A soluble autolysin has been isolated from cell walls of exponential (log) phase cultures of Streptococcus faecalis ATCC 9790 by means of gel filtration of trypsin-activated wall lysates. Trypsin and other proteolytic enzymes speed wall autolysis apparently by "activating" the autolysin. The autolysin seems to be closely associated with the wall as virtually complete wall dissolution is a prerequisite for obtaining the enzyme in solution. While the soluble autolysin can act on walls of S. faecalis and Micrococcus lysodeikticus, it is virtually inactive on walls from several other bacterial species, Borohydride reduction indicates P revious studies (Shockman, 1963b(Shockman, , 1965Shockman et al., 1958 Shockman et al., , 1961Toennies and Shockman, 1958;Conover et al., 1966) have demonstrated that cells from exponential phase (log) cultures of Streptococcus faecalis 9790 will autolyze rapidly when placed under environmental conditions which prevent the continued synthesis of cell wall peptidoglycan (mucopeptide, glycopeptide, or murein). Inhibition of peptidoglycan synthesis (resulting in autolysis) can be caused either by the addition of a specific inhibitor, such as penicillin or cycloserine, or by deprivation of a nutritionally essential precursor of wall peptidoglycan, such as L-lysine, D-alanine, aspartic and glutamic acids, or glucose.The potential for cell autolysis reached a maximum toward the end of the exponential growth phase and thereafter the cells became resistant to autolysis (Shockman, 1965). Stationary-phase, valine-deprived (Val) or threonine-deprived (Thr) cultures resist autolysis (Shockman, 1965 ; Shockman et al., 1961).Two observations implicated an enzymatic attack on the rigid and protective cell wall as the initial step in cell autolysis. First, when an osmotically protective agent, such as 0.5 M sucrose, was provided, wallfree spherical, osmotically fragile protoplasts could be obtained from log cells, but not from stationaryphase Thr cells in the absence of an exogenous lytic ~~~

Previous studies have demonstrated that cell wall synthesis may continue after exponential growth of Streptococcus faecalis has stopped. This can occur when an otherwise complete culture medium is depleted of the available supply of one or more nutritionally indispensable amino acids which are major components of protoplasmic proteins but not of the cell wall (Shockman, Kolb, and Toennies, 1958; Shockman, 1959). On the other hand, rapid lysis tends to occur when an indispensable amino acid that is a major component of the cell wall, such as L-lysine or D-alanine (in a vitamin B6-deficient medium) is exhausted from the growth medium (Toennies and Gallant, 1949a; Shockman et al., 1958). These studies suggested that a) in the absence of the continued synthesis of cell wall substance the cells of this organism are subject to autolysis and that b) cell wall synthesis may be, at least in part, independent of the synthesis of cytoplasmic proteins. The results presented here and in the following paper (Shockman et al., 1961) provide additional evidence supporting these hypotheses. MATERIALS AND METHODS Streptococcus faecalis (ATCC 9790) was grown at 38 C on the previously described highly buffered defined medium (Toennies and Gallant, 1948; Shockman et al., 1958; Toennies and

In the preceding paper (Shockman et al., 1961) the lytic properties of exponentially growing cells of Streptococcus faecalis have been described. Not only will washed and transferred exponential cells lyse but, under proper conditions, they eaD engage in either exponential growth or in postexponential cell wall synthesis. Some of the necessary conditions, and evidence for the synthesis of cell wall substances will be presented here. MATERIALS AND METHODS Methods of growth, washing, and transfer of S. faecalis (ATCC 9790) were similar to those previously described (Shockman et al., 1961). However, for the experiments reported here, an increase in turbidity was expected during the second incubation, so that a starting adjusted optical density of about 400 (0.16 mg/ml) was

The membrane fraction of Streptococcus faecalis (ATCC 9790) was isolated and purified, by a variety of procedures, from cultures that were grown under closely controlled conditions of physiological age and nutrition. The most satisfactory method required the use of lysozyme-to-cell ratios below 0.01 and the intermediate formation of protoplasts in osmotically protective media. Amino acid analyses of three of the membrane preparations indicated a characteristic and constant, but not unusual, pattern; 42% of the membranes from threonine

No abstract