This investigation was supported by a grant from the Office of Naval Research. Portions of it were presented at the 59th General Meeting of the Society of American Bacteriologists, St. Louis, Missouri, May 10 to 15, 1959. 2 This paper represents part of a thesis submitted by the senior author to the University of Illinois in August, 1959, in partial fulfillment of the requirements for the Ph.D. degree.

The membrane-bound, sugar-specific enzyme II The phosphoenolpyruvate:sugar phosphotransferase system (PTS) concomitantly catalyzes the transport and phosphorylation of mono-and disaccharides in many bacteria (23,25). The overall reaction catalyzed by the PTS requires the membrane protein enzyme II (EII) and the general spluble proteins, enzyme I (El) and HPr, and in many instances a sugar-specific, soluble protein called enzyme III, or simply III (22,23,25). The oral streptococci transport a variety of sugars via the PTS (1, 13, '26, 29), and recent studies (7,18,19,30,32,33) have been directed at the isolation and characterization of the components from these bacteria, particularly Streptococcus mutans.Over the past decade, continuous-culture studies with S. mutans Ingbritt have demonstrated that the glucose-PTS is repressed by a variety of changes in the growth environment under conditions in which glucose uptake and metabolism was either unaffected or even enhanced. For example, changes in growth conditions to acidic pHs (12), increased growth rates (4), limiting glucose to excess glucose (4), or limiting sucrose (3) resulted in repression of glucose-PTS activity when assayed with decryptified cells. These and other results (8,27) have led to the suggestion, that the organi §m possesses an alternate, non-PTS transport system supplementing the glucose-PTS, which is active at low pH, at high growth rates, and in the presence of low concentra-* Corresponding author.tions of sucrose and excess glucose. Subsequent research with S. mutans and other strains of oral streptococci have indicated that the alternate, non-PTS transport system is driven by the gradient energy in t-he proton motive force (10,14,16,17,21).We have now undertaken to study the components of the glucose-PTS in chemostat-grown cells of S. mutans Ingbritt in an effort to determine which element of the system is subject to regulation by environmental factors. In addition, experiments have been conducted with chemostat cultures of glucose-grown S. mutans DR0001 and lactose-grown Streptococcus sobrinus (formerly S. mutans 27352 [33]) to determine whether these same factors influence the components of the PTS in these organisms. Previous research with S. mutans 27352 has demonstrated that transport of lactose by the organism requires the induction of not only a specific EII for lactose (EIILaC) but also the soluble component 111Lac (33), a factor which appears to be an integral part of the lactose-PTS in a variety of strains of S. mutans (34). Furthermore, repression of EII activity for glucose and fructose has also been reported with S. mutans and S. sobrinus growing on lactose in batch cultures (34). The activity of EII was measured in isolated membrane preparations in the presence of excess' EI and HPr while the concentration of the latter components and 111Lac were determined by immunoelectrophoresis. Our results indicate that the general proteins HPr, El, and I11Lac are relatively immune to changes in environmental conditions; however, synthesi...

The effects of nutritional and environmental conditions on sporulation and the properties of the spore have been extensively studied with the hope that these may provide a clue to the mechanism of dormancy and resistance of the bacterial endospore. However, there is no general agreement and our knowledge of these is still limited. The extensive literature dealing with the various aspects of sporogenesis has been reviewed by Cook (1932); Knaysi (1948); Williams (1952); Stedman (1956); and Halvorson (1957b).

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The germination of Bacillus cereus T spore suspensions is partially prevented by several inhibitors of trypsin-like enzymes. Leupeptin, antipain, and tosyl-lysine-chloromethyl ketone are effective inhibitors, whereas chymostatin, elastatinal, and pepstatin are inactive. A synthetic substrate of trypsin, tosyl-arginine-methyl ester, also inhibits germination. Its inhibitory effect decreases as a function of incubation time in the presence of spores and is abolished by previous hydrolysis with trypsin. Germinating, but not dormant, spore suspensions hydrolyze tosyl-arginine-methyl ester; its hydrolysis is insensitive to chloramphenicol, sulfhydryl reagents, and EDTA. A crude extract of germinated B. cereus spores contains a trypsin-like enzyme whose activity, as measured by hydrolysis of benzoyl-arginine p-nitroanilide, is sensitive to germination-inhibitory compounds such as leupeptin, tosyl-arginine-methyl ester, and tosyl-lysine-chloromethyl ketone. Spore suspensions exposed to the above inhibitors under germination conditions lose only part of their heat resistance and some 10 to 30% of their dipicolinic acid content. Part of the germinating spore population becomes "phase grey" under phase optics. Based on a study of the inhibition of germination by protease inhibitors and the activity of a protease in germination spores and spore extracts, it is suggested that the activity of a trypsin-like enzyme may be involved in the mechanism of the breaking of dormancy in spores of B. cereus T.