Characterisation of the putative effector interaction site of the regulatory HbpR protein from Pseudomonas azelaica by site-directed mutagenesis
| Autor: | Jan Roelof van der Meer, Rup Lal, Christelle Vogne, Sagrario Arias, Hansi Bisht, Sofía Fraile |
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| Jazyk: | angličtina |
| Rok vydání: | 2011 |
| Předmět: |
Models
Molecular Protein Folding Subfamily DNA transcription Amino Acid Substitution/genetics Amino Acid Substitution/physiology Bacterial Proteins/chemistry Bacterial Proteins/genetics Binding Sites/genetics Computational Biology Forecasting Models Biological Mutagenesis Site-Directed/methods Organisms Genetically Modified Protein Binding/genetics Protein Interaction Mapping/methods Protein Structure Tertiary/genetics Pseudomonas/genetics Trans-Activators/chemistry Trans-Activators/genetics lcsh:Medicine Plasma protein binding Biology Microbiology chemistry.chemical_compound Bacterial Proteins Bacterial transcription RNA polymerase Pseudomonas Protein Interaction Mapping Genetics Bacterial Physiology Binding site lcsh:Science Site-directed mutagenesis Multidisciplinary Binding Sites Effector lcsh:R Bacteriology Cell biology Bacterial Biochemistry Protein Structure Tertiary chemistry Amino Acid Substitution Mutagenesis Site-Directed Trans-Activators lcsh:Q Protein folding Gene expression Research Article Protein Binding |
| Zdroj: | PLoS One, vol. 6, no. 2, pp. e16539 PLoS ONE PLoS ONE, Vol 6, Iss 2, p e16539 (2011) |
| Popis: | Bacterial transcription activators of the XylR/DmpR subfamily exert their expression control via σ(54)-dependent RNA polymerase upon stimulation by a chemical effector, typically an aromatic compound. Where the chemical effector interacts with the transcription regulator protein to achieve activation is still largely unknown. Here we focus on the HbpR protein from Pseudomonas azelaica, which is a member of the XylR/DmpR subfamily and responds to biaromatic effectors such as 2-hydroxybiphenyl. We use protein structure modeling to predict folding of the effector recognition domain of HbpR and molecular docking to identify the region where 2-hydroxybiphenyl may interact with HbpR. A large number of site-directed HbpR mutants of residues in- and outside the predicted interaction area was created and their potential to induce reporter gene expression in Escherichia coli from the cognate P(C) promoter upon activation with 2-hydroxybiphenyl was studied. Mutant proteins were purified to study their conformation. Critical residues for effector stimulation indeed grouped near the predicted area, some of which are conserved among XylR/DmpR subfamily members in spite of displaying different effector specificities. This suggests that they are important for the process of effector activation, but not necessarily for effector specificity recognition. |
| Databáze: | OpenAIRE |
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