里氏木霉木聚糖酶的分离纯化、酶学特性及降解机理的研究-生物化工专业论文.docxVIP

Purification．Characterizafion Purification．Characterizafion and Mechanism of Trichoderma reesei Xylanase Jiang Xiaohua Supervised by Associate Professor Yong Qiang (College of Chemical Engineering,Nanjing Forestry University,Nanjing，210037) June 2006 Abstract The applications of xylanase in saccharification of lignoceUulosics，pulp and paper industry feed industry,xylooligosaccharide preparation and brewing industry were attracted more interesting last decades．The purification，characterization and mechanism of Trichoderma reesei Rut C30 xylanase were investigated in this paper．The main results ale as follows： B—xylosidase(XYND)，endoxylanase I(XYN I)and endoxylanaseⅡ(XYN II)were isolated and purified from Tdchoderma reesei Rut C30 xylanase．The specific activity of XYND，XYN I and XYN 11 were 23942．86，4523．61 and 61．991U／rag，respectively． Both endoxylanases and 13-xylosidase showed apparent homogeneity in SDS—PAGE．The estimated molecular masses of XYN I，XYN II and XYND were 19．9，20．3 and 110．8kDa， whereas the molecular masses of XYN 1，XYN 11 were 6．2 and 9．1kDa determined with HiPrep Sephacryl S-100 HR gel fil口afion． The optimal reaction conditions were at 50C，pH5．5 for XYN I，at 50。C，pH4．0 for XYN II，and at 60C，pH3．5 for XYND．XYN 1 was stable over the range of pH5．5—8．0，while XYN II over the range of pH3．0—5．0．XYND showed stability over the range of pH3．0-8．0 at 50 4C．Two endoxylanases were stable at 50C for 30min，and B—xylosidase，which was stable up to 60℃，showed more thermal smbflity than that oftwo endoxylanases．The Km andⅥ毗values of XYN 1 were 12．28mg／mL and 8142．42IU／mg protein for brich xylan as substrate，the Km and Vmax values of XYN 11 were 25．42mg／mL and 2513．93IU／mg protein in the same condition． XYND had a K。of O．29I_mml／mL and VmⅡof 169．99IU／mg protein for p-ni订ophenyl-p—D— xylopyranoside as substrate． XYN I and XYNllwere endo—B一1,4一xylanases which catalyzed the hydrolysis of xylan radomly to xylooligosaccharides．The products of xylan hydrolysis with XYN 1