Effect of the arginine analog, canavanine, on the synthesis and secretion of procollagen

Abstract

Canavanine was shown to competitively inhibit the activation of arginine when tested with tRNA and synthetases prepared from whole chick embryos. The canavanine has no effect when tested with other amino acids. The K m for arginine was 2.5 μ m and the K i for canavanine was 35 μ m. When fibroblasts from embryonic chick tendons were incubated with [ 3H]arginine and increasing concentrations of canavanine, there was a progressive decrease in the incorporation of [ 3H]arginine so that at 3 m m the incorporation into nondialyzable protein was only 14% of the control. A much smaller decrease in the incorporation of other radioactive amino acids was observed. Amino acid analysis of proteins isolated from cells incubated with canavanine showed conclusively that the analog was incorporated. When the cells were incubated with [ 14C]proline or [ 3H]glycine and 3 m m canavanine, the labeled procollagen containing the canavanine was secreted more slowly than normal and accumulated intracellularly. The retained procollagen chains were normally hydroxylated, disulfide linked, and triple helical. However, slab gel electrophoresis in sodium dodecyl sulfate demonstrated that they migrated with a lower mobility than control procollagen chains. We postulate that incorporation of canavanine inhibits normal proteolytic processing of signal sequences resulting in delayed secretion of the procollagen.