The emergence of multidrug resistant strains of bacteria is a serious and growing threat to human health. One component of multidrug resistance is the presence of multidrug efflux transporters that expel drugs from the cell thereby keeping their cytosolic levels below toxic concentrations. Nearly all multidrug efflux pump genes are regulated locally by transcription factors that are induced or activated upon binding the very same structurally and chemically dissimilar drugs, which are substrates of the pumps. Thus, these transcription regulators act as intracellular multidrug sensors that increase the number of multidrug efflux pumps when the cell is threatened by an increasing drug dose. Because of their increased solubility, these proteins are outstanding systemsto understand the structural mechanisms of multidrug binding and multidrug gene regulation. In this competing renewal application, x-ray crystallographic studies on two multidrug binding transcription factors, QacR from S. aureus and BmrR from B. subtilis and two multidrug efflux pump gene regulators, MtaN from B. subtilis and MtrR from N. gonorrhoeae, will be done to dissect fully the structural mechanisms by which a single protein can bind multiple structurally and chemically dissimilar drugs as well as regulate the expression of multidrug efflux pump genes.
The specific aims are:(1) to determine the x-ray structures of QacR and a series of site directed QacR mutants bound to drugs and DNA: (2) to determine the crystal structures of a number of BmrR-drug-DNA complexes and a series of site directed BmrR mutants bound to drugs and DNA as well as the BmrR-bmr operator complex; (3) to determine the structures of site directed mutants of MtaN in order to elucidate its DNA binding mechanism; (4) to crystallize and determine the structures of MtrR bound to its operator site and off DNA.
All specific aims will be supplemented with germane DNA and drug binding studies. These data will be used in future structure-based drug design of novel drugs against pathogenic bacteria, including S. aureus, N. gonorrhoeae and potentially B. anthracis.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
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Physical Biochemistry Study Section (PB)
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Huntley, Clayton C
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University of Texas MD Anderson Cancer Center
Other Domestic Higher Education
United States
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Peters, Kate M; Brooks, Benjamin E; Schumacher, Maria A et al. (2011) A single acidic residue can guide binding site selection but does not govern QacR cationic-drug affinity. PLoS One 6:e15974
Kumaraswami, Muthiah; Newberry, Kate J; Brennan, Richard G (2010) Conformational plasticity of the coiled-coil domain of BmrR is required for bmr operator binding: the structure of unliganded BmrR. J Mol Biol 398:264-75
Kumaraswami, Muthiah; Schuman, Jason T; Seo, Susan M et al. (2009) Structural and biochemical characterization of MepR, a multidrug binding transcription regulator of the Staphylococcus aureus multidrug efflux pump MepA. Nucleic Acids Res 37:1211-24
Hassan, Karl A; Xu, Zhiqiang; Watkins, Ryan E et al. (2009) Optimized production and analysis of the staphylococcal multidrug efflux protein QacA. Protein Expr Purif 64:118-24
Schumacher, Maria A; Piro, Kevin M; Xu, Weijun et al. (2009) Molecular mechanisms of HipA-mediated multidrug tolerance and its neutralization by HipB. Science 323:396-401
Newberry, Kate J; Huffman, Joy L; Miller, Marshall C et al. (2008) Structures of BmrR-drug complexes reveal a rigid multidrug binding pocket and transcription activation through tyrosine expulsion. J Biol Chem 283:26795-804
Peters, Kate M; Schuman, Jason T; Skurray, Ronald A et al. (2008) QacR-cation recognition is mediated by a redundancy of residues capable of charge neutralization. Biochemistry 47:8122-9
Brooks, Benjamin E; Piro, Kevin M; Brennan, Richard G (2007) Multidrug-binding transcription factor QacR binds the bivalent aromatic diamidines DB75 and DB359 in multiple positions. J Am Chem Soc 129:8389-95
Schumacher, Maria A; Allen, Gregory S; Diel, Marco et al. (2004) Structural basis for allosteric control of the transcription regulator CcpA by the phosphoprotein HPr-Ser46-P. Cell 118:731-41
Newberry, Kate J; Brennan, Richard G (2004) The structural mechanism for transcription activation by MerR family member multidrug transporter activation, N terminus. J Biol Chem 279:20356-62

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