ATP-Induced Structural Remodeling in the Antiactivator FleN Enables Formation of the Functional Dimeric Form

Chanchal; Priyajit Banerjee; Deepti Jain

doi:10.1016/j.str.2016.11.022

ATP-Induced Structural Remodeling in the Antiactivator FleN Enables Formation of the Functional Dimeric Form

Structure. 2017 Feb 7;25(2):243-252. doi: 10.1016/j.str.2016.11.022. Epub 2017 Jan 5.

Authors

Chanchal¹, Priyajit Banerjee², Deepti Jain³

Affiliations

¹ 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.
² Transcription Regulation Lab, Regional Centre for Biotechnology, NCR Biotech Science Cluster, 3(rd) Milestone, Faridabad-Gurgaon Expressway, Faridabad, Haryana 121001, India.
³ 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.

PMID: 28065505
DOI: 10.1016/j.str.2016.11.022

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.

Keywords: FleN; Pseudomonas aeruginosa flagella; antiactivator; crystal structure; transcription.

MeSH terms

Adenosine Triphosphate / chemistry*
Adenosine Triphosphate / metabolism
Amino Acid Sequence
Apoproteins / chemistry*
Apoproteins / genetics
Apoproteins / metabolism
Bacterial Proteins / chemistry*
Bacterial Proteins / genetics
Bacterial Proteins / metabolism
Binding Sites
Cloning, Molecular
Crystallography, X-Ray
Escherichia coli / genetics
Escherichia coli / metabolism
Flagella / genetics
Flagella / metabolism
Gene Expression
Gene Expression Regulation, Bacterial*
Hydrolysis
Kinetics
Models, Molecular
Mutation
Protein Binding
Protein Conformation, alpha-Helical
Protein Conformation, beta-Strand
Protein Interaction Domains and Motifs
Protein Multimerization
Pseudomonas aeruginosa / genetics*
Pseudomonas aeruginosa / metabolism
Recombinant Proteins / chemistry
Recombinant Proteins / genetics
Recombinant Proteins / metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Substrate Specificity
Trans-Activators / chemistry*
Trans-Activators / genetics
Trans-Activators / metabolism

Substances

Apoproteins
Bacterial Proteins
FleQ protein, Pseudomonas aeruginosa
Recombinant Proteins
Trans-Activators
fleN protein, Pseudomonas aeruginosa
Adenosine Triphosphate