Processive kinetics in the three-step lanosterol 14α-demethylation reaction catalyzed by human cytochrome P450 51A1.

Cytochrome P450 (P450, CYP) family 51 enzymes catalyze the 14α-demethylation of sterols, leading to critical products used for membranes and the production of steroids, as well as signaling molecules. In mammals, P450 51 catalyzes the 3-step, 6-electron oxidation of lanosterol to form (4ß,5α)-4,4-dimethyl-cholestra-8,14,24-trien-3-ol (FF-MAS). P450 51A1 can also use 24,25-dihydrolanosterol (a natural substrate in the Kandutsch-Russell cholesterol pathway). 24,25-Dihydrolanosterol and the corresponding P450 51A1 reaction intermediates, the 14α-alcohol and -aldehyde derivatives of dihydrolanosterol, were synthesized to study the kinetic processivity of the overall 14α-demethylation reaction of human P450 51A1. A combination of steady-state kinetic parameters, steady-state binding constants, dissociation rates of P450-sterol complexes, and kinetic modeling of the time course of oxidation of a P450-dihydrolanosterol complex showed that the overall reaction is highly processive, with koff rates of P450 51A1-dihydrolanosterol and the 14α-alcohol and 14α-aldehyde complexes being 1 to 2 orders of magnitude less than the forward rates of competing oxidations. epi-Dihydrolanosterol (the 3α-hydroxy analog) was as efficient as the common 3ß-hydroxy isomer in the binding and formation of dihydro FF-MAS. The common lanosterol contaminant dihydroagnosterol was found to be a substrate of human P450 51A1, with roughly one-half the activity of dihydrolanosterol. Steady-state experiments with 14α-methyl deuterated dihydrolanosterol showed no kinetic isotope effect, indicating that C-14α C-H bond breaking is not rate-limiting in any of the individual steps. The high processivity of this reaction generates higher efficiency and also renders the reaction less sensitive to inhibitors.

Role of Electronic Structure on Nitrate Reduction to Ammonium: A Periodic Journey.

Electrocatalysis is a promising approach to convert waste nitrate to ammonia and help close the nitrogen cycle. This renewably powered ammonia production process sources hydrogen from water (as opposed to methane in the thermal Haber-Bosch process) but requires a delicate balance between a catalyst's activity for the hydrogen evolution reaction (HER) and the nitrate reduction reaction (NO3RR), influencing the Faradaic efficiency (FE) and selectivity to ammonia/ammonium over other nitrogen-containing products. We measure ammonium FEs ranging from 3.6 ± 6.6% (on Ag) to 93.7 ± 0.9% (on Co) across a range of transition metals (TMs; Ti, Fe, Co, Ni, Ni0.68Cu0.32, Cu, and Ag) in buffered neutral media. To better understand these competing reaction kinetics, we develop a microkinetic model that captures the voltage-dependent nitrate rate order and illustrates its origin as competitive adsorption between nitrate and hydrogen adatoms (H*). NO3RR FE can be described via competition for electrons with the HER, decreasing sharply for TMs with a high work function and a correspondingly high HER activity (e.g., Ni). Ammonium selectivity nominally increases as the TM d-band center energy (Ed) approaches and overcomes the Fermi level (EF), but is exceptionally high for Co compared to materials with similar Ed. Density functional theory (DFT) calculations indicate Co maximizes ammonium selectivity via (1) strong nitrite binding enabling subsequent reduction and (2) promotion of nitric oxide dissociation, leading to selective reduction of the nitrogen adatom (N*) to ammonium.

Iodine and Carbonate Species Monitoring in Molten NaOH-KOH Eutectic Scrubber via Dual-Phase In Situ Raman Spectroscopy.

Molten hydroxide scrubbing of off-gas vapors is a potential process to improve safety during the operation of generation IV molten salt nuclear reactors (MSRs). MSRs produce off-gases that can be vented by the reactor core and treated via off-gas scrubbers. Molten hydroxide scrubbers focus on capturing volatile iodine radionuclides, and they can also be used to capture aerosols and particulates and to neutralize acidic species. The performance of these scrubbers depends on the chemical interactions of the scrubbing medium with the off-gas species. Knowledge of the concentration and speciation of scrubbed or target species, as well as process and environmental interferents, can enable advanced operation of MSR off-gas treatment systems. Optical online monitoring is an excellent technology to provide this information in real time, while limiting the need for operators to interact with radioactive samples through hands-on interrogation. Raman spectroscopy can provide crucial chemical information on the state of the molten eutectic during treatment in the molten phase, as well as the gas phase. In this work, Raman spectroscopy is used to detect iodine species, specifically iodate, in the molten phase of a NaOH-KOH eutectic and to construct a calibration curve of the Raman signal of those species. Additionally, a carbonate interferent is followed from the gas phase to the liquid phase as a basis for reaching a Raman-aided mass balance of the molten hydroxide eutectic scrubber system.