title: Improving the secretion of a methyl parathion hydrolase in Pichia pastoris by modifying its N-terminal sequence.
abstract: Pichia pastoris is commonly used to express and secrete target proteins, although not all recombinant proteins can be successfully produced. In this study, we used methyl parathion hydrolase (MPH) from Ochrobactrum sp. M231 as a model to study the importance of the N-terminus of the protein for its secretion. While MPH can be efficiently expressed intracellularly in P. pastoris, it is not secreted into the extracellular environment. Three MPH mutants (N66-MPH, D10-MPH, and N9-MPH) were constructed through modification of its N-terminus, and the secretion of each by P. pastoris was improved when compared to wild-type MPH. The level of secreted D10-MPH was increased to 0.21 U/mL, while that of N9-MPH was enhanced to 0.16 U/mL. Although N66-MPH was not enzymatically active, it was secreted efficiently, and was identified by SDS-PAGE. These results demonstrate that the secretion of heterologous proteins in P. pastoris may be improved by modifying their N-terminal structures.
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Ping
authors.initials: P
authors.lastname: Wang
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Lu
authors.initials: L
authors.lastname: Huang
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Hu
authors.initials: H
authors.lastname: Jiang
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Jian
authors.initials: J
authors.lastname: Tian
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Xiaoyu
authors.initials: X
authors.lastname: Chu
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Ningfeng
authors.initials: N
authors.lastname: Wu
mesh_terms: D000595:Amino Acid Sequence; D015971:Gene Expression Regulation, Enzymologic; D008743:Methyl Parathion / Q000378:metabolism; D042041:Ochrobactrum / Q000201:enzymology; D010744:Phosphoric Monoester Hydrolases / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D010843:Pichia / Q000201:enzymology / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D008743:Methyl Parathion; D010744:Phosphoric Monoester Hydrolases