The goal of this Project is to develop assays and identify compounds that will be useful as research tools for studies of Mycobacterium tuberculosis (Mtb), and as potential drugs against tuberculosis (TB). It is anticipated that this approach will provide assay and chemical inputs into the pipeline for the development of new lead molecules for the treatment of TB infection, and for Mtb research. Compounds identified through the assay development and screening efforts of this Project, will be useful as research tools, providing important insights about the biological functions and structures of targeted Mtb membrane proteins, and will may be useful for therapeutic developments. This Project will focus on an Initial Target List of essential Mtb membrane proteins, selected for their functional importance, developed for this Program Project.
The specific aims are (1) to develop biochemical assays for Mtb membrane proteins and perform high throughput screening of small molecule libraries, (2) to perform NMR screening of small molecule libraries with the Mtb proteins in micelles, and (3) to develop and perform NMR screening of small molecule libraries for the Mtb proteins in bilayers and bicelles. Biochemical assays and NMR-based methods for screening will be developed. The biochemical assays will facilitate further functional characterization, and enable the screening of compounds that interfere with activity. NMR will be used to screen for small molecules that bind to Mtb proteins even before a protein function is fully annotated. The identification of weak binders by this method is very useful for both the function and structure determination processes in Projects 1 and 3, and has the potential of identifying drug- like lead compounds for therapeutic development. NMR screening is well developed for globular proteins in aqueous solutions, but it has not been applied to membrane proteins in micelles or membrane proteins in lipid bilayers, and developing NMR screening for membrane proteins in lipid environments is a major goal of this Project.
|Gong, Xiao-Min; Ding, Yi; Yu, Jinghua et al. (2015) Structure of the Na,K-ATPase regulatory protein FXYD2b in micelles: implications for membrane-water interfacial arginines. Biochim Biophys Acta 1848:299-306|
|Jones, Christopher M; Wells, Ryan M; Madduri, Ashoka V R et al. (2014) Self-poisoning of Mycobacterium tuberculosis by interrupting siderophore recycling. Proc Natl Acad Sci U S A 111:1945-50|
|Tian, Ye; Lu, George J; Marassi, Francesca M et al. (2014) Structure of the membrane protein MerF, a bacterial mercury transporter, improved by the inclusion of chemical shift anisotropy constraints. J Biomol NMR 60:67-71|
|Kolocouris, Antonios; Tzitzoglaki, Christina; Johnson, F Brent et al. (2014) Aminoadamantanes with persistent in vitro efficacy against H1N1 (2009) influenza A. J Med Chem 57:4629-39|
|Wu, Chin H; De Angelis, Anna A; Opella, Stanley J (2014) Magic angle Lee-Goldburg frequency offset irradiation improves the efficiency and selectivity of SPECIFIC-CP in triple-resonance MAS solid-state NMR. J Magn Reson 246:1-3|
|Danilchanka, Olga; Sun, Jim; Pavlenok, Mikhail et al. (2014) An outer membrane channel protein of Mycobacterium tuberculosis with exotoxin activity. Proc Natl Acad Sci U S A 111:6750-5|
|Murray, Dylan T; Griffin, James; Cross, Timothy A (2014) Detergent optimized membrane protein reconstitution in liposomes for solid state NMR. Biochemistry 53:2454-63|
|Jean-Francois, Frantz L; Dai, Jian; Yu, Lu et al. (2014) Binding of MgtR, a Salmonella transmembrane regulatory peptide, to MgtC, a Mycobacterium tuberculosis virulence factor: a structural study. J Mol Biol 426:436-46|
|Siegrist, M Sloan; Steigedal, Magnus; Ahmad, Rushdy et al. (2014) Mycobacterial Esx-3 requires multiple components for iron acquisition. MBio 5:e01073-14|
|Das, Bibhuti B; Zhang, Hua; Opella, Stanley J (2014) Dipolar Assisted Assignment Protocol (DAAP) for MAS solid-state NMR of rotationally aligned membrane proteins in phospholipid bilayers. J Magn Reson 242:224-32|
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