title: Expression, purification and characterization of pectin methylesterase inhibitor from kiwi fruit in Escherichia coli.
abstract: A significant problem in production of fruit juices for human consumption is auto-clarification, where enzyme catalyzes pectin demethylation resulting in loss of the ''natural" cloudy appearance of juices. To overcome this problem, a plant inhibitor protein which blocks the action of pectin methylesterase has been used. In this paper, expression of recombinant kiwi pectin methylesterase inhibitor (PMEI) was carried out in Escherichia coli, and the target protein was expressed in the form of inclusion bodies. The expression level reached 46% of total cell protein. Then the fusion protein was purified by nickel ion metal affinity chromatography, and the purity was finally up to 98%. After refolding in GSH/GSSG redox system, recombinant PMEI not only could efficiently inhibit PMEs from eight different plants, but could remain effective inhibitor activity in the pH 3.0-10.0 and 20-40 degrees C. Thus, recombinant PMEI has potential application in the production of fruit juices product industry.
authors.affiliation: College of Food Science and Nutritional Engineering, China Agricultural University, 17 Qing Hua East Road, Hai Dian District, Beijing 100083, PR China.
authors.forename: Yanling
authors.initials: Y
authors.lastname: Hao
authors.forename: Xinyi
authors.initials: X
authors.lastname: Huang
authors.forename: Xiaohong
authors.initials: X
authors.lastname: Mei
authors.forename: Ruoyu
authors.initials: R
authors.lastname: Li
authors.forename: Zhengyuan
authors.initials: Z
authors.lastname: Zhai
authors.forename: Sheng
authors.initials: S
authors.lastname: Yin
authors.forename: Ying
authors.initials: Y
authors.lastname: Huang
authors.forename: Yunbo
authors.initials: Y
authors.lastname: Luo
mesh_terms: D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005638:Fruit / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D013696:Temperature
chemical_list: C412789:PMEI protein, Actinidia chinensis; D010940:Plant Proteins; D011994:Recombinant Proteins