Optimization of pro-urokinase secretion from recombinant Saccharomyces cerevisiae.
Secretion of a nonglycosylated form of human pro-urokinase, also known as single-chain urinary plasminogen activator (scu-PA), from Saccharomyces cerevisiae is described. A "supersecreting" yeast strain harboring multiple copies of integrated plasmids was grown batchwise and at constant respiratory quotient (RQ) in 20-L fermenters. Because the promoters used to drive expression of the pro-urokinase genes are not tightly regulated, secretion into the culture supernatant was growth associated. Although the final cell density achieved in the perturbed-batch fermentation (45 g dry wt/L) was less than that observed in the RQ-controlled culture (77 g dry wt/L), the scu-PA titer in the perturbed-batch fermentation (1863 IU/mL) was nearly twice that attained at constant RQ (1108 IU/mL). The effects on cell growth and scu-PA titer of other process variables (pH, temperature, phosphate concentration, and medium composition) are also discussed.
Turner
B G
BG
Collaborative Research, Inc., Two Oak Park, Bedford, Massachusetts 01730, USA.
Avgerinos
G C
GC
Melnick
L M
LM
Moir
D T
DT