title: Enrichment and proteome analysis of a hyperthermostable protein set of archaeon Thermococcus onnurineus NA1.
abstract: Thermococcus onnurineus NA1 is a hyperthermophilic archaeon that can be used for the screening of thermophilic enzymes. Previously, we characterized the metabolic enzymes of the cytosolic proteome by two-dimensional electrophoresis/tandem mass spectrometry (2-DE/MS-MS). In this study, we identified a subset of hyperthermostable proteins in the cytosolic proteome using enrichment by in vitro heat treatment and protein identification. After heat treatment at 100°C for 2 h, 13 and 149 proteins were identified from the soluble proteome subset by 2-DE/MS-MS and 1-DE/MS-MS analysis, respectively. Representative proteins included intracellular protease I, thioredoxin reductase, triosephosphate isomerase, putative hydroperoxide reductase, proteasome, and translation initiation factors. Intracellular protease, deblocking aminopeptidases, and fructose-1,6-bisphosphatase were overexpressed in Escherichia coli and biological activity above 85°C was confirmed. The folding transition temperature (Tm) of identified proteins was analyzed using the in silico prediction program TargetStar. The proteins enriched with the heat treatment have higher Tm than the homologous proteins from mesophilic strains. These results suggested that the heat-stable protein set of hyperthermophilic T. onnurineus NA1 can be effectively fractionated and enriched by in vitro heat treatment.
authors.affiliation: Division of Life Science, Korea Basic Science Institute, Daejeon, Korea.
authors.forename: Sung-Ho
authors.initials: SH
authors.lastname: Yun
authors.forename: Chi-Won
authors.initials: CW
authors.lastname: Choi
authors.forename: Sang Oh
authors.initials: SO
authors.lastname: Kwon
authors.forename: Yeol Gyun
authors.initials: YG
authors.lastname: Lee
authors.forename: Young-Ho
authors.initials: YH
authors.lastname: Chung
authors.forename: Hoi Jong
authors.initials: HJ
authors.lastname: Jung
authors.forename: Yun-Jae
authors.initials: YJ
authors.lastname: Kim
authors.forename: Jung-Hyun
authors.initials: JH
authors.lastname: Lee
authors.forename: Jong-Soon
authors.initials: JS
authors.lastname: Choi
authors.forename: Soohyun
authors.initials: S
authors.lastname: Kim
authors.forename: Seung Il
authors.initials: SI
authors.lastname: Kim
mesh_terms: D019843:Archaeal Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006358:Hot Temperature; D055550:Protein Stability; D020543:Proteome / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D019712:Thermococcus / Q000235:genetics / Q000378:metabolism*
chemical_list: D019843:Archaeal Proteins; D020543:Proteome; D011994:Recombinant Proteins