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Title
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1H, 15N and 13C resonance assignments and secondary structure determination of the RNA-binding domain of E.coli rho protein.
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Authors
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D.M.Briercheck,
T.J.Allison,
J.P.Richardson,
J.F.Ellena,
T.C.Wood,
G.S.Rule.
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Ref.
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J Biomol Nmr, 1996,
8,
429-444.
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PubMed id
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Abstract
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Protein fragments containing the RNA-binding domain of Escherichia coli rho
protein have been over-expressed in E. coli. NMR spectra of the fragment
containing residues 1-116 of rho protein (Rho116) show that a region of this
protein is unfolded in solution. Addition of (dC)10 to this fragment stabilizes
the folded form of the protein. The fragment comprising residues 1-130 of rho
protein (Rho130) is found to be stably folded, both in absence and presence of
nucleic acid. NMR studies of the complex of Rho130 with RNA and DNA
oligonucleotides indicate that the binding-site size, affinity, and specificity
of Rho130 are similar to those of intact rho protein; therefore, Rho130 is a
suitable model of the RNA-binding domain of Rho protein. NMR line widths as well
as titration experiments of Rho130 complexed with oligonucleotides of various
lengths suggests that Rho130 forms oligomers in the presence of longer
oligonucleotides. 1H, 15N and 13C resonance assignments were facilitated by the
utilization of two pulse sequences, CN-NOESY and CCH-TOCSY. The secondary
structure of unliganded Rho130 has been determined by NMR techniques, and it is
clear that the RNA-binding domain of rho is more structurally similar to the
cold shock domain than to the RNA recognition motif.
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