Dynamics on multiple timescales in the cystoviral RNA-directed RNA polymerase
Item
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Title
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Dynamics on multiple timescales in the cystoviral RNA-directed RNA polymerase
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Identifier
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d_2009_2013:2861c8c96d74:11584
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identifier
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12106
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Creator
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Ren, Zhen,
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Contributor
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Ranajeet Ghose
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Date
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2012
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Language
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English
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Publisher
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City University of New York.
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Subject
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Biochemistry | NMR | RNA directed RNA polymerase | slow dynamics
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Abstract
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The RNA-dependent RNA polymerase P2 from cystovirus ϕ6 directs the replication and transcription of the double-stranded RNA genomes. In spite of the availability of multiple crystal structures at various points along its catalytic pathway, the dynamics role involving in the catalytic cycle and fidelity control remain unclear. Isoleucine residues are distributed over the three-dimensional fold of P2. By using the delta1 positions of 25 Ile residues as probes, we measured the dynamic modes and their alterations along P2 catalytic cycle with CPMG-based multiple quantum relaxation dispersion experiments. The results indicate that P2 is dynamic on the fast (ps-ns) and slow (mus-ms) timescale. The characteristics of several motional modes are altered in the presence of substrate analogs and single-stranded RNA templates. The timescale of the lower frequency motional modes that involve several conserved functional motifs coincides with the catalytic timescale (1-2 ms), which was determined from kinetic analyses of representative RdRPs. We further investigated the influence of the extreme 3'-end sequence of the single-stranded RNA templates and the nature of the substrate nucleotide triphosphates on the slow motional modes using multiple-quantum relaxation dispersion. We found that P2, in the presence of templates bearing the proper genomic 3'-ends or the preferred initiation nucleotide (GTP), displays unique dynamic signatures that are different from those in the presence of nonphysiological templates or substrates. This further suggests that dynamics may play a role in the fidelity of recognition of the correct substrates and template sequences to initiate RNA polymerization.
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Type
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dissertation
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Source
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2009_2013.csv
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degree
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Ph.D.
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Program
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Biochemistry
