SEQUENTIAL STEPS IN SYNTHESIS OF HYDROXYLSINE AND GLYCOSYLATION OF HYDROXYLYSINE DURING BIOSYNTHESIS OF COLLAGEN

Cartilage from chick embryos was pulse-labeled with [14C]lysine, and the incorporation of [14C]lysine, synthesis of [14C]hydroxylysine and synthesis of glycosylated [14C]hydroxylysine were followed. The results indicated that the synthesis of [14C]hydroxylysine lagged behind the incorporation of [14C]lysine into protein. Incubation with puromycin or cycloheximide inhibited the incorporation of [14C]-lysine, but there was a more marked effect on the synthesis of [14C]hydroxylysine, and non-dialyzable peptides containing [14C]lysine and little [14C]hydroxylysine were recovered from the tissue. Pulse-label experiments also indicated that the synthesis of glycosylated [14C]hydroxylysine lagged behind the synthesis of [14C]hydroxylysine. Assays of the glycosylated [14C]hydroxylysine in tissues pulse-labeled for 5, 10, or 20 min indicated that the ratio of glucosylgalactosyl[14C]hydroxylysine to galactosyl[14C]hydroxylysine gradually increased. Incubation of the tissues with 2-deoxyglucose resulted in the synthesis of fully hydroxylated but non-glycosylated collagen. The results indicate that the synthesis of hydroxylysine is similar to the synthesis of hydroxyproline during collagen biosynthesis in that most or all of the hydroxylation occurs after protocollagen polypeptides are released from ribosomal complexes. In addition, they indicate that the glycosylation of the hydroxylysine in collagen also occurs in discrete, sequential steps. © 1969.