Interaction surface of the transcription terminator Rho required to form a complex with the C-terminal domain of the antiterminator NusG.
Rho-dependent transcription termination in bacteria requires an interaction between the terminator Rho and the antiterminator NusG. The interaction surface of the Rho-NusG complex is unknown. Here we provide direct evidence that the β-sheet bundle of the C-terminal domain of NusG (NusG-CTD) has the binding determinants for Rho, proving the hypothesis described earlier [Mooney, R. A., Schweimer, K., Rosch, P., Gottesman, M., & Landick, R., (2009). Two structurally independent domains of E. coli NusG create regulatory plasticity via distinct interactions with RNA polymerase and regulators. J. Mol. Biol., 391, 341-358.]. Disulfide bridges can be engineered from NusG-CTD with the surface-exposed amino acids 217 and 224 of Rho, which belong to its P-loop ATPase domain. Mutational analyses of this region of Rho revealed that a hydrophobic pocket, located behind these amino acids of Rho, is the docking site for NusG-CTD. The proximity of this region of Rho to NusG-CTD in the Rho-NusG complex was also confirmed by an efficient fluorescence resonance energy transfer between residue K224 of Rho and residue A168 of NusG-CTD. The identification of the Rho-NusG interaction surface will be useful not only in understanding the role of NusG in the termination process but also in explaining the molecular basis of the involvement of NusG-CTD in recruiting Rho and the ribosome to the same transcription machinery.
Center for DNA Fingerprinting and Diagnostics, Nampally, Hyderabad 500 001, India.
Jisha
J
Chalissery
Ghazala
G
Muteeb
Nisha C
NC
Kalarickal
Shalini
S
Mohan
V
V
Jisha
Ranjan
R
Sen
D001665:Binding Sites; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010445:Peptide Elongation Factors / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*
D029968:Escherichia coli Proteins; D050505:Mutant Proteins; C070291:NusG protein, E coli; D010445:Peptide Elongation Factors; C534627:Rho protein, E coli; D014157:Transcription Factors