title: Discovery of an intermolecular disulfide bond required for the thermostability of a heterodimeric protein from the thermophile Hydrogenobacter thermophilus.
abstract: Factors that increase protein thermostability are of considerable interest in both scientific and industrial fields. Disulfide bonds are one of such factors that increase thermostability, but are rarely found in intracellular proteins because of the reducing environment of the cytosol. Here, we report the first example of an intermolecular disulfide bond between heteromeric subunits of a novel-type phosphoserine phosphatase from a thermophilic bacterium Hydrogenobacter thermophilus, which contributes to the protein thermostability at the physiological temperature. Comparison of remaining soluble proteins between wild-type and cysteine-deleted mutant using SDS-PAGE revealed that the disulfide bond increases the thermostability of the whole protein by tightly connecting a subunit with low solubility to the partner with higher solubility. Furthermore, it was strongly suggested that the disulfide bond is formed and contributes to the stability in vivo. This finding will open new avenues for the design of proteins with increased thermostability.
authors.affiliation: a Department of Biotechnology, Graduate School of Agricultural and Life Sciences , University of Tokyo , Tokyo , Japan.
authors.forename: Keug Tae
authors.initials: KT
authors.lastname: Kim
authors.affiliation: b Faculty of Life and Environmental Science , University of Tsukuba , Tsukuba , Japan.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Chiba
authors.affiliation: a Department of Biotechnology, Graduate School of Agricultural and Life Sciences , University of Tokyo , Tokyo , Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Arai
authors.affiliation: a Department of Biotechnology, Graduate School of Agricultural and Life Sciences , University of Tokyo , Tokyo , Japan.
authors.forename: Masaharu
authors.initials: M
authors.lastname: Ishii
mesh_terms: D001419:Bacteria / Q000737:chemistry* / Q000201:enzymology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry*; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006358:Hot Temperature; D009154:Mutation; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D055503:Protein Multimerization; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012995:Solubility
chemical_list: D001426:Bacterial Proteins; D004220:Disulfides; D021122:Protein Subunits; D011994:Recombinant Proteins; D010744:Phosphoric Monoester Hydrolases; C022394:phosphoserine phosphatase
keywords: heterodimer; intermolecular disulfide bond; phosphoserine phosphatase; protein solubility; protein thermostability