产甲壳素酶菌株HD001的发酵条件研究及酶的分离纯化

Fermentation Conditions and Purification of the Chitinase from Strain HD001

通过对降解甲壳素菌株HD001的发酵条件研究,确定了其产甲壳素酶最适培养基组分为(w/v):(NH4)2SO42.0%,NaCl0.5%,K2HPO40.07%,KH2PO40.03%,MgSO4·7H2O0.05%以及葡萄糖0.1%、酵母粉0.3%和胶体甲壳素1.5%;最适产酶培养条件为:培养基起始pH值6.0,培养温度30℃,培养时间120h,发酵液酶活力达376.2U/mL。采用硫酸铵分级盐析、DEAE-纤维素离子交换层析、SephacrylS-100凝胶过滤层析对该酶进行纯化后,SDS-PAGE电泳显示单一条带,其相对分子质量约为85.7kDa。

The optimal compositions of the medium for strain HD001 to produce chitinase were as follows （unit： %, w/v）： （NH4）2SO4 2.0, NaCl 0.5, K2HPO4 0.07, KH2PO4 0.03, MgSO4·7H2O 0.05, glucose 0.1, yeast extract 0.3. powder chitin 1.5. When incubated at 30℃ for 120h in the liquid medium with initial pH6.0, the maximal chitinase activity could reach 376.2 U/mL. The chitinase is extracted by ammonium sulfate precipitation, and B purified by ion-exchange chromatography （DEAE- cellulose column） and gel filtration （Sephacryl S-100 column）. SDS-PAGE shows a single band for. the purified chitinase and the molecular weight is estimated to be about 85.7 kDa.