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Title: Structural basis for salt-dependent folding of ribonuclease H1 from halophilic archaeon Halobacterium sp. NRC-1
Authors: Dong Ju You
Nujarin Jongruja
Elias Tannous
Clement Angkawidjaja
Yuichi Koga
Shigenori Kanaya
Osaka University
Korea Basic Science Institute
Mahidol University
Keywords: Biochemistry, Genetics and Molecular Biology;Medicine
Issue Date: 1-Jan-2014
Citation: Journal of Structural Biology. Vol.187, No.2 (2014), 119-128
Abstract: RNase H1 from extreme halophilic archaeon Halobacterium sp. NRC-1 (Halo-RNase H1) requires ≥2M NaCl, ≥10mM MnCl2, or ≥300mM MgCl2for folding. To understand the structural basis for this salt-dependent folding of Halo-RNase H1, the crystal structure of Halo-RNase H1was determined in the presence of 10mM MnCl2. The structure of Halo-RNase H1 highly resembles those of metagenome-derived LC11-RNase H1 and Sulfolobus tokodaii RNase H1 (Sto-RNase H1), except that it contains two Mn2+ions at the active site and has three bi-aspartate sites on its surface. To examine whether negative charge repulsion at these sites are responsible for low-salt denaturation of Halo-RNase H1, a series of the mutant proteins of Halo-RNase H1 at these sites were constructed. The far-UV CD spectra of these mutant proteins measured in the presence of various concentrations of NaCl suggest that these mutant proteins exist in an equilibrium between a partially folded state and a folded state. However, the fraction of the protein in a folded state is nearly 0% for the active site mutant, 40% for the bi-aspartate site mutant, and 70% for the mutant at both sites in the absence of salt. The active site mutant requires relatively low concentration (~0.5M) of salt for folding. These results suggest that suppression of negative charge repulsion at both active and bi-aspartate sites by salt is necessary to yield a folded protein. © 2014 Elsevier Inc.
ISSN: 10958657
Appears in Collections:Scopus 2011-2015

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