C-terminal Protein Engineering of α-amylase from Vibrio parahaemolyticus

• Main Authors: Ming-Shang Fang, 方銘盛
• Other Authors: Fu-Pang Lin
• Format: Others
• Language: zh-TW
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/29732608401118185292

碩士 === 國立臺灣海洋大學 === 生物科技研究所 === 100 === The α-amylase gene from Vibrio parahaemolyticus BCRC13026 (VpAmy) encoding a protein with MW of 53kDa was cloned and expressed in Escherichia coli-pET20b(+) expression system with α-amylase activity. Protease factor Xa cleavage site of Ile-Glu-Gly-Arg was inserted at K424 amino acid of VpAmy by PCR techniques (VpAmyK424/Xa). In order to characterize the VpAmyK424 mutant enzyme, the recombinant VpAmyK424 was attempted to be obtained by the proteolytic cleavage of VpAmyK424/Xa with protease factor Xa in vitro. Results indicated that the VpAmyK424/Xa was not expressed from the E.coli expression system and was confirmed by peptide mass finger printing analysis (PMF).