CN107384903A - 一种海藻糖合酶突变体及其在制备海藻糖中的应用 - Google Patents
一种海藻糖合酶突变体及其在制备海藻糖中的应用 Download PDFInfo
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- CN107384903A CN107384903A CN201710776847.2A CN201710776847A CN107384903A CN 107384903 A CN107384903 A CN 107384903A CN 201710776847 A CN201710776847 A CN 201710776847A CN 107384903 A CN107384903 A CN 107384903A
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Abstract
本发明公开了一种海藻糖合酶突变体及其在制备海藻糖中的应用，属于基因工程和酶工程领域。本发明将Thermobifida fusca YX的海藻糖合酶活性中心附近第289位的谷氨酸突变成甘氨酸得到突变体E289G；将第295位的组氨酸突变成天冬酰胺突变体H295N；将第344位的甲硫氨酸突变成赖氨酸突变体M344K；将第367位的甲硫氨酸突变成亮氨酸突变体M367L，并在H295N基础上进行双突变得到突变体H295N/E289G、H295N/M344K、H295N/M367L、H295N/M344K/M367L。突变体实现了底物中即使含有一定的葡萄糖，海藻糖合酶制备海藻糖的转化效率依然不会受到很大的影响，具有较高的工业价值。
Description
本申请是申请号为：201510210023.X，申请日为：2015年4月28日，申请名称为：一种海藻糖合酶的突变体及其制备方法和应用的分案申请。
技术领域
本发明涉及一种海藻糖合酶突变体及其在制备海藻糖中的应用，属于基因工程和酶工程领域。
背景技术
海藻糖是一种安全无毒以1,1-糖苷键构成的非还原性二糖，有三种异构体即(α,α)、异海藻糖(β,β)、新海藻糖(α,β)，一般以二水化合物的形式存在。与蛋白质或者氨基酸共同加热，不会产生美拉德反应，并且在酸性、碱性、高温、超低温环境都能保持一定的稳定性。其独特的生物活性，使得海藻糖得到广泛的应用。大量的研究表明，海藻糖是单细胞生物、动物组织和器官、蛋白质、生物膜、医药制剂等的保护剂，能够抑制脂质酸化、淀粉老化、蛋白质变性，具有矫味矫臭功能、搞玻璃转化温度、低吸湿性、低甜度等性质，能够将其应用到医药业、农业、生化制品业、化妆品行业、食品加工业。
早期商业化的海藻糖是从酵母中提取的。1990年价格约700美元/kg，提取率过低，成本过高。1995年日本利用双酶法实现工业化生产，使得海藻糖价格由原来的2万日元/kg大幅降到1997年的280日元/kg。中国2002年首次以双酶法实现海藻糖的产业化，价格79元/kg。双酶法以淀粉为原料，在麦芽寡糖基海藻糖水解酶和麦芽寡糖基海藻糖合成酶的作用下生成海藻糖，此法生产工艺复杂，难以推广，目前全世界只有几个公司能生产。而海藻糖合酶以麦芽糖为底物，一步转化生成海藻糖，是相对经济的生产方法，但仍有很多问题需要研究解决，其中海藻糖合酶是关键。因此，挖掘适合生产海藻糖的海藻糖合酶对于推动海藻糖的大规模工业化、降低产业成本具有重大的意义。
麦芽糖可以通过水解淀粉得到，生产过程中会产生一定的葡萄糖，工业化生产海藻糖使用纯的麦芽糖成本过高。海藻糖合酶除了转苷作用外还有微弱的水解反应，由此生成副产物葡萄糖。本发明所使用的来源于Thermobifida fusca YX海藻糖合酶若以工业级的麦芽糖(含有1％-10％的葡萄糖)为底物，海藻糖合酶酶转化率会受到一定的影响。本发明通过定点突变对海藻糖合酶进行改造，削弱葡萄糖对酶转化的抑制作用。
发明内容
本发明所要解决的一个技术问题是提供一种海藻糖合酶的突变体，其活性中心位点附近氨基酸发生突变，得到受葡萄糖抑制影响减弱的海藻糖合酶。所述突变体包括含有一个、两个或三个相对于Thermobifida fusca YX海藻糖合酶活性氨基酸残基的取代。
所述海藻糖合酶的亲本的氨基酸序列与NCBI数据库中的Thermobifida fusca YX海藻糖合酶一致(登录号WP_011291031.1)。
所述突变体是将亲本海藻糖合酶的第289位的谷氨酸(Glu)突变为甘氨酸(Gly)，命名为E289G；或将第295位的组氨酸(His)突变成天冬酰胺(Asn)，所得突变体命名为H295N；或将第344位的甲硫氨酸(Met)突变成赖氨酸(Lys)，所得突变体命名为M344K；或将第367位的甲硫氨酸(Met)突变成亮氨酸(Leu)，所得突变体命名为M367L。
所述突变体还可以是将单突变体酶H295N中第289位的谷氨酸(Glu)突变为甘氨酸(Gly)，所得突变体命名为H295N/E289G；或将单突变体酶H295N中第344位的甲硫氨酸(Met)突变成赖氨酸(Lys)，所得突变体命名为H295N/M344K；或将单突变体酶H295N基因中第367位的甲硫氨酸(Met)突变成亮氨酸(Leu)，所得突变体命名为H295N/M367L。
所述突变体还可以是将双突变体酶H295N/M344K基因中第367位的甲硫氨酸(Met)突变成亮氨酸(Leu)，所得突变体命名为H295N/M344K/M367L。
本发明所要解决的另一个技术问题是提供一种受葡萄糖抑制影响减弱的海藻糖合酶突变体的制备方法，包括如下步骤：
(1)在Thermobifida fusca YX海藻糖合酶氨基酸序列的基础上确定突变位点；设计定点突变的突变引物，以携带海藻糖合酶基因的载体为模板进行定点突变；构建含突变体的质粒载体；
(2)将突变体质粒转化进宿主细胞；
(3)挑选阳性克隆进行发酵培养，并纯化获得海藻糖合酶突变体。
在本发明的一种实施方式中，所述质粒载体为pUC系列，pET系列，或pGEX中的任意一种。
在本发明的一种实施方式中，所述宿主细胞为细菌或真菌细胞。
在本发明的一种实施方式中，所述的细菌为革兰氏阴性菌或革兰氏阳性菌。
本发明削弱了以工业级麦芽糖(含葡萄糖10％)为底物时葡萄糖对海藻糖合酶生产海藻糖的抑制作用，以工业级麦芽糖(含葡萄糖10％)为底物，野生酶生产海藻糖转化率为62.2％，而突变体E289G、H295N、M344K、M367L、H295N/E289G、H295N/M344K、H295N/M367L、H295N/M344K/M367L，生产海藻糖的转化率分别达到69.7％、70.5％、70.3％、69.6％、70.4％、70.9％、72.3％、73.7％，达到野生酶使用纯麦芽糖为底物时生产海藻糖的转化率(70.7％)。
具体实施方式
实施例1：重组菌构建
实验室保藏有前期构建的含有编码海藻糖合酶的基因的质粒TreS/pMD 18T。用于构建大肠杆菌的质粒是pET24a(+)，带有T7启动子。将pET24a(+)质粒和含有TreS基因的质粒分别进行NdeⅠ和Hind Ⅲ双酶切，酶切产物割胶回收后，再用T4连接酶连接，连接产物转化E.coli JM109感受态细胞，经37℃培养培养8h，挑转化子在含有30mg/L卡那霉素液体的LB培养基中震荡培养，提取质粒，酶切验证得到表达质粒TreS/pET24a(+)。
将质粒TreS/pET24a(+)转化E.coli BL21(DE3)宿主菌，涂布含卡那霉素(30mg/L)的LB平板，37℃培养8h，命名为TreS/pET24a(+)/E.coli BL21(DE3)。挑单菌落到含有30mg/L卡那霉素液体LB培养基中，37℃培养过夜，保存甘油管。
实施例2：突变体的制备
(1)单突变
来源于Thermobifida fusca YX的海藻糖合酶的突变体酶E289G、H295N、M344K、M367L。
根据Thermobifida fusca YX的海藻糖合酶的基因序列，分别设计并合成引入E289G、H295N、M344K、M367L突变的引物，对海藻糖合酶基因进行定点突变，测定DNA编码序列，分别鉴别出第289位的Glu密码子变成Gly密码子，第295位的His密码子变成Asn密码子，第344位Met密码子变成Lys密码子，第367位Met密码子变成Leu密码子。将突变体基因置于适当的表达载体并导入大肠杆菌中进行表达，得到单突变海藻糖合酶。单突变E289G、H295N、M344K、M367L的定点突变：利用快速PCR技术，以表达载体TreS/pET24a(+)为模板，
引入E289G突变的定点突变引物为：
正向引物：5’-GAATCCGGCGGCGACGGTTGCCACATGAACT-3’(下划线为突变碱基)
反向引物：5’-AGTTCATGTGGCAACCGTCGCCGCCGGATTCG-3’(下划线为突变碱基)
引入H295N突变的定点突变引物为：
正向引物：5’-ATGCCACATGAACTTCAACTTCCCGCTGATGCC-3’(下划线为突变碱基)
反向引物：5’-GGCATCAGCGGGAAGTTGAAGTTCATGTGGCA-3’(下划线为突变碱基)
引入M344K突变的定点突变引物为：
正向引物：5’-CGAGCTGACCTTGGAGAAAGTCAGCGATGAAG-3’(下划线为突变碱基)
反向引物：5’-TCTTCATCGCTGACTTTCTCCAAGGTCAGCTCG-3’(下划线为突变碱基)
引入M367L突变的定点突变引物为：
正向引物：5’-GCGGATGCGCGCCAACTTAGGGATCCGCCGCCGGC-3’(下划线为突变碱基)
反向引物：5’-GCCGGCGGCGGATCCCTAAGTTGGCGCGCATCCGC-3’(下划线为突变碱基)
PCR反应体系均为：5×PS buffer 10μL，dNTPs Mix(2.5mM)4μL，正向引物(10μM)1μL，反向引物(10μM)1μL，模板DNA 1μL，PrimerStar HS(5U/μL)0.5μL，加入双蒸水至50μL。
PCR扩增条件为：94℃预变性4min；随后30个循环(98℃10s，55℃5s，72℃8min)；72℃继续延伸10min。
PCR产物经Dpn Ⅰ消化，转化大肠杆菌JM109感受态，感受态细胞在LB固体培养基(含30μg/mL卡那霉素)培养过夜后，挑克隆于LB液体培养基(含30μg/mL卡那霉素)中培养后提取质粒，将突变质粒转化表达宿主大肠杆菌BL21(DE3)感受态细胞，所有突变质粒均测序正确。
发酵产酶
(2)双突变
Thermobifida fusca YX的海藻糖合酶的双突变体酶H295N/E289G、H295N/M344K、H295N/M367L：将单突变体酶H295N基因中第289位的谷氨酸(Glu)突变成甘氨酸(Gly)，或者第344位的甲硫氨酸(Met)突变成赖氨酸(Lys)，或者第367位的甲硫氨酸(Met)突变成亮氨酸(Leu)，分别命名为H295N/E289G、H295N/M344K、H295N/M367L。双突变体酶的制备方法，以单突变体H295N编码基因为模板，分别设计并合成引入E289G、M344K、M367L突变的引物，对单突变体H295N编码基因进行定点突变，测定序列，鉴别出第289位的Glu变成Gly密码子，第344位的Met密码子变成Lys密码子，或者第367位的Met变成Leu密码子，将突变体基因置于适当的表达载体并导入大肠杆菌中进行表达，得到双突变海藻糖合酶突变体。
双突变H295N/E289G、H295N/M344K、H295N/M367L的定点突变：利用快速PCR技术，以表达载体H295N/pET24a(+)为模板，
引入E289G突变的定点突变引物为：
正向引物：5’-GAATCCGGCGGCGACGGTTGCCACATGAACT-3’(下划线为突变碱基)
反向引物：5’-AGTTCATGTGGCAACCGTCGCCGCCGGATTCG-3’(下划线为突变碱基)
引入M344K突变的定点突变引物为：
正向引物：5’-CGAGCTGACCTTGGAGAAAGTCAGCGATGAAG-3’(下划线为突变碱基)
反向引物：5’-TCTTCATCGCTGACTTTCTCCAAGGTCAGCTCG-3’(下划线为突变碱基)
引入M367L突变的定点突变引物为：
正向引物：5’-CGGATGCGCGCCAACTTAGGGATCCGCCGCCG-3’(下划线为突变碱基)
反向引物：5’-CCGGCGGCGGATCCCTAAGTTGGCGCGCATCC-3’(下划线为突变碱基)
PCR反应体系、反应条件及突变基因的测序方法同单突变体的方法。
(3)三突变
Thermobifida fusca YX的海藻糖合酶的双突变体酶H295N/M344K/M367L：将双突变体酶H295N/M344K基因中第367位的甲硫氨酸(Met)突变成亮氨酸(Leu)，命名为H295N/M344K/M367L。三突变体酶的制备方法，以双突变体H295N/M344K编码基因为模板，设计并合成引入M367L突变的引物，对双突变体H295N/M344K编码基因进行定点突变，测定序列，鉴别出第367位的Met变成Leu密码子，将突变体基因置于适当的表达载体并导入大肠杆菌中进行表达，得到三突变海藻糖合酶突变体。
引入M367L突变的定点突变引物为：
正向引物：5’-CGGATGCGCGCCAACTTAGGGATCCGCCGCCG-3’(下划线为突变碱基)
反向引物：5’-CCGGCGGCGGATCCCTAAGTTGGCGCGCATCC-3’(下划线为突变碱基)
PCR反应体系、反应条件及突变基因的测序方法同单突变体的方法。
(4)突变体酶的发酵与纯化
挑取转入表达宿主大肠杆菌BL21(DE3)的阳性克隆于LB液体培养基(含30μg/mL卡那霉素)生长8～10h，按5％接种量将种子发酵液接到TB培养基(含30μg/mL卡那霉素)中，在37℃摇床中培养48h后，将发酵液于4℃、8000rpm离心10min除菌体，收集离心上清液。
实施例3：HPLC检测海藻糖的产量
在反应器中加入麦芽糖300g/L(含葡萄糖10％)，加入一定量实例2中获得的野生酶和突变体的浓缩酶液，用20％的氢氧化钠水溶液将pH调节到8.0，在30℃、150rpm的水浴摇床中反应30-50小时定时取样，煮沸10min终止反应后将样品12000rpm离心10min，取上清液适度稀释后用0.45μm超滤膜过滤，并进行HPLC分析。色谱条件如下：示差折光检测器，NH2柱(APS-2HYPERSIL，Thermo Scientific)，流动相(水:乙腈＝1:4)，流速：0.8mL·min-1，柱温：40℃。
表1以工业级麦芽糖为底物生产海藻糖的转化率
| 酶 | 转化率(％) |
| 野生酶 | 62.2％ |
| E289G | 69.7％ |
| H295N | 70.5％ |
| M344K | 70.3％ |
| M367L | 69.6％ |
| H295N/E289G | 70.4％ |
| H295N/M344K | 70.9％ |
| H295N/M367L | 71.3％ |
| H295N/M344K/M367L | 73.7％ |
结果见表1，突变体表达获得的突变酶与野生酶相比，可以发现，突变体实现了海藻糖合酶制备海藻糖转化效率的提高。野生酶生产海藻糖转化率为62.2％，而突变体E289G、H295N、M344K、M367L、H295N/E289G、H295N/M344K、H295N/M367L、H295N/M344K/M367L，生产海藻糖的转化率分别达到69.7％、70.5％、70.3％、69.6％、70.4％、70.9％、72.3％、73.7％，达到野生酶使用纯麦芽糖为底物时生产海藻糖的转化率(70.7％)。
<110> 湖南汇升生物科技有限公司
<120> 一种海藻糖合酶突变体及其在制备海藻糖中的应用
<160> 17
<170> PatentIn version 3.3
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<212> PRT
<213> Thermobifida fusca
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545 550 555 560
Gly His Gly Phe Tyr Trp Phe Gln Leu Pro Pro Val Ala Glu Glu Gln
565 570 575
Pro Leu Ala Gln Pro Val Thr Thr Val Pro Ala Ala Pro Gln Pro Pro
580 585 590
Ala Pro Ala Asp Arg Pro Ala Ser Asp Pro Thr Gln Arg Ser
595 600 605
<210> 2
<211> 31
<212> DNA
<213> 人工序列
<400> 2
gaatccggcg gcgacggttg ccacatgaac t 31
<210> 3
<211> 32
<212> DNA
<213> 人工序列
<400> 3
agttcatgtg gcaaccgtcg ccgccggatt cg 32
<210> 4
<211> 33
<212> DNA
<213> 人工序列
<400> 4
atgccacatg aacttcaact tcccgctgat gcc 33
<210> 5
<211> 32
<212> DNA
<213> 人工序列
<400> 5
ggcatcagcg ggaagttgaa gttcatgtgg ca 32
<210> 6
<211> 32
<212> DNA
<213> 人工序列
<400> 6
cgagctgacc ttggagaaag tcagcgatga ag 32
<210> 7
<211> 33
<212> DNA
<213> 人工序列
<400> 7
tcttcatcgc tgactttctc caaggtcagc tcg 33
<210> 8
<211> 35
<212> DNA
<213> 人工序列
<400> 8
gcggatgcgc gccaacttag ggatccgccg ccggc 35
<210> 9
<211> 35
<212> DNA
<213> 人工序列
<400> 9
gccggcggcg gatccctaag ttggcgcgca tccgc 35
<210> 10
<211> 31
<212> DNA
<213> 人工序列
<400> 10
gaatccggcg gcgacggttg ccacatgaac t 31
<210> 11
<211> 32
<212> DNA
<213> 人工序列
<400> 11
agttcatgtg gcaaccgtcg ccgccggatt cg 32
<210> 12
<211> 32
<212> DNA
<213> 人工序列
<400> 12
cgagctgacc ttggagaaag tcagcgatga ag 32
<210> 13
<211> 33
<212> DNA
<213> 人工序列
<400> 13
tcttcatcgc tgactttctc caaggtcagc tcg 33
<210> 14
<211> 32
<212> DNA
<213> 人工序列
<400> 14
cggatgcgcg ccaacttagg gatccgccgc cg 32
<210> 15
<211> 32
<212> DNA
<213> 人工序列
<400> 15
ccggcggcgg atccctaagt tggcgcgcat cc 32
<210> 16
<211> 32
<212> DNA
<213> 人工序列
<400> 16
cggatgcgcg ccaacttagg gatccgccgc cg 32
<210> 17
<211> 32
<212> DNA
<213> 人工序列
<400> 17
ccggcggcgg atccctaagt tggcgcgcat cc 32
Claims (7)
1.一种海藻糖合酶突变体，其特征在于，相对于海藻糖合酶亲本，将第367位的甲硫氨酸突变成亮氨酸；或将第367位的甲硫氨酸突变成亮氨酸，同时将第295位的组氨酸突变成天冬酰胺；所述海藻糖合酶亲本的氨基酸序列如SEQ ID NO.1所示。
2.制备权利要求1所述突变体的方法，其特征在于，包括如下步骤：
(1)在亲本海藻糖合酶氨基酸序列的基础上确定突变位点；设计定点突变的突变引物，以携带海藻糖合酶基因的载体为模板进行定点突变并构建含突变体的质粒载体；
(2)将携带编码突变体的基因的质粒转化进宿主细胞；
(3)挑选阳性克隆进行发酵培养，并纯化得到海藻糖合酶突变体。
3.根据权利要求2所述的方法，其特征在于，所述质粒载体为pUC系列，pET系列，或pGEX中的任意一种。
4.权利要求1所述突变体在制备海藻糖中的应用。
5.编码权利要求1所述突变体的基因。
6.携带权利要求5所述基因的细胞。
7.权利要求6所述细胞在制备海藻糖中的应用。
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| CN103421761A (zh) * | 2013-08-23 | 2013-12-04 | 南宁中诺生物工程有限责任公司 | 一种耐热海藻糖合成酶融合酶突变体基因及其应用 |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103421761A (zh) * | 2013-08-23 | 2013-12-04 | 南宁中诺生物工程有限责任公司 | 一种耐热海藻糖合成酶融合酶突变体基因及其应用 |
Non-Patent Citations (1)
| Title |
|---|
| 朱绮霞 等: "谷氨酸棒杆菌（Corynebacterium glutamicum）合成酶的定点突变及其酶学性质研究", 《广西科学》 * |
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