Cloning of pectin methylesterase inhibitor from kiwi fruit and its high expression in Pichia pastoris
A new cDNA fragment from kiwi (Actinidia deliciosa) was cloned by RT-PCR and successfully expressed in Pichia pastoris KM71. Like the pectin methylesterase inhibitor (PMEI) purified from kiwi fruit, the recombinant protein (designated as kWPMEI) was able to inhibit pectin methylesterases of some plants. Therefore, kWPMEI was a new isoform of the native PMEI. By G418 resistance screening, six recombinant strains containing multiple copies were obtained. The results of recombinant protein expression indicated that the six strains produced high-level kWPMEI. The highest yield of kWPMEI reached 0.2 g/L. In addition, kWPMEI was purified by single gel filtration chromatography, which greatly simplified the procedure of protein purification. The success of production of the PMEI in P. pastoris significantly broadened its applications in juice industry.
Journal: Enzyme and Microbial Technology - Volume 40, Issue 5, 3 April 2007, Pages 1001–1005