title: Conformational change and inactivation of arginine kinase from shrimp Feneropenaeus chinensis in oxidized dithiothreitol solutions.
abstract: The effect of oxidized dithiothreitol (DTT) on the conformation and function of arginine kinase from shrimp Feneropenaeus chinensis was investigated with the methods of intrinsic fluorescence, ANS fluorescence, size exclusion chromatography (SEC), sodium dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE), and activity assay. The excess molecular oxidized dithiothreitol could result in a loss of activity and conformational change of arginine kinase. The oxidized arginine kinase was characterized by monitoring the changes of fluorescence emission wavelength (excitation wavelength: 295 nm) and the intensity of 1-anilino-8-naphthalenesulfonate (ANS) binding (excitation wavelength: 380 nm) to the protein. The results of fluorescence spectra showed that the presence of oxidized DTT could result in a marked change in the enzyme tertiary structure. The conformational changes of native and oxidized arginine kinase are induced by the presence of the full set of transition state analog (TSA) components. The results of size exclusion chromatography and SDS-PAGE indicated that no disulfide bond was formed among the protein molecules in the oxidized-DTT solution.
authors.affiliation: Department of Biological Science and Biotechnology, Tsinghua University, Beijing 100084, People's Republic of China.
authors.forename: Ji-Cheng
authors.initials: JC
authors.lastname: Pan
authors.forename: Zhen-Hang
authors.initials: ZH
authors.lastname: Yu
authors.forename: En-Fu
authors.initials: EF
authors.lastname: Hui
authors.forename: Hai-Meng
authors.initials: HM
authors.lastname: Zhou
mesh_terms: D000818:Animals; D001122:Arginine Kinase / Q000737:chemistry*; D033364:Decapoda / Q000201:enzymology*; D004229:Dithiothreitol / Q000737:chemistry*; D004795:Enzyme Stability; D010084:Oxidation-Reduction; D017434:Protein Structure, Tertiary; D013050:Spectrometry, Fluorescence
chemical_list: D001122:Arginine Kinase; D004229:Dithiothreitol