| 产新琼四糖的海洋微泡菌属AG1热稳定性β-琼胶酶的过量表达与鉴定 |
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| 引用本文： | 朱艳冰,高贺,李鹤宾,倪辉,姜泽东,李利君,肖安风.产新琼四糖的海洋微泡菌属AG1热稳定性β-琼胶酶的过量表达与鉴定[J].海洋学报(英文版),2019,38(2):96-106. |
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| 作者姓名： | 朱艳冰 高贺 李鹤宾 倪辉 姜泽东 李利君 肖安风 |
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| 作者单位： | 集美大学食品与生物工程学院, 厦门 361021;福建省食品微生物与酶工程重点实验室, 厦门 361021;厦门市食品与生物工程技术研究中心, 厦门 361021;厦门南方海洋研究中心经济海藻资源化利用与深加工重点实验室, 厦门 361021,集美大学食品与生物工程学院, 厦门 361021,厦门医学院药学系, 厦门 361023,集美大学食品与生物工程学院, 厦门 361021;福建省食品微生物与酶工程重点实验室, 厦门 361021;厦门市食品与生物工程技术研究中心, 厦门 361021;厦门南方海洋研究中心经济海藻资源化利用与深加工重点实验室, 厦门 361021,集美大学食品与生物工程学院, 厦门 361021;福建省食品微生物与酶工程重点实验室, 厦门 361021;厦门市食品与生物工程技术研究中心, 厦门 361021;厦门南方海洋研究中心经济海藻资源化利用与深加工重点实验室, 厦门 361021,集美大学食品与生物工程学院, 厦门 361021;福建省食品微生物与酶工程重点实验室, 厦门 361021;厦门市食品与生物工程技术研究中心, 厦门 361021;厦门南方海洋研究中心经济海藻资源化利用与深加工重点实验室, 厦门 361021,集美大学食品与生物工程学院, 厦门 361021;福建省食品微生物与酶工程重点实验室, 厦门 361021;厦门市食品与生物工程技术研究中心, 厦门 361021;厦门南方海洋研究中心经济海藻资源化利用与深加工重点实验室, 厦门 361021 |
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| 基金项目： | The Natural Science Foundation of Fujian Province of China under contract No. 2016J01162; the Program for New Century Excellent Talents in Fujian Province University, China under contract No. B15139. |
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| 摘 要： | 从微泡菌属AG1（Microbulbifer sp. AG1）克隆得到1302 bp大小的琼胶酶基因，该基因编码产物为一个成熟蛋白（413个氨基酸残基）外加一个信号肽（20个残基）。将不含信号肽片段的琼胶酶在E. coli BL21 （DE3）中进行了异源表达和纯化。使用琼脂糖作为底物，该重组琼胶酶的最适反应温度和pH分别为60℃和7.5。该重组酶表现出优良的热稳定性，在50℃和60℃下处理1 h，重组琼胶酶仍能分别保持67%和19%的残余酶活力。除了SDS，重组琼胶酶对于其他测试的抑制剂、去垢剂和尿素变性剂有着较好的抗性。利用薄层色谱和以对硝基苯-α/β-D-吡喃半乳糖苷为底物的酶活力分析结果表明，该重组琼胶酶为β型琼胶酶，它水解琼脂糖的主要终产物为新琼四糖，而且不同聚合度的酶解产物具有抗氧化活性。
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| 关 键 词： | 热稳定 β-琼胶酶 新琼四糖 微泡菌属 |
| 收稿时间： | 2017/1/14 0:00:00 |
Overexpression and characterization of a thermostable β-agarase producing neoagarotetraose from a marine isolate Microbulbifer sp. AG1 |
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| ZHU Yanbing,GAO He,LI Hebin,NI Hui,JIANG Zedong,LI Lijun and XIAO Anfeng.Overexpression and characterization of a thermostable β-agarase producing neoagarotetraose from a marine isolate Microbulbifer sp. AG1[J].Acta Oceanologica Sinica,2019,38(2):96-106. |
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| Authors: | ZHU Yanbing GAO He LI Hebin NI Hui JIANG Zedong LI Lijun and XIAO Anfeng |
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| Institution: | College of Food and Biological Engineering, Jimei University, Xiamen 361021, China;Fujian Provincial Key Laboratory of Food Microbiology and Enzyme Engineering, Xiamen 361021, China;Research Center of Food Biotechnology of Xiamen City, Xiamen 361021, China;Key Laboratory of Systemic Utilization and In-depth Processing of Economic Seaweed, Xiamen Southern Ocean Technology Center of China, Xiamen 361021, China,College of Food and Biological Engineering, Jimei University, Xiamen 361021, China,Department of Pharmacy, Xiamen Medical College, Xiamen 361023, China,College of Food and Biological Engineering, Jimei University, Xiamen 361021, China;Fujian Provincial Key Laboratory of Food Microbiology and Enzyme Engineering, Xiamen 361021, China;Research Center of Food Biotechnology of Xiamen City, Xiamen 361021, China;Key Laboratory of Systemic Utilization and In-depth Processing of Economic Seaweed, Xiamen Southern Ocean Technology Center of China, Xiamen 361021, China,College of Food and Biological Engineering, Jimei University, Xiamen 361021, China;Fujian Provincial Key Laboratory of Food Microbiology and Enzyme Engineering, Xiamen 361021, China;Research Center of Food Biotechnology of Xiamen City, Xiamen 361021, China;Key Laboratory of Systemic Utilization and In-depth Processing of Economic Seaweed, Xiamen Southern Ocean Technology Center of China, Xiamen 361021, China,College of Food and Biological Engineering, Jimei University, Xiamen 361021, China;Fujian Provincial Key Laboratory of Food Microbiology and Enzyme Engineering, Xiamen 361021, China;Research Center of Food Biotechnology of Xiamen City, Xiamen 361021, China;Key Laboratory of Systemic Utilization and In-depth Processing of Economic Seaweed, Xiamen Southern Ocean Technology Center of China, Xiamen 361021, China and College of Food and Biological Engineering, Jimei University, Xiamen 361021, China;Fujian Provincial Key Laboratory of Food Microbiology and Enzyme Engineering, Xiamen 361021, China;Research Center of Food Biotechnology of Xiamen City, Xiamen 361021, China;Key Laboratory of Systemic Utilization and In-depth Processing of Economic Seaweed, Xiamen Southern Ocean Technology Center of China, Xiamen 361021, China |
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| Abstract: | An agarase gene containing 1 302 bp was cloned from Microbulbifer sp. AG1. It encoded a mature protein of 413 amino acids plus a 20-residue signal peptide. The recombinant enzyme without the signal peptide was expressed and purified from Escherichia coli BL21 (DE3). When agarose was used as a substrate, the optimal temperature and pH for the enzyme were 60℃ and 7.5, respectively. The recombinant agarase showed excellent thermostability with 67% and 19% of residual activities after incubation at 50℃ and 60℃ for 1 h, respectively. Except SDS, the recombinant agarase had a relatively good resistance against the detected inhibitors, detergents and urea denaturant. Thin layer chromatography analysis and enzyme assay using p-nitrophenyl-α/β-D-galactopyranoside revealed that the recombinant agarase was a β-agarase that degraded agarose into neoagarotetraose as the main end product. The enzymatic hydrolysis products with different degree of polymerization exhibited the antioxidant activities. |
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| Keywords: | thermostable β-agarase neoagarotetraose Microbulbifer sp |
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