effect of N-terminal sequence of RSuS2 on its enzyme activity

• Main Authors: Cheng-Tze Liang, 梁成芝
• Other Authors: Ai-Yu Wang
• Format: Others
• Language: zh-TW
• Published: 2007
• Online Access: http://ndltd.ncl.edu.tw/handle/98511212457519155497

碩士 === 臺灣大學 === 微生物與生化學研究所 === 95 === Sucrose synthase catalyzes the reversible conversion of sucrose and UDP into fructose and UDPG. In rice, the enzyme may be encoded by six RSus genes. The expression product of RSus2 gene has been heterologously expressed and characterized in Pichia pastoris in the previous study. However, a point mutation was found in the 5’-end of RSus2 open reading frame. The mutation may cause the deletion of 39 a.a. residues of the recombinant RSuS2 protein. In order to investigate whether the biochemical properties of the recombinant RSuS2 protein are affected by its N-terminal amino acid sequence, the expression plasmid carrying the wild-type RSus2 open reading frame was constructed and transformed into P. pastoris X33 for expression. The wild-type recombinant RSuS2 protein was purified by DEAE SephacelTM ion exchange chromatography, SephacrylTM S-300 gel filtration chromatography and Ni-NTA affinity chromatography, and used for biochemical characterization. The results of enzyme kinetic analysis revealed that the N-terminal part of RSuS2 may involve in sucrose binding.