title: [Optimization of two-dimensional electrophoresis conditions for proteomics of Issatchenkia orientalis in degrading dyes].
abstract: Total protein of the yeast Issatchenkia orientalis was extracted and separated by two-dimensional electrophoresis (2-DE) before and after the dye Reactive Brilliant Red K-2BP was degraded by this yeast, respectively. Different protein extraction methods, different volumes of sample loaded and different staining techniques were tested and compared for the 2-DE. Among three different protein extraction methods, the final protein concentrations of 3.4 mg/mL, 1.8 mg/mL and 5.6 mg/mL were obtained by single ultrasonication, ultrasonication-TCA/acetone,and ultrasonication-ammonium sulfate precipitation, respectively. The best electrophoresis pattern could be gotten by loading 150 microg protein samples from the method of ultrasonication-ammonium sulfate precipitation, using IPG strips of pH 4-7 for the first dimensional electrophoresis and staining with silver nitrate. This electrophoresis pattern had high resolution and good repetition. It was detected to have 730 +/- 30 protein points by preliminary image analysis. This research results provided a technical support for screening dye-degrading enzymes from the yeast of I. orientalis.
authors.affiliation: College of Resources and Environment, Graduate University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Yu
authors.forename: Zhi-Sheng
authors.initials: ZS
authors.lastname: Yu
authors.forename: Zhi-Hui
authors.initials: ZH
authors.lastname: Bai
authors.forename: Hong-Xun
authors.initials: HX
authors.lastname: Zhang
authors.forename: Ling
authors.initials: L
authors.lastname: Zhang
mesh_terms: D001673:Biodegradation, Environmental; D004396:Coloring Agents / Q000302:isolation & purification* / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D005656:Fungal Proteins / Q000032:analysis / Q000302:isolation & purification; D007220:Industrial Waste / Q000517:prevention & control*; D020543:Proteome / Q000032:analysis / Q000302:isolation & purification*; D015003:Yeasts / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*
chemical_list: D004396:Coloring Agents; D005656:Fungal Proteins; D007220:Industrial Waste; D020543:Proteome