting assembly if added during the lag ph ase and early propagation phase but mot 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 th e ol(I) chain. A fragment of nine amino acids with sequences in the al pha 2-telopeptide was effective in inhibiting fibril assembly. Mutatin g two aspartates in the 9-mer peptide to serine had no effect on inhib ition of fibril assembly, but mutating two tyrosine residues and one p henylalanine residue abolished the inhibitory action. Molecular modeli ng of the binding sites demonstrated favorable hydrophobic and electro static interactions between the alpha 2-telopeptide and residues 781-7 94 of the alpha(I) chain.