Autolytic defective mutant of Streptococcus faecalis

Abstract

Properties of a variant of Streptococcus faecalis ATCC 9790 with defective cellular autolysis are described. The mutant strain was selected as a survivor from a mutagenized cell population simultaneously challenged with two antibiotics which inhibit cell wall biosynthesis, penicillin G and cycloserine. Compared to the parental strain, the mutant strain exhibited: (i) a thermosensitive pattem of cellular autolysis; (ii) an autolytic enzyme activity that had only a slightly increased thermolability when tested in solution in the absence of wall substrate; and (iii) an isolated autolysin that had hydrolytic activity on isolated S. faecalis wall substrate indistinguishable from that of the parental strain, but that was inactive when tested on walls of Micrococcus lysodeikticus as a substrate. These data indicate an alteration in the substrate specificity of the autolytic enzyme of the mutant which appears to result from the synthesis of an altered form of autolytic enzyme. 'o.2 S.A.

Evidence for the presence of a second peptidoglycan hydrolase in Streptococcus faecium

“…Similarly, evidence indicating the presence of two murein transglycosylases in E. coli has been recently reported [19]. Further studies of the autolysis-defective mutants at hand [9][10][11] and attempts to obtain conditional, autolysis-defective, second-stage mutants of Lyt-14 should be revealing.…”

“…Units of activity [4,10]. In contrast, although ConA-Sepharosepurified enzyme rapidly hydrolyzed and dissolved its homologous wall substrate, it dissolved walls of M. luteus only extremely slowly (Table 1).…”

“…Since muramidase-1 is a glycoenzyme (Kawamura and Shockman, in preparation), it is possible that PG hydrolase-2 is a non-glycosylated form of muramidase-1 with a different substrate specificity profile. Because of (i) the several differences in the two activities summarized above, and the previous isolation of a lytic defective mutant, Lyt-14 [10], from which extracted crude autolysin activity dissolved walls of S. faecium but failed to dissolve walls of M. luteus, we believe that muramidase-I and PG hydrolase-2 are separate gene products. The data presented here clearly show the presence of two PG hydrolase activities in S. faecium.…”

“…A large body of evidence [4][5][6][7][8], including detailed analyses of the products of autolytic dissolution of cell walls of Streptococcus faecium ATCC9790, is consistent with the presence and action of only PG hydrolase activity that cleaves the fl-l,4 linkage between N-acetylmuramic acid and N-acetylglucosamine (an N-acetylmuramoylhydrolase; muramidase). However, several efforts to obtain truly lytmutants [9][10][11][12][13] of S. faecium were unsuccessful. Recently, while isolating, purifying and characterizing the autolytic muramidase (muramidase-l) of S. faecium (Kawamura and Shockman, in preparation) we discovered a second PG hydrolase (PG hydrolase-2) in S. faecium.…”

Variation between penicillin-tolerant and penicillin-sensitive states in Streptococcus faecium ATCC9790

“…The biochemical basis for tolerance in the S and T derivatives requires further study. They differ from the lytic defective derivatives isolated by mutagenesis of the 9790-S strain [5][6][7]. First, the Aut-1 and Lyt-14 derivatives had substantial rates of lysis in phosphate buffer [6], whereas our derivatives do not (Table 3).…”