食品与发酵工业2024，Vol.50Issue(12)：243-250,中插8,9.DOI:10.13995/j.cnki.11-1802/ts.036810
一株产凝乳酶变色栓菌鉴定及其凝乳酶的分离纯化与酶学性质研究
Identification of a milk-clotting enzyme-producing strain Trametes versicolor and enzyme purification and characterization
摘要
Abstract
Microbial milk-clotting enzymes present an economically viable alternative to traditional calf rennet,attributed to their ex-pedited production cycles and facile fermentation processes.The objective of this study was to screen for novel microbial strains that pro-duce milk-clotting enzymes with both high activity and minimal proteolytic effects.Molds isolated from traditional sufu served as the start-ing material for this selection process.Strains demonstrating high milk-clotting activity and low proteolytic activity were identified through ITS and mt SSU rDNA amplification sequencing for molecular characterization.Enzyme purification was conducted using a sequence of ammonium sulfate fractionation,DEAE-Sepharose Fast Flow anion exchange chromatography,and Sephadex G100 gel filtration chromatog-raphy.Subsequent analysis of the enzymatic properties of the purified milk-clotting enzyme was performed.Strain CQ3 emerged as a prom-inent candidate,showing substantial milk-clotting activity with reduced proteolytic effects,and was identified as Trametes versicolor.SDS-PAGE analysis of the purified enzyme from strain CQ3 revealed a single band corresponding to a molecular weight of 61 kDa.The enzyme exhibited optimal activity at 45 ℃ and a pH of 6.5,with Ca2+markedly enhancing its activity.Stability tests demonstrated consistent en-zyme performance within a pH range of 4.5-7.0 and at temperatures below 50℃.Protease inhibitor assays classified the enzyme as an aspartic protease.These results document a novel source of fungal strains for milk-clotting enzyme production,laying a theoretical ground-work for further assessment of Trametes versicolor enzymes in cheese manufacturing applications.关键词
凝乳酶/变色栓菌/酶的分离纯化/内源转录间隔区/线粒体小亚基核糖体DNAKey words
milk-clotting enzyme/Trametes versicolor/isolation and purification of enzyme/ITS/mt SSU rDNA引用本文复制引用
黄武营,李云,朱慧,周飞..一株产凝乳酶变色栓菌鉴定及其凝乳酶的分离纯化与酶学性质研究[J].食品与发酵工业,2024,50(12):243-250,中插8,9.基金项目
韩山师范学院科研项目(XN202030) （XN202030）
潮州市科技专项计划项目(202102GY19) （202102GY19）
韩山师范学院科研项目(QD20191030) （QD20191030）
