NMR Study of Hydrogen Exchange in the DNA Decamer Duplexes Containing the -10 galP1 Promoter Sequence

Abstract

Most Escherichia coli promoters are consists of two 6-bp sequence elements which are centered at ~10 and 35 nucleotides upstream of the transcription initiation point. The conserved region 4.2 and 2.4 of the RNA polymerase σ subunit recognize −10 and −35 promoter elements, respectively. The open complex between σ subunit and promoter elements, in which the promoter DNA is locally melted, is competent to initiate RNA synthesis according to the DNA sequence of template strand. Promoter opening is temperature-dependent; the most −10/−35 promoter complexes display closed forms below 15 C, whereas promoter complexes on the extended −10 galP1 promoter remain open at 5 C. The reason for this unusual behavior of the −10 galP1 promoter is not completely understood. The conserved A·T base pair at the −11 position of the promoters plays very important role in the initiation of E. coli transcription. The in vitro transcription with the −10 galP1 promoter was not initiated when this A base was substituted to other bases. To understand the function of the A-11 base in the transcription initiation process, the imino proton exchange rates were measured for the DNA decamer duplex containing the wild-type −10 galP1 promoter sequence (position: −14 ~ −5) (referred to as wt galP1, Fig. 1). The exchange rate constants of the imino protons for the wt galP1 duplex were compared with those of the modified galP1 promoters at the −11 position (see Fig. 1). This comparison indicates that some differences in the base pair dynamics are correlated with function of the −10 galP1 promoter during transcription initiation.