Differential inhibition of abortive transcription initiation at different promoters catalysed by E. coli RNA polymerase. Effect of rifampicin on purine or pyramidine-initiated phosphodiester synthesis.
The action of rifampicin on the RNA chain initiation catalysed by E. coli RNA polymerase over different templates has been studied. The steady-state formation of dinucleoside tetraphosphate under the condition of abortive initiation reaction was assayed. It was observed that rifampicin shows a spectrum of inhibitory effects on transcription initiation at different promoters. At two different promoters with a pyrimidine nucleotide at the 5'-initiation site, e.g. rrnB P2 having CTP and lacP2 having UTP, the effect of rifampicin on the abortive synthesis of the first phosphodiester bond was found to be total, even at low concentrations of the antibiotic. On the other hand, in most cases the effect of rifampicin on the abortive synthesis with a purine nucleotide at the 5'-initiation site was found to be only partial, with the exception of the T7A2 promoter, where rifampicin stimulates the abortive synthesis of pppGpC. It was also noticed that if there was a purine nucleotide at the second position of a dinucleotide which had already been synthesised by the enzyme, then further addition of the third nucleotide was not blocked in the presence of rifampicin. It appeared that a purine nucleotide at the initiation site or at the product terminus site of a translocated dinucleotide behaved similarly towards rifampicin. In the same way, if this position was occupied by a pyrimidine, rifampicin would inhibit further phosphodiester synthesis, even at a very low concentration. The stimulatory effect of rifampicin at the T7A2 promoter was presumably because here a ternary complex containing the promoter, enzyme and the abortive transcript pppGpC was initially stable, but dissociated upon addition of rifampicin, resulting in the rapid turn-over of the product.
Kumar
K P
KP
Centre for Cellular and Molecular Biology, Hyderabad, India.
Chatterji
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D001483:Base Sequence; D002384:Catalysis; D004269:DNA, Bacterial; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology*; D008969:Molecular Sequence Data; D009843:Oligoribonucleotides / Q000096:biosynthesis / Q000235:genetics; D011401:Promoter Regions, Genetic; D011687:Purines / Q000378:metabolism; D011743:Pyrimidines / Q000378:metabolism; D012335:RNA, Ribosomal / Q000235:genetics; D012293:Rifampin / Q000494:pharmacology*; D013604:T-Phages / Q000187:drug effects / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects*
D004269:DNA, Bacterial; D009843:Oligoribonucleotides; D011687:Purines; D011743:Pyrimidines; D012335:RNA, Ribosomal; D012321:DNA-Directed RNA Polymerases; D012293:Rifampin