Protein network centralities as descriptor for QM region construction in QM/MM simulations of enzymes.

The construction of a suitable QM region is the most crucial step in setting up hybrid quantum mechanics/molecular mechanics (QM/MM) simulations for enzymatic reactions. The QM region should ideally include all important amino acids residues, while being as small as possible to save computational effort. Most available methods for systematic QM region construction are based either on the distance of single amino acids to the active site or on their electrostatic effect. Such approaches might miss non-electrostatic and long-range allosteric interactions. Here, we present a proof of concept study for the application of protein network analysis to tackle this problem. Specifically, we explore the use of the protein network centralities as descriptor for QM region construction. We find that protein network centralities, in particular the betweenness centrality, can be a useful descriptor for systematic QM region construction. We show that in combination with our previously developed point charge variation analysis, they can be used to identify important residues that are missed in purely electrostatic approaches.

Efficient automatic construction of atom-economical QM regions with point-charge variation analysis.

The setup of QM/MM calculations is not trivial since many decisions have to be made by the simulation scientist to achieve reasonable and consistent results. The main challenge to be tackled is the construction of the QM region to make sure to take into account all important parts of the adjacent environment and exclude less important ones. In our previous work [F. Brandt and Ch. R. Jacob, Systematic QM Region Construction in QM/MM Calculations Based on Uncertainty Quantification, J. Chem. Theory Comput., 2022, 18, 2584-2596.], we introduced the point charge variation analysis (PCVA) as a simple and reliable tool to systematically construct QM regions based on the sensitivity of the reaction energy with respect to variations of the MM point charges. Here, we assess several simplified variants of this PCVA approach for the example of catechol O-methyltransferase and apply PCVA for another system, the triosephosphate isomerase. Furthermore, we extend its scope by applying it to a DNA system. Our results indicate that PCVA offers an efficient and versatile approach of the automatic construction of atom-economical QM regions, but also identify possible pitfalls and limitations.

Biased Borate Esterification during Nucleoside Phosphorylase-Catalyzed Reactions: Apparent Equilibrium Shifts and Kinetic Implications.

Biocatalytic nucleoside (trans-)glycosylations catalyzed by nucleoside phosphorylases have evolved into a practical and convenient approach to the preparation of modified nucleosides, which are important pharmaceuticals for the treatment of various cancers and viral infections. However, the obtained yields in these reactions are generally determined exclusively by the innate thermodynamic properties of the nucleosides involved, hampering the biocatalytic access to many sought-after target nucleosides. We herein report an additional means for reaction engineering of these systems. We show how apparent equilibrium shifts in phosphorolysis and glycosylation reactions can be effected through entropically driven, biased esterification of nucleosides and ribosyl phosphates with inorganic borate. Our multifaceted analysis further describes the kinetic implications of this in situ reactant esterification for a model phosphorylase.

Electrocatalytic Activation of Donor-Acceptor Cyclopropanes and Cyclobutanes: An Alternative C(sp3 )-C(sp3 ) Cleavage Mode.

We describe the first electrochemical activation of D-A cyclopropanes and D-A cyclobutanes leading after C(sp3 )-C(sp3 ) cleavage to the formation of highly reactive radical cations. This concept is utilized to formally insert molecular oxygen after direct or DDQ-assisted anodic oxidation of the strained carbocycles, delivering ß- and γ-hydroxy ketones and 1,2-dioxanes electrocatalytically. Furthermore, insights into the mechanism of the oxidative process, obtained experimentally and by additional quantum-chemical calculations are presented. The synthetic potential of the reaction products is demonstrated by diverse derivatizations.

EARLY STARVATION1 specifically affects the phosphorylation action of starch-related dikinases.

Starch phosphorylation by starch-related dikinases glucan, water dikinase (GWD) and phosphoglucan, water dikinase (PWD) is a key step in starch degradation. Little information is known about the precise structure of the glucan substrate utilized by the dikinases and about the mechanisms by which these structures may be influenced. A 50-kDa starch-binding protein named EARLY STARVATION1 (ESV1) was analyzed regarding its impact on starch phosphorylation. In various in vitro assays, the influences of the recombinant protein ESV1 on the actions of GWD and PWD on the surfaces of native starch granules were analyzed. In addition, we included starches from various sources as well as truncated forms of GWD. ESV1 preferentially binds to highly ordered, α-glucans, such as starch and crystalline maltodextrins. Furthermore, ESV1 specifically influences the action of GWD and PWD at the starch granule surface. Starch phosphorylation by GWD is decreased in the presence of ESV1, whereas the action of PWD increases in the presence of ESV1. The unique alterations observed in starch phosphorylation by the two dikinases are discussed in regard to altered glucan structures at the starch granule surface.

Uptake of Ra during the recrystallization of barite: a microscopic and time of flight-secondary ion mass spectrometry study.

A combined macroscopic and microanalytical approach was applied on two distinct barite samples from Ra uptake batch experiments using time of flight-secondary ion mass spectrometry (ToF-SIMS) and detailed scanning electron microscopy (SEM) investigations. The experiments were set up at near to equilibrium conditions to distinguish between two possible scenarios for the uptake of Ra by already existent barite: (1) formation of a Ba1-xRaxSO4 solid solution surface layer on the barite or (2) a complete recrystallization, leading to homogeneous Ba1-xRaxSO4 crystals. It could be clearly shown that Ra uptake in all barite particles analyzed within this study is not limited to the surface but extends to the entire solid. For most grains a homogeneous distribution of Ra could be determined, indicating a complete recrystallization of barite into a Ba1-xRaxSO4 solid solution. The maxima of the Ra/Ba intensity ratio distribution histograms calculated from ToF-SIMS are identical with the expected Ra/Ba ratios calculated from mass balance assuming a complete recrystallization. In addition, the role of Ra during the recrystallization of barite was examined via detailed SEM investigations. Depending on the type of barite used, an additional coarsening effect or a strong formation of oriented aggregates was observed compared to blank samples without Ra. In conclusion, the addition of Ra to a barite at close to equilibrium conditions has a major impact on the system leading to a fast re-equilibration of the solid to a Ba1-xRaxSO4 solid solution and visible effects on the particle size distribution, even at room temperature.

Nucleoside Phosphorylases make N7-xanthosine.

Modern, highly evolved nucleoside-processing enzymes are known to exhibit perfect regioselectivity over the glycosylation of purine nucleobases at N9. We herein report an exception to this paradigm. Wild-type nucleoside phosphorylases also furnish N7-xanthosine, a "non-native" ribosylation regioisomer of xanthosine. This unusual nucleoside possesses several atypical physicochemical properties such as redshifted absorption spectra, a high equilibrium constant of phosphorolysis and low acidity. Ultimately, the biosynthesis of this previously unknown natural product illustrates how even highly evolved, essential enzymes from primary metabolism are imperfect catalysts.

Systematic QM Region Construction in QM/MM Calculations Based on Uncertainty Quantification.

While QM/MM studies of enzymatic reactions are widely used in computational chemistry, the results of such studies are subject to numerous sources of uncertainty, and the effect of different choices by the simulation scientist that are required when setting up QM/MM calculations is often unclear. In particular, the selection of the QM region is crucial for obtaining accurate and reliable results. Simply including amino acids by their distance to the active site is mostly not sufficient as necessary residues are missing or unimportant residues are included without evidence. Here, we take a first step toward quantifying uncertainties in QM/MM calculations by assessing the sensitivity of QM/MM reaction energies with respect to variations of the MM point charges. We show that such a point charge variation analysis (PCVA) can be employed to judge the accuracy of QM/MM reaction energies obtained with a selected QM region and devise a protocol to systematically construct QM regions that minimize this uncertainty. We apply such a PCVA to the example of catechol O-methyltransferase and demonstrate that it provides a simple and reliable approach for the construction of the QM region. Our PCVA-based scheme is computationally efficient and requires only calculations for a system with a minimal QM region. Our work highlights the promise of applying methods of uncertainty quantification in computational chemistry.

Caesium and iodine release from spent mixed oxide fuels under repository relevant conditions: Initial leaching results.

Autoclave leaching experiments are conducted on three well-characterised, irradiated, and cladded mixed oxide fuel-rod segments with burnups ranging from 29 GWd/tHM to 52 GWd/tHM to investigate the instant release fraction of fission gases and long-lived fission products and to assess the long-term fuel matrix corrosion. The segments are exposed to bicarbonate solutions as reference groundwater at neutral pH and a synthetic young cementitious water at pH 13.5 under reducing atmosphere (4 vol% H2 in Ar at 40 bar pressure), since 2018. The initial leaching results for the fission products caesium and iodine as representative elements of the instant release fraction were found to depend on the leachate composition as well as on the fuel burnup.

Chromium Doped UO2-Based Ceramics: Synthesis and Characterization of Model Materials for Modern Nuclear Fuels.

Cr-doped UO2 as a modern nuclear fuel type has been demonstrated to increase the in-reactor fuel performance compared to conventional nuclear fuels. Little is known about the long-term stability of spent Cr-doped UO2 nuclear fuels in a deep geological disposal facility. The investigation of suitable model materials in a step wise bottom-up approach can provide insights into the corrosion behavior of spent Cr-doped nuclear fuels. Here, we present new wet chemical approaches providing the basis for such model systems, namely co-precipitation and wet coating. Both were successfully tested and optimized, based on detailed analyses of all synthesis steps and parameters: Cr-doping method, thermal treatment, reduction of U3O8 to UO2, green body production, and pellet sintering. Both methods enable the production of suitable model systems with a similar microstructure and density as a reference sample from AREVA. In comparison with results from the classical powder route, similar trends upon grain size and lattice parameter were determined. The results of this investigation highlight the significance of subtly different synthesis routes on the properties of Cr-doped UO2 ceramics. They enable a reproducible tailor-made well-defined microstructure, a homogeneous doping, for example, with lanthanides or alpha sources, the introduction of metallic particles, and a dust-free preparation.

Dissolution Kinetics of International Simple Glass and Formation of Secondary Phases at Very High Surface Area to Solution Ratio in Young Cement Water.

Static dissolution experiments were carried out with the reference International Simple Glass under hyperalkaline pH at 70 °C and very high SA/V ratio. Three aspects of glass dissolution behavior were investigated, (1) the rate drop regime and the residual rate (stage II), (2) the formation of secondary phases including thermodynamic aspects, and (3) the microstructure of the interface of altered glass and secondary phases. A very low residual rate of 6 × 10-6 g/m2d was determined based on boron release, which was several orders of magnitude lower than the initial rate established between the start of the experiments and the first sampling on day 59. The presence of a porous layer with a thickness varying between 80 nm and 250 nm and a pore size between 10 nm and 50 nm was observed. CSH phases with a low Ca/Si ratio of 0.3-0.4 and zeolites were also visible at the surface of the altered glass grains, but no glass alteration resumption occurred, probably due to an important pH decrease already at day 59. Thermodynamic calculations assuming congruent glass dissolution and precipitation of the dissolved aqueous species confirmed the precipitation of CSH phases and zeolites.

Insight Into Disorder, Stress and Strain of Radiation Damaged Pyrochlores: A Possible Mechanism for the Appearance of Defect Fluorite.

We have examined the irradiation response of a titanate and zirconate pyrochlore-both of which are well studied in the literature individually-in an attempt to define the appearance of defect fluorite in zirconate pyrochlores. To our knowledge this study is unique in that it attempts to discover the mechanism of formation by a comparison of the different systems exposed to the same conditions and then examined via a range of techniques that cover a wide length scale. The conditions of approximately 1 displacement per atom via He2+ ions were used to simulate long term waste storage conditions as outlined by previous results from Ewing in a large enough sample volume to allow for neutron diffraction, as not attempted previously. The titanate sample, used as a baseline comparison since it readily becomes amorphous under these conditions behaved as expected. In contrast, the zirconate sample accumulates tensile stress in the absence of detectable strain. We propose this is analogous to the lanthanide zirconate pyrochlores examined by Simeone et al. where they reported the appearance of defect fluorite diffraction patterns due to a reduction in grain size. Radiation damage and stress results in the grains breaking into even smaller crystallites, thus creating even smaller coherent diffraction domains. An (ErNd)2(ZrTi)2O7 pyrochlore was synthesized to examine which mechanism might dominate, amorphization or stress/strain build up. Although strain was detected in the pristine sample via Synchrotron X-ray diffraction it was not of sufficient quality to perform a full analysis on.

On the change in UO2 redox reactivity as a function of H2O2 exposure.

To assess the long-term leaching behaviour of UO2, the main constituent of spent nuclear fuel, the oxidative dissolution of UO2 pellets was studied at high H2O2 exposures ranging from 0.33 mol m-2 to 1.36 mol m-2. The experiments were performed in aqueous media containing 10 mM HCO3- where the pellets were exposed to H2O2 three consecutive times. The results indicate that the dissolution yield (amount of dissolved uranium per consumed H2O2) at high H2O2 exposures is significantly lower compared to previous studies of both pellets and powders and decreases for each H2O2 addition for a given pellet. This implies a change in redox reactivity, which is attributed to irreversible alteration of the pellet surface. Surface characterization after the exposure to H2O2, by SEM, XRD and Raman spectroscopy shows, that the surface of all pellets is significantly oxidized.

Position 123 of halohydrin dehalogenase HheG plays an important role in stability, activity, and enantioselectivity.

HheG from Ilumatobacter coccineus is a halohydrin dehalogenase with synthetically useful activity in the ring opening of cyclic epoxides with various small anionic nucleophiles. This enzyme provides access to chiral ß-substituted alcohols that serve as building blocks in the pharmaceutical industry. Wild-type HheG suffers from low thermostability, which poses a significant drawback for potential applications. In an attempt to thermostabilize HheG by protein engineering, several single mutants at position 123 were identified which displayed up to 14 °C increased apparent melting temperatures and up to three-fold higher activity. Aromatic amino acids at position 123 resulted even in a slightly higher enantioselectivity. Crystal structures of variants T123W and T123G revealed a flexible loop opposite to amino acid 123. In variant T123G, this loop adopted two different positions resulting in an open or partially closed active site. Classical molecular dynamics simulations confirmed a high mobility of this loop. Moreover, in variant T123G this loop adopted a position much closer to residue 123 resulting in denser packing and increased buried surface area. Our results indicate an important role for position 123 in HheG and give first structural and mechanistic insight into the thermostabilizing effect of mutations T123W and T123G.

Direct Measurement of Surface Dissolution Rates in Potential Nuclear Waste Forms: The Example of Pyrochlore.

The long-term stability of ceramic materials that are considered as potential nuclear waste forms is governed by heterogeneous surface reactivity. Thus, instead of a mean rate, the identification of one or more dominant contributors to the overall dissolution rate is the key to predict the stability of waste forms quantitatively. Direct surface measurements by vertical scanning interferometry (VSI) and their analysis via material flux maps and resulting dissolution rate spectra provide data about dominant rate contributors and their variability over time. Using pyrochlore (Nd2Zr2O7) pellet dissolution under acidic conditions as an example, we demonstrate the identification and quantification of dissolution rate contributors, based on VSI data and rate spectrum analysis. Heterogeneous surface alteration of pyrochlore varies by a factor of about 5 and additional material loss by chemo-mechanical grain pull-out within the uppermost grain layer. We identified four different rate contributors that are responsible for the observed dissolution rate range of single grains. Our new concept offers the opportunity to increase our mechanistic understanding and to predict quantitatively the alteration of ceramic waste forms.