EFFECTS OF VARIOUS ESTERS OF TRANS-4-GUANIDINOMETHYLCYCLOHEXANECARBOXYLIC ACID, TRYPSIN-INHIBITORS, ON THE GROWTH OF BACILLUS-SUBTILIS

Various aromatic esters of trans-4-guanidinomethylcyclohexanecarboxyli c acid (GMCHA), trypsin inhibitors, strongly inhibited the growth of B acillus subtilis 558 and their effects were markedly affected by the s pecies of substitution on the phenyl nucleus of the GMCHA phenyl ester s. 4-tert-Butylphenyl ester of GMCHA (GMCHA-OPh(t)Bu), a representativ e of various GMCHA esters, dose-dependently inhibited the growth of B. subtilis and DNA, RNA and protein syntheses in the cells. The growth inhibition was preceded by suppressive effect of GMCHA-OPh(t)Bu on DNA synthesis. These results suggested the possible involvement of a tryp sin-like proteinase in DNA synthesis. A trypsin-like proteinase was pa rtially purified from B. subtilis 558 by DEAE-cellulose column chromat ography, ammonium sulfate fractionation and successive chromatographie s on Sephadex G-200, phenyl-Sepharose CL-4B, L-arginine-Sepharose 4B a nd Sephadex G-200 columns. The properties were compared with those of proteinase In, which momentarily appears just before the onset of DNA synthesis and seems to participate in the initiation of DNA replicatio n, and which was purified from E. coli K-12 IAM 1264. The properties o f the proteinase from B. subtilis 558 were similar to proteinase In, h owever, the molecular mass (110000) was different from that of the lat ter (66000). Various GMCHA esters strongly inhibited the proteinase ac tivity and the order of the effects was closely correlated with that o n the cell growth. The proteinase was tentatively called subtilis prot einase In.