Autolytic Enzyme System of Streptococcus faecalis III. Localization of the Autolysin at the Sites of Cell Wall Synthesis

Abstract

Cell walls from exponential-phase cultures of S. faecalis ATCC 9790 autolyzed in dilute buffers. Walls were isolated from cultures grown in the presence of l4C-lysine for about 10 generations and then on 12c-lysine for 0.1-0.8 of a generation (prelabeled). These walls released 14C to the soluble fraction more slowly than they lost turbidity during the initial stages of autolysis. Walls isolated from cultures grown in the presence of l4C-lysine for only the last 0.1-0.4 of a generation (postlabeled) released 14C to the supernatant fluid more rapidly than they lost turbidity. Autolysin in both pre- and postlabeled walls was inactivated, and such walls were then incubated in the presence of unlabeled walls containing active autolysin. The inactivated walls lost their l4C label only very slowly until autolysis of the unlabeled walls was virtually complete and release of soluble autolysin was expected. When this experiment was done in the presence of trypsin, a 4-fold increase in the autolysis rate resulted; but the same pattern of 14C release was observed. A parallel release of l4C and loss of turbidity from pre- or postlabeled walls was observed upon trypsin "activation" and by addition of isolated soluble autolysin to inactivated walls. We conclude that the wall-bound autolysin acts 1st on the more recently synthesized portion of the wall. Trypsin appears to speed wall autolysis by activating additional latent autolysin in situ at sites in the older portion of the wall.