Differences in Contacts of RNA Polymerases from Escherichia coli and Thermus aquaticus with lacUV5 Promoter Are Determined by Core-Enzyme of RNA Polymerase

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A. V. Kulbachinskiy^*, V. G. Nikiforov, and K. L. Brodolin

Institute of Molecular Genetics, Russian Academy of Sciences, pl. Kurchatova 2, 123182 Moscow, Russia; fax: (7-095) 196-0221; E-mail: akulb@img.ras.ru

^* To whom correspondence should be addressed.

Received December 27, 2004; Revision received February 14, 2005

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The interaction of RNA polymerases from Escherichia coli and Thermus aquaticus with lacUV5 promoter was studied at various temperatures. Using DNA-protein cross-linking induced by formaldehyde, it was demonstrated that each RNA polymerase formed a unique pattern of contacts with DNA in the open promoter complex. In the case of E. coli RNA polymerase, beta´ and sigma subunits were involved into formation of cross-links with the promoter, whereas in the case of T. aquaticus RNA polymerase its beta subunit formed the cross-links with the promoter. A cross-linking pattern in promoter complexes of a hybrid holoenzyme comprised of the core-enzyme of E. coli and sigma subunit of T. aquaticus was similar to that of the E. coli holoenzyme. This suggests that DNA-protein contacts in the promoter complex are primarily determined by the core-enzyme of RNA polymerase. However, temperature-dependent behavior of contact formation is determined by the sigma subunit. Results of the present study indicate that the method of formaldehyde cross-linking can be employed for elucidation of differences in the structure of promoter complexes of RNA polymerases from various bacteria.

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KEY WORDS: RNA polymerase, promoter complex, formaldehyde cross-linking

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