Incorporation of proline analogues into collagen polypeptides Effects on the production of extracellular procollagen and on the stability of the triple-helical structure of the molecule

Abstract

The effects of several proline analogues on collagen biosynthesis were examined in freshly isolated cells from embryonic tendon. Four analogues which were previously shown to be incorporated into collagen were found to decrease the amount of newly synthesized 14C-labeled procollagen recovered in the medium after incubation of the cells for 2–3 h. The decreased production of extracellular procollagen was not explained by inhibition of protein synthesis, failure of the procollagen to form interchain disulfide bonds, or any effects of the free analogues on hydroxylation of 14C-labeled protocollagen or secretion of 14C-labeled procollagen which did not contain the analogues. The specific effects of the four proline analogues on collagen could be explained by the hypothesis that the presence of the analogues in the polypeptide chains prevents them from assuming the normal triple-helical conformation of collagen and as a result the polypeptides are degraded. This hypothesis was supported by the observation that the concentration of the effective analogues which blocked the production of extracellular 14C-labeled procollagen also decreased the amount of intracellular 14C-labeled procollagen which was in a helical conformation as judged by its resistance to proteolytic digestion. With the same concentrations of analogues there was an increase in the amount of small 14C-labeled peptides in the medium and an increase in the rate of degradation of collagen 14C-labeled polypeptides. The protein containing the proline analogues did not become helical on cooling to 15 C under conditions in which protocollagen became helical. Therefore the decreased tendency of the polypeptides to become triple-helical could not be explained simply by their apparently low content of trans-hydroxyproline and the