Cycling of ribonucleic acid polymerase to produce oligonucleotides during initiation in vitro at the lac UV5 promoter - PubMed (original) (raw)
Cycling of ribonucleic acid polymerase to produce oligonucleotides during initiation in vitro at the lac UV5 promoter
A J Carpousis et al. Biochemistry. 1980.
Abstract
High-resolution gel electrophoresis has been used to detect and quantitate promoter-specific oligonucleotides produced during initiation of transcription in vitro at the lactose operon (lac) UV5 promoter. The resolved products are RNA species of various lengths which correspond to the initial lac mRNA sequence. Quantitation shows that many oligonucleotides can be formed per preinitiation complex, including species as long as hexanucleotide. Synthesis occurs without dissociation of the enzyme, as evidenced by levels of synthesis in the presence of heparin, a selective inhibitor of free RNA polymerase. Thus, RNA polymerase cycles at this promoter in vitro producing oligonucleotides reiteratively. In general, the yield of oligonucleotides decreases when the total concentration of all four substrates is increased or when a missing nucleoside triphosphate substrate is added. Nevertheless, oligonucleotide synthesis persists under all conditions tested. Strikingly, the dinucleotide always represents 50% of the total of all oligonucleotides, even when conditions are manipulated to cause a 100-fold variation in this total. This shows that, after formation of the first phosphodiester bond at the lac UV5 promoter, dissociation of the dinucleotide is as likely as formation of the second phosphodiester bond. As discussed above, after release of a small RNA, RNA polymerase may then begin another RNA chain, which is again subject to premature release. These considerations lead to a model in which RNA polymerase cycles to produce oligonucleotides during initiation of transcription at the lac UV5 promoter in vitro. Production of a long RNA transcript is then essentially an escape from this cycling reaction. The drug rifampicin, which drastically inhibits escape to produce RNA, limits, but dose not prevent, the cycling reaction.
Similar articles
- Productive and abortive initiation of transcription in vitro at the lac UV5 promoter.
Gralla JD, Carpousis AJ, Stefano JE. Gralla JD, et al. Biochemistry. 1980 Dec 9;19(25):5864-9. doi: 10.1021/bi00566a031. Biochemistry. 1980. PMID: 6450614 - Probing contacts of phosphate groups of oligonucleotides from the non-template strand of lac UV5 promoter with E. coli RNA polymerase using regioselective cross-linking.
Rudakova EA, Ivanovskaya MG, Kozlov MV, Khoretonenko MV, Oretskaya TS, Nikiforov VG. Rudakova EA, et al. Biochemistry (Mosc). 2000 Jun;65(6):640-50. Biochemistry (Mosc). 2000. PMID: 10887281 - The nucleotide sequence of the lactose messenger ribonucleic acid transcribed from the UV5 promoter mutant of Escherichia coli.
Maizels NM. Maizels NM. Proc Natl Acad Sci U S A. 1973 Dec;70(12):3585-9. doi: 10.1073/pnas.70.12.3585. Proc Natl Acad Sci U S A. 1973. PMID: 4587256 Free PMC article.
Cited by
- Intrinsic RNA Targeting Triggers Indiscriminate DNase Activity of CRISPR-Cas12a.
Zhang J, Li Z, Guo C, Guan X, Avery L, Banach D, Liu C. Zhang J, et al. Angew Chem Int Ed Engl. 2024 May 13;63(20):e202403123. doi: 10.1002/anie.202403123. Epub 2024 Apr 10. Angew Chem Int Ed Engl. 2024. PMID: 38516796 - Digital Twin Fundamentals of mRNA In Vitro Transcription in Variable Scale Toward Autonomous Operation.
Hengelbrock A, Schmidt A, Strube J. Hengelbrock A, et al. ACS Omega. 2024 Feb 5;9(7):8204-8220. doi: 10.1021/acsomega.3c08732. eCollection 2024 Feb 20. ACS Omega. 2024. PMID: 38405539 Free PMC article. - Rhodobacter sphaeroides CarD Negatively Regulates Its Own Promoter.
Henry KK, Ross W, Gourse RL. Henry KK, et al. J Bacteriol. 2021 Aug 9;203(17):e0021021. doi: 10.1128/JB.00210-21. Epub 2021 Aug 9. J Bacteriol. 2021. PMID: 34152199 Free PMC article. - Region 4 of the RNA polymerase σ subunit counteracts pausing during initial transcription.
Brodolin K, Morichaud Z. Brodolin K, et al. J Biol Chem. 2021 Jan-Jun;296:100253. doi: 10.1074/jbc.RA120.016299. Epub 2021 Jan 8. J Biol Chem. 2021. PMID: 33380428 Free PMC article. - The Context-Dependent Influence of Promoter Sequence Motifs on Transcription Initiation Kinetics and Regulation.
Jensen D, Galburt EA. Jensen D, et al. J Bacteriol. 2021 Mar 23;203(8):e00512-20. doi: 10.1128/JB.00512-20. Print 2021 Mar 23. J Bacteriol. 2021. PMID: 33139481 Free PMC article. Review.
Publication types
MeSH terms
Substances
LinkOut - more resources
Other Literature Sources
Molecular Biology Databases
Research Materials