[Kinetics of DNA-dependent RNA synthesis: coupled synthesis of di- and trinucleotides in the presence of a minimum complement of substrate]

Abstract

The qualitative and quantitative characteristics of the synthesis of the short oligonucleotides by Escherichia coli RNA polymerase on A1 promoter of the bacteriophage T7 deletion mutant delta D111 DNA in the presence of the incomplete set of nucleoside triphosphates were studied. It was shown, that in conformity with the structure of A1 promoter the oligonucleotides pppApU, pppApUpC were synthesized in the presence of ATP, UTP, CTP; the oligonucleotides pApU, pApUpC-in the presence of AMP, UTP, CTP and oligonucleotides pApU, pApUpC, pApUpCpG-in the presence of AMP, UTP, CTP, GTP. The curves of di- and trinucleotide syntheses as the functions of the substrate concentrations were obtained. The analytical formulas for the rates of the coupled synthesis were derived from these curves. A kinetic scheme that is in conformity with the experimental data was proposed. This scheme includes the stage of the reversible, random and release of di- and trinucleotides from the enzyme-template complex.