[Reaction of pyrophosphorolysis catalyzed by Escherichia coli RNA polymerase].
E. coli RNA polymerase is shown to be capable of catalyzing consecutive DNA-dependent pyrophosphorolysis of RNA in the presence of inorganic pyrophosphate and Mg2+. Active ternary complex of the enzyme with DNA and nascent RNA is needed for the reaction, the mixure of all the components can not carry out pyrophosphorolysis. The reaction was realized in the absence of added nucleoside triphosphates. Nucleoside triphosphates are low molecular mass products of the reaction. The rate of pyrophosphorolysis of the RNA synthesised for the AI promoter of the DNA of wild type T7 phage and delta D III T7 mutant phage was followed as a function of primary structure of the DNA, temperature, ionic strength and inorganic pyrophosphate concentration. The relative rate pyrophosphorolysis for particular nucleotides in different regions of the RNA can differ by several orders of magnitude depending on the primary structure of the RNA that undergoes pyrophosphorolysis. Ternary complex containing partially pyrophosphorilised RNA is active on the RNA synthesis when pyrophosphate is removed and nucleoside triphosphates are added to the reaction mixture. RNA as short as 70-8 nucleotides long can be produced at the conditions used. It seems that efficient dissociation in this region of RNA limits the pyrophosphorolysis to proceed up to the 5' end of RNA. Ternary complex of RNA polymerase with nascent RNA and DNA is shown to undergo site specific dissociation. The specificity of the dissociation is shown to be a function of the primary structure of RNA and the direction of the reaction. Dissociation occurs at different places along RNA sequence when the RNA is synthesised and when it is pyrophosphorilised.
D001483:Base Sequence; D004279:DNA, Viral; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D011756:Diphosphates / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008970:Molecular Weight; D013604:T-Phages
D004279:DNA, Viral; D011756:Diphosphates; D012321:DNA-Directed RNA Polymerases; D008274:Magnesium
Reaktsiia pirofosforoliza, kataliziruemaia RNK-polimerazoĭ Escherichia coli.