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J Biotechnol. 1996 Feb 28;45(2):149-64.

Growth energetics and metabolic fluxes in continuous cultures of Penicillium chrysogenum.

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Center for Process Biotechnology, Technical University of Denmark, Lyngby.


Continuous cultures of the penicillin producing fungus Penicillium chrysogenum have been analyzed with respect to the macromolecular composition of the mycelium. All cultivations were carried out using a chemically defined medium with glucose as the growth limiting component. Biomass was harvested at steady state and analyzed for proteins, lipids, RNA, DNA, and carbohydrates. Carbohydrates present in the cell wall, i.e., glucans and chitin, and carbohydrates serving as storage materials, i.e., glycogen, were measured. It was observed that the levels of DNA and lipids are relative constant, whereas the proteins and stable RNA levels increase with the specific growth rate and the total amount of carbohydrates decreases with the specific growth rate. Glycogen is only present in small amounts, decreasing with the specific growth rate. As an average the measured macromolecules account for 77 +/- 2% (w/w) of the biomass. On the basis of estimations of the metabolic costs for biosynthesis and polymerization of the different macromolecules the total ATP and NADPH requirements for cell biosynthesis from glucose and inorganic salts, i.e., YxATP,growth and YxNADPH, have been quantified. The biosynthesis of 1 g biomass was calculated to require 39.9 mmol of ATP and 7.5 mmol of NADPH when cytosolic acetyl-CoA is formed from citrate by citrate lyase and oxaloacetate is recycled back into the TCA cycle. Other pathways of acetyl-CoA biosynthesis have been considered. The calculations show that the different biosynthetic routes for generating cytosolic acetyl-CoA have a significant influence on the theoretical value of ATP and NADPH requirements for cell biosynthesis. Combining a detailed stoichiometric model for growth and product formation of P. chrysogenum with experimental data on the macromolecular composition of P. chrysogenum and precise measurements of substrate uptake and product formation the intracellular flux distribution was calculated for different cultivation conditions.

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