Engineering Pichia pastoris for biocatalysis: co-production of two active enzymes.
High levels of active glycolate oxidase from spinach (GO) and active catalase T from Saccharomyces cerevisiae (catT) have been co-produced in the methylotrophic yeast Pichia pastoris (Pp). In sequential rounds of transformation using two selectable markers, multiple copies of the genes encoding GO and catT were integrated into the Pp chromosome under control of the methanol inducible alcohol oxidase I promoter, resulting in a strain designated MSP8.6. MSP8.6 is a second-generation biocatalyst used for the conversion of glycolate to glyoxylate in the presence of a reaction component which inhibits endogenous Pp catalase. This work demonstrates a significant advance in the utility of recombinant Pp for commercial bioprocess development.
Payne
M S
MS
DuPont Central Research and Development, Glasgow Site 301, Newark, DE 19714-6101, USA. paynems@esvax.dnet.dupont.com
Petrillo
K L
KL
Gavagan
J E
JE
DiCosimo
R
R
Wagner
L W
LW
Anton
D L
DL
D000429:Alcohol Oxidoreductases / Q000096:biosynthesis* / Q000235:genetics; D002374:Catalase / Q000096:biosynthesis* / Q000235:genetics; D002384:Catalysis; D003001:Cloning, Molecular / Q000379:methods; D004789:Enzyme Activation; D005656:Fungal Proteins / Q000096:biosynthesis* / Q000235:genetics; D005818:Genetic Engineering; D010843:Pichia / Q000235:genetics*; D010940:Plant Proteins / Q000096:biosynthesis* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D018724:Spinacia oleracea; D014170:Transformation, Genetic
D005656:Fungal Proteins; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; D000429:Alcohol Oxidoreductases; C020821:glycollate oxidase; D002374:Catalase