Multiple RNA Polymerase Conformations and GreA: Control of the Fidelity of Transcription

• Published in: Science (American Association for the Advancement of Science) 1993-11, Vol.262 (5135), p.867-873
• Main Authors: Erie, Dorothy A., Hajiseyedjavadi, Omid, Young, Mark C., von Hippel, Peter H.
• Format: Article
• Language: English
• Subjects: Base Sequence; Biochemistry; Biological and medical sciences; Corporations; DNA; DNA-Directed RNA Polymerases - chemistry; DNA-Directed RNA Polymerases - metabolism; Endoribonucleases - metabolism; Enzymes; Escherichia coli; Escherichia coli - enzymology; Escherichia coli Proteins; Fundamental and applied biological sciences. Psychology; Gels; Genetic transcription; Kinetics; Microbiology; Models, Genetic; Molecular Sequence Data; Mycology; Nucleotides; Nucleotides - metabolism; Pathogenicity, host-agent relations, miscellaneous strains, epidemiology; Peptide Elongation Factors - metabolism; Protein Conformation; Radioactive decay; Rationing; RNA; RNA polymerases; RNA, Messenger - biosynthesis; RNA, Messenger - metabolism; Templates, Genetic; Transcription (Genetics); Transcription Factors - metabolism; Transcription, Genetic; Uridine Triphosphate - metabolism
• ISSN: 0036-8075; 1095-9203
• DOI: 10.1126/science.8235608

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.