Amino acid residues in the α-subunit C-terminal domain of Escherichia coli RNA polymerase involved in activation of transcription from the mtr promoter

J. Yang, K. Murakami, H. Camakaris, N. Fujita, A. Ishihama, A. J. Pittard

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Abstract

To examine the role of the amino acid residues (between positions 258 and 275 and positions 297 and 298) of the α-subunit of RNA polymerase in TyrR-mediated activation of the mtr promoter, we have carried out in vitro transcription experiments using a set of mutant RNA polymerases with a supercoiled mtr template. Decreases in factor-independent transcription in vitro by mutant RNA polymerases L262A, R265A, and K297A suggested the presence of a possible UP element associated with the mtr promoter. Mutational studies have revealed that an AT-rich sequence centered at -41 of the mtr promoter (SeqA) functions like an UP element. In vivo and in vitro analyses using a mutant mtr promoter carrying a disrupted putative UP element showed that this AT-rich sequence is responsible for interactions with the α-subunit which influence transcription in the absence of TyrR protein. However, the putative UP element is not needed for activator-dependent activation of the mtr promoter by TyrR and phenylalanine. The results from in vitro studies indicated that the α-subunit residues leucine-262, arginine- 265, and lysine-297 are critical for interaction with the putative UP element of the mtr promoter and play major roles in TyrR-dependent transcription activation. The residues at positions 258, 260, 261, 268, and 270 also play important roles in TyrR-dependent activation. Other residues, at positions 259, 263, 264, 266, 269, 271, 273, 275, and 298, appear to play less significant roles or no role in activation of mtr transcription.

Original languageEnglish (US)
Pages (from-to)6187-6191
Number of pages5
JournalJournal of bacteriology
Volume179
Issue number19
DOIs
StatePublished - Jan 1 1997

All Science Journal Classification (ASJC) codes

  • Microbiology
  • Molecular Biology

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