降解β-胡萝卜素产香菌株的分离鉴定及降解酶酶学性质

Isolation and Identification of A β-carotenes-degradating Strain and Characterization of the Degrading Enzyme

采取平板划线法从红茶中筛选分离高效降解β-胡萝卜素菌株,通过形态学特征、生理生化和16S r DNA等方法对其进行鉴定,对该菌株所产β-胡萝卜素降解酶分离纯化,并研究其酶学性质。结果表明:筛选得到一株高效降解β-胡萝卜素菌株HC-3,该菌株对β-胡萝卜素的降解率达86.82%,主要降解产物为5,6-环氧-β-紫罗兰酮、二氢猕猴桃内酯和反式-β-紫罗兰酮等香味物质。根据形态特征、生理生化性质和16S r DNA系统进化树分析,初步鉴定该菌株为肠杆菌(HC-3)。通过硫酸铵分级沉淀和阴离子交换柱等步骤分离纯化得到该菌株所产β-胡萝卜素降解酶,SDS-PAGE分析表明该酶分子质量约53 ku。该酶最适反应温度45℃,最适p H值8.0;金属离子Ca2+和Cu2+对该酶有激活作用,Fe2+和Mn2+对该酶有抑制作用;β-胡萝卜素为该酶的最适作用底物。

A high efficient β-carotenes-degradating strain HC-3 was isolated and identified from black tea. Classical morphology, physiological and biochemical characrteristics and 16 S r DNA sequencing were performed to identify the strain. The degrading enzyme from this strain was purified and characterized. The results demonstrated that one efficient degradation strain was screened and obtained. The degradation rate of β-carotene reached 86.62%, and the main degradation products included 5, 6-epoxy-β-ionone, dihydroactinidiolide, trans-β-ionone and other flavouring substances. The strain was identified as Enterobacter sp. HC-3 through classical morphology, physiological and biochemical characrteristics and 16 S r DNA sequence of phylogenetic tree. The degrading enzyme from the strain HC-3 was purified to homogeneity after ammonium sulfate precipitation and anion exchange chromatography. The relative molecular mass of purified enzyme was approximately 53 ku on SDS-PAGE. The enzyme displayed optimal activity at 45 ℃ and 8.0. The enzyme was activated by Ca^2＋and Cu^2 to some extent and inhibited by Fe^2＋and Mn^2＋. The β-carotene was the optimum substrate for the degrading enzyme.