title: The α10 helix of DevR, the Mycobacterium tuberculosis dormancy response regulator, regulates its DNA binding and activity.
abstract: The crystal structures of several bacterial response regulators provide insight into the various interdomain molecular interactions potentially involved in maintaining their 'active' or 'inactive' states. However, the requirement of high concentrations of protein, an optimal pH and ionic strength buffers during crystallization may result in a structure somewhat different from that observed in solution. Therefore, functional assessment of the physiological relevance of the crystal structure data is imperative. DevR/DosR dormancy regulator of Mycobacterium tuberculosis (Mtb) belongs to the NarL subfamily of response regulators. The crystal structure of unphosphorylated DevR revealed that it forms a dimer through the α5/α6 interface. It was proposed that phosphorylation may trigger extensive structural rearrangements in DevR that culminate in the formation of a DNA-binding competent dimeric species via α10-α10 helix interactions. The α10 helix-deleted DevR protein (DevR∆α10 ) was hyperphosphorylated but defective with respect to in vitro DNA binding. Biophysical characterization reveals that DevR∆α10 has an open but less stable conformation. The combined cross-linking and DNA-binding data demonstrate that the α10 helix is essential for the formation and stabilization of the DNA-binding proficient DevR structure in both the phosphorylated and unphosphorylated states. Genetic studies establish that Mtb strains expressing DevR∆α10 are defective with respect to dormancy regulon expression under hypoxia. The present study highlights the indispensable role of the α10 helix in DevR activation and function under hypoxia and establishes the α10-α10 helix interface as a novel target for developing inhibitors against DevR, a key regulator of hypoxia-triggered dormancy.
authors.affiliation: Department of Biotechnology, All India Institute of Medical Sciences, New Delhi, India.|Experimental Animal Facility, National JALMA Institute of Leprosy and Other Mycobacterial Diseases, Tajganj, Agra, India.
authors.forename: Atul
authors.initials: A
authors.lastname: Vashist
authors.affiliation: Department of Biotechnology, All India Institute of Medical Sciences, New Delhi, India.
authors.forename: D
authors.initials: D
authors.lastname: Prithvi Raj
authors.affiliation: Experimental Animal Facility, National JALMA Institute of Leprosy and Other Mycobacterial Diseases, Tajganj, Agra, India.
authors.forename: Umesh Datta
authors.initials: UD
authors.lastname: Gupta
authors.affiliation: School of Biotechnology, Jawaharlal Nehru University, New Delhi, India.
authors.forename: Rajiv
authors.initials: R
authors.lastname: Bhat
authors.affiliation: Department of Biotechnology, All India Institute of Medical Sciences, New Delhi, India.
authors.forename: Jaya Sivaswami
authors.initials: JS
authors.lastname: Tyagi
mesh_terms: D000693:Anaerobiosis; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D002942:Circular Dichroism; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins; D015964:Gene Expression Regulation, Bacterial; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D017510:Protein Folding; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017433:Protein Structure, Secondary; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; C542863:DosR protein, Mycobacterium tuberculosis; D011494:Protein Kinases
keywords: DevR/DosR; M. tuberculosis; biophysical characterization; dormancy regulon; signal transduction