Promoter recognition by Escherichia coli RNA polymerase: Effects of the UPelement on open complex formation and promoter clearance

Escherichia coli promoters for transcription of ribosomal and tRNAs are gre atly activated by an A+T-rich "UP" element upstream of the -35 region. Thes e same promoters have also been found to otherwise deviate in several respe cts from the consensus promoter sequence. Here we present the results of a kinetic characterization of the interaction of Escherichia coli RNA polymer ase with UP element-containing promoters which by virtue of consensus or ne ar-consensus sequence features should be among the most optimal that can be encountered by Escherichia coli RNA polymerase. We show that fur such prom oters, (1) the second-order rate constant describing formation of the initi al (closed) complex is close to that expected for a diffusion-limited proce ss, (2) the extent of activation by the UP clement is temperature-sensitive , (3) the UP element accelerates a process after DNA binding by RNA polymer ase, and (4) the presence of the UP element delays promoter clearance upon addition of nucleoside triphosphates to preformed RNA polymerase-promoter c omplexes. Finally, we provide evidence in support of models which describe the DNA melting process accompanying open complex formation as initiating i n the -10 promoter region and progressing in the downstream direction.