Investigation Of The Catalase Produced By Serratia Marcescens SYBCT02

As one kind of oxidases, catalase (CAT, EC 1.11.1.6) can be found in most of the animals, plants and microorganisms. Catalase can catalyze the decomposition of the cellular hydrogen peroxide, protect membrane lipid against peroxidation and can be diversely applied in food, pulp, and textile etc. field. Thus, there are certain theoretical significance and potential applicable potential to investigate the screen of catalase producing strains, fermentation for catalase fermentation, and catalase characterization.In this study, 9 bacteria were screened from many soil samples, which can produce the catalase. The activity of catalase in the 9 bacteria were compared under the conventional condition, the crude catalase of these strains was characterized respectively and their optimal temperature, the optimal pH and the half-life was obtained respectively; according to analysis of data comparison, strain 9 was selected as the candidate material .Morphological aspects, physiological and biochemical charactors and 16 S rRNA gene sequence were analyzed for the strain classification. The strain was designated as Serratia marcescens SYBCT02.The optimized fermentation medium contained (g/L): citrate 20, glucose 5, cornmeal 20, pH 7.0. The optimized ferment condition was: 7 h of seed age, and 33°C of the fermentation temperature. The catalase activity was increased 60 times from 162.03 U/mL to 9662.22 U/mL after optimization.The catalase produced by Serratia marcescens SYBCT02 was purified by (NH4)2SO4 fractionation, chromatography separation on a DEAE-cellulose-52 column and a Sephadex G-200 column.. Isozyme CAT1 had been purified from this strain, with a three-step procedure consisting of ammonium sulfate presipitation, ion exchange chromatography and gel filtration . The purification and yield of CAT1 were 71.52-fold and 32.15% respectively. The purification and yield of CAT2 were 84.51-fold and 32.78% respectively.The optimum temperature for the activity of CAT1 was founded to be 40°C, CAT1 had good stability when temperature was below 50°C. The optimum pH for the activity of the laccase was 6 with relative activity of about 80% at the pH rang between 7.0 and 10.0.The molecular mass of CAT1 was estimated to be 51 KDa by gel filtration chromatography, and it is a dipolymer comprising of four identical subunits. The apparent Km and Vmax value of CAT1 was 42.17 mmol/L and 57472 U/mg respectively. No peroxidase activity of this enzyme was detected when using guaiacol as electron donor. Therefore, it was concluded that CAT1 belonged to the typical catalases. The optimum temperature for the activity of CAT2 was founded to be 50°C, CAT2 had good stability when temperature was below 50°C. The optimum pH for the activity of the laccase was 8 with the best relative activity at the pH 9.The molecular mass of CAT2 was estimated to be 34.0 KDa by gel filtration chromatography, and it is a dipolymer comprising of six identical subunits.The apparent Km and Vmax value of CAT2 was 38.17 mmol/L and 64103 U/mg respectively No peroxidase activity of this enzyme was detected when using guaiacol as electron donor. Therefore, it was concluded that CAT 2 also belonged to the typical catalases.