NADH-Regulated Metabolic Model for Growth of Methylosinus trichosporium OB3b. Cometabolic Degradation of Trichloroethene and Optimization of Bioreactor System Performance

Abstract

A metabolic model describing growth of Methylosinus trichosporium OB3b and cometabolic contaminant conversion is used to optimize trichloroethene (TCE) conversion in a bioreactor system. Different process configurations are compared: a growing culture and a nongrowing culture to which TCE is added at both constant and pulsed levels. The growth part of the model, presented in the preceding article, gives a detailed description of the NADH regeneration required for continued TCE conversion. It is based on the metabolic pathways, includes Michaelis-Menten type enzyme kinetics, and uses NADH as an integrating and controlling factor. Here the model is extended to include TCE transformation, incorporating the kinetics of contaminant conversion, the related NADH consumption, toxic effects, and competitive inhibition between TCE and methane. The model realistically describes the experimentally observed negative effects of the TCE conversion products, both on soluble methane monooxygenase through the explicit incorporation of the activity of this enzyme and on cell viability through the distinction between dividing and nondividing cells. In growth-based systems, the toxicity of the TCE conversion products causes rapid cell death, which leads to wash-out of suspended cultures at low TCE loads (below microM inlet concentrations). Enzyme activity, which is less sensitive, is hardly affected by the toxicity of the TCE conversion products and ensures high conversions (>95%) up to the point of wash-out. Pulsed addition of TCE (0.014-0.048 mM) leads to a complete loss of viability. However, the remaining enzyme activity can still almost completely convert the subsequently added large TCE pulses (0.33-0.64 mM). This emphasizes the inefficient use of enzyme activity in growth-based systems. A comparison of growth-based and similar non-growth-based systems reveals that the highest TCE conversions per amount of cells grown can be obtained in the latter. Using small amounts of methane (negligible compared to the amount needed to grow the cells), NADH limitation in the second step of this two-step system can be eliminated. This results in complete utilization of enzyme activity and thus in a very effective treatment system.

Conventional methanotrophs are responsible for atmospheric methane oxidation in paddy soils

“…2a ). PHB represents an endogenous source of reducing power to methanotrophs 23 when cells are subjected to environmental stress, such as starvation, nutrient deprivation or a lack of reducing equivalents (for example, NADH, reduced nicotinamide adenine dinucleotide) 24 . PHB accumulation often occurs under conditions of carbon excess in combination with nitrogen limitation 23 25 .…”

Modeling Electron Competition among Nitrogen Oxides Reduction and N2O Accumulation in Denitrification

“…C tot was assumed to be 0.01 mmol/(gVSS) based on literature-reported values for intracellular NAD + NADH concentrations. 35,41 K Mox was assumed to be 1% of C tot to ensure that the reduction of Mox would not be the rate limiting step during carbon oxidation.…”

“…Mred (reduced mediator) and Mox (oxidized mediator), defined as the reduced and oxidized forms of electron carriers, respectively, are included in the model as two new state variables. Because the electron carrier pool (with the NADH/NAD + pool as an example) is relatively small compared with its turnover rate, the continued availability of Mox for carbon oxidation and Mred for nitrogen oxides reduction relies on their concomitant regeneration. , In modeling, this recirculation loop could be reflected by an increase in Mred being balanced by a decrease in the Mox and vice versa ( Mred ⇋ Mox + 2 e – + 2H + ), with the total level of Mred and Mrox being held constant, i.e., S Mred + S Mox = C tot , , where C tot is the total concentration of electron carriers.…”

Genome-scale metabolic reconstructions and theoretical investigation of methane conversion in Methylomicrobium buryatense strain 5G(B1)

“…Metabolic modeling became a useful tool for in silico experiments with whole-cell metabolic phenotyping and engineering [ 12 , 13 ]. While the genomics and biochemistry of microbial C 1 -metabolism are relatively well established, only a few mathematical descriptions of methane or methanol utilization have been developed [ 14 – 16 ]. It has been assumed that growth of C 1 -compounds is reducing power limited [ 16 , 17 ].…”