Increasing glycolytic flux in Torulopsis glabrata by redirecting ATP production from oxidative phosphorylation to substrate-level phosphorylation

    loading  Checking for direct PDF access through Ovid

Abstract

Aims

This study aimed at further increasing the pyruvate productivity of a multi-vitamin auxotrophic yeast Torulopsis glabrata by redirecting ATP production from oxidative phosphorylation to substrate-level phosphorylation.

Methods and Results

We examined two strategies to decrease the activity of F0F1-ATPase. The strategies were to inhibit F0F1-ATPase activity by addition of oligomycin, or to disrupt F0F1-ATPase by screening neomycin-resistant mutant. The addition of 0·05 mmol l−1 oligomycin to the culture broth of T. glabrata CCTCC M202019 resulted in a significantly decreased intracellular ATP level (35·7%) and a significantly increased glucose consumption rate (49·7%). A neomycin-resistant mutant N07 was screened and selected after nitrosoguanidine mutagenesis of the parent strain T. glabrata CCTCC M202019. Compared with the parent strain, the F0F1-ATPase activity of the mutant N07 decreased about 65%. As a consequence, intracellular ATP level of the mutant N07 decreased by 24%, which resulted in a decreased growth rate and growth yield. As expected, glucose consumption rate and pyruvate productivity of the mutant N07 increased by 34% and 42·9%, respectively. Consistently, the activities of key glycolytic enzymes of the mutant N07, including phosphofructokinase, pyruvate kinase and glyceraldyde-3-phosphate dehydrogenase, increased by 63·7%, 28·8% and 14·4%, respectively. In addition, activities of the key enzymes involved in electron transfer chain of the mutant N07 also increased.

Conclusions

Impaired oxidative phosphorylation in T. glabrata leads to a decreased intracellular ATP production, thereby increasing the glycolytic flux.

Significance and Impact of the Study

The strategy of redirecting ATP production from oxidative phosphorylation to substrate-level phosphorylation provides an alternative approach to enhance the glycolytic flux in eukaryotic micro-organisms.

Related Topics

    loading  Loading Related Articles