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Bridge helix and trigger loop perturbations generate superactive RNA polymerases.

Title: Bridge helix and trigger loop perturbations generate superactive RNA polymerases.
Authors: Tan, L
Wiesler, S
Trzaska, D
Carney, HC
Weinzierl, RO
Item Type: Journal Article
Abstract: BACKGROUND: Cellular RNA polymerases are highly conserved enzymes that undergo complex conformational changes to coordinate the processing of nucleic acid substrates through the active site. Two domains in particular, the bridge helix and the trigger loop, play a key role in this mechanism by adopting different conformations at various stages of the nucleotide addition cycle. The functional relevance of these structural changes has been difficult to assess from the relatively small number of static crystal structures currently available. RESULTS: Using a novel robotic approach we characterized the functional properties of 367 site-directed mutants of the Methanocaldococcus jannaschii RNA polymerase A subunit, revealing a wide spectrum of in vitro phenotypes. We show that a surprisingly large number of single amino acid substitutions in the bridge helix, including a kink-inducing proline substitution, increase the specific activity of RNA polymerase. Other superactivating substitutions are located in the adjacent base helices of the trigger loop. CONCLUSION: The results support the hypothesis that the nucleotide addition cycle involves a kinked bridge helix conformation. The active center of RNA polymerase seems to be constrained by a network of functional interactions between the bridge helix and trigger loop that controls fundamental parameters of RNA synthesis.
Issue Date: 31-Jan-2008
URI: http://hdl.handle.net/10044/1/15524
DOI: http://dx.doi.org/10.1186/jbiol98
Start Page: 40
Journal / Book Title: J Biol
Volume: 7
Issue: 10
Copyright Statement: © 2008 Tan et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Conference Place: England
Appears in Collections:Faculty of Natural Sciences



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