[Purification and properties of thermostable catalase in engineered E. coli].
A thermostable catalase in engineered bacterium E. coli was purified to electrophoretic homogenenity by heat treatment, ammonium sulfate fractionation precipitation, DEAE-A50 ion exchange chromatography, HiPrep 16/10 Phenyl hydrophobic interaction chromatography and Superdex200 HR 10/30 size exclusion chromatography with 187.2-fold purification and 9.8% recovery. The optimum reaction temperature and pH of this recombinant catalase were 70 degrees C and 7.0 respectively. The catalase is stable below 60 degrees C and at pH range 3-8. The residual activity of the catalase was about 60% after treated at 70 degrees C for 60 minutes and 80 degrees C for 10 minutes. The apprant Km and Vmax value of the catalase were 7.75 mmol/L and 27.8 mmol.min-1.mg-1 respectively. The affects of some metal ions and compounds on this enzyme were shown. Zn2+, Ba2+, Mn2+ of 1 mmol/L could completely inactivate the enzyme, EDTA of 50 mmol/L had no affect on activity.
Wang
Fanqing
F
Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Southern Yangtze University, Wuxi 214036, China.
Wang
Zhengxiang
Z
Shao
Weilan
W
Liu
Jiquan
J
Xu
Chengyong
C
Zhuge
Jian
J
D001464:Barium / Q000494:pharmacology; D002374:Catalase / Q000096:biosynthesis / Q000302:isolation & purification* / Q000378:metabolism; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D001411:Geobacillus stearothermophilus / Q000201:enzymology / Q000235:genetics; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D010957:Plasmids; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D013696:Temperature; D014170:Transformation, Genetic; D015032:Zinc / Q000494:pharmacology
D011994:Recombinant Proteins; D001464:Barium; D002374:Catalase; D015032:Zinc