Purification and Characterization of Thermostable Cellulase Free Xylanase from Pseudomonas Sp. XPB-6

Pawan Kumar Sharma, Duni Chand

Department of Biotechnology, Himachal Pradesh University, Shimla, India.

DOI: 10.4236/aim.2012.21003                

Abstract

Extracellular cellulase free xylanase from Pseudomonas sp. XPB-6 was purified to homogeneity with 2.15 fold of purity and 60.31% yield, respectively by ammonium sulfate precipitation and DEAE ion exchange chromatography. The protein was found to be a homotrimer consisting of three subunits of 41 kDa each and the native molecular weight of 123 kDa. The pH and temperature optimum for enzyme were 7.5 in 100 mM sodium phosphate buffer at 60°C and was stable to a high and broader range of pH and to a temperature up to 75°C. Kinetic experiments at 60°C gave V_max and K_m values of 144.92 U/mg and 0.60 mg·ml^-1 respectively for birch wood xylan. The enzyme had no apparent requirement for cofactors, and its activity was strongly inhibited 1 mM Hg²⁺ and Cu²⁺ while it was slightly inactivated by 1 mM Mg²⁺ and Pb²⁺. Shelf life of xylanase of Pseudomonas sp. XPB-6 was studied both at room temperature and 4°C. The enzyme was stable both at room temperature and at 4°C for 90 and 28 days respectively. The half-life of the enzyme was approximately 40 days at room temperature.

Keywords

Xylanase; Pseudomonas sp. XPB-6; Purification; DEAE Chromatography

Conflicts of Interest

The authors declare no conflicts of interest.

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