ANALYSIS OF RNA CLEAVAGE BY RNA POLYMERASES FROM <i>E. coli </i>AND <i>D. radiodurans</i>
RNA polymerase can both synthesize and cleave RNA. Both reactions occur at the same catalytic center that contains two magnesium ions bound to three aspartic acid residues of the absolutely conserved NADFDGD motif of the RNA polymerase β' subunit. We demonstrated that RNA polymerase from <i>D. radiodurans </i>possesses much higher rate of intrinsic RNA cleavage than RNA polymerase from <i>E. coli </i>(the difference in the rates is about 15-fold at 20°C). At the same time, these RNA polymerases do not differ in the rates of RNA synthesis. Comparison of the RNA polymerase sequences adjacent to the NADFDGD motif reveals the only amino acid substitution in this region (Glu751 in <i>D. radiodurans vs. </i>Ala455 in <i>E. coli) </i>that is located in the secondary enzyme channel and can potentially affect the rate of RNA cleavage. Introduction of the corresponding substitution in the <i>E. coli </i>RNA polymerase leads to a slight (about 2-3 fold) increase in the cleavage rate but does not affect RNA synthesis. Thus, the differences in the RNA cleavage rates between <i>E. coli </i>and <i>D. radiodurans </i>RNA polymerases are likely determined by multiple amino acid substitutions that do not affect the rate of RNA synthesis and are located in several regions of the active center.