[The decrease of the activity of electron transfer chain of Torulopsis glabrata enhanced pyruvate productivity].

Abstract

Torulopsis glabrata CCTCC M202019 was mutated by ethidium bromide to screen for respiratory-deficient mutants. Seven mutants that produced pyruvate higher than that of the parent were subjected to the tests of the capability assimilating fermentable substrate (glucose) and non-fermentable substrates (glycerol and acetate) to characterize true respiratory-deficient mutants. Mutants RD-16, RD-17 and RD-18 were unable to assimilate acetate or glycerol and were therefore identified as respiratory-deficient mutants. Compared to the parent strain, the growth the intracellular ATP content of those mutants decreased by 21% - 29% and 15% - 21%, respectively, while the glucose consumption per cell and the pyruvate production per cell of those mutants were enhanced by 20.7% - 30.7% and 30.7% - 55.5%, respectively. Qualitative analysis of cytochromes involved in electron transfer chain showed that mutants RD-16 and RD-18 lacked both cytochrome aa3 and b, while mutant RD-17 lacked cytochrome b. Enzymes analysis indicated that the activities of ATPase, succinate-cytochrome c reductase (complex I ), complex I + III , complex II + III, and complex IV of those mutants decreased by 14.6% - 22.2%, 34% - 41%, 38.6% - 52.6%, 21% - 25%, and 150% - 630%, respectively. However, increased glucose consumption per cell was not observed in those mutants, which might be due to that the NADH generated in glycolysis can not be completely oxidized via electron transfer chain. To avoid the accumulation of NADH, 2.1 mmol/L acetaldehyde was added to the culture broth of mutant RD-17 at 26h of fermentation. Using this strategy, the amount of pyruvate produced increased by 21.6% while the fermentation time was shortened from 62h to 48h.