Loss of glucokinase function relieves carbon catabolite repression in basidiomycetous yeast Pseudozyma antarctica.
TL;DR
Results indicate that disruption of the glucokinase gene considerably reduces the cost of the PaE-producing medium and facilitates the efficient production of useful proteins in basidiomycetous yeasts.
Abstract
The basidiomycetous yeast Pseudozyma antarctica produces the biodegradable plastic-degrading enzyme PaE. The PaE overexpression strain, constructed by replacing the promoter of the PaE gene with the xylanase promoter, exhibited high-level production of PaE in the presence of xylose. However, this production is suppressed by carbon catabolite repression (CCR) in the presence of glucose. Although CCR is a major issue involving the decreased expression of hydrolytic genes from their promoters in ascomycetes, little information is available in basidiomycetous yeasts. In the present study, we demonstrated that glucokinase is involved in the regulation of CCR in P. antarctica. Ultraviolet irradiation of P. antarctica resulted in mutant strains that exhibited resistance to the toxic glucose analog 2-deoxyglucose and reduced the CCR of the xylanase and PaE driven by their own promoters. These strains had missense or frameshift mutation within the glucokinase gene (PaGLK1). The restoration of CCR through intact PaGLK1 complementation in these strains indicated that loss-of-function mutations in PaGLK1 contribute to CCR defects. Deletion of PaGLK1 in the PaE overexpression strain showed only a limited reduction in PaE production, even when 3% of the 4% xylose in the liquid medium was replaced with glucose. These results indicate that disruption of the glucokinase gene considerably reduces the cost of the PaE-producing medium and facilitates the efficient production of useful proteins in basidiomycetous yeasts.