title: Enhanced secretion of a methyl parathion hydrolase in Pichia pastoris using a combinational strategy.
abstract: BACKGROUND
  Although Pichia pastoris has been successfully used to produce various recombinant heterologous proteins, the efficiency varies. In this study, we used methyl parathion hydrolase (MPH) from Ochrobactrum sp. M231 as an example to study the effect of protein amino acid sequence on secretion from P. pastoris.
  
  
  RESULTS
  The results indicated that the protein N-terminal sequence, the endoplasmic reticulum (ER) retention signal (KKXX) at the protein C-terminus, and the acidic stability of the protein could affect its secretion from P. pastoris. Mutations designed based on these sequence features markedly improved secretion from P. pastoris. In addition, we found that the secretion properties of a protein can be cumulative when all of the above strategies are combined. The final mutant (CHBD-DQR) designed by combining all of the strategies greatly improved secretion and the secreted MPH activity of CHBD-DQR was enhanced up to 195-fold compared with wild-type MPH without loss of catalytic efficiency.
  
  
  CONCLUSIONS
  These results demonstrate that the secretion of heterologous proteins from P. pastoris could be improved by combining changes in multiple protein sequence features.
authors: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, 100081, People's Republic of China. wangping84040@163.com.
authors: Ping
authors: P
authors: Wang
authors: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, 100081, People's Republic of China. Lu.huang@sdstate.edu.
authors: Lu
authors: L
authors: Huang
authors: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, 100081, People's Republic of China. jhzy0315@foxmail.com.
authors: Hu
authors: H
authors: Jiang
authors: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, 100081, People's Republic of China. tianjian@caas.cn.
authors: Jian
authors: J
authors: Tian
authors: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, 100081, People's Republic of China. sadylee@163.com.
authors: Xiaoyu
authors: X
authors: Chu
authors: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, 100081, People's Republic of China. wuningfeng@caas.cn.
authors: Ningfeng
authors: N
authors: Wu
mesh_terms: D018628:Gene Dosage; D005818:Genetic Engineering; D030781:Organisms, Genetically Modified; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D010843:Pichia / Q000235:genetics / Q000378:metabolism*; D021382:Protein Sorting Signals; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D020539:Sequence Analysis, Protein
chemical_list: D021382:Protein Sorting Signals; D012333:RNA, Messenger; D011994:Recombinant Proteins; D010744:Phosphoric Monoester Hydrolases