Characterization of a bifunctional alginate lyase as a new member of the polysaccharide lyase family 17 from a marine strain BP-2
Characterization of a bifunctional alginate lyase as a new member of the polysaccharide lyase family 17 from a marine strain BP-2
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来自海洋菌株 BP-2 的多糖裂解酶家族 17 新成员双功能藻酸盐裂解酶的表征
DOI：
10.1007/s10529-019-02722-1
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发表时间：
2019-10-01
影响因子：
2.7
通讯作者：
Huang, Shushi
中科院分区：
文献类型：
--
作者：
Huang, Guiyuan;Wen, Shunhua;Huang, Shushi
目的利用双功能海藻酸酯裂解酶对海藻酸酯生物质进行高效糖化，制备功能性海藻酸酯低聚糖。结果从腐烂的马尾藻中筛选出一株产海藻酸解酶的BP-2菌株。通过冷冻干燥、透析、离子交换色谱等方法从BP-2发酵液中分离得到一种新的海藻酸解酶Alg17B，属于多糖解酶家族17。对纯化的裂解酶的酶学性质进行了研究。Alg17B的分子量约为77 kDa，最适反应温度为40 ~ 45℃，最适反应pH为7.5 ~ 8.0。该酶在pH 7.0-8.0，温度25-35℃范围内相对稳定，纯化酶的比活性达到4036 U/mg。低浓度Na+刺激Alg17B酶活性，Ca2+、Zn2+等金属离子抑制Alg17B酶活性。底物特异性分析、薄层色谱和质谱分析表明，Alg17B是一种海藻酸解酶，可催化海藻酸钠、聚甘露醛酸（polyM）和聚高糖醛酸水解生成单糖和低分子量低聚糖。Alg17B也具有双功能，对海藻酸盐具有内溶和外溶活性，并且对底物的利用范围很广，对polyM有偏好。结论salg17b可用于褐藻燃料乙醇生产中主要碳水化合物海藻酸盐的糖化，也可用于低聚糖的制备和研究。
ObjectivesBifunctional alginate lyase can efficiently saccharify alginate biomass and prepare functional oligosaccharides of alginate.ResultsA new BP-2 strain that produces alginate lyase was screened and identified from rottedSargassum. A new alginate lyase, Alg17B, belonging to the polysaccharide lyase family 17, was isolated and purified from BP-2 fermentation broth by freeze-drying, dialysis, and ion exchange chromatography. The enzymatic properties of the purified lyase were investigated. The molecular weight of Alg17B was approximately 77 kDa, its optimum reaction temperature was 40–45 °C, and its optimum reaction pH was 7.5–8.0. The enzyme was relatively stable at pH 7.0–8.0, with a temperature range of 25–35 °C, and the specific activity of the purified enzyme reached 4036 U/mg. A low Na+concentration stimulated Alg17B enzyme activity, but Ca2+, Zn2+, and other metal ions inhibited it. Substrate specificity analysis, thin-layer chromatography, and mass spectrometry showed that Alg17B is an alginate lyase that catalyses the hydrolysis of sodium alginate, polymannuronic acid (polyM) and polyguluronic acid to produce monosaccharides and low molecular weight oligosaccharides. Alg17B is also bifunctional, exhibiting both endolytic and exolytic activities toward alginate, and has a wide substrate utilization range with a preference for polyM.ConclusionsAlg17B can be used to saccharify the main carbohydrate, alginate, in the ethanolic production of brown algae fuel as well as in preparing and researching oligosaccharides.