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Nucleic Acids Research, 1991, Vol. 19, No. 11 2947-2953
© 1991

MOLECULAR BIOLOGY

Effect of DNA bending in various regions of a consensuslike Escherichia coli promoter on its strength in vivo and structure of the open complex in vitro

Tomasz Lozinski, Katarzyna Adrych-Rozek1, Wojciech T. Markiewicz1 and Kazimierz L. Wierzchowski*

Institute of Biochemistry and Biophysics, Polish Academy of Sciences ul. Rakowiecka 36, 02–532 Warszawa 1Institute of Bioorganic Chemistry, Polish Academy of Sciences ul. Noskowskiego 12/14, Poznan, Poland

*To whom correspondence should be addressed

Received February 22, 1991. Revised April 4, 1991. Accepted April 4, 1991.

A series of E. coli promoters made of the consensus –35 and –10 hexamers separated by 17 bp spacer with variously located bending dTn-dAn, n = 5 or 6, sequences was constructed and cloned into the plasmid pDS3. Electrophoretic gel mobilities of restriction fragments containing these promoters correlated with the number of the T tracts encoded in the promoter sequences. The open complexes formed by E. coli RNA polymerase on promoters containing the T5(–34... –38) tract exhibited gel retardation indicative of their different gross geometry. The strength of these promoters measured in vivo in relation to an internal transcriptional standard was shown to be significantly lower than that of the group without the T5(–34...–38) tract. Within both these groups the promoters with two T6 tracts in the spacer, aligned in phase with the B-DNA helix repeat, had lower transcriptional activity, while the T6 tract encoded in the –7 ... –2 promoter region apparently had no influence on the strength of the respective promoters.


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