A SPORE-LYTIC ENZYME RELEASED FROM BACILLUS-CEREUS SPORES DURING GERMINATION

The exudate of fully germinated spores of Bacillus cereus IFO 13597 in 0.25 M sodium phosphate buffer, ph 7.0, was found to contain a spore-lytic enzyme. This enzyme was found to cause loss of absorbance in coat-stripped spore suspensions and phase-darkening of the spores but had minimal activity on isolated peptidoglycan substrates. The enzyme was purified in an active form and identified as a 24 kDa protein which is either an amidase or a peptidase. The amino-terminal 19 residues had the following sequence: FSNQVIQRGASGEKVIELQ. The spore-lytic enzyme retained its activity in a medium of a relatively high ionic strength containing a non-ionic surfactant such as nonaethyleneglycol n-dodecyl ether. This activity was optimum at a salt concentration of about 30 mM in assay buffer at neutral ph. In contrast to the enzyme in a spore-bound form, the enzyme in solution was shown to be heat-sensitive and was readily inactivated by thiol reagents.