Production and secretion patterns of cloned glucoamylase in plasmid-harboring and chromosome-integrated recombinant yeasts employing an SUC2 promoter

To understand the differences in production and secretion patterns between plasmid-harboring and chromosome-integrated recombinant yeasts, the two rec ombinant Saccharomyces cerevisiae yeasts, containing the structural glucoam ylase STA gene and the SUC2 promoter, were investigated. Both systems were regulated by glucose concentration in the culture broth. First, the glucoam ylase activity per gene copy number of the chromosome-integrated recombinan t yeast was 2.8- to 5.6-fold higher than that of the plasmid-harboring reco mbinant yeast. Overburdened owing to high copy number, the plasmid-harborin g recombinant yeast gave lower glucoamylase activity per gene copy number. Second, the efficiency of signal sequence was compared; the secretion effic iency of glucoamylase in the plasmid-harboring recombinant yeast was higher than that in the chromosome-integrated recombinant yeast at 96 h of cultiv ation (74 vs 65%). We postulated that the higher level of secretion efficie ncy of the plasmid-harboring recombinant yeast resulted because the product ion level did not reach the capacity of the secretory apparatus of the host yeast. However, the specific secretion rate was much higher in the chromos ome-integrated recombinant yeast even though the final secretion efficiency was lower. The lower secretion rate in the plasmid-harboring recombinant y east could be explained by an adverse effect caused by higher production ra te. Finally, the optimal glucose concentration for glucoamylase production in the chromosome-integrated recombinant yeast culture was lower than that in the plasmid-harboring recombinant yeast culture owing to gene dosage eff ect.