Kinetic, dynamic, and pathway studies of glycerol metabolism byKlebsiella pneumoniae in anaerobic continuous culture: III. Enzymes and fluxes of glycerol dissimilation and 1,3-propanediol formation

Abstract

The initial steps of glycerol dissimilation and 1,3‐propanediol (1,3‐PD) formation by Klebsiella pneumoniae anaerobically grown on glycerol were studied by quantifying the in vitro and in vivo activities of enzymes in continuous culture under conditions of steady state and oscillation and during transient phases. The enzymes studied included glycerol dehydrogenase (GDH), glycerol dehydratase (GDHt), and 1,3‐propanediol oxidoreductase (PDOR). Three conclusions can be drawn from the steady‐state results. First, glycerol concentration in the culture is a key parameter that inversely affects the in vitro activities (concentrations) of all three enzymes, but has a positive effect on their in vivo activities. Growth rate significantly affects the ratio of in vitro and in vivo enzyme activities under low glycerol concentrations, but not under glycerol excess. Second, whereas the flux through the oxidative pathway of glycerol dissimilation is governed mainly by the regulation of in vivo enzyme activity on a metabolic level, the flux through the reductive pathway is largely controlled by the synthesis of enzymes. Third, GDHt is a major rate‐liming enzyme for the consumption of glycerol and the formation of 1,3‐PD in K. pneumoniae at high glycerol concentrations. Results from oscillating cultures revealed that both in vitro and in vivo activities of the enzymes oscillated. The average values of the in vitro activities during an oscillation cycle agreed well with their corresponding values for nonoscillating cultures under similar environmental conditions. Experiments with step changes in the feed concentration of glycerol demonstrated that growth and product formation are very sensitive to changes of substrate concentration in the culture. This sensitivity is due to the dynamic responses of the genetic and metabolic networks. They should be considered when modeling the dynamics of the culture and attempting to improve the formation of 1,3‐PD. © 1998 John Wiley & Sons, Inc. Biotechnol Bioeng 59: 544–552, 1998.

1,3-Propanediol Production by Clostridium Butyricum in Various Fed- Batch Feeding Strategies

“…It is at present used in the chemical synthesis of polyesters and polyurethanes, agent of tobacco elaboration, the lubricant of type polyalcohol, solvent, and as food additive [2]. The synthesis of 1,3-Propanediol can be realized with chemical way from the hydro carbonation of the oxide of ethylene or of hydration of acrolein in hydroxypropanol followed by a catalytic reduction [3]. These expensive processes implement toxic products (acrolein) and generate intermediaries compound and secondary not useful whish make them little profitable.…”

Exotic glycerol dehydrogenase expressing Escherichia coli increases yield of 2,3-butanediol

“…However, the intracellular GDH activity was determined at room temperature by measuring the reduction of NAD + to the substrate-dependent absorbance change of NAD(H) at 340 nm (ε340 = 6.22 mM −1 cm −1 ) using the method described by Ahrens et al (1998) with slight modification (Rahman et al 2015). Briefly, 1 mL of the reaction mixture contains 50 mM potassium phosphate buffer (pH 8.0), 30 mM ammonium sulfate, 0.2 M glycerol and 1.2 mM NAD.…”

1,3-Propanediol Dehydrogenase fromKlebsiella pneumoniae: Decameric Quaternary Structure and Possible Subunit Cooperativity

“…In these organisms, glycerol is metabolized by two different parallel pathways. The first is an oxidative route, in which glycerol is converted by a NAD-dependent glycerol oxidoreductase to dihydroxyacetone, that is further converted to glyceraldehyde-3-phosphate, phosphoenolpyruvate, and pyruvate to be inserted in the Krebs cycle (1). The reduced cofactors generated in the conversion of glycerol to dihydroxyacetone can be recycled by using a second reductive pathway.…”