초록
Engineering cellulolytic ability into the yeast Saccharomyces cerevisiae to create an organism for consolidated bioprocessing (CBP) will require the simultaneous production and secretion of a number of heterologous cellulases. In addition, the generally low secretion titers achieved by this yeast will have to be overcome. To this end two native S. cerevisiae genes, PSE1 and SOD1, were individually overexpressed by placing each gene under the transcriptional control of the constitutive PGK1 promoter. The effect of these genes on heterologous protein secretion of three cellulases - an exoglucanase encoded by cel6A of Neocallimastix patriciarum, a β-glucosidase encoded by cel3A of Saccharomycopsis fibuligera and an endoglucanase encoded by cel7B of Trichoderma reesei was investigated by integrating the PGK1<SUB>P/T</SUB>-PSE1 and PGK1<SUB>P/T</SUB>-SOD1 cassettes into S. cerevisiae strains producing the relevant cellulases. Transformants were obtained that showed significantly higher secreted protein yield, with a resulting heterologous protein activity that ranged between 10% and 373% higher compared to the parental strains when grown in complex media. When both PSE1 and SOD1 were overexpressed in the yeast that produced Cel3A, a dramatic 447% increase in β-glucosidase activity was observed. This study shows that cellulase secretion in S. cerevisiae could be greatly improved with strain engineering. However, it also demonstrated that such strain engineering may have very enzyme specific effects as the induction of Cel3A secretion was far greater than that of the other cellulases investigated. Identifying cellulases amenable to expression in S. cerevisiae and engineering strains to maximize heterologous protein secretion may be imperative to creating optimal strains for CBP and may have wider implications for heterologous protein secretion in S. cerevisiae in general.