Principles of Protein Mimicry of DNA

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DESCRIPTION (provided by applicant): This revised application of a new proposal describes a comprehensive biochemical and biophysical investigation into the `DNA mimicry' of a recently discovered member of the Pentapeptide Repeat Protein (PRP) family of proteins. The atomic resolution structure of the MfpA PRP from Mycobacterium tuberculosis revealed a new protein fold; MfpA's right-handed 2-helical coils yield an elongated structure that is a remarkable mimic of the size, shape and electrostatic surface of DNA. That DNA mimicry is important to the biological role of MfpA is suggested by its ability to confer resistance to fluoro- quinolones by binding DNA gyrase thereby inhibiting its function. We will test the hypothesis that mimicry of the physical properties DNA by MfpA is the heart of its ability to confer drug resistance. Our long term goal is to understand the molecular mechanism by which MfpA and other PRP mediate antibiotic resistance and potentially regulate the enzymatic processing of DNA. MfpA will be developed as a model system for the exploration of DNA mimicry by this newly-discovered and widely-distributed class of proteins. Fluoroquinolones are extremely important antibiotics due to their efficacy against Gram- positive, Gram-negative and mycobacteria. Fluoroquinolones act on DNA gyrase with a unique mechanism of action. They are remarkable for their absence of toxicity and a therapeutic index that is the highest of all currently prescribed antibiotics. The rapid emergence, and spread, of transmissible forms of fluoroquinolone resistance due to the expression of the plasmid-encoded proteins threatens the clinical utility of these antibiotics. The proposed studies seek to understand how the physical properties of the Peptapeptide Repeat Proteins MfpA and Qnr generate resistance to fluoroquinolones. This understanding would provide new targets for therapeutics complementary to fluoroquinolone that would counter the PRP-mediated resistance. Conversely, an understanding of the mechanism by which these proteins function may allow the PRP fold to be used as a platform for the development of novel protein or peptide therapeutics.