Multiple RNA polymerase conformations and GreA: control of the fidelity of transcription.
Pre-steady state kinetics of misincorporation were used to investigate the addition of single nucleotides to nascent RNA by Escherichia coli RNA polymerase during transcription elongation. The results were fit with a branched kinetic mechanism that permits conformational switching, at each template position, between an activated and an unactivated enzyme complex, both of which can bind nucleotide triphosphates (NTPs) from solution. The complex exists most often in the long-lived activated state, and only becomes unactivated when transcription is slowed. This model permits multiple levels of nucleotide discrimination in transcription, since the complex can be "kinetically trapped" in the unactivated state in the absence of the correct NTP or if the 3' terminal residue is incorrectly matched. The transcription cleavage factor GreA (or an activity enhanced by GreA) increased the fidelity of transcription by preferential cleavage of transcripts containing misincorporated residues in the unactivated state of the elongation complex. This cleavage mechanism by GreA may prevent the formation of "dead-end" transcription complexes in vivo.
Erie
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DA
Institute of Molecular Biology, University of Oregon, Eugene 97403.
Hajiseyedjavadi
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Young
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von Hippel
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PH
D001483:Base Sequence; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry* / Q000378:metabolism; D004722:Endoribonucleases / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins; D007700:Kinetics; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009711:Nucleotides / Q000378:metabolism; D010445:Peptide Elongation Factors / Q000378:metabolism*; D011487:Protein Conformation; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000378:metabolism; D013698:Templates, Genetic; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014544:Uridine Triphosphate / Q000378:metabolism
D029968:Escherichia coli Proteins; C079668:GreA protein, E coli; D009711:Nucleotides; D010445:Peptide Elongation Factors; D012333:RNA, Messenger; D014157:Transcription Factors; D012321:DNA-Directed RNA Polymerases; D004722:Endoribonucleases; D014544:Uridine Triphosphate