THE BIOSYNTHESIS OF GRAMICIDIN S IN A CELL-FREE SYSTEM

Abstract

A cell-free system from Bacillus brevis cells, harvested in the late phase of growth and consisting of the 11000g supernatant, incorporates into gramicidin-S the 5 constituent amino-acids. The resuits are consistent with complete synthesis and not merely a completion of pre-existing intermediate peptides. The incorporation of C14-labelled amino-acids by the 11000g supernatant into gramicidin-S requires an energy source. Omission of phosphoenolpyruvate and pyruvate kinase from the incubation mixture prevents incorporation into gramicidin-S. The cell-free system incorporates [C14]-leucine, -pro-line and -phenylalanine over a period of 4 hr. With [C14]leucine, incorporation into gramicidin-S takes place in the range pH 6-9 with maximum incorporation at pH 7.0. High concentrations of chloram-phenicol or puromycin decreased the incorporation into gramicidin-S by only about 20%. The 50 OOOg supernatant did not decrease the ability to incorporate [C14]valine into gramicidin-S as compared with the llOOOg supernatant. About 40% of the incorporating ability remained in the 105 OOOg supernatant after 3 hr. centrifugation. When recombining the 105000g sediment with the 105000g supernatant, some increase in incorporation over that obtained with the supernatant alone was obtained. Gramicidin-S may be synthesized in a different manner from that of proteins.