Mutational analysis of the active center of plant fructosyltransferases: Festuca 1-SST and barley 6-SFT
- 作者：Denise Altenbach ; Eveline Nü ; esch ; Tita Ritsema ; Thomas Boller and Andres Wiemken
- 关键词：Fructosyltransferase ; Invertase ; Enzyme specificity ; Fructan ; Pichia pastoris
- 刊名：FEBS Letters
- 出版年：2005
- 期刊代码：70_00145793
- 类别：bio
- 出版时间：2005
- 卷：579
- 期：21
- 页码：4647-4653
- 文件大小：316 K
摘要
The active center of the glycoside hydrolase family 32 contains the three characteristic motifs (N/S)DPNG, RDP, and EC. We replaced the N-terminal region including the (N/S)DPNG motif of barley 6-SFT (sucrose:fructan 6-fructosyltransferase) by the corresponding region of Festuca 1-SST (sucrose:sucrose 1-fructosyltransferase). The chimeric enzyme, expressed in Pichia, retained the specificity of 6-SFT. Attempts to replace a larger piece at the N-terminus including also the RDP motif failed. A point mutation introduced in the RDP motif of 1-SST abolished enzymatic activity. Interestingly, point mutations of the EC-motif resulted in an enzyme which had lost the capability to form 1-kestose and glucose from sucrose but still accepted 1-kestose, producing fructose and sucrose as well as nystose.