EXPRESSION OF L-N-CARBAMOYLASE GENE IN PICHIA PASTORIS / 微生物学通报

ABSTRACT

N-carbamoylase is a part of hydantoinase operon which can transform N-carbamoylamino acid to corresponding ammo acids. The L-N-carbamoylase of Arthrobacter BT801, codied by the HyuC gene, is the rate-limiting and the only stereoselective enzyme. HyuC DNA fragment was amplified by PCR from the plasmid of pUC18-169. The target fragment was introduced into pPIC3. 5K plasmid to construct the pPIC3. 5K-hyuC expressing vector which was then transduced into Pichia pastoris GS115 cells after being linearized by BglⅡ digestion. Multi-copies insertion transformants were screened on G418 plates. The recombinant protein was proved to have biological activity of hydrolyzing N-carbamoylphenylalanine into phenylalanine through enzyme activity determination.