Book of Abstracts: Albany 2011
June 14-18 2011
©Adenine Press (2010)
Electrostatic Properties of Bacteriophage T7 Early Promoters Recognized by E. coli RNA Polymerase.
During T7 bacteriophage infection, its transcription is closely regulated by two different RNA polymerases. RNA polymerase of E. coli transcribes the early (class I) genes of T7, and newly made T7 RNA polymerase transcribes the late (class II and class III) genes. The both enzymes are very specific for their own promoters. RNA polymerase of E. coli recognized three strong (A1, A2, and A3) and five week (B,C, D, E, F) promoters in T7 DNA. All these promoters have been earlier characterized by their biochemical properties. Their functional characteristics were shown to differ from “consensus sequence rule” behavior thus stimulating a search of new promoter determinants.
Here electrostatic properties of the promoters were studied, and this has been recently reported to play an important role in protein-nucleic acid recognition (1, 2). Electrostatic potential distribution around 300 bp- fragments containing the individual promoters was calculated by Coulomb method (3) using the computer program of Sorokin A. A. Electrostatic profiles of three strong and one weak promoters are shown in Fig. 1(a-d). Electrostatic properties in the far upstream region corresponding to -75 - -100 bp position are of most interest for our task since this region is known to be involved in electrostatic interaction with E. coli RNA polymerase α-subunit (4).
Electrostatic profile of PA1 (Fig. 1a) can be characterized by the presence of the most negatively charged element at -70 - -100 bp and more positive flanking sites. The promoter shares these specific electrostatic features with two E. coli ribosomal promoters (5). Although electrostatic profiles of PA2 (Fig. 1b) and PA3 (Fig. 1c) are not identical, they reveal some common features in the far upstream region and they share these features with T4 early promoters P114.6 and P73.0 (6). The important feature of their patterns is a continuous rise of electrostatic potential at -80 - -100 bp with an extended positive peak at -90 bp Thus, PA2 and PA3 are characterized by the presence of the positively charged element in the functionally important region of promoter DNA. At the same time, this region for PA1 is mostly negative having no pronounced positively charged area. The difference in electrostatic properties of the tandem T7 promoters can result in a difference in their functional behavior allowing a high level of gene 1 transcription to be maintained in different conditions during the phage infection.
Electrostatic properties of all weak T7 promoters (a representative example in Fig. 1d) differ from that of the strong PA1, PA2 and PA3. Their electrostatic up-elements are less structurally expressed, some of them being barely distinguishable from ordinary coding regions.
Thus, there is a good correlation between electrostatic properties of early T7 promoters and their strength. All the results support that the far upstream region of T7 early promoters can be involved in modulation their activities by acting through electrostatic interaction with E. coli RNA polymerase α-subunit.
1Institute of Biophysics
Moscow Region 142290
Fig. 1. Distribution of electrostatic potential around PA1(a), PA2(b), PA3(c) and PB(d)