Glucosylation of galactosylhydroxylysyl residues in collagen in vitro by collagen glucosyltransferase. Inhibition by triple-helical conformation of the substrate.
Glucosylation of galactosylhydroxylysyl residues in various collagen polypeptide chains and in small peptides prepared from collagen was studied in vitro using collagen glucosyltransferase purified about 200 to 500-fold from extract prepared from chick embryos. When various denatured polypeptide or peptide chains were compared as substrates for the enzyme, no significant differences were found between citrate-soluble collagens from normal or lathyritic rats and isolated alpha1 and alpha2 chains. In contrast, gelatinized insoluble calf skin collagen, and peptides prepared from collagen and having an average molecular weight of about 500 were clearly less effective substrates as judged from their Km and V values. A marked difference was found between native and heat-denatured citrate-soluble collagen in that no synthesis of glucosylgalactosylhydroxylysine was observed with the native collagen when the reaction was studied at 30 degrees C with different times, enzyme concentrations, and substrate concentrations. When the reaction was studied as a function of temperature, little glucosylation of native collagen was observed below 37 degrees C, but there was a sharp transition in the rate of glucosylation of native collagen at temperatures above 37 degrees C, similar to that observable in the melting curve of collagen. The data suggest that triple-helical conformation of collagen prevents that glucosylation of galactosylhydroxylysyl residues.
D000818:Animals; D002417:Cattle; D002642:Chick Embryo; D002951:Citrates; D003094:Collagen / Q000378:metabolism*; D005690:Galactose; D005964:Glucosyltransferases / Q000378:metabolism*; D006358:Hot Temperature; D006901:Hydroxylysine; D007700:Kinetics; D007842:Lathyrism / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism; D011487:Protein Conformation; D011489:Protein Denaturation; D051381:Rats; D013696:Temperature
D002951:Citrates; D010446:Peptide Fragments; D006901:Hydroxylysine; D003094:Collagen; D005964:Glucosyltransferases; D005690:Galactose