Reverse transcriptase pauses at N2-methylguanine during in vitro transcription of Escherichia coli 16S ribosomal RNA.
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.
D001483:Base Sequence; D004926:Escherichia coli; D006150:Guanine Nucleotides / Q000378:metabolism; D007700:Kinetics; D012329:RNA, Bacterial / Q000378:metabolism; D012335:RNA, Ribosomal / Q000378:metabolism*; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism*; D013379:Substrate Specificity; D013698:Templates, Genetic; D014158:Transcription, Genetic
D006150:Guanine Nucleotides; D012329:RNA, Bacterial; D012335:RNA, Ribosomal; D012194:RNA-Directed DNA Polymerase