This application is for support of a project focused on the acid-base physiology of plaque bacteria, especially the mutans streptococci, but also Lactobacillus casei and arginine-deiminase positive streptococci. The proposed work arises from previous studies the acid tolerance with particular attention on proton-translocating F-ATPases. A basic hypothesis is that the more acid-tolerant bacteria in plaque are also the more cariogenic. To be cariogenic, a bacterium must be able to colonize plaque, become a significant part of the flora, and produce acid. However, since the challenge to the tooth is exponentially related to the extent of acidification of plaque, those organisms which can lower the plaque pH to values as low as .4 are particularly damaging. The results of our prior work indicated two adaptations in F-ATPases for acid tolerance. More tolerant organisms have higher F-ATPase activities per unit of cell weight, and the enzymes have lower pH optima for activity. Plans are to extend these results with other organisms, especially the acid-tolerant, non-mutants, plaque streptococci. The proposed work will now consider proton-translocating efficiencies of the enzymes in terms of coupling ratios i.e., how many protons are transported per ATP hydrolyzed, means to inhibit the enzyme and to affect the efficiency of coupling, and the possible involvement of lactate transport in proton currents. Membrane vesicles isolated from Streptococcus mutans GS-5 will be used extensively for the proposed work, as will bacteria grown in biofilms under a variety of conditions. In addition, isolated genes for the atp operon of S. mutans will be used for studies of regulation of F-ATPase synthesis. In the overall acid-base cycles of plaque, ammonia production by arginine-deiminase-positive bacteria is considered important for protecting less acid-tolerant bacteria against acid damage and for reducing cariogenicity. Studies are proposed on regulation of the arginine deiminase system (ADS) by redox potential in S. sanguis and S. rattus, on ADS-negative mutants, on cells grown in biofilms and on transmembrane transport of arginine peptides. The results of our previous work and that of others has indicated overlap in the adaptations of plaque streptococci for protection against acid damage and against oxidative damage caused, for example, by hydroperoxides. We proposed to define in more detail the nature of acid damage, especially to the glycolytic system, and the relationship between acid damage and oxidative damage. A major objective will be to develop means, for example, with enhancers of oxidative damage, to reduce acid tolerance and cariogenicity.
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