Protein Science
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Protein Science (2008), 17:401-408. Published by Cold Spring Harbor Laboratory Press. Copyright © 2008 The Protein Society
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Allen, W. J.
Right arrow Articles by Waksman, G.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Allen, W. J.
Right arrow Articles by Waksman, G.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?

An intramolecular FRET system monitors fingers subdomain opening in Klentaq1

William J. Allen1,2, Paul J. Rothwell1,2, and Gabriel Waksman1,2,3

1 Institute of Structural Molecular Biology, UCL and Birkbeck, London WC1E 7HX, United Kingdom
2 School of Crystallography, Birkbeck, London WC1E 7HX, United Kingdom
3 Research Department of Structural and Molecular Bioscience, University College London, London WC1E 6BT, United Kingdom

(RECEIVED October 23, 2007; FINAL REVISION November 22, 2007; ACCEPTED November 26, 2007)

A major goal of polymerase research is to determine the mechanism through which a nucleotide complementary to a templating DNA base is selected and delivered to the polymerase active site. Structural evidence suggests a large open-to-closed conformational change affecting the fingers subdomain as being crucial to the process. We previously designed a FRET system capable of measuring the rate of fingers subdomain closure in the presence of correct nucleotide. However, this FRET system was limited in that it could not directly measure the rate of fingers subdomain opening by FRET after polymerization or in the absence of DNA. Here we report the development of a new system capable of measuring both fingers subdomain closure and reopening by FRET, and show that the rate of fingers subdomain opening is limited only by the rate of polymerization. We anticipate that this system will scale down to the single molecule level, allowing measurement of fingers subdomain movements in the presence of incorrect nucleotide and in the absence of DNA.

Keywords: polymerase; Klentaq1; subdomain movement; FRET; stopped-flow; kinetics


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




HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Copyright © 2008 by The Protein Society.