title: Role of the recognition helix of response regulator WalR from Bacillus anthracis in DNA binding and specificity.
abstract: WalRK two-component system of Bacillus anthracis potentially regulates multiple genes spanning diverse cellular functions. Its constituent response regulator (RR), WalR belongs to the OmpR/PhoB family which possesses a winged helix-turn-helix motif for DNA binding. An in silico knowledge based model of WalR C-terminal DNA binding domain in complex with its ftsE promoter region binding motif was used to identify specific residues of the recognition helix important for DNA binding. The model was validated by mutagenesis in conjunction with in vitro DNA binding analysis. The ftsE promoter region DNA binding motif was also varied. Optimal binding of WalR to DNA required the presence of both half-sites in its binding motif. Substitution of invariant bases of WalR DNA binding motif abrogated the binding whereas changes at variable motif positions governed affinity. D199 was not in direct contact with the DNA but its substitution modified the WalR-DNA specificity indicating the importance of contact avoidance by this residue for DNA specificity. This represents the first in-depth study of RR-DNA interaction from B. anthracis.
authors.affiliation: Laboratory of Molecular Biology and Genetic Engineering, School of Biotechnology, Jawaharlal Nehru University, New Delhi, India.
authors.forename: Alisha
authors.initials: A
authors.lastname: Dhiman
authors.affiliation: Laboratory of Molecular Biology and Genetic Engineering, School of Biotechnology, Jawaharlal Nehru University, New Delhi, India.
authors.forename: Amit
authors.initials: A
authors.lastname: Rahi
authors.affiliation: Laboratory of Molecular Biology and Genetic Engineering, School of Biotechnology, Jawaharlal Nehru University, New Delhi, India.
authors.forename: Monisha
authors.initials: M
authors.lastname: Gopalani
authors.affiliation: GE Healthcare Life Sciences, Gurgaon, Haryana, India.
authors.forename: Sailesh
authors.initials: S
authors.lastname: Bajpai
authors.affiliation: Computational and Structural Biology Lab., Division of Biotechnology, Netaji Subhas Institute of Technology, Dwarka, New Delhi, India. Electronic address: ecc999@gmail.com.
authors.forename: Sonika
authors.initials: S
authors.lastname: Bhatnagar
authors.affiliation: Laboratory of Molecular Biology and Genetic Engineering, School of Biotechnology, Jawaharlal Nehru University, New Delhi, India. Electronic address: rakeshbhatnagar@jnu.ac.in.
authors.forename: Rakesh
authors.initials: R
authors.lastname: Bhatnagar
mesh_terms: D001408:Bacillus anthracis; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D001483:Base Sequence; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D000072417:Protein Domains; D013379:Substrate Specificity; D020080:Tandem Repeat Sequences
chemical_list: D001426:Bacterial Proteins; C119939:YycF protein, Bacteria; D004247:DNA
keywords: Recognition helix; Two component system; WalR