PROTEIN-PRODUCTION USING RECOMBINANT YEAST IN AN IMMOBILIZED-CELL-FILM AIRLIFT BIOREACTOR

A recombinant Saccharomyces cerevisiae C468/pGAC9 (ATCC 20690), which expresses Aspergillus awamori glucoamylase gene under the control of t he yeast enolase I (ENO1) promoter and secretes glucoamylase into the extracellular medium, was used as a model system to investigate the ef fect of cell immobilization on bioreactor culture performance. Free su spension cultures in stirred-tank and airlift bioreactors confirmed in herent genetic instability of the recombinant yeast. An immobilized-ce ll-film airlift bioreactor was developed by employing cotton cloth she ets to immobilize the yeast cells by attachment. Enhanced enzyme produ ctivity and production stability in the immobilized-cell system were o bserved. Experimental data indicated that the immobilized cells mainta ined a higher proportion of plasmid-bearing cells for longer periods u nder continuous operation. The higher plasmid maintenance with immobil ized cells is possibly due to reduced specific growth rate and increas ed plasmid copy number. Double-selection pressure was used to select a nd maintain the recombinant yeast. The selected strain showed better p roduction performance than the original strain. (C) 1997 John Wiley & Sons, Inc.