The ability to adapt to acidic conditions is a primary mechanism that Streptococcus mutans employs to survive the low pH environments that can occur in dental plaque. As part of that adaptive ability, the organism increases production of an enzyme, known as an exonuclease, which begins the repair of its DNA damaged during growth under acid conditions. Homologues of the enzyme, in other bacteria, are known to play a major role in the removal and repair of DNA damaged by oxidative agents, such as hydrogen peroxide. The investigators, and others, have shown that growth at low pH increases the ability of S. mutans to resist hydrogen peroxide. Thus, there is a link in S. mutans between resistance to acid conditions and the repair of damage caused by oxidative agents. Since many healthcare products now contain hydrogen peroxide, it is of interest to determine the mechanisms and abilities that S. mutans has available to resist its effects. Moreover, it appears that the organism has evolved mechanisms that are cross-protective, or over-lap, in their regulatory schemes. The objectives of the present application are to clarify how the organism becomes resistant to oxidative attack when it is growing in acidic conditions. The investigators will determine the regulatory mechanism and the biochemical abilities of the exonuclease to protect S. mutans from acidic and oxidative attack. The investigators goals will be accomplished by developing additional mutant strains of the bacteria; by developing site-specific mutations in the exonuclease; and by determining the mechanism by which S. mutans regulates exonuclease production at the level of transcriptional control.
Specific Aims for the project are as follows: 1. Determination of how acidic conditions and oxidative damage act to induce, exonuclease synthesis and a determination of the identity of other participants, in addition to exonuclease, in the inducible repair system; 2. Definition of the kinds of damage that the exonuclease can recognize in DNA and the nature of the exonuclease structure that permits the enzyme its flexibility; 3. Elucidation of the regulatory mechanism for the S. mutans exonuclease, including a description of the genetic elements that participate in regulating synthesis of the enzyme and the protein(s) that function to regulate transcription of the exonuclease gene.
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