title: ATP-Induced Structural Remodeling in the Antiactivator FleN Enables Formation of the Functional Dimeric Form.
abstract: FleN, a P loop ATPase is vital for maintaining a monotrichous phenotype in Pseudomonas aeruginosa. FleN exhibits antagonistic activity against FleQ, the master transcriptional regulator of flagellar genes. Crystal structures of FleN in the apo form (1.66 Å) and in complex with β,γ-imidoadenosine 5'-triphosphate (1.55 Å) reveal that it undergoes drastic conformational changes on ATP binding to attain a structure capable of dimerization. Mutations of the residues that stabilize the binding of ATP were defective in their ability to dimerize and do not inhibit ATP hydrolysis by FleQ. Conversely, the catalytic mutant of FleN, was an efficient inhibitor. These observations posit that the dimer is the functional form of FleN and it is nucleotide binding and not hydrolysis by FleN that is necessary to exert an antagonistic effect against FleQ. Our study shows that ATP-induced dimerization may be a strategy to achieve reversible inhibition of FleQ to fine-tune the function of this activator to an optimal level.
authors.affiliation: Transcription Regulation Lab, Regional Centre for Biotechnology, NCR Biotech Science Cluster, 3(rd) Milestone, Faridabad-Gurgaon Expressway, Faridabad, Haryana 121001, India; Manipal University, Manipal, Karnataka, 576104, India.
authors.lastname: Chanchal
authors.affiliation: Transcription Regulation Lab, Regional Centre for Biotechnology, NCR Biotech Science Cluster, 3(rd) Milestone, Faridabad-Gurgaon Expressway, Faridabad, Haryana 121001, India.
authors.forename: Priyajit
authors.initials: P
authors.lastname: Banerjee
authors.affiliation: Transcription Regulation Lab, Regional Centre for Biotechnology, NCR Biotech Science Cluster, 3(rd) Milestone, Faridabad-Gurgaon Expressway, Faridabad, Haryana 121001, India. Electronic address: deepti@rcb.res.in.
authors.forename: Deepti
authors.initials: D
authors.lastname: Jain
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry* / Q000378:metabolism; D000595:Amino Acid Sequence; D001059:Apoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005407:Flagella / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D015964:Gene Expression Regulation, Bacterial; D006868:Hydrolysis; D007700:Kinetics; D008958:Models, Molecular; D009154:Mutation; D011485:Protein Binding; D000072756:Protein Conformation, alpha-Helical; D000072757:Protein Conformation, beta-Strand; D054730:Protein Interaction Domains and Motifs; D055503:Protein Multimerization; D011550:Pseudomonas aeruginosa / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D015534:Trans-Activators / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D001059:Apoproteins; D001426:Bacterial Proteins; C108098:FleQ protein, Pseudomonas aeruginosa; D011994:Recombinant Proteins; D015534:Trans-Activators; C403326:fleN protein, Pseudomonas aeruginosa; D000255:Adenosine Triphosphate
keywords: FleN; Pseudomonas aeruginosa flagella; antiactivator; crystal structure; transcription