title: Divalent metal ion-induced folding mechanism of RNase H1 from extreme halophilic archaeon Halobacterium sp. NRC-1.
abstract: RNase H1 from Halobacterium sp. NRC-1 (Halo-RNase H1) is characterized by the abundance of acidic residues on the surface, including bi/quad-aspartate site residues. Halo-RNase H1 exists in partially folded (I) and native (N) states in low-salt and high-salt conditions respectively. Its folding is also induced by divalent metal ions. To understand this unique folding mechanism of Halo-RNase H1, the active site mutant (2A-RNase H1), the bi/quad-aspartate site mutant (6A-RNase H1), and the mutant at both sites (8A-RNase H1) were constructed. The far-UV CD spectra of these mutants suggest that 2A-RNase H1 mainly exists in the I state, 6A-RNase H1 exists both in the I and N states, and 8A-RNase H1 mainly exists in the N state in a low salt-condition. These results suggest that folding of Halo-RNase H1 is induced by binding of divalent metal ions to the bi/quad-aspartate site. To examine whether metal-induced folding is unique to Halo-RNase H1, RNase H2 from the same organism (Halo-RNase H2) was overproduced and purified. Halo-RNase H2 exists in the I and N states in low-salt and high-salt conditions respectively, as does Halo-RNase H1. However, this protein exists in the I state even in the presence of divalent metal ions. Halo-RNase H2 exhibits junction ribonuclease activity only in a high-salt condition. A tertiary model of this protein suggests that this protein does not have a quad-aspartate site. We propose that folding of Halo-RNase H1 is induced by binding of divalent metal ion to the quad-aspartate site in a low-salt condition.
authors.affiliation: Department of Material and Life Science, Graduate School of Engineering, Osaka University, Suita, Osaka, Japan.
authors.forename: Elias
authors.initials: E
authors.lastname: Tannous
authors.affiliation: Department of Material and Life Science, Graduate School of Engineering, Osaka University, Suita, Osaka, Japan.
authors.forename: Shigenori
authors.initials: S
authors.lastname: Kanaya
mesh_terms: D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics; D001224:Aspartic Acid / Q000737:chemistry; D020134:Catalytic Domain; D002413:Cations, Divalent; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006217:Halobacterium / Q000737:chemistry* / Q000201:enzymology; D008274:Magnesium / Q000737:chemistry*; D008345:Manganese / Q000737:chemistry*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D016914:Ribonuclease H / Q000737:chemistry* / Q000235:genetics; D016415:Sequence Alignment; D012965:Sodium Chloride / Q000737:chemistry
chemical_list: D019843:Archaeal Proteins; D002413:Cations, Divalent; D011994:Recombinant Proteins; D001224:Aspartic Acid; D008345:Manganese; D012965:Sodium Chloride; D016914:Ribonuclease H; C078856:ribonuclease HI; D008274:Magnesium