同步分泌高效纤维素酶和木聚糖酶菌株YB的鉴定及其酶学性质研究

筛选和鉴定可降解木质纤维素的真菌,并研究其产酶特征。采用刚果红平板涂布法,从荔枝腐叶中筛选具有木质纤维素降解能力的真菌,结合ITS-rDNA序列分析进行鉴定,初步测定其产酶条件,然后采用DEAE Sepharose Fast Flow阴离子交换层析与Sephadex G-100凝胶层析对硫酸铵沉淀的粗酶液进行分离纯化,对其开展酶学性质研究。结果显示,筛选出一株可降解木质纤维素降解的菌株YB,鉴定为绿木霉(Trichoderma virens)。在发酵过程中,纤维素酶和木聚糖酶的最大活力分别为313.53±26.78 U/mL和18 120.87±500.37 U/mL。分离纯化得到纤维素酶(CMC酶)Ⅰb、Ⅳ和木聚糖酶Ⅰa;通过SDS-PAGE检测,其分子量分别为58.5 kD、22.8 kD和44.5 kD。3种酶的最适酶促反应条件均为:50℃,pH 5.0。其中,木聚糖酶能有效降解玉米芯木聚糖为木糖和多种木寡糖。菌株Trichoderma virens YB可分泌高效木质纤维素降解酶,具有应用于木聚糖酶和木寡糖生产的潜力。

Identification and Enzymatic Properties of Strain YB Simultaneously Secreting Highly Efficient Cellulase and Xylanase

The objective of this work is to screen and identify fungi that can degrade lignocellulose and to study its enzyme production characteristics. The Congo red plate coating method was used to screen the fungi with ability of degrading lignocellulose. Then the ITS-rDNA sequence analysis was combined to identify the enzyme production conditions,and the DEAE Sepharose Fast Flow anion exchange and Chromatography and Sephadex G-100 gel chromatography were employed to separate and purify the crude enzyme solution precipitated by ammonium sulfate,and finally the enzymatic properties were studied. A strain YB degrading lignocellulose was screened and identified as Trichoderma virens. The maximum viability of cellulase and xylanase during fermentation was 313.53±26.78 U/mL and 18120.87±500.37 U/mL,respectively. The cellulase(CMC enzyme)Ib,Ⅳ and xylanase Ⅰa were isolated and purified,and their molecular weights were 58.5 kD,22.8 kD and 44.5 kD,respectively,as determined by SDS-PAGE. The optimal enzymatic reaction conditions for the three enzymes were :50℃,pH5.0. Among them,xylanase effectively degraded corncob xylan into xylose and various xylooligosaccharides. In conclusion,the strain Trichoderma virens YB can secrete lignocellulose efficiently degrading enzyme and thus has potential for application in xylanase and xylooligosaccharide production.