Abstract
Increasing the growth rate of the industrial host Corynebacterium glutamicum is a promising target to rise productivities of growth coupled product formation. As a prerequisite, detailed knowledge about the tight regulation network is necessary for identifying promising metabolic engineering goals. Here, we present comprehensive metabolic and transcriptional analysis of C. glutamicum ATCC 13032 growing under glucose limited chemostat conditions with μ = 0.2, 0.3, and 0.4 h–1. Intermediates of central metabolism mostly showed rising pool sizes with increasing growth. 13C-metabolic flux analysis (13C-MFA) underlined the fundamental role of central metabolism for the supply of precursors, redox, and energy equivalents. Global, growth-associated, concerted transcriptional patterns were not detected giving rise to the conclusion that glycolysis, pentose-phosphate pathway, and citric acid cycle are predominately metabolically controlled under glucose-limiting chemostat conditions. However, evidence is found that transcriptional regulation takes control over glycolysis once glucose-rich growth conditions are installed.
Highlights
Corynebacterium glutamicum, discovered in 1957 as a soil bacterium, has gained prominent acceptance in industry for the production of amino acids, organic acids, and even proteins (Hermann, 2003; Wendisch et al, 2006; Freudl, 2017)
After about 7.5 h batch phase [μmax = (0.44 ± 0.01) h−1] the continuous mode was started with a dilution rate D = 0.2 h−1, which was increased to 0.3 h−1 after ca. 28 h and to D = 0.4 h−1 after ca. 49 h
Three independent samples for OD, cell dry weight (CDW), filtrate TC/total inorganic carbon (TIC), metabolome, and transcriptome were withdrawn after five residence time (RT) within 1.5 hintervals to monitor the respective metabolic steady-states at D = 0.2, 0.3, and 0.4 h−1
Summary
Corynebacterium glutamicum, discovered in 1957 as a soil bacterium, has gained prominent acceptance in industry for the production of amino acids, organic acids, and even proteins (Hermann, 2003; Wendisch et al, 2006; Freudl, 2017). Silberbach et al (2005) were among the first to perform systems biology analysis on C. glutamicum ATCC 13032 by analyzing the cell’s transcriptional and proteomic response to ammonium limitation at two different growth rates They identified growth rate dependent expression of ribosomal proteins and of F0F1 subunits of ATP-synthase. Under the tested glucose-limited growth conditions, Noack et al (2017) concluded that metabolic fluxes within C. glutamicum’s central carbon metabolism are likely more affected by concentrations of local substrates and allosteric metabolite effectors than by the concentration of enzymes Another systems biology study was performed in traditional non-substrate limited batch cultivations to investigate determinants of global regulation orchestrating the growth of C. glutamicum (Haas et al, 2019). In other words we wonder: How similar are regulatory patterns of C. glutamicum growing with equal rates but under different nutrient availability?
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