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Identification of suitable promoter for sigma factor of Mycobacterium tuberculosis

Abstract

The promoter binding specificity is one of the important characters of transcription by Mycobacterium tuberculosis (Mtb) sigma (σ) factors, which remains unexplored due to limited structural evidences. Our previous study on structural features of Mtb-SigH, consisting of three alpha helices, and its interaction with core RNA polymerase are extended in this work to determine the little known DNA sequence recognition pattern involving its cognate promoters. Here, high resolution X-ray crystallographic structures of the protein-DNA complexes are inspected to determine the tentative DNA-binding helix of σ factor. The binding interface in the available crystal structures is found to be populated mainly with specific residues, such as Arg, Asn, Lys, Gln, Ser. We uncover the helix 3 of Mtb-SigH containing most of these amino acids, which ranges from Arg 64 to Arg 75 forming the predicted active site. The complex of Mtb-SigH:DNA are modelled with 20 promoter sequences. The binding affinity is predicted by scoring these protein-DNA complexes through proximity and interaction parameters obtained by molecular dynamics simulations. The promoters are ranked considering hydrogen bonding, energy of interaction, buried surface area and distance between centers of masses in interaction with the protein. The ranking is validated through in vitro transcription assay. The trends of these selected promoter interactions have shown variation parallel to the experimental evaluation, emphasizing the success of the active site determination along with screening of the promoter strength. The promoter interaction of Mtb-SigH can be highly beneficial in understanding the regulation of gene expression of the pathogen and also extends a solid platform to predict promoters for other bacterial σ factors as well.

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Publication details

The article was received on 30 May 2017, accepted on 11 Sep 2017 and first published on 12 Sep 2017


Article type: Paper
DOI: 10.1039/C7MB00317J
Citation: Mol. BioSyst., 2017, Accepted Manuscript
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    Identification of suitable promoter for sigma factor of Mycobacterium tuberculosis

    A. M. Gupta, S. Mukherjee, A. Dutta, J. Mukhopadhyay, D. Bhattacharyya and S. Mandal, Mol. BioSyst., 2017, Accepted Manuscript , DOI: 10.1039/C7MB00317J

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