Expression of the human blood coagulation protein factor XIIIa in Saccharomyces cerevisiae: dependence of the expression levels from host-vector systems and medium conditions.
The human blood coagulation protein Factor XIIIa (FXIIIa) was expressed in Saccharomyces cerevisiae employing Escherichia coli-yeast shuttle vectors based on a 2-mu plasmid. Several factors affecting high production yield of recombinant FXIIIa were analysed. The use of the regulatable GAL-CYC1 hybrid promoter resulted in higher FXIIIa expression when compared with the constitutive ADCI promoter. Screening for suitable yeast strains for expression of FXIIIa under the transcriptional control of the GAL-CYC1 hybrid promoter revealed a broad spectrum of productivity. No obvious correlation between the expression rate and the genetic markers of the strains could be identified. The medium composition markedly influenced the FXIIIa expression rates. The expression of FXIIIa was strictly regulated by the carbon source. Glucose as the only sugar and energy source repressed the synthesis of FXIIIa, whereas addition of galactose induced FXIIIa expression. Special feeding schemes resulted in a productivity of up to 100 mg FXIIIa/l in shake flasks.
Research Laboratories of Behringwerke AG, Marburg, Federal Republic of Germany.
M
M
Bröker
O
O
Bäuml
A
A
Göttig
J
J
Ochs
M
M
Bodenbenner
E
E
Amann
D015825:Chromosomes, Fungal / Q000502:physiology; D003001:Cloning, Molecular; D003470:Culture Media; D004271:DNA, Fungal / Q000737:chemistry; D005690:Galactose / Q000235:genetics; D005696:Galactosidases / Q000096:biosynthesis / Q000235:genetics; D015870:Gene Expression; D008894:Milk Proteins / Q000235:genetics; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D011503:Transglutaminases / Q000096:biosynthesis / Q000235:genetics*; D000067816:Whey Proteins
D003470:Culture Media; D004271:DNA, Fungal; D008894:Milk Proteins; D000067816:Whey Proteins; D011503:Transglutaminases; D005696:Galactosidases; D005690:Galactose