Reverse transcriptase pauses at N2-methylguanine during in vitro transcription of Escherichia coli 16S ribosomal RNA
- PMID: 91169
- PMCID: PMC383911
- DOI: 10.1073/pnas.76.8.3751
Reverse transcriptase pauses at N2-methylguanine during in vitro transcription of Escherichia coli 16S ribosomal RNA
Abstract
A restriction fragment strand complementary to a sequence near the 3' end of Escherichia coli 16S rRNA has been used to prime reverse transcriptase (avian myeloblastosis virus RNA-directed DNA nucleotidyltransferase; deoxynucleosidetriphosphate:DNA deoxynucleotidyltransferase, EC 2.7.7.7). In addition to transcripts that were extended to the 5' end of the RNA, two major transcription intermediates were observed. These discrete-sized cDNA intermediates are the result of a kinetic barrier imposed by monomethylation of the amino group on guanine that participates in base-pairing. Both major transcription intermediates correspond to attenuation at the known positions of N2-methylguanine (m2G) in the rRNA sequence. The relaxation time for elongation of the cDNA through m2G is approximately 3 min. No other major kinetic pauses were observed in the 1340 bases transcribed.
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