Determination of structure-function relationships and role in virulence of a MerR-type regulator that mediates zinc tolerance in Streptococcus mutans

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Abstract Dental caries is the most prevalent chronic infectious disease, with an estimated treatment cost of ~$70 billion/year. While a multifactorial disease, Streptococcus mutans is recognized as a keystone caries pathogen because of its capacity to modulate the oral biofilm in a way that promotes the establishment of a highly acidogenic (cariogenic) microbiota. The trace metals iron, manganese, and zinc (Zn) play structural, catalytic, and regulatory roles in proteins and so are essential to all forms of life. Conversely, these same metals are toxic when in excess such that the ability to maintain metal homeostasis is a critical aspect of host-pathogen interactions. The toxicity of Zn derives from its top position in the Irving-Williams series of metal stability, which allows Zn to avidly bind to non-cognate metalloproteins rendering them nonfunctional. Because it has both antimicrobial and immunomodulatory properties and relatively low toxicity to mammalian cells, Zn has been used for decades in medicine, including incorporation into oral health products for the treatment of gingivitis, halitosis, and prevention of calculus formation. Recently, it was discovered that S. mutans is intrinsically and substantially more tolerant to toxic levels of Zn than all other streptococci and that this high tolerance is mediated by a unique P-type ATPase exporter that has been named ZccE. Expression of zccE is controlled by a unique MerR-type regulator, ZccR for zccE regulator, that strongly activates zccE transcription in response to high Zn stress. Because both ZccE and ZccR are unique to S. mutans, the Lemos lab proposes that both can be targeted for the development of a Zn-based therapy tailored against S. mutans. While other projects in the Lemos lab are exploring the antimicrobial potential of ZccE, the specific aims of this application center around the ZccR regulator. The goals of this application are to determine the structure-function relationships of ZccR and explore its potential as an antimicrobial target. To accomplish these goals, the PI will (i) use crystallography and computational-based approaches to determine the structure of ZccR and identify its critical functional residues, and (ii) utilize the rat model to determine the contribution of ZccR to oral biofilm colonization and caries development alone or in combination with topical Zn treatment. Knowledge gained from this study will facilitate the structure-guided design of small molecule inhibitors for ZccR and reveal the potential of targeting mechanisms of Zn homeostasis to combat S. mutans infections, with the long-term objective of developing novel therapies for the prevention of dental caries and treatment of systemic S. mutans infections (i.e., infective endocarditis). This work will be conducted in a highly supportive and collaborative research environment with Aim 1 establishing a collaboration between the Lemos and McKenna labs. Moreover, the comprehensive training plan provided will accelerate the PI's academic and scientific growth through participation in formal course work and workshops, opportunities to learn new methodologies, mentoring students, establishing new collaborations, and improving scientific writing and presentation skills.