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Alteration in template recognition by Escherichia coli RNA polymerase lacking the \omega subunit: A mechanistic analysis through gel retardation and foot-printing studies

Mukherjee, Kakoli and Chatterji, Dipankar (1999) Alteration in template recognition by Escherichia coli RNA polymerase lacking the \omega subunit: A mechanistic analysis through gel retardation and foot-printing studies. In: Journal of Biosciences, 24 (4). pp. 453-459.

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Abstract

The \omega subunit of Escherichia coli RNA polymerase is a 91 amino acid polypeptide which co-purifies with the enzyme and is thought to help in maturing the rest of the enzyme to its full functionality. Purified \omega when added externally was found to inhibit general transcriptional activity of \omega-less RNA polymerase as well as promoter-specific single-round transcriptional activity at all the promoters tested. In this study we have tried to analyse the observed inhibition of transcription using gel retardation assays and KMnO4 foot- printing. Further, through protein foot-printing we have attempted to identify alterations in the interaction of the \omega-less core enzyme with the s70 subunit. Our results suggest that the \omega-less holoenzyme has lesser affinity towards the DNA template and external addition of \omega destabilizes the open complex for both the wild-type and \omega- less enzyme. The \omega-less core enzyme interacts with the s70 subunit to expose the - 35 recognition domain (domain 4.2) unlike that observed in the wild-type interaction. Thus the absence of the omega subunit leads to the formation of an enzyme which has altered DNA binding and s70 binding properties. Circular dichroic measurements also indicate a major conformational alteration of both hole and core RNA polymerase in the presence and absence of the \omega subunit.

Item Type: Journal Article
Additional Information: Copyright for this article belongs to Indian Academy of Sciences.
Keywords: escherichia coli;RNA polymerase;enzyme;amino acid polypeptide
Department/Centre: Division of Biological Sciences > Molecular Biophysics Unit
Date Deposited: 06 Jan 2005
Last Modified: 19 Sep 2010 04:15
URI: http://eprints.iisc.ernet.in/id/eprint/1507

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