HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 278, Issue 41, 39755-39761, October 10, 2003

This Article Full Text Full Text (PDF) All Versions of this Article: 278/41/39755    most recent M305995200v2 M305995200v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Liu, M. Articles by Garges, S. Search for Related Content PubMed PubMed Citation Articles by Liu, M. Articles by Garges, S. Social Bookmarking                      What's this?

Kinetics of Transcription Initiation at lacP1

MULTIPLE ROLES OF CYCLIC AMP RECEPTOR PROTEIN^*

Mofang Liu  , Geeta Gupte , Siddhartha Roy  ¶, Rajiv P. Bandwar ||, Smita S. Patel || and Susan Garges 

From the Laboratory of Molecular Biology, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892-4264, the ^¶Department of Biophysics, Bose Institute, P-1/12, C.I.T. Scheme VII M, Calcutta 700 054, India, and the ^||Department of Biochemistry, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854

The cyclic AMP receptor protein (CRP) acts as a transcription activator at many promoters of Escherichia coli. We have examined the kinetics of open complex formation at the lacP1 promoter using tryptophan fluorescence of RNA polymerase and DNA fragments with 2-aminopurine substituted at specific positions. Apart from the closed complex formation and promoter clearance, we were able to detect three steps. The first step after the closed complex formation leads to a rapid increase of 2-aminopurine fluorescence. This was followed by another rapid step in which quenching of tryptophan fluorescence of RNA polymerase was observed. The slowest step detected by 2-aminopurine fluorescence increase is assigned to the final open complex formation. We have found that CRP not only enhances RNA polymerase binding at the promoter, but also enhances the slowest isomerization step by about 2-fold. Furthermore, potassium permanganate probing shows that the conformation of the open complex in the presence of CRP appears qualitatively and quantitatively different from that in the absence of CRP, suggesting that contact with RNA polymerase is maintained throughout the transcription initiation.

Received for publication, June 6, 2003 , and in revised form, July 21, 2003.

^* This work was supported by National Institutes of Health Grant GM51966 (to S. S. P.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

To whom correspondence should be addressed: Laboratory of Molecular Biology, NCI, 37 Convent Dr., Rm. 5138, Bethesda, MD 20892-4264. Tel.: 301-451-8820; Fax: 301-496-2212; E-mail: liumo{at}pop.nci.nih.gov.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				M. Liu, S. Garges, and S. Adhya lacP1 Promoter with an Extended - 10 Motif: PLEIOTROPIC EFFECTS OF CYCLIC AMP PROTEIN AT DIFFERENT STEPS OF TRANSCRIPTION INITIATION J. Biol. Chem., December 24, 2004; 279(52): 54552 - 54557. [Abstract] [Full Text] [PDF]

				M. Liu, M. Tolstorukov, V. Zhurkin, S. Garges, and S. Adhya A mutant spacer sequence between -35 and -10 elements makes the Plac promoter hyperactive and cAMP receptor protein-independent PNAS, May 4, 2004; 101(18): 6911 - 6916. [Abstract] [Full Text] [PDF]

				S. D. Seredick and G. B. Spiegelman The Bacillus subtilis Response Regulator Spo0A Stimulates {sigma}A-Dependent Transcription Prior to the Major Energetic Barrier J. Biol. Chem., April 23, 2004; 279(17): 17397 - 17403. [Abstract] [Full Text] [PDF]