Free Energy Profiles Research Articles

First-Principles Insights into the Mechanism of CO2 Hydrogenation Reactions by Fe-PNP Pincer Complex.

Using the state of the art theoretical methods, we have provided a comprehensive mechanistic understanding of the CO2 hydrogenation into HCOOH, H2CO, and CH3OH by 2,6-bis(diisopropylphosphinomethyl)pyridine (PNP)-ligated Fe pincer complex, featuring one CO and two H as co-ligands. For the computational investigation, a verified structural model containing methyl groups in place of the experimental isopropyl groups was used. Three catalytic conversions involving hydrogenation of CO2 into formic acid (HCOOH), HCOOH into formaldehyde and methanol were studied in different solvent medium. Our modelled complex appears to be a viable base-free catalyst for the conversion of CO2 into HCOOH and HCOOH into H2CO, based on the free energy profiles, which show apparent activation energy barriers of 16.28 kcal/mol and 23.63 kcal/mol for the CO2 to HCOOH and HCOOH to H2CO conversion, respectively. However, the computed results show that, due to the huge energy span of H2CO to CH3OH conversion, complete hydrogenation of CO2 into methanol could not occur under moderate conditions. Morpholine co-catalyst, which can lower the hydrogenation barrier by taking part in a simultaneous H-atom donation-acceptance process, could have assisted in completing this step.

Studies on the compositional dependent structural and electrical properties of CaTiO3-modified K0.5Na0.5NbO3 piezoelectric system

Lead-free piezoelectric ceramics of (1 − x)K0.5Na0.5NbO3-xCaTiO3 were fabricated, and their crystal structure, microstructure, and electrical properties were systematically studied. Rietveld refinement of the x-ray diffraction data and Raman spectroscopic analyses revealed a composition-dependent structural phase transition: three phase transitions, namely, from a pure orthorhombic phase for x ≤ 0.02 to a mixed phase of orthorhombic and tetragonal phases (0.03 ≤ x ≤ 0.08) and finally another mixed phase of tetragonal + cubic for x = 0.10 and 0.15 at room temperature (RT). The morphological study reveals a decrease in grain size along with a more uniform distribution of grains as the concentration of CaTiO3 (CT) increases; notably, a homogeneous distribution of grains is observed for x = 0.05. The temperature-dependent dielectric properties show two phase transitions, from orthorhombic to tetragonal (TO-T) and tetragonal to cubic (TC), for unmodified K0.5Na0.5NbO3 (KNN). However, both the phase transition temperatures (TO-T and TC) decrease, and the transition peaks broaden with an increase in CT substitution, and for x > 0.06, the TO-T shifted below RT. The broadening of the transition peak at TO-T may be due to the relaxation behavior. Among the prepared samples, the 5 mol. % CT-modified KNN shows the optimum electrical properties (d33 = 114 pC/N, ɛr = 412, and 2Pr = 15.25 μC/cm2) at RT. The enhanced electrical properties for x = 0.05 are due to the coexistence of orthorhombic and tetragonal phases, facilitating easy polarization rotation and flattening of the free energy profile. A phase diagram has been constructed based on the information gathered from the temperature-dependent dielectric measurements, RT x-ray diffraction, and Raman spectroscopy data and is discussed in detail.

Exploring the photophysics of cinnamoyl-coumarin derivatives in cucurbit [7]uril complexes and assessing phototoxicity in HeLa cells

Cinnamoyl-coumarin derivatives containing 5 or 12 carbon aliphatic chains were evaluated as photosensitizers and their photophysical properties were studied in detail. Moreover, these compounds were encapsulated in cucurbit[7]uril macrocycle with relatively high binding affinities in aqueous media. Fluorescence quantum yields, fluorescence lifetimes and singlet oxygen quantum yields were measured upon encapsulation. Computational DFT calculations and molecular dynamics simulations allowed the visualization of the binding modes of both derivatives inside the macrocycle. In particular, the free energy profiles for the insertion of the molecules into the macrocycle were determined to assess the most probable conformations of the complexes in water. In vitro phototoxicity assays were carried out in HeLa cells showed significant decreases in cell viability upon illumination of the 5 carbon aliphatic chain derivative at 460 nm with an LED source at different concentrations and enhanced by cucurbit[7]uril. However, the one with 12 carbon aliphatic chain did not display any phototoxicity regardless of the presence of cucurbit[7]uril. This behavior is related to the high lipophilicity introduced by a 12 carbon aliphatic chain conditioning its distribution on hydrophobic compartments and decreasing its phototoxicity. The enhanced phototoxicity elicited by cucurbit[7]uril on the 5 carbon aliphatic chain derivative but not in the 12 carbon derivative remark that there is a balance between hydrophobicity of these probes with the carrier properties of cucurbit[7]uril to be reach for effective phototoxicity properties.

Benzylic Trifluoromethyl Accelerates 1,6-Elimination Toward Rapid Probe Activation.

Activity-based detection of hydrogen sulfide in live cells can expand our understanding of its reactivity and complex physiological effects. We have discovered a highly efficient method for fluorescent probe activation, which is driven by H2S-triggered 1,6-elimination of an α-CF3-benzyl to release resorufin. In detecting intracellular H2S, 4-azido-(α-CF3)-benzyl resorufin offers significantly faster signal generation and improved sensitivity compared to 4-azidobenzyl resorufin. Computed free energy profiles for the 1,6-elimination process support the hypothesis that a benzylic CF3 group can reduce the activation energy barrier toward probe activation. This novel probe design allows for near-real-time detection of H2S in HeLa cells under stimulation conditions.

Combining Transition Path Sampling with Data-Driven Collective Variables through a Reactivity-Biased Shooting Algorithm.

Rare event sampling is a central problem in modern computational chemistry research. Among the existing methods, transition path sampling (TPS) can generate unbiased representations of reaction processes. However, its efficiency depends on the ability to generate reactive trial paths, which in turn depends on the quality of the shooting algorithm used. We propose a new algorithm based on the shooting success rate, i.e., reactivity, measured as a function of a reduced set of collective variables (CVs). These variables are extracted with a machine learning approach directly from TPS simulations, using a multitask objective function. Iteratively, this workflow significantly improves the shooting efficiency without any prior knowledge of the process. In addition, the optimized CVs can be used with biased enhanced sampling methodologies to accurately reconstruct the free energy profiles. We tested the method on three different systems: a two-dimensional toy model, conformational transitions of alanine dipeptide, and hydrolysis of acetyl chloride in bulk water. In the latter, we integrated our workflow with an active learning scheme to learn a reactive machine learning-based potential, which allowed us to study the mechanism and free energy profile with an ab initio-like accuracy.

Mechanism of Peroxynitrite Interaction with Ferric M. tuberculosis Nitrobindin: A Computational Study.

Nitrobindins (Nbs) are all-β-barrel heme proteins present along the evolutionary ladder. They display a highly solvent-exposed ferric heme group with the iron atom being coordinated by the proximal His residue and a water molecule at the distal position. Ferric nitrobindins (Nb(III)) play a role in the conversion of toxic peroxynitrite (ONOO-) to harmless nitrate, with the value of the second-order rate constant being similar to those of most heme proteins. The value of the second-order rate constant of Nbs increases as the pH decreases; this suggests that Nb(III) preferentially reacts with peroxynitrous acid (ONOOH), although ONOO- is more nucleophilic. In this work, we shed light on the molecular basis of the ONOO- and ONOOH reactivity of ferric Mycobacterium tuberculosis Nb (Mt-Nb(III)) by dissecting the ligand migration toward the active site, the water molecule release, and the ligand binding process by computer simulations. Classical molecular dynamics simulations were performed by employing a steered molecular dynamics approach and the Jarzynski equality to obtain ligand migration free energy profiles for both ONOO- and ONOOH. Our results indicate that ONOO- and ONOOH migration is almost unhindered, consistent with the exposed metal center of Mt-Nb(III). To further analyze the ligand binding process, we computed potential energy profiles for the displacement of the Fe(III)-coordinated water molecule using a hybrid QM/MM scheme at the DFT level and a nudged elastic band approach. These results indicate that ONOO- exhibits a much larger barrier for ligand displacement than ONOOH, suggesting that water displacement is assisted by protonation of the leaving group by the incoming ONOOH.

Large-area ultrathin 2D Co(OH)2 nanosheets: a bifunctional electrode material for supercapacitor and water oxidation

Ultrathin film of active materials having thickness <5 nm emerged as a promising candidate for miniaturized energy storage and conversion devices. However, the small lateral size restricts its real device applications. Herein, we report large area, 2D, ultrathin (thickness: 4.3 ± 0.3 nm) Co(OH)2 nanosheets as bifunctional electrode material for supercapacitor and oxygen evolution reaction developed by ionic layer epitaxy method at the water-air interface. The symmetric supercapacitor device exhibited excellent volumetric capacitance of 2313 F/cm3 at 0.4 mA/cm2 current density. Additionally, it exhibited a remarkable volumetric energy density of 0.205 Wh/cm3 at a power density of 0.145 W/cm3, which is better than reported 2D electrode materials. Furthermore, the cobalt hydroxide (Co(OH)2) ultrathin-film electrodes showed improved oxygen evolution reaction electrocatalytic activity with low overpotential (ƞ10, 330 mV) and Tafel slope (47 mV/dec). The structural and morphological investigations after long-term operations show substantial stability of the electrode material. The theoretical investigations of electronic structures, quantum capacitance, and free energy profiles for the oxygen evolution reaction mechanism corroborate the experimental results.

Cryptotanshinone-Induced Permeabilization of Model Phospholipid Membranes: A Biophysical Study.

The Danshen terpenoid cryptotanshinone (CPT) is gaining enormous interest in light of its various outstanding biological activities. Among those, CPT has been shown to interact with cell membranes and, for instance, to have antibacterial activity. Several works have shown that CPT alone, or in combination with other drugs, can effectively act as an antibiotic against various infectious bacteria. Some authors have related the mechanism underlying this action to CPT-membrane interaction. This work shows that CPT readily partitions into phosphatidylcholine membranes, but there is a limiting capacity of accommodation of ca. 1 mol CPT to 3 mol phospholipid. The addition of CPT to unilamellar liposomes composed of 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) causes membrane permeabilization, as shown by fluorescent probe leakage. This process has been kinetically studied, as well as its modulation by incorporation of phosphatidylethanolamine or phosphatidylglycerol, as a model for pathogenic cell membranes. The thermotropic behavior of 1,2-dimyristoylphosphatidylcholine (DMPC) model membranes is weakly affected by CPT, but the terpenoid causes significant dehydration of the polar region of the bilayer and weak disordering of the acyl chain palisade, as observed in Fourier-transform infrared spectroscopy (FTIR) results. Small-angle X-ray scattering (SAXS) shows that CPT increases DMPC bilayer thickness, which could be due to localization near the phospholipid/water interface. Molecular dynamics (MD) simulations show that the lateral diffusion coefficient of the phospholipid increases with the presence of CPT. CPT extends from the polar head region to the center of the bilayer, being centered between the carbonyl groups and the unsaturated region of the POPC, where there is greater overlap. Interestingly, the free energy profiles of a water molecule crossing the lipid membrane show that the POPC membrane becomes more permeable in the presence of CPT. In summary, our results show that CPT perturbs the physicochemical properties of the phospholipid membrane and compromises its barrier function, which could be of relevance to explain part of its antimicrobial or anticancer activities.

A novel Imidazo[1,2-a]pyridine derivative modulates active KRASG12D through off-like conformational shifts in switch-I and switch-II regions, mimicking inactive KRASG12D

KRASG12D are the most prevalent oncogenic mutations and a promising target for solid tumor therapies. However, its inhibition exhibits tremendous challenge due to the necessity of high binding affinity to obviate the need for covalent binders. Here we report the evidence of a novel class of Imidazo[1,2-a]pyridine derivative as potentially significant novel inhibitors of KRASG12D, discovered through extensive ligand-based screening against 2-[(2R)-piperidin-2-yl]-1H-indole, an important scaffold for KRASG12D inhibition via switch-I/II (S-I/II) pocket. The proposed compounds exhibited similar binding affinities and overlapped pose configurations to 2-[(2R)-piperidin-2-yl]-1H-indole, serving as a reliable starting point for drug discovery. Comparative free energy profiles demonstrated that C4 [2-methyl-3-((5-phenyl-1H-1,2,4-triazol-3-yl)methyl)imidazo[1,2-a]pyridine] effectively shifted the protein to a stable low-energy conformation via a prominent transition state. The conformational changes across the transition revealed the conformational shift of switch-I and II to a previously known off-like conformation of inactive KRASG12D with rmsd of 0.91 Å. These conformations were even more prominent than the privileged scaffold 2-[(2R)-piperidin-2-yl]-1H-indole. The representative structure overlay of C4 and another X-ray crystallography solved BI-2852 bound inactive KRASG12D revealed that Switch-I and II exhibited off-like conformations. The cumulative variance across the first eigenvalue that accounted for 57 % of the collective variance validated this on-to-off transition. In addition, the relative interaction of C4 binding showed consistent patterns with BI-2852. Taken together, our results support the inhibitory activity of [2-methyl-3-((5-phenyl-1H-1,2,4-triazol-3-yl)methyl)imidazo[1,2-a]pyridine] by shifting active KRASG12D to an inactive conformation.

Enhanced Sampling Molecular Dynamics Simulations Reveal Transport Mechanism of Glycoconjugate Drugs through GLUT1.

Glucose transporters GLUT1 belong to the major facilitator superfamily and are essential to human glucose uptake. The overexpression of GLUT1 in tumor cells designates it as a pivotal target for glycoconjugate anticancer drugs. However, the interaction mechanism of glycoconjugate drugs with GLUT1 remains largely unknown. Here, we employed all-atom molecular dynamics simulations, coupled to steered and umbrella sampling techniques, to examine the thermodynamics governing the transport of glucose and two glycoconjugate drugs (i.e., 6-D-glucose-conjugated methane sulfonate and 6-D-glucose chlorambucil) by GLUT1. We characterized the specific interactions between GLUT1 and substrates at different transport stages, including substrate recognition, transport, and releasing, and identified the key residues involved in these procedures. Importantly, our results described, for the first time, the free energy profiles of GLUT1-transporting glycoconjugate drugs, and demonstrated that H160 and W388 served as important gates to regulate their transport via GLUT1. These findings provide novel atomic-scale insights for understanding the transport mechanism of GLUT1, facilitating the discovery and rational design of GLUT1-targeted anticancer drugs.

Energetics of the Transmembrane Peptide Sorting by Hydrophobic Mismatch.

Hydrophobic mismatch between a lipid membrane and embedded transmembrane peptides or proteins plays a role in their lateral localization and function. Earlier studies have resolved numerous mechanisms through which the peptides and membrane proteins adapt to mismatch, yet the energetics of lateral sorting due to hydrophobic mismatch have remained elusive due to the lack of suitable computational or experimental protocols. Here, we pioneer a molecular dynamics simulation approach to study the sorting of peptides along a membrane thickness gradient. Peptides of different lengths tilt and diffuse along the membrane to eliminate mismatch with a rate directly proportional to the magnitude of mismatch. We extract the 2-dimensional free energy profiles as a function of local thickness and peptide orientation, revealing the relative contributions of sorting and tilting, and suggesting their thermally accessible regimes. Our approach can readily be applied to study other membrane systems of biological interest where hydrophobic mismatch, or membrane thickness in general, plays a role.

Binding of Phosphate Species to Ca2+ and Mg2+ in Aqueous Solution.

Phosphate derivatives and their interaction with metal cations are involved in many important biological phenomena, so an accurate characterization of the phosphate-metal interaction is necessary to properly understand the role of phosphate-metal contacts in mediating biological function. Herein, we improved the standard 12-6 Lennard-Jones (LJ) potential via the usage of the 12-6-4 LJ model, which incorporates ion-induced dipole interactions. Via parameter scanning, we fine-tuned the 12-6-4 LJ polarizability values to obtain accurate absolute binding free energies for the phosphate anions H2PO4-, HPO42-, PO43- coordinating with Ca2+ and Mg2+. First, we modified the phosphate 12-6-4 LJ parameters to reproduce the solvation free energies of the series of phosphate anions using the thermodynamic integration (TI) method. Then, using the potential mean force (PMF) method, the polarizability of the metal-phosphate interaction was obtained. We show that the free energy profiles of phosphate ions coordinated to Ca2+ and Mg2+ generally show similar trends at longer metal-phosphate distances, while the absolute binding energy values increased with deprotonation. The resulting parameters demonstrate the flexibility of the 12-6-4 LJ-type nonbonded model and its usefulness in accurately describing cation-anion interactions.

Analytic Gradients for the Electrostatic Embedding QM/MM Model in Periodic Boundary Conditions Using Particle-Mesh Ewald Sums and Electrostatic Potential Fitted Charge Operators.

Long-range electrostatic effects are fundamental for describing chemical reactivity in the condensed phase. Here, we present the methodology of an efficient quantum mechanical/molecular mechanical (QM/MM) model in periodic boundary conditions (PBC) compatible with QM/MM boundaries at chemical bonds. The method combines electrostatic potential fitted charge operators and electrostatic potentials derived from the smooth particle-mesh Ewald (PME) sum approach. The total energy and its analytic first derivatives with respect to QM, MM, and lattice vectors allow QM/MM molecular dynamics (MD) in the most common thermodynamic ensembles. We demonstrate the robustness of the method by performing a QM/MM MD equilibration of methanol in water. We simulate the cis/trans isomerization free-energy profiles in water of proline amino acid and a proline-containing oligopeptide, showing a correct description of the reaction barrier. Our PBC-compatible QM/MM model can efficiently be used to study the chemical reactivity in the condensed phase and enzymatic catalysis.

The role of memory-dependent friction and solvent viscosity in isomerization kinetics in viscogenic media

Molecular isomerization kinetics in liquid solvent depends on a complex interplay between the solvent friction acting on the molecule, internal dissipation effects (also known as internal friction), the viscosity of the solvent, and the dihedral free energy profile. Due to the absence of accurate techniques to directly evaluate isomerization friction, it has not been possible to explore these relationships in full. By combining extensive molecular dynamics simulations with friction memory-kernel extraction techniques we consider a variety of small, isomerising molecules under a range of different viscogenic conditions and directly evaluate the viscosity dependence of the friction acting on a rotating dihedral. We reveal that the influence of different viscogenic media on isomerization kinetics can be dramatically different, even when measured at the same viscosity. This is due to the dynamic solute-solvent coupling, mediated by time-dependent friction memory kernels. We also show that deviations from the linear dependence of isomerization rates on solvent viscosity, which are often simply attributed to internal friction effects, are due to the simultaneous violation of two fundamental relationships: the Stokes-Einstein relation and the overdamped Kramers prediction for the barrier-crossing rate, both of which require explicit knowledge of friction.

Deciphering the Cofilin Oligomers via Intermolecular Disulfide Bond Formation: A Coarse-Grained Molecular Dynamics Approach to Understanding Cofilin's Regulation on Actin Filaments.

Cofilin, a key actin-binding protein, orchestrates the dynamics of the actomyosin network through its actin-severing activity and by promoting the recycling of actin monomers. Recent experiments suggest that cofilin forms functionally distinct oligomers via thiol post-translational modifications (PTMs) that promote actin nucleation and assembly. Despite these advances, the structural conformations of cofilin oligomers that modulate actin activity remain elusive because there are combinatorial ways to oxidize thiols in cysteines to form disulfide bonds rapidly. This study employs molecular dynamics simulations to investigate human cofilin 1 as a case study for exploring cofilin dimers via disulfide bond formation. Utilizing a biasing scheme in simulations, we focus on analyzing dimer conformations conducive to disulfide bond formation. Additionally, we explore potential PTMs arising from the examined conformational ensemble. Using the free energy profiling, our simulations unveil a range of probable cofilin dimer structures not represented in current Protein Data Bank entries. These candidate dimers are characterized by their distinct population distributions and relative free energies. Of particular note is a dimer featuring an interface between cysteines 139 and 147 residues, which demonstrates stable free energy characteristics and intriguingly symmetrical geometry. In contrast, the experimentally proposed dimer structure exhibits a less stable free energy profile. We also evaluate frustration quantification based on the energy landscape theory in the protein-protein interactions at the dimer interfaces. Notably, the 39-39 dimer configuration emerges as a promising candidate for forming cofilin tetramers, as substantiated by frustration analysis. Additionally, docking simulations with actin filaments further evaluate the stability of these cofilin dimer-actin complexes. Our findings thus offer a computational framework for understanding the role of thiol PTM of cofilin proteins in regulating oligomerization, and the subsequent cofilin-mediated actin dynamics in the actomyosin network.

Development of Membrane Permeability Coefficient by Means of Novel Molecular Dynamics Methods

The membrane permeability, and its evaluation, is crucial factor in the process of uptake of compounds from outside to inside the cell and in the inhibition of the activity of disease-causing target proteins. Although molecular dynamics (MD) simulations have been shown to be able to reproduce the conformational changes of compounds occurring during membrane permeation, it is still challenging to extract the membrane permeability at an affordable computational workload solely by conventional MD. Indeed, the time scale accessible by MD is far below the one characterizing the actual permeation process. Phenomena occurring in living organisms escaping the reach of standard MD are generally referred to as biological rare events, and the membrane permeation process is one of them. To overcome this time-scale problem, several enhanced sampling methods have been proposed over the years to improve conformational sampling. In this review, a hybrid sampling method that combines the parallel cascade selection MD (PaCS-MD) and the outlier flooding method (OFLOOD), introduced and developed by our group, is proposed as a tool to study the membrane permeation from structural sampling (rare-event sampling). The obtained trajectories are used to estimate the free energy profiles for the membrane permeation and to compute the membrane permeation coefficients. Moreover, we present an example of application of the free energy reaction network method as a versatile way for incorporating explicitly into reaction coordinates the degrees of freedom related to internal motion.

Exploring the potential of silymarin-loaded nanovesicles as an effective drug delivery system for cancer therapy: in vivo, in vitro, and in silico experiments.

We aimed to perform a comprehensive study on the development and characterization of silymarin (Syl)-loaded niosomes as potential drug delivery systems. The results demonstrate significant novelty and promising outcomes in terms of morphology, size distribution, encapsulation efficiency, in vitro release behavior, free energy profiles of Syl across the niosome bilayer, hydrogen bonding interactions, antimicrobial properties, cytotoxicity, and in vivo evaluations. The physical appearance, size, and morphology assessment of free niosomes and Syl-loaded niosomes indicated stable and well-formed vesicular structures suitable for drug delivery. Transmission electron microscopy (TEM) analysis revealed spherical shapes with distinct sizes for each formulation, confirming uniform distribution. Dynamic light scattering (DLS) analysis confirmed the size distribution results with higher polydispersity index for Syl-loaded niosomes. The encapsulation efficiency of Syl in the niosomes was remarkable at approximately 91%, ensuring protection and controlled release of the drug. In vitro release studies showed a sustained release profile for Syl-loaded niosomes, enhancing therapeutic efficacy over time. Free energy profiles analysis identified energy barriers hindering Syl permeation through the niosome bilayer, emphasizing challenges in drug delivery system design. Hydrogen bonding interactions between Syl and niosome components contributed to energy barriers, impacting drug permeability. Antimicrobial assessments revealed significant differences in inhibitory effects against S. aureus and E. coli. Cytotoxicity evaluations demonstrated the superior tumor-killing potential of Syl-loaded niosomes compared to free Syl. In vivo studies indicated niosome formulations' safety profiles in terms of liver and kidney parameters compared to bulk Syl, showcasing potential for clinical applications. Overall, this research highlights the promising potential of Syl-loaded niosomes as effective drug delivery systems with enhanced stability, controlled release, and improved therapeutic outcomes.

Multiple-time scale integration method based on an interpolated potential energy surface for abinitio path integral molecular dynamics.

A new multiple-time scale integration method is presented that propagates abinitio path integral molecular dynamics (PIMD). This method uses a large time step to generate an approximate geometrical configuration whose energy and gradient are evaluated at the level of an abinitio method, and then, a more precise integration scheme, e.g., the Bulirsch-Stoer method or velocity Verlet integration with a smaller time step, is used to integrate from the previous step using the computationally efficient interpolated potential energy surface constructed from two consecutive points. This method makes the integration of PIMD more efficient and accurate compared with the velocity Verlet integration. A Nosé-Hoover chain thermostat combined with this new multiple-time scale method has good energy conservation even with a large time step, which is usually challenging in velocity Verlet integration for PIMD due to the very small chain mass when a large number of beads are used. The new method is used to calculate infrared spectra and free energy profiles to demonstrate its accuracy and capabilities.

Regioselective amination of 4-methylene-5,7-dinitroquinazoline: a mechanistic consideration on non-conventional N-H—π interactions between amine and ethylene moiety

ABSTRACT Pi (π) interactions originating from the N-H bond of amine and ethylene moiety have been explored on the mechanism of aromatic nucleophilic substitution (ArSN) of the nitro group of 4-methylene-5,7-dinitroquinazoline with methylamine in the gas phase and solvent media within DFT framework. The free energy profiles confirmed that the amination should take place at peri-position and calculations support the one-step mechanism through a transition state with no intermediate in the reaction route. Stabilisation of peri-transition state by intramolecular weak interaction akin to hydrogen bonding N-H—CH2 = C leads to the regioselective amination at peri-position of 4-methylene-5,7-dinitroquinazoline. The intramolecular hydrogen bond interactions at N-H—CH2 = C in the peri-transition state are strongly supported by a red shift in N-H stretching vibrations in simulated IR spectra and are confirmed by studying energetics of amination of structural/electronic analogues of 4-methylene-5,7-dinitroquinazoline. The present study established that ethylene plays an important role as an efficient H-bond acceptor in fixing the regioselectivity at peri-position in the amination of 4-methylene-5,7-dinitroquinazoline. This is the first-ever report for the exploration of the role of ethylene moiety as a hydrogen bond acceptor in mediating the regioselectivity of organic reactions.

Molecular dynamics in multidimensional space explains how mutations affect the association path of neomycin to a riboswitch

Riboswitches are messenger RNA (mRNA) fragments binding specific small molecules to regulate gene expression. A synthetic N1 riboswitch, inserted into yeast mRNA controls the translation of a reporter gene in response to neomycin. However, its regulatory activity is sensitive to single-point RNA mutations, even those distant from the neomycin binding site. While the association paths of neomycin to N1 and its variants remain unknown, recent fluorescence kinetic experiments indicate a two-step process driven by conformational selection. This raises the question of which step is affected by mutations. To address this, we performed all-atom two-dimensional replica-exchange molecular dynamics simulations for N1 and U14C, U14C[Formula: see text], U15A, and A17G mutants, ensuring extensive conformational sampling of both RNA and neomycin. The obtained neomycin association and binding paths, along with multidimensional free-energy profiles, revealed a two-step binding mechanism, consisting of conformational selection and induced fit. Neomycin binds to a preformed N1 conformation upon identifying a stable upper stem and U-turn motif in the riboswitch hairpin. However, the positioning of neomycin in the binding site occurs at different RNA-neomycin distances for each mutant, which may explain their different regulatory activities. The subsequent induced fit arises from the interactions of the neomycin's N3 amino group with RNA, causing the G9 backbone to rearrange. In the A17G mutant, the critical C6-A17/G17 stacking forms at a closer RNA-neomycin distance compared to N1. These findings together with estimated binding free energies coincide with experiments and elucidate why the A17G mutation decreases and U15A enhances N1 activity in response to neomycin.