Monday, July 27, 2009 - 9:30 AM
S20

Characterization of the malic enzyme deletion in Saccharomyces cerevisiae BY4742 grown on glucose and galactose using 13C metabolic flux analysis and optical online dissolved oxygen measurement in shake flasks

Konstantin Schneider1, Elmar Heinzle1, Tae Hoon Yang2, and Gernot Thomas John3. (1) Biochemical Engineering Institute, Saarland University, Campus A 1.5, Saarbrücken, Germany, (2) Genomatica, Inc., San Diego, CA, (3) Presens Precision Sensing GmbH, Regensburg, Germany

Malic enzyme, catalyzing the oxidative decarboxylation of malate to pyruvate and CO2, was characterized by Boles et al. For the cells growing on acetate or ethanol malic enzyme is one source of  glycolytic metabolite generation from C4 metabolites of TCA. Malic enzyme is not essential for the  growth on fermentable carbon sources, yet its activity has been  observed during the growth on glucose. The aim of the present work was to investigate the influence of mae1 deletion on S. cerevisiae growing respectively on glucose and galactose as carbon sources, especially, on cell metabolism and oxygen utilization. 13C metabolic flux analysis was carried out by assuming a pseudo-steady state, confirmed by exponential cell growth and constant yields. An opto-electronic device was utilized to measure dissolved oxygen as well as pH in shake flask clutures. Optical measurements of dissolved oxygen in shake flasks gave distinctive patterns between the two different carbon sources for the both cases with wild-type and mutant. During the growth on galactose the deletion strain gave rise to a 50% reduced specific oxygen uptake rate compared to the wild-type. In contrast, the specific rates during growth on glucose were comparable. According to 13CMFA, a higher flux through the pentose phosphate pathway was observed for the growth of mae1 deletion mutation on galactose and a reduction in TCA cycle activity linked to the decreased oxygen consumption. Thus combination of 13CMFA and onlineDO measurement draws a complete picture of the central carbon metabolism in respect to the role of malic enzyme.

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