Enhancement of pyruvate productivity in Torulopsis glabrata: Increase of NAD + availability

• Published in: Journal of biotechnology 2006-11, Vol.126 (2), p.173-185
• Main Authors: Liu, Liming, Li, Yin, Shi, Zhongping, Du, Guocheng, Chen, Jian
• Format: Article
• Language: English
• Subjects: alcohol dehydrogenase; Biological and medical sciences; Biological Availability; biosynthesis; Biotechnology; Candida glabrata - genetics; Candida glabrata - metabolism; Cell Culture Techniques - methods; Fundamental and applied biological sciences. Psychology; Genetic Enhancement - methods; metabolism; metabolites; mutants; NAD (coenzyme); NAD + available; NAD - metabolism; niacin; nutrient availability; Productivity; Pyruvate; pyruvic acid; Pyruvic Acid - metabolism; Torulopsis glabrata
• ISSN: 0168-1656; 1873-4863
• DOI: 10.1016/j.jbiotec.2006.04.014

This study aimed at increasing the pyruvate productivity from a multi-vitamin auxotrophic yeast Torulopsis glabrata, by increasing the availability of NAD +. We examined two strategies for increasing availability of NAD +. To supplement nicotinic acid (NA), the precursor of NAD +; and to increase the activity of alcohol dehydrogenase integrating with addition acetaldehyde as exterior electron acceptor. The addition of 8 mg l −1 NA to the fermentation medium resulted in a significant increase in the glucose consumption rate (48.4%) and the pyruvate concentration (29%). An ethanol-utilizing mutant WSH-13 was screened and selected after nitrosoguanidine mutagenesis of the parent strain T. glabrata CCTCC M202019. Compared with the parent strain, the alcohol dehydrogenase activity of the mutant WSH-13 increased about 110% and the mutant could utilize ethanol as the sole carbon source for growth (1.8 g l −1 dry cell weight). When growing with glucose, the addition of 4 mg l −1 acetaldehyde to the mutant WSH-13 culture broth led to a significant increase in the glucose consumption rate (26.3%) and pyruvate production (22.5%), but the ratio of NADH/NAD + decreased to 0.22. Acetaldehyde did not affect the glucose and energy metabolism at high dissolved oxygen (DO) concentration. However, at lower DO concentration (20%), maintaining the acetaldehyde concentration in the mutant culture broth at 4 mg l −1 caused an increased NAD + concentration but a decreased NADH concentration. As a consequence, the pyruvate production rate, the pyruvate yield on glucose and the pyruvate concentration were 68, 44 and 45% higher, respectively, than the corresponding values of the control (without acetaldehyde). The strategy for increasing the glycolytic flux and the pyruvate productivity in T. glabrata by increasing the availability of NAD + may provide an alternative approach to enhance the metabolites productivity in yeast.