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Papers In Press, published online ahead of print August 16, 2002
Microbiology and Immunology, SUNY Downstate Medical Center, Brooklyn, NY 11203-2098
Corresponding Author: pogo51{at}aol.com
To examine changes that occur during the transition from an initiation complex (IC1) to an elongation complex (EC) in T7 RNA polymerase (RNAP) we used nucleic acid:protein crosslinking methods to probe interactions of the RNAP with RNA and DNA in a halted EC. As the RNA is displaced from the RNA:DNA hybrid ~9 bp upstream from the active site (at -9) it interacts with a region within the specificity loop (residues 744-750) and is directed towards a positively charged surface that surrounds residues K303 and K303. Surprisingly, the template (T) and non-template (NT) strands of the DNA at the upstream edge of the hybrid (near the site where the RNA is displaced) interact with a region in the N-terminal domain of the RNAP (residues 172-191) that is far away from the specificity loop before isomerization (in the IC). In order to bring these two regions of the RNAP into proximity, major conformational changes must occur during the transition from an IC to an EC. The observed nucleic acid:protein interactions help to explain the behavior of a number of mutant RNAPs that are affected at various stages in the initiation process, and in termination.
J. Biol. Chem, 10.1074/jbc.M206658200
Submitted on July 3, 2002
Revised on August 14, 2002
Accepted on August 16, 2002
Major conformational changes occur during the transition from an initiation complex to an elongation complex by T7 RNA polymerase
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