Pilot scale production of a heterologous Trichoderma reesei cellulase by Saccharomyces cerevisiae.
Cellobiohydrolase II of Trichoderma reesei was produced in laboratory and pilot scale using a transformant strain of Saccharomyces cerevisiae harbouring a multicopy expression plasmid. Different strategies were compared for concentration and partial purification of the enzyme produced in a 200 1 pilot cultivation. After efficient separation of biomass and sub-cellular particulate matter, a combination of ultrafiltration and adsorbent treatment for removal of protein impurities was used to provide a concentrate for chromatographic purification. Effective purification of the CBH II protein was obtained by passing the concentrate through a column of DEAE Sepharose, on which almost all the yeast proteins were adsorbed. The purified enzyme reacted with antibodies prepared against T. reesei CBH II and catalyzed partial solubilization of crystalline cellulose to soluble sugars.
Zurbriggen
B
B
VTT, Biotechnical Laboratory, Espoo, Finland.
Bailey
M J
MJ
Penttilä
M E
ME
Poutanen
K
K
Linko
M
M
D001709:Biotechnology; D043366:Cellulose 1,4-beta-Cellobiosidase; D006026:Glycoside Hydrolases / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D003904:Mitosporic Fungi / Q000201:enzymology*; D010865:Pilot Projects; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D014242:Trichoderma / Q000201:enzymology*
D006026:Glycoside Hydrolases; D043366:Cellulose 1,4-beta-Cellobiosidase