N protein from lambdoid phages transforms NusA into an antiterminator by modulating NusA-RNA polymerase flap domain interactions.
Bottom Line: Here we showed that mutations in β-flap domain specifically defective for N antitermination exhibited altered NusA-nascent RNA interaction and have widened RNA exit channel indicating an intricate role of flap domain in the antitermination.The presence of N reoriented the RNAP binding surface of NusA-NTD, which changed its interaction pattern with the flap domain.We propose that in addition to affecting the RNA exit channel and the active center of the EC, β-flap domain rearrangement is also a mechanistic component in the N antitermination process.
Affiliation: Laboratory of Transcription, Center for DNA Fingerprinting and Diagnostics, Tuljaguda Complex, 4-1-714 Mozamjahi Road, Nampally, Hyderabad 500 001, India Graduate Studies, Manipal University, India.Show MeSH
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Mentions: Earlier, we have isolated a RNAP β-subunit flap domain mutant, G1045D, together with β′-subunit mutants located in and around the RNA exit channel (P251S, P254L and R270C) that were defective for N-mediated antitermination (21). In the same study, a suppressor, L108F, in the CTD (the RNAP-binding domain) of H-19B N was also found to specifically suppress the defects of the RNAP β′-subunit mutants. Interestingly, it was unable to suppress the antitermination defect of G1045D mutant. Also in the model structure of the EC of the E. coli RNAP (Figure 2A), G1045D bearing part of the flap-domain is located away from the exit channel, which is the proposed area through where N approaches the interior of the EC (25). Therefore, G1045D mutation most likely does not directly affect the binding of C-terminal domain of the N protein to the RNAP.
Affiliation: Laboratory of Transcription, Center for DNA Fingerprinting and Diagnostics, Tuljaguda Complex, 4-1-714 Mozamjahi Road, Nampally, Hyderabad 500 001, India Graduate Studies, Manipal University, India.