其他题名Cloning,expression,purification and characterization of thermostable and acid-stable α-amylase from Thermococuus HJ21
王淑军1; 吕明生1; 秦松2
; 陆兆新3; 房耀维1; 邓祥元2; 林谦3; 刘红飞1发表期刊海洋学报
ISSN0253-4193
2011
卷号33.0期号:003页码:158-164
关键词深海古菌高温酸性α-淀粉酶基因克隆表达酶学性质
英文摘要通过简并引物扩增热球菌（Thermococcus siculi）HJ21高温α-淀粉酶保守区域之间的序列,再利用Site-finding技术获得两端未知序列。构建了在N端添加了His6标签的表达载体pEt-28a-His6-THJA后转化E.coli,在IPTG诱导下表达。进一步纯化后SDS-PAGE电泳检测达到电泳纯,重组酶分子量为50 KDa。重组酶最适作用温度和pH值分别为90℃,pH5.0。重组酶在pH为5.5时最稳定;K＋,Sr2＋,Mg2＋,Na＋对α-淀粉酶活力的促进作用不显著,Cu2＋,Pb2＋,Hg2＋,Zn2＋,N-溴代丁酰亚胺和三氯乙酸对该酶有显著抑制作用。
中文摘要The gene sequence between the conserved regions was acquired by PCR using the primers designed based on the sequence of that of Thermococcus siculi.HJ21 deposited in the GenBank.The upstream and downstream of the HJ21 α-amylase gene were acquired by sitefinding PCR.The expression plasmid pEt-28a-His6-THJA was structured.The plasmid was transformed into E.coli stain TOP 10F′ and expressed.The His6-α-amylase was further purified.Its molecular weight was about 54.5 KDa detected by SDS-PAGE.The His6-α-amylase was further purified.The optimal temperature and pH of His6-α-amylase are 90 ℃ and 5.0 respectively.K＋,Sr2＋,Mg2＋,Na＋ improved the activity of His6-α-amylase while Cu2＋,Pb2＋,Hg2＋,Zn2＋ reduced activity of His6-α-amylase.N-bromosuccinimide and TCA significantly inhibited the activity of His6-α-amylase.
收录类别CSCD
语种中文
CSCD记录号CSCD:4212077
引用统计
文献类型期刊论文
条目标识符http://ir.yic.ac.cnhttp://ir.yic.ac.cn/handle/133337/28168
专题海岸带生物学与生物资源利用重点实验室
海岸带生物学与生物资源利用重点实验室_海岸带生物学与生物资源保护实验室
作者单位1.淮海工学院海洋学院
2.中国科学院海岸带研究所
3.南京农业大学食品科技学院
推荐引用方式
GB/T 7714王淑军,吕明生,秦松,等. 热球菌HJ21高温α-淀粉酶基因克隆、表达及性质研究[J]. 海洋学报,2011,33.0(003):158-164.
APA王淑军.,吕明生.,秦松.,陆兆新.,房耀维.,...&刘红飞.(2011).热球菌HJ21高温α-淀粉酶基因克隆、表达及性质研究.海洋学报,33.0(003),158-164.
MLA王淑军,et al."热球菌HJ21高温α-淀粉酶基因克隆、表达及性质研究".海洋学报 33.0.003(2011):158-164.
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