Hydroxylation of proline after the release of proline-rich polypeptides from ribosomal complexes during uninhibited collagen biosynthesis.
- Additional Document Info
- View All
Pulse-labeling techniques were used to study the hydroxylation of proline in isolated embryonic cartilage which synthesizes collagen at a rapid rate in vitro. In cartilage incubated with [14C]proline for short periods of time, the synthesis of [14C]hydroxyproline consistently lagged behind the incorporation of 14C into protein. The ratio of [14C]hydroxyproline to total 14C in non-dialyzable fractions was less than 1 % after incubation with [14C]proline for 1 min, and it gradually approached a maximal value of 20 to 25 % in 10 to 30 min. Incubation of extracts of pulse-labeled tibiae with purified protocollagen hydroxylase suggested that a large part of the [14C]proline incorporated in 1 min was in polypeptides resembling collagen. Gel-filtration chromatography of extracts of pulse-labeled cartilage indicated that after short incubation periods with [14C]proline, significant amounts of [14C]proline were incorporated into molecules which were of about the same size as the complete polypeptide chains of -collagen, but which did not contain significant amounts of [14C]hydroxyproline. Analysis of the elution patterns of 14C made it possible to estimate the time required to synthesize complete polypeptides de novo by a technique which apparently has not been used previously. The synthesis time calculated with this technique was about 1 min. Since maximal hydroxylation of collagen polypeptides in the system required over 10 min, the results indicated that cells synthesizing collagen under relatively normal conditions contain a significant intracellular pool of complete polypeptide precursors of collagen which are incompletely hydroxylated. A few proline residues may be hydroxylated while polypeptides are still attached to ribosomes, but the results suggested that most of the synthesis of collagen hydroxyproline occurs after completed polypeptides are released from ribosomal complexes. 1967.
author list (cited authors)
Rosenbloom, J., Bhatnagar, R. S., & Prockop, D. J.
complete list of authors
Rosenbloom, J||Bhatnagar, RS||Prockop, DJ