title: The interaction surface of a bacterial transcription elongation factor required for complex formation with an antiterminator during transcription antitermination.
abstract: The bacterial transcription elongation factor, NusA, functions as an antiterminator when it is bound to the lambdoid phage derived antiterminator protein, N. The mode of N-NusA interaction is unknown, knowledge of which is essential to understand the antitermination process. It was reported earlier that in the absence of the transcription elongation complex (EC), N interacts with the C-terminal AR1 domain of NusA. However, the functional significance of this interaction is obscure. Here we identified mutations in NusA N terminus (NTD) specifically defective for N-mediated antitermination. These are located at a convex surface of the NusA-NTD, situated opposite its concave RNA polymerase (RNAP) binding surface. These NusA mutants disrupt the N-nut site interactions on the nascent RNA emerging out of a stalled EC. In the N/NusA-modified EC, a Cys-53 (S53C) from the convex surface of the NusA-NTD forms a specific disulfide (S-S) bridge with a Cys-39 (S39C) of the NusA binding region of the N protein. We conclude that when bound to the EC, the N interaction surface of NusA shifts from the AR1 domain to its NTD domain. This occurred due to a massive away-movement of the adjacent AR2 domain of NusA upon binding to the EC. We propose that the close proximity of this altered N-interaction site of NusA to its RNAP binding surface, enables N to influence the NusA-RNAP interaction during transcription antitermination that in turn facilitates the conversion of NusA into an antiterminator.
authors.affiliation: From the Laboratory of Transcription, Centre for DNA Fingerprinting and Diagnostics, Tuljaguda Complex, 4-1-714 Mozamjahi Road, Nampally, Hyderabad-500001, India.
authors.forename: Saurabh
authors.initials: S
authors.lastname: Mishra
authors.forename: Shalini
authors.initials: S
authors.lastname: Mohan
authors.forename: Sapna
authors.initials: S
authors.lastname: Godavarthi
authors.forename: Ranjan
authors.initials: R
authors.lastname: Sen
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism*; D001435:Bacteriophages / Q000378:metabolism; D001665:Binding Sites; D003545:Cysteine / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000378:metabolism*; D009154:Mutation; D010445:Peptide Elongation Factors / Q000378:metabolism*; D017354:Point Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D016601:RNA-Binding Proteins / Q000378:metabolism*; D016914:Ribonuclease H / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D035602:Transcriptional Elongation Factors / Q000378:metabolism*; D054334:Viral Regulatory and Accessory Proteins / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C059417:N protein, Bacteriophage lambda; D010445:Peptide Elongation Factors; D016601:RNA-Binding Proteins; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; D054334:Viral Regulatory and Accessory Proteins; C064872:antiterminator proteins, Bacteria; C068727:nusA protein, E coli; D012321:DNA-Directed RNA Polymerases; D016914:Ribonuclease H; D003545:Cysteine
keywords: Bacterial Transcription; Bacteriophage; Cysteine-mediated Cross-linking; Footprinting; N; NusA; RNA Polymerase; Transcription Antitermination; Transcription Termination
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