Context and conformation dictate function of a transcription antitermination switch.
In bacteriophage l, transcription elongation is regulated by the N protein, which binds a nascent mRNA hairpin (termed boxB) and enables RNA polymerase to read through distal terminators. We have examined the structure, energetics and in vivo function of a number of N-boxB complexes derived from in vitro protein selection. Trp18 fully stacks on the RNA loop in the wild-type structure, and can become partially or completely unstacked when the sequence context is changed three or four residues away, resulting in a recognition interface in which the best binding residues depend on the sequence context. Notably, in vivo antitermination activity correlates with the presence of a stacked aromatic residue at position 18, but not with N-boxB binding affinity. Our work demonstrates that RNA polymerase responds to subtle conformational changes in cis-acting regulatory complexes and that approximation of components is not sufficient to generate a fully functional transcription switch.
D001426:Bacterial Proteins / Q000378:metabolism*; D029968:Escherichia coli Proteins; D007700:Kinetics; D010445:Peptide Elongation Factors / Q000378:metabolism; D010455:Peptides / Q000378:metabolism; D011487:Protein Conformation; D012313:RNA / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000502:physiology*; D035602:Transcriptional Elongation Factors
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D010445:Peptide Elongation Factors; D010455:Peptides; D016601:RNA-Binding Proteins; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; C064872:antiterminator proteins, Bacteria; C068727:nusA protein, E coli; D012313:RNA