Cloning, expression, purification and characterization of thermostable and acid-stable α-amylase from <em>Thermococuus</em> HJ21

WANG Shu-jun; LU Ming-sheng; QIN Song; LU Zhao-xin; FANG Yao-wei; DENG Xiang-yuan; LIN Qian; LIU Hong-fei

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Article Navigation > Haiyang Xuebao > 2011 > 33(3): 158-164

WANG Shu-jun, LU Ming-sheng, QIN Song, LU Zhao-xin, FANG Yao-wei, DENG Xiang-yuan, LIN Qian, LIU Hong-fei. Cloning, expression, purification and characterization of thermostable and acid-stable α-amylase from Thermococuus HJ21[J]. Haiyang Xuebao, 2011, 33(3): 158-164.

Citation:

WANG Shu-jun, LU Ming-sheng, QIN Song, LU Zhao-xin, FANG Yao-wei, DENG Xiang-yuan, LIN Qian, LIU Hong-fei. Cloning, expression, purification and characterization of thermostable and acid-stable α-amylase from Thermococuus HJ21[J]. Haiyang Xuebao, 2011, 33(3): 158-164.

Citation:

WANG Shu-jun, LU Ming-sheng, QIN Song, LU Zhao-xin, FANG Yao-wei, DENG Xiang-yuan, LIN Qian, LIU Hong-fei. Cloning, expression, purification and characterization of thermostable and acid-stable α-amylase from Thermococuus HJ21[J]. Haiyang Xuebao, 2011, 33(3): 158-164.

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Cloning, expression, purification and characterization of thermostable and acid-stable α-amylase from Thermococuus HJ21

1.
School of Marine Science and Technology, Huaihai Institute of Technology, Lianyungang 222005, China
2.
Yantai Instisute of Coastal Zone Research, Chinese Academy of Sciences, Yantai 264003, China
3.
College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, China;4.Jiangsu Marine Resources Develepment Research Insititute,Liangyuangang 222005,China

Received Date: 2010-08-09

Abstract

Abstract

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^＋,Sr^2＋,Mg^2＋,Na^＋ improved the activity of His₆-α-amylase while Cu^2＋,Pb^2＋,Hg^2＋,Zn^2＋ reduced activity of His₆-α-amylase. N-bromosuccinimide and TCA significantly inhibited the activity of His₆-α-amylase.
- archaeon in deep sea,
- thermostable and acid-stable α-amylase,
- gene cloning,
- expression,
- characterization

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References(13)

References

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