NusG alters rho-dependent termination of transcription in vitro independent of kinetic coupling.
To complement the recent discovery that rho-dependent termination in E. coli requires nusG protein in vivo, we have tested the effect of purified nusG protein on rho-dependent termination in vitro. With the well-characterized trp t' terminator of E. coli, and no other proteins than E. coli RNA polymerase and rho factor, nusG causes a proximal shift in the terminated RNA endpoints, compared to the endpoints generated by rho alone. The presence of nusG also enhances rho-mediated termination on partially defective mutant trp t' templates. We rule out explanations such as a change in the kinetic coupling between rho and RNA polymerase or a nusG-mediated increase in the affinity of rho for RNA. We also detect no difference in the helicase rate of rho in the presence of nusG. Even assays with completely stalled and isolated ternary complexes indicate that rho is able to effect the release of RNA with the assistance of nusG at points preceding the most proximal release sites observed in the absence of nusG. Our observations support a model in which nusG acts as a component of the transcription complex, possibly interacting with both rho and RNA polymerase as it governs accessibility to the nascent transcript.
Nehrke
K W
KW
Department of Biochemistry, University of Rochester Medical Center, NY 14642.
Zalatan
F
F
Platt
T
T
D001426:Bacterial Proteins / Q000378:metabolism*; D001483:Base Sequence; D004269:DNA, Bacterial; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins; D007700:Kinetics; D008969:Molecular Sequence Data; D010445:Peptide Elongation Factors / Q000378:metabolism*; D017354:Point Mutation; D012329:RNA, Bacterial / Q000378:metabolism; D012234:Rho Factor / Q000378:metabolism*; D013489:Suppression, Genetic; D013728:Terminator Regions, Genetic; D014157:Transcription Factors; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; C070291:NusG protein, E coli; D010445:Peptide Elongation Factors; D012329:RNA, Bacterial; D012234:Rho Factor; D014157:Transcription Factors; D012321:DNA-Directed RNA Polymerases