Structural biophysics of the NusB:NusE antitermination complex.
In prokaryotic transcription regulation, several host factors form a complex with RNA polymerase and the nascent mRNA. As part of a process known as antitermination, two of these host factors, NusB and NusE, bind to form a heterodimer, which interacts with a specific boxA site on the RNA. The NusB/NusE/boxA RNA ternary complex interacts with the RNA polymerase transcription complex, stabilizing it and allowing transcription past premature termination points. The NusB protein also binds boxA RNA individually and retains all specificity for boxA. However, NusE increases the affinity of RNA to NusB in the ternary complex, which contributes to efficient antitermination. To understand the molecular mechanism of the process, we have determined the structure of NusB from the thermophilic bacterium Aquifex aeolicus and studied the interaction of NusB and NusE. We characterize this binding interaction using NMR, isothermal titration calorimetry, gel filtration, and analytical ultracentrifugation. The binding site of NusE on NusB was determined using NMR chemical shift perturbation studies. We have also determined the NusE binding site in the ternary Escherichia coli NusB/NusE/boxA RNA complex and show that it is very similar to that in the NusB/NusE complex. There is one loop of residues (from 113 to 118 in NusB) affected by NusE binding in the ternary complex but not in the binary complex. This difference may be correlated to an increase in binding affinity of RNA for the NusB/NusE complex.
Das
Ranabir
R
Structural Biophysics Laboratory, National Cancer Institute, Frederick, MD 21702, USA.
Loss
Sandra
S
Li
Jess
J
Waugh
David S
DS
Tarasov
Sergey
S
Wingfield
Paul T
PT
Byrd
R Andrew
RA
Altieri
Amanda S
AS
D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D012269:Ribosomal Proteins / Q000378:metabolism; D040681:Structural Homology, Protein; D013728:Terminator Regions, Genetic; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C074952:NusB protein, E coli; D016601:RNA-Binding Proteins; D012269:Ribosomal Proteins; D014157:Transcription Factors; C064872:antiterminator proteins, Bacteria; C029537:ribosomal protein S10