Active Site Model Research Articles

Identification of a CTRP9 C-Terminal polypeptide capable of enhancing bone-derived mesenchymal stem cell cardioprotection through promoting angiogenic exosome production

BackgroundMesenchymal stem cell therapy improves ischemic heart failure via incompletely understood mechanisms. C1q-TNFα related protein-9 (CTRP9) is a novel anti-oxidative cardiokine capable of improving the local microenvironment and cell survival by its c-terminal active globular domain (gCTRP9). The current study attempted to: 1) identify active gCTRP9 c-terminal polypeptides with stem cell protective function; 2) determine whether a lead polypeptide may enable/enhance cortical bone-derived mesenchymal stem cell (CBSC) cardioprotection against post-myocardial infarction (post-MI) remodeling; and 3) define the responsible underlying cellular/molecular mechanisms. Methods and resultsUtilizing I-TASSER structure prediction and 3-D active site modeling, we cloned and purified 3 gCTRP9 fragments (CTRP9-237, CTRP9-277, and CTRP9-281). Their activation of cell salvage kinase was compared against gCTRP9. Among the three fragments, CTRP9-281 (a 45 residue-containing polypeptide) exerted comparable or greater ERK1/2 activation compared to gCTRP9. Treatment with CTRP9-281 or gCTRP9 significantly increased CBSC proliferation and migration, and attenuated oxidative stress-induced CBSC apoptosis. CTRP9-281 and gCTRP9 comparably upregulated SOD2 and SOD3 expression. However, CTRP9-281, not gCTRP9, upregulated FGF2 and VEGFA expression/secretion in an ERK1/2 dependent manner. Administration of gCTRP9 or CTRP9-281 alone attenuated post-MI cardiac dysfunction and improved CBSC retention in the infarcted heart in similar fashion. However, CTRP9-281 exerted greater synergistic effect with CBSC than gCTRP9 related to pro-angiogenic, anti-fibrotic, and anti-remodeling effects. Mechanistically, CTRP9-281 significantly increased SOD2-rich and VEGFA-rich exosome production by CBSC. Exosomes from CTRP9-281 treated CBSC significantly attenuated oxidative stress-induced cardiomyocyte apoptosis in vitro. An exosome generation inhibitor attenuated CTRP9-281 enhancement of CBSC cardioprotection in vivo. ConclusionWe identified a CTRP9 polypeptide that upregulates SOD2/SOD3 expression and improves CBSC survival/retention, similar to gCTRP9. Moreover, CTRP9-281 stimulates VEGFA-rich exosome production by CBSC, exerting superior pro-angiogenic, anti-fibrotic, and cardioprotective actions.

How Do Electrostatic Perturbations of the Protein Affect the Bifurcation Pathways of Substrate Hydroxylation versus Desaturation in the Nonheme Iron-Dependent Viomycin Biosynthesis Enzyme?

The viomycin biosynthesis enzyme VioC is a nonheme iron and α-ketoglutarate-dependent dioxygenase involved in the selective hydroxylation of l-arginine at the C3-position for antibiotics biosynthesis. Interestingly, experimental studies showed that using the substrate analogue, namely, l-homo-arginine, a mixture of products was obtained originating from C3-hydroxylation, C4-hydroxylation, and C3-C4-desaturation. To understand how the addition of one CH2 group to a substrate can lead to such a dramatic change in selectivity and activity, we decided to perform a computational study using quantum mechanical (QM) cluster models. We set up a large active-site cluster model of 245 atoms that includes the oxidant with its first- and second-coordination sphere influences as well as the substrate binding pocket. The model was validated against experimental work from the literature on related enzymes and previous computational studies. Thereafter, possible pathways leading to products and byproducts were investigated for a model containing l-Arg and one for l-homo-Arg as substrate. The calculated free energies of activation predict product distributions that match the experimental observation and give a low-energy C3-hydroxylation pathway for l-Arg, while for l-homo-Arg, several barriers are found to be close in energy leading to a mixture of products. We then analyzed the origins of the differences in product distributions using thermochemical, valence bond, and electrostatic models. Our studies show that the C3-H and C4-H bond strengths of l-Arg and l-homo-Arg are similar; however, external perturbations from an induced electric field of the protein affect the relative C-H bond strengths of l-Arg dramatically and make the C3-H bond the weakest and guide the reaction to a selective C3-hydroxylation channel. Therefore, the charge distribution in the protein and the induced electric dipole field of the active site of VioC guides the l-Arg substrate activation to C3-hydroxylation and disfavors the C4-hydroxylation pathway, while this does not occur for l-homo-Arg. Tight substrate positioning and electrostatic perturbations from the second-coordination sphere residues in VioC also result in a slower overall reaction for l-Arg; however, they enable a high substrate selectivity. Our studies highlight the importance of the second-coordination sphere in proteins that position the substrate and oxidant, perturb charge distributions, and enable substrate selectivity.

HNO to NO Conversion Mechanism with Copper Zinc Superoxide Dismutase, Comparison with Heme Protein Mediated Conversions, and the Origin of Questionable Reversibility.

The interconversion of NO and HNO, via copper zinc superoxide dismutase (CuZnSOD), is important in biomedicine and for HNO detection. Many mechanistic questions, including the decades-long debate on reversibility, were resolved in this work. Calculations of various active-site and full-protein models show that the basic mechanism is proton-coupled electron transfer with a computed barrier of 10.98 kcal mol-1 , which is in excellent agreement with experimental results (10.62 kcal mol-1 ), and this nonheme protein-mediated reaction has many significant mechanistic differences compared with the conversions mediated by heme proteins due to geometric and electronic factors. The reasons for the irreversible nature of this conversion and models with the first thermodynamically favorable and kinetically feasible mechanism for the experimental reverse reaction were discovered. Such results are the first for nonheme enzyme mediated HNO to NO conversions, which shall facilitate other related studies and HNO probe development.

Automating, Analyzing, and Databasing Quantum Mechanical Enzyme Simulations

Enzyme simulations are notoriously challenging as their large size prevents modeling the whole enzyme with highly accurate but computationally expensive quantum mechanical (QM) methods. Currently, only the chemically important “active site” residues/substrates/cofactors are modeled with QM-methods. However, determining which biological sub-units necessarily should be included in the QM-model remains ambiguous, and current practices for model construction typically suffer from human bias and unclear methodology. My lab has been developing software called RINRUS (Residue Interaction Network ResidUe Selector) designed to use protein topology to construct residue interaction networks for the prediction of appropriate QM-only models of enzyme active sites. This methodology has been applied successfully to several enzymes including threonyl-tRNA synthetase, human carbonic anhydrase II, DD-peptidase, and catechol-O-methyltransferase. The use of a standardized model construction through RINRUS opens the possibility for databasing and data mining computational enzyme models by their features (e.g. model composition, enzyme identity, accuracy of properties compared to experimental data). As an example, we detail the automation and analysis of over 500 unique models of the enzyme catechol-O-methyltransferase. Ongoing efforts to further improve the RINRUS selection algorithm have examined incorporating random forest algorithms in the process of ranking residue importance. Current strides towards improving the automation of enzyme model construction, experiences with both collating and analyzing feature data for hundreds of models, and the outlook towards amassing a public database for atomic-level protein simulations will be discussed.

Discovery of beta-lactamase CMY-10 inhibitors for combination therapy against multi-drug resistant Enterobacteriaceae.

β-lactam antibiotics are the most widely used antimicrobial agents since the discovery of benzylpenicillin in the 1920s. Unfortunately, these life-saving antibiotics are vulnerable to inactivation by continuously evolving β-lactamase enzymes that are primary resistance determinants in multi-drug resistant pathogens. The current study exploits the strategy of combination therapeutics and aims at identifying novel β-lactamase inhibitors that can inactivate the β-lactamase enzyme of the pathogen while allowing the β-lactam antibiotic to act against its penicillin-binding protein target. Inhibitor discovery applied the Site-Identification by Ligand Competitive Saturation (SILCS) technology to map the functional group requirements of the β-lactamase CMY-10 and generate pharmacophore models of active site. SILCS-MC, Ligand-grid Free Energy (LGFE) analysis and Machine-learning based random-forest (RF) scoring methods were then used to screen and filter a library of 700,000 compounds. From the computational screens 74 compounds were subjected to experimental validation in which β-lactamase activity assay, in vitro susceptibility testing, and Scanning Electron Microscope (SEM) analysis were conducted to explore their antibacterial potential. Eleven compounds were identified as enhancers while 7 compounds were recognized as inhibitors of CMY-10. Of these, compound 11 showed promising activity in β-lactamase activity assay, in vitro susceptibility testing against ATCC strains (E. coli, E. cloacae, E. agglomerans, E. alvei) and MDR clinical isolates (E. cloacae, E. alvei and E. agglomerans), with synergistic assay indicating its potential as a β-lactam enhancer and β-lactamase inhibitor. Structural similarity search against the active compound 11 yielded 28 more compounds. The majority of these compounds also exhibited β-lactamase inhibition potential and antibacterial activity. The non-β-lactam-based β-lactamase inhibitors identified in the current study have the potential to be used in combination therapy with lactam-based antibiotics against MDR clinical isolates that have been found resistant against last-line antibiotics.

Bioinspired mononuclear Mn complexes for O2 activation and biologically relevant reactions.

A general interest in harnessing the oxidizing power of dioxygen (O2) continues to motivate research efforts on bioinspired and biomimetic complexes to better understand how metalloenzymes mediate these reactions. The ubiquity of Fe- and Cu-based enzymes attracts significant attention and has resulted in many noteworthy developments for abiotic systems interested in direct O2 reduction and small molecule activation. However, despite the existence of Mn-based metalloenzymes with important O2-dependent activity, there has been comparatively less focus on the development of these analogues relative to Fe- and Cu-systems. In this Perspective, we summarize important contributions to the development of bioinspired mononuclear Mn complexes for O2 activation and studies on their reactivity, emphasizing important design parameters in the primary and secondary coordination spheres and outlining mechanistic trends.

New copper(II) µ-Alkoxo-µ-Carboxylato Double Bridged Complexes As Models for Active Site of Catechol Oxidase: Synthesis, Spectral Characterization and DFT Calculations

A series of four copper(II) µ-alkoxo-µ-carboxylato double bridged complexes, [{Cu 2 (L)} 2 ][(µ-O 2 C-CO 2 ] 1 , [{Cu 2 (L)} 2 (µ-O 2 C-(CH 2 )CO 2 ] 2 , [{Cu 2 (L)} 2 (µ-O 2 C-CH 2 -CO 2 ] 3 and [{Cu 2 (L)} 2 (µ-O 2 C-C 6 H 4 -CO 2 ] 4 (H 3 L = 4-bromo-2-((E)-((3-(((E)-5-chloro-2-hydroxybenzylidene) amino)-2-hydroxypropyl) imino) methyl)-6-methoxyphenol and µ-dicarboxylate ions = oxalate, malonate, succinate and terephthalate) have been synthesized and characterized using several physicochemical technique. The tridentate nature of H 3 L is interpreted from IR spectra. The Epr spectra of these complexes are characteristic of the quintet state (S = 2) in central features and the triplet state (S = 1) of the tetranuclear complexes. The electrochemical potential of these complexes was investigated using cyclic Voltammetry (CV) and differential pulse Voltammetry (DPV). All complexes exhibited quasi-reversible reduction peaks in the cathodic region. To explore the stability of these complexes, quantum chemical parameters like electronegativity, ionization potential, electron affinity, global hardness and softness and electrophilicity were estimated and discussed. The synthesized complexes have been designed as structural and functional models of the catechol oxidase enzymes to investigate the catecholase activity. Additionally, superoxide dismutase activity data of all complexes have also been evaluated and compared with known SOD mimics.

A bioinspired oxoiron(IV) motif supported on a N2S2 macrocyclic ligand.

A mononuclear oxoiron(iv) complex 1-trans bearing two equatorial sulfur ligations is synthesized and characterized as an active-site model of the elusive sulfur-ligated FeIV[double bond, length as m-dash]O intermediates in non-heme iron oxygenases. The introduction of sulfur ligands weakens the Fe[double bond, length as m-dash]O bond and enhances the oxidative reactivity of the FeIV[double bond, length as m-dash]O unit with a diminished deuterium kinetic isotope effect, thereby providing a compelling rationale for nature's use of the cis-thiolate ligated oxoiron(iv) motif in key metabolic transformations.

First-Principles Calculations on Ni,Fe-Containing Carbon Monoxide Dehydrogenases Reveal Key Stereoelectronic Features for Binding and Release of CO2 to/from the C-Cluster.

In view of the depletion of fossil fuel reserves and climatic effects of greenhouse gas emissions, Ni,Fe-containing carbon monoxide dehydrogenase (Ni-CODH) enzymes have attracted increasing interest in recent years for their capability to selectively catalyze the reversible reduction of CO2 to CO (CO2 + 2H+ + 2e– CO + H2O). The possibility of converting the greenhouse gas CO2 into useful materials that can be used as synthetic building blocks or, remarkably, as carbon fuels makes Ni-CODH a very promising target for reverse-engineering studies. In this context, in order to provide insights into the chemical principles underlying the biological catalysis of CO2 activation and reduction, quantum mechanics calculations have been carried out in the framework of density functional theory (DFT) on different-sized models of the Ni-CODH active site. With the aim of uncovering which stereoelectronic properties of the active site (known as the C-cluster) are crucial for the efficient binding and release of CO2, different coordination modes of CO2 to different forms and redox states of the C-cluster have been investigated. The results obtained from this study highlight the key role of the protein environment in tuning the reactivity and the geometry of the C-cluster. In particular, the protonation state of His93 is found to be crucial for promoting the binding or the dissociation of CO2. The oxidation state of the C-cluster is also shown to be critical. CO2 binds to Cred2 according to a dissociative mechanism (i.e., CO2 binds to the C-cluster after the release of possible ligands from Feu) when His93 is doubly protonated. CO2 can also bind noncatalytically to Cred1 according to an associative mechanism (i.e., CO2 binding is preceded by the binding of H2O to Feu). Conversely, CO2 dissociates when His93 is singly protonated and the C-cluster is oxidized at least to the Cint redox state.

Alkaline Phosphatases: in Silico Study on the Catalytic Effect of Conserved Active Site Residues Using Human Placental Alkaline Phosphatase (PLAP) As a Model Protein.

The metalloenzymes from the alkaline phosphatase (AP) superfamily catalyze the hydrolysis and transphosphorylation of phosphate monoesters. The role of several amino acids highly conserved in the active site of this family of enzymes was examined, using human placental AP (PLAP) as a model protein. By employing an active-site model based on the X-ray crystal structure of PLAP, mutations of several key residues were modeled by quantum mechanical methods in order to determine their impact on the catalytic activity. Kinetic and thermodynamic estimations were achieved for each reaction step of the catalytic mechanism by characterization of the intermediates and transition states on the reaction pathway, and the effects of mutations on the activation barriers were analyzed. A good accordance was observed between the present computational results and experimental measurements reported in the literature.

Kinetic modeling of dynamic changing active sites in a Mars-van Krevelen type reaction: Ethylene oxychlorination on K-doped CuCl2/Al2O3

A kinetic model was developed by taking into account the dynamic nature of the active sites in Mars–van Krevelen type catalytic reactions to predict the evolution of the reactant and product composition in the gas phase and the CuCl2 concentration in the solid catalyst. The kinetic model at the steady-state of ethylene oxychlorination was obtained by combining transient experiments of the two half-reactions in the redox cycle, namely CuCl2 reduced to CuCl by ethylene and CuCl oxidation by oxygen on the K-promoted CuCl2/γ-Al2O3 catalyst. The dynamic transitions between CuCl2 and CuCl of the active sites during the reactions are also modeled, and the contributions of two active sites, namely Cu coordination numbers of 4 and 3 in CuCl2 were distinguished and included in the kinetic model. The kinetic models describe well the transient response of the reduction and oxidation steps as well as the reaction at the steady-state at different reaction conditions. Moreover, by combining the reactor modeling through a steady-state approach, the spatial-time resolved CuCl2 profile and the C2H4 reaction rate can be well predicted in comparison with the experimental results. The approach of both transient and steady-state kinetic modeling and simulation is supposed to have general relevance for a better understanding of Mars–van Krevelen type reactions.

AFM Research in Catalysis and Medicine

Background: In this study, we show that the AFM method not only allows monitoring the morphological changes in biological structures fixed on the surface due to H-bonds, but also makes it possible to study the self-organization of metal complexes by simulating the active center of enzymes due to intermolecular H-bonds into stable nanostructures; the sizes of which are much smaller than the studied biological objects. The possible role of intermolecular hydrogen bonds in the formation of stable supramolecular metal complexes, which are effective catalysts for the oxidation of alkyl arenes to hydroperoxides by molecular oxygen and mimic the selective active sites of enzymes, was first studied by AFM. Methods and Results: The formation of supramolecular structures due to intermolecular hydrogen bonds and, possibly, other non-covalent interactions, based on homogenous catalysts and models of active centers enzymes, heteroligand nickel and iron complexes, was proven by AFM-technique. AFM studies of supramolecular structures were carried out using NSG30 cantilever with a radius of curvature of 2 nm, in the tapping mode. To form nanostructures on the surface of a hydrophobic, chemically modified silicon surface as a substrate, the sample was prepared using a spin-coating process from solutions of the nickel and iron complexes. The composition and the structure of the complex Ni2(acac)(OAc)3·NMP·2H2O were determined in earlier works using various methods: mass spectrometry, UV- and IR-spectroscopy, elemental analysis, and polarography. Self-assembly of supramolecular structures is due to intermolecular interactions with a certain coordination of these interactions, which may be a consequence of the properties of the components themselves, the participation of hydrogen bonds and other non-covalent interactions, as well as the balance of the interaction of these components with the surface. Using AFM, approaches have been developed for fixing on the surface and quantifying parameters of cells. Conclusion: This study summarizes the authors' achievements in using the atomic force microscopy (AFM) method to study the role of intermolecular hydrogen bonds (and other non-covalent interactions) and supramolecular structures in the mechanisms of catalysis. The data obtained from AFM based on nickel and iron complexes, which are effective catalysts and models of active sites of enzymes, indicate a high probability of the formation of supramolecular structures in real conditions of catalytic oxidation, and can bring us closer to understanding enzymes activity. With a sensitive AFM method, it is possible to observe the self-organization of model systems into stable nanostructures due to H-bonds and possibly other non-covalent interactions, which can be considered as a step towards modeling the active sites of enzymes. Methodical approaches of atomic force microscopy for the study of morphological changes of cells have been developed.

Novel ribonucleotide discrimination in the RNA polymerase-like two-barrel catalytic core of Family D DNA polymerases.

Family D DNA polymerase (PolD) is the essential replicative DNA polymerase for duplication of most archaeal genomes. PolD contains a unique two-barrel catalytic core absent from all other DNA polymerase families but found in RNA polymerases (RNAPs). While PolD has an ancestral RNA polymerase catalytic core, its active site has evolved the ability to discriminate against ribonucleotides. Until now, the mechanism evolved by PolD to prevent ribonucleotide incorporation was unknown. In all other DNA polymerase families, an active site steric gate residue prevents ribonucleotide incorporation. In this work, we identify two consensus active site acidic (a) and basic (b) motifs shared across the entire two-barrel nucleotide polymerase superfamily, and a nucleotide selectivity (s) motif specific to PolD versus RNAPs. A novel steric gate histidine residue (H931 in Thermococcus sp. 9°N PolD) in the PolD s-motif both prevents ribonucleotide incorporation and promotes efficient dNTP incorporation. Further, a PolD H931A steric gate mutant abolishes ribonucleotide discrimination and readily incorporates a variety of 2′ modified nucleotides. Taken together, we construct the first putative nucleotide bound PolD active site model and provide structural and functional evidence for the emergence of DNA replication through the evolution of an ancestral RNAP two-barrel catalytic core.

Effect of Promoter M (M = Au, Ag, Cu, Ce, Fe, Ni, Co, Zn) on the Activity of Pd–M/Al2O3 Catalysts of Ethanol Conversion into α-Alcohols

Pd/Al2O3 and Pd–M/Al2O3 catalysts (M = Au, Ag, Cu, Ce, Fe, Ni, Co, Zn) were obtained by ion exchange and impregnation. Pd/Al2O3 had high initial activity in the conversion of ethanol into α-alcohols, but lost 90% of its activity after 10 h of operation because of deactivation caused by the chemisorption of the by-product (CO) on Pd atoms. Modification of Pd with gold or silver led to an increase in the rate of Pd deactivation. As a result, the Pd–Au and Pd–Ag systems were less active and stable. In contrast, the Pd–Fe, Pd–Co, Pd–Ni, Pd–Cu, Pd–Zn, and Pd–Ce systems exhibited higher resistance to CO poisoning than Pd and demonstrated high activity and stability. The observed tendencies in the catalytic action of the mono- and bimetallic systems were explained within the framework of the d band model proposed by Hammer and Norskov. Pd–Cu/Al2O3 was most effective in the target process; it is not poisoned by CO and allows ethanol conversion into α-alcohols at 95% selectivity, while the time of its stable operation is at least 100 h. The structure of the Pd–Cu catalytic system was studied by TEM, EDA, XPS, TPR-H2, and TPD-NH3. A model of active catalyst sites was proposed.

Reaction mechanism catalyzed by the dissimilatory adenosine 5′-phosphosulfate reductase. Adenosine 5′-monophosphate inhibitor and key role of arginine 317 in switching the course of catalysis

The present research is a continuation of our work on dissimilatory reduction pathway of sulfate - involved in biogeochemical sulfur turnover. Adenosine 5′-phosphosulfate reductase (APSR) is the second enzyme in the dissimilatory pathway of the sulfate to sulfide reduction. It reversibly catalyzes formation of the sulfite anion (HSO3−) from adenosine 5′-phosphosulfate (APS) - the activated form of sulfate provided by ATP sulfurylase (ATPS). Two electrons required for this redox reaction derive from reduced FAD cofactor, which is suggested to be involved directly in the catalysis by formation of FADH-SO3− intermediate. The present work covers quantum-mechanical (QM) studies on APSR reaction performed for eight models of APSR active site. The cluster models were constructed based on two crystal structures (PDB codes: 2FJA and 2FJB), differing in conformation of Arg317 active site residue. The described results indicated the most feasible mechanism of APSR forward reaction, including formation of FADHN-SO3− adduct (with proton on N5 atom of isoalloxazine), tautomerization of FADHN-SO3− to FADHO-SO3− (with proton on CO moiety of isoalloxazine), and its reductive cleavage to oxidized FAD and sulfite anion. The reverse reaction proceeds in the backward direction. It is suggested that it requires two AMP molecules, one acting as a substrate and another as an inhibitor of forward reaction, which forces change of Arg317 conformation from “arginine in” (2FJA) to “arginine out” (2FJB). Important role of Arg317 in switching the course of the APSR catalytic reaction is revealed by changing the direction of thermodynamic driving force. The presented research also shows the importance of the protonation pattern of the reduced FAD cofactor and protein residues within the active site.

Lignin Biodegradation by a Cytochrome P450 Enzyme: A Computational Study into Syringol Activation by GcoA.

A recently characterized cytochrome P450 isozyme GcoA activates lignin components through a selective O‐demethylation or alternatively an acetal formation reaction. These are important reactions in biotechnology and, because lignin is readily available; it being the main component in plant cell walls. In this work we present a density functional theory study on a large active site model of GcoA to investigate syringol activation by an iron(IV)‐oxo heme cation radical oxidant (Compound I) leading to hemiacetal and acetal products. Several substrate‐binding positions were tested and full energy landscapes calculated. The study shows that substrate positioning determines the product distributions. Thus, with the phenol group pointing away from the heme, an O‐demethylation is predicted, whereas an initial hydrogen‐atom abstraction of the weak phenolic O‐H group would trigger a pathway leading to ring‐closure to form acetal products. Predictions on how to engineer P450 GcoA to get more selective product distributions are given.

Chiral discrimination in a mutated IDH enzymatic reaction in cancer: a computational perspective.

Chiral discrimination in biological systems, such as L-amino acids in proteins and d-sugars in nucleic acids, has been proposed to depend on various mechanisms, and chiral discrimination by mutated enzymes mediating cancer cell signaling is important in current research. We have explored how mutated isocitrate dehydrogenase (IDH) catalyzes the oxidative decarboxylation of isocitrate to α-ketoglutarate which in turn is converted to d-2-hydroxyglutatrate (d-2HG) as a preferred product instead of l-2-hydroxyglutatrate (l-2HG) according to quantum chemical calculations. Using transition state structure modeling, we delineate the preferred product formation of d-2HG over l-2HG in an IDH active site model. The mechanisms for the formation of d-2HG over l-2HG are assessed by identifying transition state structures and activation energy barriers in gas and solution phases. The calculated reaction energy profile for the formation of d-2HG and l-2HG metabolites shows a 29 times higher value for l-2HG as compared to d-2HG. Results for second-order Møller-Plesset perturbation theory (MP2) do not alter the observed trend based on Density Functional Theory (DFT). The observed trends in reaction energy profile explain why the formation of D-2HG is preferred over l-2HG and reveal why mutation leads to the formation of d-2HG instead of l-2HG. For a better understanding of the observed difference in the activation barrier for the formation of the two alternative products, we performed natural bond orbital analysis, non-covalent interactions analysis and energy decomposition analysis. Our findings based on computational calculations clearly indicate a role for chiral discrimination in mutated enzymatic pathways in cancer biology.

Effect of Oxygen on the Oxidation of Methane with Hydrogen Peroxide to Methanol in the Presence of Glutathione-Stabilized Gold Nanoclusters

The catalytic activity of glutathione-stabilized gold thiolate nanoclusters (1–2 nm) in the oxidation of methane with H2O2 and air oxygen in aqueous medium at 70°C in 30 atm CH4 was studied. The ratio and yield of the oxidation products CH3OH and CH3OOH depends on the content of H2O2 and oxygen in the reactor. At increased partial pressure of oxygen, the total yield of products increases compared with the total yield of independent oxidations with H2O2 or O2 at 5 atm of air, but decreases at 10 atm. A molecular mechanism of methane oxidation was proposed, which describes well the kinetic curves of accumulation and consumption of CH3OH and CH3OOH and the effect of oxygen on their yield. A molecular model of active site was proposed based on the literature data about the structure of glutathione-stabilized Au25 clusters and the results of quantum chemical modeling. The experiments with re-introduction of the gas phase and renewal of the H2O2 content demonstrated 100% conservation of the catalytic activity. The yield of CH3OH reached 60 mol per mole of Au25 clusters.

Mechanisms of Solid–Gas Reactions: Reduction of Air Pollutants on Carbons

Ozone is a strong oxidizer and sulfur dioxide is a precursor to acid rain, both are air pollutants that can damage the respiratory tissues of animals and plants making them hazardous to the environment. They are isoelectronic valence O = X = O (X = S, O) molecules and their mechanism of reduction on carbons is similar. The solid–gas kinetics were studied in a flow system with a tubular reactor under differential and steady state conditions. Initial rates and product distribution were used to determine the stoichiometry of the reaction. The reduction of XO2 on carbons proceeds through a common primary mechanism with oxidized and reduced intermediates. The reactivity of the intermediates that were inserted on carbons (graphite, activated carbon, graphene oxide) modified by SO2 is selective with respect to aminolysis and thiolysis. A theoretical study of the chemisorption of SO2 on dehydrogenated pyrene as graphite active site model showed that at 900 °C the chemisorption occurs mainly on the diradical zigzag edge. Computational quantum chemistry calculations were carried out for the reduction of SO2 on graphite to produce elemental sulfur and CO2 using tetradehydrogenated-benzo[α]anthracene (TBA) as model. The mechanisms of the decarboxylation and sulfur transport steps were postulated. Ozonation of graphite showed that the 1,2,3-trioxolane decomposes to an oxyrene, eliminating O2. Both reactions, the SO2 and O3 with graphite, have the same experimental free energy of activation for the decarboxylation reaction. The results show that for SO2 the desulfurization has a much lower energetic demand than the decarboxylation route raising the important possibility of using the reaction of reduction of SO2 on carbons to reduce the acid rain, producing elemental sulfur as the main product, without increasing the greenhouse effect due to the formation of CO2.

ProdaMatch: A fast and accurate active site matching algorithm for de novo enzyme design

Developing enzymes with high catalytic activity solely based on computation is still a great challenge in de novo enzyme design. Based on the complex active site model in which the catalytic residues, in addition to interacting with the transition state, impose geometrical constraints on each other, a fast and accurate active-site matching algorithm ProdaMatch was developed to search the whole protein structure database and create fully preorganized active sites in selected scaffolds. The current version of ProdaMatch runs ten times as fast as previous versions while maintaining sub-angstrom matching accuracy for transition states and 90% of the catalytic residues based on tests on the active site recapitulation of 10 enzyme-transition state complex structures. The ProdaMatch method was validated computationally using parallel computation to identify scaffolds from a structure database for Kemp elimination and create fully connected active sites in the scaffolds. The catalytic residues were anchored in geometrically compatible orientations in the scaffolds, and the transition states were positioned in catalytically competent conformations. ProdaMatch can help create fully preorganized enzymes for the de novo design of enzymes with high starting catalytic activities.