Studies On Xylanases From Aspergillus Usamii

The xylanase production was investigated with solid fermentation by Aspergillususamii E001. Multivariant statistical approaches were employed to evaluate theeffects of several variables (carbon and nitrogen source,initial pH and temperature)on xylanase production. Xylanase activity reached 7442 IU/g dry medium under theoptimized conditions at 28℃ for 72 h.Cross enzyme showed an optimum temperature and pH of 50℃ and pH 4.6,respectively. Xylanase was stable below 50℃ and within pH 5.0~10.0. Its activitywas increased by Ca2+ ion and strongly inhibited by Mn2+,Mg2+ and Ba2+ ions.Two xylanases Xyn I and Xyn II from solid-state fermentation culture werepurified by extracting with water, ammonium sulfate precipitation, and Phenyl-Sepharose CL-4B, Sephadex G-75 and DEAE-Sepharose fast flow column chromato-graphies. Two purified xylanases Xyn I and Xyn II were homogeneous on SDS-PAGE,PAGE,HPLC as well as zymogram on RBB-xylan, respectively. Themolecular weights of Xyn I and Xyn II were determined as 24.4 kDa and 26.8 kDarespectively on SDS-PAGE and as 25.0 kDa on gel filtration, indicating that they werea monomer. They didn't contained carbohydrate by phenol-vitriol method. Theisoelectric points of Xyn I and Xyn II were estimated to be 5.0 and 4.2 by isoelectricfocusing. Analysis of amino acid composition showed that content of acid amino acidwas high. The sequence of 15 amino acids at N-terminus of Xyn II was NH2-Ser-Ala-Gly-Ile-Asn-Tyr-Val-Gln-Asn-Tyr-Asn-Gly-Asn-Leu-Gly-。The optimum temperatures of two xylanases were all 50℃. The pH values ofXyn I and Xyn II were pH 5.0 and pH 4.5 respectively. Xyn I was stable below 45℃and within pH 5~9;while Xyn II was stable below 55℃ and within pH 3~9. Kineticscoefficients Km of Xyn I and Xyn II were 0.83 mg/mL and 9.2 mg/mL with birchxylan as substrate;while were 3.8 mg/mL and 13.6 mg/mL with oat spelt xylan assubstrate respectively. Analysis of hydrolysis products by HPLC showed that twoenzymes may be endo-xylanase.