文献类型：期刊文献

英文题名：Effect of glycosylation on biochemical characterization of recombinant phytase expressed in Pichia pastoris

作者：Guo, Meijin[1,2];Hang, Haifeng[1];Zhu, Taicheng[1];Zhuang, Yingping[1];Chu, Ju[1];Zhang, Siliang[1]

机构：[1]E China Univ Sci & Technol, State Key Lab Bioreactor Engn, Shanghai 200237, Peoples R China;[2]Jiangxi Agr Univ, Dept Biotechnol, Nanchang 330045, Peoples R China

年份：2008

卷号：42

期号：4

起止页码：340

外文期刊名：ENZYME AND MICROBIAL TECHNOLOGY

收录：;WOS:【SCI-EXPANDED(收录号:WOS:000254728500006)】；

语种：英文

外文关键词：phytase; glycosylation; biochemical characterization; thermostability; micro-heterogeneity

摘要：Recombinant phytase expressed in Pichia pastoris FPHY34 is a typical glycoprotein, whose gene possesses 10 potential N-glycosylation sites without O-glycosylation. Effects of glycosylation on phytase's biochemical characterization were investigated. Secreted phytase's microheterogeniousglycosylation phenomenon was observed and verified by SDS-PAGE, N-terminal amino acid sequencing and mass spectrum analyses. Deglycosylation of recombinant phytase reduced the molecular weight from 83 to 63 kDa, which indicates the presence of total carbohydrate content of approximately 24.1%. According to image analysis of 2-dimension gel electrophoresis, the range of isoeclectric point of phytase covers nearly I pH value rather than a specific point, which might be due to heavy and complex glycosylation. The phytase expressed by P pastoris still retained 40% and 30% activities at 80 and 90 degrees C for 10 min, respectively. However, upon cleglycosylation in vitro by peptide-N-glycohydrolase F (PNGase F). thermostability of deglycosylated phytase significantly declined after 10 min treatment at 40 and 50 degrees C. Optimum pH of phytase tested was shifted from pH 5.0 to 2.5 by cleglycosylation, although there exhibit bi-humps pH optima at both before and after PNGase F digestion cases. (c) 2007 Elsevier Inc. All rights reserved.