Crystal Structures of Multidrug Binding Protein TtgR in Complex with Antibiotics and Plant Antimicrobials
| Autor: | Wilson Terán, Yilmaz Alguel, María-Trinidad Gallegos, Cuixiang Meng, Xiaodong Zhang, Tino Krell, Juan L. Ramos |
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| Rok vydání: | 2007 |
| Předmět: |
crystal structure
antibiotic resistance Protein Conformation Antibiotic resistance Phloretin Molecular Sequence Data Molecular Conformation Calorimetry Crystallography X-Ray Ligands Article chemistry.chemical_compound Protein structure Anti-Infective Agents Bacterial Proteins Structural Biology multidrug binding Amino Acid Sequence Binding site Molecular Biology Binding Sites Sequence Homology Amino Acid biology Binding protein Pseudomonas Cooperative binding Drug Resistance Microbial Isothermal titration calorimetry Plants ITC biology.organism_classification Protein Structure Tertiary Repressor Proteins Multidrug binding chemistry Biochemistry protein–ligand interaction Crystal structures Protein–ligand interaction Efflux Carrier Proteins |
| Zdroj: | Digital.CSIC. Repositorio Institucional del CSIC instname Journal of Molecular Biology |
| ISSN: | 0022-2836 1746-6326 |
| DOI: | 10.1016/j.jmb.2007.03.062 |
| Popis: | 12 pages, 5 figures, 2 tables.-- PMID: 17466326 [PubMed].-- Printed version published Jun 8, 2007. Supporting information (Suppl. figures S1-S2) available at Appendix A: http://dx.doi.org/10.1016/j.jmb.2007.03.062 All the structural data have been deposited to the RCSB Protein Data Bank through European Bioinformatics Institute and are available under accession codes 2UXP, 2UXO, 2UXI, 2UXU, 2UXH. Antibiotic resistance is a widely spread phenomenon. One major mechanism that underlies antibiotic resistance in bacteria is the active extrusion of toxic compounds through the membrane-bound efflux pumps that are often regulated at the transcriptional level. TtgR represses the transcription of TtgABC, a key efflux pump in Pseudomonas putida, which is highly resistant to antibiotics, solvents and toxic plant secondary products. Previously we showed that TtgR is the only reported repressor that binds to different classes of natural antimicrobial compounds, which are also extruded by the efflux pump. We report here five high-resolution crystal structures of TtgR from the solvent-tolerant strain DOT-T1E, including TtgR in complex with common antibiotics and plant secondary metabolites. We provide structural basis for the unique ligand binding properties of TtgR. We identify two distinct and overlapping ligand binding sites; the first one is broader and consists of mainly hydrophobic residues, whereas the second one is deeper and contains more polar residues including Arg176, a unique residue present in the DOT-T1E strain but not in other Pseudomonas strains. Phloretin, a plant antimicrobial, can bind to both binding sites with distinct binding affinities and stoichiometries. Results on ligand binding properties of native and mutant TtgR proteins using isothermal titration calorimetry confirm the binding affinities and stoichiometries, and suggest a potential positive cooperativity between the two binding sites. The importance of Arg176 in phloretin binding was further confirmed by the reduced ability of phloretin in releasing the mutant TtgR from bound DNA compared to the native protein. The results presented here highlight the importance and versatility of regulatory systems in bacterial antibiotic resistance and open up new avenues for novel antimicrobial development. We are grateful to the Human Frontier Science Programme for its generous support to X.Z. and M.T.G. that allowed the project to initiate. We also thank the Royal Society, which supported the collaborative visits between X.Z. and M.T.G. X.Z. acknowledges the Medical Research Council for its financial support. M.T.G. acknowledges the Spanish Ministry of Education and Science for its financial support. |
| Databáze: | OpenAIRE |
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