title: Change in structure and ligand binding properties of hyperstable cytochrome c555 from Aquifex aeolicus by domain swapping.
abstract: Cytochrome c555 from hyperthermophilic bacteria Aquifex aeolicus (AA cyt c555 ) is a hyperstable protein belonging to the cyt c protein family, which possesses a unique long 310 -α-310 helix containing the heme-ligating Met61. Herein, we show that AA cyt c555 forms dimers by swapping the region containing the extra 310 -α-310 helix and C-terminal α-helix. The asymmetric unit of the crystal of dimeric AA cyt c555 contained two dimer structures, where the structure of the hinge region (Val53-Lys57) was different among all four protomers. Dimeric AA cyt c555 dissociated to monomers at 92 ± 1°C according to DSC measurements, showing that the dimer was thermostable. According to CD measurements, the secondary structures of dimeric AA cyt c555 were maintained at pH 2.2-11.0. CN(-) and CO bound to dimeric AA cyt c555 in the ferric and ferrous states, respectively, owing to the flexibility of the hinge region close to Met61 in the dimer, whereas these ligands did not bind to the monomer under the same conditions. In addition, CN(-) and CO bound to the oxidized and reduced dimer at neutral pH and a wide range of pH (pH 2.2-11.0), respectively, in a wide range of temperature (25-85°C), owing to the thermostability and pH tolerance of the dimer. These results show that the ligand binding character of hyperstable AA cyt c555 changes upon dimerization by domain swapping.
authors.affiliation: Graduate School of Materials Science, Nara Institute of Science and Technology, Ikoma, Nara, 630-0192, Japan.
authors.forename: Masaru
authors.initials: M
authors.lastname: Yamanaka
authors.forename: Satoshi
authors.initials: S
authors.lastname: Nagao
authors.forename: Hirofumi
authors.initials: H
authors.lastname: Komori
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Higuchi
authors.forename: Shun
authors.initials: S
authors.lastname: Hirota
mesh_terms: D001419:Bacteria / Q000201:enzymology*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D045304:Cytochromes c / Q000737:chemistry* / Q000378:metabolism*; D004795:Enzyme Stability; D006418:Heme / Q000737:chemistry / Q000378:metabolism; D006860:Hydrogen Bonding; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D017434:Protein Structure, Tertiary; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D006418:Heme; D045304:Cytochromes c
keywords: cytochrome c; domain swapping; ligand binding; pH tolerance; thermostable