类芽胞杆菌碱性果胶裂解酶的纯化与酶学性质表征  

Purification and characterization of alkaline pectate lyase from Paenibacillus sp.WZ008

作　　者：熊斌[1] 应向贤[1] 陈丽娜[1] 谢丽萍[1] 魏春[1] 汪钊[1] 

机构地区：[1]浙江工业大学生物与环境工程学院,杭州310014

出　　处：《生物加工过程》2013年第6期42-46,共5页Chinese Journal of Bioprocess Engineering

基　　金：浙江省自然科学基金(LY12B06010);教育部留学回国人员科研启动基金(第40批)

摘　　要：从类芽胞杆菌Paenibacillus sp.WZ008的发酵上清液中纯化得到一个高活力碱性果胶裂解酶,经SDS-PAGE电泳估算其亚基相对分子质量为4.5×104。通过对该酶进行酶学性质研究发现:该酶能催化裂解果胶酸、低酯果胶和高酯果胶;酶催化反应最适温度范围为55～60℃,最适pH为9.6,在最适条件下以低酯果胶为底物酶的比酶活达3 021.6 U/mg;Ca2+能增强该酶的活力,而Mn2+,Ba2+和EDTA强烈抑制该酶活力;当没有Ca2+存在时,高度酯化的果胶是该酶的最适底物,在4 mmol/L Ca2+存在时,该酶以果胶酸为底物比酶活最高(25 467 U/mg)。该酶N端序列比对分析发现与类芽胞杆菌Paenibacillus amylolyticus strain 27c64果胶裂解酶高度同源。An extracellular pectate lyase was purified from the culture supernatant of Paenibacillus sp. WZ008 grown in the pectin-containing medium. The subunit molecular weight of the purified enzyme was determined to be around 4. 5 × 10^4 by SDS-PAGE analysis. It showed activity on both polygalacturonic acid and methylated pectins. The optimum activity of the purified enzyme was demonstrated in a temperature range of 55 ℃ to 60℃ and at pH 9.6. The specific activity of pectate lyase was up to 3 021.6 U/mg using 20% methylated pectin as substrate under the optimized conditions. The Ca2＋ enhanced the enzyme activity but Mn2＋ , Ba2＋ , and EDTA strongly inhibited it. Highly methylated pectin was the optimum substrate without Ca2 ＋ addition while the enzyme exhibited the maximal activity（25 467 U/mg） on polygalacturonic acid in the presence of 4 mmol/L Ca2＋. The amino-terminal sequence of the enzyme was highly identical to that of a pectate lyase from Paenibacillus amylolyticus strain 27c64.

关 键 词：酶学性质 酯化果胶 碱性果胶酶 纯化 

分 类 号：Q814[生物学—生物工程]