Low-energy Conformations Research Articles

The adsorption of drugs on nanoplastics has severe biological impact

Micro- and nanoplastics can interact with various biologically active compounds forming aggregates of which the effects have yet to be understood. To this end, it is vital to characterize these aggregates of key compounds and micro- and nanoplastics. In this study, we examined the adsorption of the antibiotic tetracycline on four different nanoplastics, made of polyethylene (PE), polypropylene (PP), polystyrene (PS), and nylon 6,6 (N66) through chemical computation. Two separate approaches were employed to generate relevant conformations of the tetracycline-plastic complexes. In the first approach, we folded the plastic particle from individual polymer chains in the presence of the drug through multiple separate simulated annealing setups. In the second, more biased, approach, the neat plastic was pre-folded through simulated annealing, and the drug was placed at its surface in multiple orientations. The former approach was clearly superior to the other, obtaining lower energy conformations even with the antibiotic buried inside the plastic particle. Quantum chemical calculations on the structures revealed that the adsorption energies show a trend of decreasing affinity to the drug in the order of N66> PS> PP> PE. In vitro experiments on tetracycline-sensitive cell lines demonstrated that, in qualitative agreement with the calculations, the biological activity of tetracycline drops significantly in the presence of PS particles. Preliminary molecular dynamics simulations on two selected aggregates with each plastic served as first stability test of the aggregates under influence of temperature and in water. We found that all the selected cases persisted in water indicating that the aggregates may be stable also in more realistic environments. In summary, our data show that the interaction of micro- and nanoplastics with drugs can alter drug absorption, facilitate drug transport to new locations, and increase local antibiotic concentrations, potentially attenuating antibiotic effect and at the same time promoting antibiotic resistance.

Photochemical and Collision-Induced Cross-Linking in Stereochemically Distinct Scaffolds of Peptides and Nitrile Imines in Gas-Phase Ions.

Intramolecular cross-linking between peptides and nitrile-imine intermediates was studied in stereochemically distinct conjugates in which the reacting components were mounted on cis-1,2-cyclohexane and trans-1,4-cyclohexane scaffolds that we call 1,2-s-peptides and 1,4-s-peptides, respectively. The nitrile-imine intermediates were generated by N2 loss from 2,5-diaryltetrazole tags upon UV-photodissociation at 213 and 250 nm or by collision-induced dissociation, and further interrogated by CID and UVPD-MS3. Peptide fragment ion series originating from linear structures and macrocyclic cross-links were distinguished and used to quantify the cross-linking yields. The yields in MS2 varied between 27% for AAAG conjugates to 78% for GAAAK conjugates, depending on the peptide sequence. The CID-MS3 yields were in a 57-97% range, depending on the peptide sequence. Structures of 1,2-s-peptide and 1,4-s-peptide ions as well as several of their nitrile-imine intermediates and cross-links were investigated by high-resolution cyclic ion mobility in combination with Born-Oppenheimer molecular dynamics and density functional theory calculations. Matches between the experimental and calculated collision cross sections and ion relative Gibbs energies were used to assign peptide structures. Peptide conjugates C-terminated with Gly and Lys residues underwent cross-linking by the carboxyl group, as established by MS3 sequencing and corroborated by carboxyl blocking experiments that lowered the cross-linking yields. Peptide conjugates C-terminated with Arg also cross-linked via the side-chain guanidine group. A notable feature of the 1,4-s-peptide ions was the participation of low-energy twist-boat cyclohexane conformers that was enforced by strong hydrogen bonds between the peptide and nitrile imine.

Electronic Structures and Spectra of Donor-Acceptor Conjugated Oligomers.

Narrow band gap donor-acceptor conjugated polymers present excellent paradigms in photonics and optoelectronics due to their chemical tunability, correlated electronic structures, and tunable open-shell electronic configurations. However, rational design for enhancing the properties of these molecular systems remains challenging. In this study, we employed density functional theory (DFT) calculations to investigate prototypical narrow band gap donor-acceptor conjugated oligomers, consisting of alternating cyclopentadithiophene (CPDT) donors paired with benzothiadiazole (BT), benzoselenadiazole (BSe), benzobisthiadiazole (BBT), and thiadiazoloquinoxaline (TQ) acceptors. Analyses of structures, singlet-triplet gaps, and absorption spectra of oligomers with up to ten repeat units have shown that when incorporating the BT, BSe, and TQ acceptors, the backbone curvature resulted in spiral structures that were energetically favored over their linear counterparts, causing differences in the calculated circular dichroism spectra. Oligomers with BBT-based acceptors preferred, however, a linear geometry, consistent with an open-shell electronic structure. Calculated singlet-triplet splittings demonstrated the importance of long chains and specific structures for consistency with the experiment, while effects of the solvent were also quantified. Based on the predicted low-energy conformations, one-photon absorption spectra for the considered oligomers have shown that using the Tamm-Dancoff approximation within time-dependent DFT for the large systems offers good agreement with the first absorption maxima in measured experimental spectra, thus validating the method for large donor-acceptor oligomers. Natural transition orbital analyses provided insights into the excited-state characteristics. Two-photon absorption maxima were accurately predicted, but the cross-sections were overestimated or underestimated, as dependent on the level of theory employed, to be addressed in future work.

A novel application of hydrophilic interaction liquid chromatography for the identification of compounds with intramolecular hydrogen bonds

The incorporation of intramolecular hydrogen bonds (IMHB) into small molecules constitutes an interesting optimization strategy to afford potential drug candidates with enhanced solubility as well as permeability and consequently improved bioavailability (if metabolic stability is high). Common methods to assess IMHB rely on spectroscopic or diffraction techniques, which, however, have limited throughput when screening for hit compounds in early phases of drug discovery. Inspired by literature findings using supercritical fluid chromatography (SFC) as an indirect method for IMHB identification in a screening context, we aimed at developing a secondary chromatographic methodology taking advantage of commonly used HPLC-MS instrumentation. In this work, we explored hydrophilic interaction liquid chromatography (HILIC) and developed a method for discriminating compounds based on their hydrogen bonding features. By quantifying retention of different matched molecular pairs (MMP) and using information about their low energy conformations from quantum-mechanical calculations, we defined a hydrogen bonding-driven adsorption (kads) chromatographic parameter to assess a compound’s propensity to forming IMHB. In addition to the MMP analysis, we found that the kads parameter allows for the differentiation of analytes forming IMHB regardless of the comparison with control compounds.

Structural dissection of the CMP-pseudaminic acid synthetase, PseF.

Pseudaminic acid is a non-mammalian sugar found in the surface glycoconjugates of many bacteria, including several human pathogens, and is a virulence factor thought to facilitate immune evasion. The final step in the biosynthesis of the nucleotide activated form of the sugar, CMP-Pse5Ac7Ac is performed by a CMP-Pse5Ac7Ac synthetase (PseF). Here we present the biochemical and structural characterization of PseF from Aeromonas caviae (AcPseF), with AcPseF displaying metal-dependent activity over a broad pH and temperature range. Upon binding to CMP-Pse5Ac7Ac, AcPseF undergoes dynamic movements akin to other CMP-ulosonic acid synthetases. The enzyme clearly discriminates Pse5Ac7Ac from other ulosonic acids, through active site interactions with side-chain functional groups and by positioning the molecule in a hydrophobic pocket. Finally, we show that AcPseF binds the CMP-Pse5Ac7Ac side chain in the lowest energy conformation, a trend that we observed in the structures of other enzymes of this class.

Identification of novel brain penetrant GSK-3β inhibitors toward Alzheimer’s disease therapy by virtual screening, molecular docking, dynamic simulation, and MMPBSA analysis

One of the significant therapeutic targets for Alzheimer’s disease (AD) is Glycogen Synthase Kinase-3β (GSK-3β). Inhibition of GSK-3β can prevent hyperphosphorylation of tau, and thus prevent formation and accumulation of neurofibrillary tangles and neuropil threads that block intracellular transport, trigger unfolded protein response, and increase oxidative stress, cumulatively leading to neurodegeneration. In this study, we have performed structure-based virtual screening of two small-molecule libraries from ChemDiv CNS databases using AutoDock Vina to identify hit molecules based on their binding affinities compared to that of an established GSK-3β inhibitor, indirubin-3’-monoxime (IMO). Pharmacoinformatic screening on SwissADME and pkCSM servers enabled identification of lead molecules with favorable pharmacoinformatic properties for drug likeliness, including blood brain barrier (BBB) permeability. Further, molecular dynamic simulations identified six candidate lead molecules that show stable complex formation with GSK-3β in dynamic state under physiological conditions. Principal component analysis of the dynamic state was used to plot Free Energy Landscapes (FELs) of GSK-3β-ligand complexes. STRIDE secondary structure analysis of the lowest energy conformations identified from FEL plots, and binding free energy calculations by Molecular Mechanics Poisson-Boltzmann Surface Area ((ΔGbind )MM-PBSA) of the simulation trajectories led to the identification of two ligands as potential lead molecules having favorable free energy landscape profiles as well as significantly lower (ΔGbind )MM-PBSA in dynamic state compared to that of reference inhibitor IMO. Hence, this study identifies two novel brain penetrant GSK-3β inhibitors that are likely to have therapeutic potential against Alzheimer’s disease.

Infrared Spectrum of the Adduct 2-Chloro-2-hydroperoxybut-3-ene [(C2H3)CCl(CH3)OOH] of the Reaction between the Criegee Intermediate Methyl Vinyl Ketone Oxide [C2H3C(CH3)OO] and HCl.

Methyl vinyl ketone oxide (MVKO, C2H3C(CH3)OO) is an important Criegee intermediate produced from ozonolysis of isoprene, which is the most abundant nonmethane hydrocarbon emitted into the atmosphere. Reactions between Criegee intermediate and hydrogen chloride (HCl) are important because of their large rate coefficients. In this work, we photolyzed a mixture of (Z)-(CH2I)HC═C(CH3)I/HCl/O2 at 248 nm to produce MVKO to carry out the reaction MVKO + HCl and recorded infrared spectra of transient species with a step-scan Fourier-transform infrared absorption spectrometer. Eleven bands near 1415, 1381, 1350, 1249, 1178, 1118, 1103, 1065, 978, 931, and 895 cm-1 were observed and assigned to the hydrogen-transferred adduct (C2H3)CCl(CH3)OOH (2-chloro-2-hydroperoxybut-3-ene, CHPB) according to the predicted IR spectrum using the B3LYP/aug-cc-pVTZ method; the conformation could not be definitively determined. According to calculations, most low-energy conformers can interconvert, and the most stable conformers anti-trans-CHPB and syn-trans-CHPB might have the major contributions. Four bands near 1423, 1360, 1220, and 1080 cm-1 were observed and tentatively assigned to the OH-decomposition products (C2H3)CCl(CH3)O (1-chloro-1-methyl-2-propenyloxy, CMP); both trans-CMP and cis-CMP might contribute. Unlike in the case of CH2OO + HCl and CH3CHOO + HCl, in which secondary reactions of the OH-decomposition products reacted readily with O2 to produce the dehydrated products HC(O)Cl and CH3C(O)Cl, respectively, the secondary reaction of CMP with O2 was not observed because there is no feasible H atom in CMP for O2 to abstract.

Expanding Automated Multiconformer Ligand Modeling to Macrocycles and Fragments.

Small molecule ligands exhibit a diverse range of conformations in solution. Upon binding to a target protein, this conformational diversity is generally reduced. However, ligands can retain some degree of conformational flexibility even when bound to a receptor. In the Protein Data Bank (PDB), a small number of ligands have been modeled with distinct alternative conformations that are supported by X-ray crystallography density maps. However, the vast majority of structural models are fit to a single ligand conformation, potentially ignoring the underlying conformational heterogeneity present in the sample. We previously developed qFit-ligand to sample diverse ligand conformations and to select a parsimonious ensemble consistent with the density. While this approach indicated that many ligands populate alternative conformations, limitations in our sampling procedures often resulted in non-physical conformations and could not model complex ligands like macrocycles. Here, we introduce several improvements to qFit-ligand, including the use of routines within RDKit for stochastic conformational sampling. This new sampling method greatly enriches low energy conformations of small molecules and macrocycles. We further extended qFit-ligand to identify alternative conformations in PanDDA-modified density maps from high throughput X-ray fragment screening experiments. The new version of qFit-ligand improves fit to electron density and reduces torsional strain relative to deposited single conformer models and our previous version of qFit-ligand. These advances enhance the analysis of residual conformational heterogeneity present in ligand-bound structures, which can provide important insights for the rational design of therapeutic agents.

Mechanisms of peptide agonist dissociation and deactivation of adhesion G-protein-coupled receptors.

Adhesion G protein-coupled receptors (ADGRs) belong to Class B2 of GPCRs and are involved in a wide array of important physiological processes. ADGRs contain a GPCR autoproteolysis-inducing (GAIN) domain that is proximal to the receptor N-terminus and undergoes autoproteolysis during biosynthesis to generate two fragments: the N-terminal fragment (NTF) and C-terminal fragment (CTF). Dissociation of NTF reveals a tethered agonist to activate CTF of ADGRs for G protein signaling. Synthetic peptides that mimic the tethered agonist can also activate the ADGRs. However, mechanisms of peptide agonist dissociation and deactivation of ADGRs remain poorly understood. In this study, we have performed all-atom enhanced sampling simulations using a novel Protein-Protein Interaction-Gaussian accelerated Molecular Dynamics (PPI-GaMD) method on the ADGRG2-IP15 and ADGRG1-P7 complexes. The PPI-GaMD simulations captured dissociation of the IP15 and P7 peptide agonists from their target receptors. We were able to identify important low-energy conformations of ADGRG2 and ADGRG1 in the active, intermediate, and inactive states, as well as exploring different states of the peptide agonists IP15 and P7 during dissociation. Therefore, our PPI-GaMD simulations have revealed dynamic mechanisms of peptide agonist dissociation and deactivation of ADGRG1 and ADGRG2, which will facilitate rational design of peptide regulators of the two receptors and other ADGRs.

H-atom abstraction reactions of C1-C4 alkanes by ketenyl radical: Kinetic investigation and analysis

The kinetics of 16 H-atom abstraction reactions of C1-C4 alkanes by ketenyl radical (HCCO) are studied to enhance the existing combustion mechanisms. Firstly, the obtain the lowest-energy conformer, vibration frequencies and single point energies for the reactants, prepolymers, transition states, and products are determined using the CCSD(T)/cc-pVQZ//B3LYP/6-311G+(d,p) theory level. Subsequently, temperature-dependent reaction rate constants for the major channels are predicted by the TST method. Finally, the H-atom abstraction reactions are validated under multiple temperature conditions, and the effects of the reactions on auto-ignition process and CO emission characteristics of n-heptane/NG mixture are researched. The results indicate that the H-atom abstraction reactions by Cα-atom play a significant role for the diesel/NG combustion. To combine the reaction pathways and rate constants into the existing kinetic mechanisms resulted in more accurate predictions of the ignition delay time. Furthermore, these kinetics data are valuable for revealing the CO emissions of the dual-fuel engines.

Computational Study on the Conformational Flexibility-Mediated Intramolecular Oxidative Spirocyclization of Procyanidin B4.

Procyanidins, found widely in foods and beverages, are prone to oxidation, yet the chemical structures of their oxidation products and the mechanisms involved remain unclear. Herein, we report that the conformation of procyanidin B4 influences its oxidation products and their stereochemistry. Eight spirocyclized oxidation products were obtained from procyanidin B4 and classified as S- or R-forms based on the configuration of the spiro carbons. The ratios of S- and R-forms derived from the compact and extended rotamers of procyanidin B4, respectively, varied with the solvent. DFT calculations suggested that the four lowest-energy conformers of procyanidin B4 are diverged by interflavan bond rotation and heterocyclic ring inversion. Conformations with an axial-oriented B-ring were estimated as reactive conformations showing proximity between reaction sites on the B- and D-rings. Moreover, the extended rotamer bearing the axially oriented B-ring showed greater stabilization by noncovalent interactions (NCIs), such as OH-π interactions, compared to the counterpart of the compact rotamer. This NCI-based stabilization accounts for a higher production of the R-form despite the predominant presence of the compact rotamer in H2O. These findings highlight the conformational effects that bias the stereoselectivity of oxidative spirocyclization in procyanidin B4, advancing our understanding of procyanidin oxidation mechanisms and product stereochemistry.

A comprehensive analysis of the spectroscopic and drug-like properties of dimethyl 5-hydroxybenzene-1,3-dicarboxylate: insights from DFT and MD simulations

ABSTRACT An in-depth investigation was undertaken to explore the ground-state molecular and electronic structure of Dimethyl 5-hydroxybenzene-1,3-dicarboxylate (D5HD). The potential energy scan of D5HD at various dihedral angles identified the lowest energy conformer, which was re-optimised at the higher basis set. The recorded FT-Raman and FT-IR spectra were compared with corresponding theoretical spectra, demonstrating a high degree of coherence. A comprehensive Natural Bond Orbital analysis was done for D5HD to gain insights for the overall electronic distribution within the molecule. Additionally, nonlinear optical (NLO) properties along with various DFT-based descriptors were calculated. Molecular docking studies against human acetylcholinesterase enzyme (PDB 2JF0), D5HD displayed good binding affinity. The study revealed that hydrogen bonded as well as van der Waals interactions exist between D5HD and different residues in the binding cavity of the target protein. The 100 ns MD simulation confirms the conformational stability of protein (2JF0)–ligand (D5HD) complex. The binding energy and residue decomposition analysis performed using MM-GBSA approach showed positive results with binding energy of −17.76 kcal/mol. Furthermore, D5HD effectively passed all pharmacokinetic filters, establishing its potential as a promising candidate in the quest for novel acetylcholinesterase inhibitors. Highlights Systematic exploration of the structure and spectroscopic properties of D5HD. Molecular docking between D5HD and acetylcholinesterase (PDB 2JF0) reveals the establishment of hydrogen bonds and Van der Waals interactions. D5HD successfully complies with all relevant pharmacokinetic filters. MD simulation analysis on 2JF0–D5HD complex confirms the conformational stability. Energy and residue decomposition based on MMGBSA approach.

EC-Conf: A ultra-fast diffusion model for molecular conformation generation with equivariant consistency

Despite recent advancement in 3D molecule conformation generation driven by diffusion models, its high computational cost in iterative diffusion/denoising process limits its application. Here, an equivariant consistency model (EC-Conf) was proposed as a fast diffusion method for low-energy conformation generation. In EC-Conf, a modified SE (3)-equivariant transformer model was directly used to encode the Cartesian molecular conformations and a highly efficient consistency diffusion process was carried out to generate molecular conformations. It was demonstrated that, with only one sampling step, it can already achieve comparable quality to other diffusion-based models running with thousands denoising steps. Its performance can be further improved with a few more sampling iterations. The performance of EC-Conf is evaluated on both GEOM-QM9 and GEOM-Drugs sets. Our results demonstrate that the efficiency of EC-Conf for learning the distribution of low energy molecular conformation is at least two magnitudes higher than current SOTA diffusion models and could potentially become a useful tool for conformation generation and sampling.Scientific ContributionsIn this work, we proposed an equivariant consistency model that significantly improves the efficiency of conformation generation in diffusion-based models while maintaining high structural quality. This method serves as a general framework and can be further extended to more complex structure generation and prediction tasks, including those involving proteins, in future steps.

Bis(amino acidato)copper(II) compounds in blood plasma: a review of computed structural properties and amino acid affinities for Cu2+ informing further pharmacological research.

Neutral bis(amino acidato)copper(II) [Cu(aa)2] coordination compounds are the physiological species of copper(II) amino acid compounds in blood plasma taking the form of bis(l-histidinato)copper(II) and mixed ternary copper(II)-l-histidine complexes, preferably with l-glutamine, l-threonine, l-asparagine, and l-cysteine. These amino acids have three functional groups that can bind metal ions: the common α-amino and carboxylate groups and a side-chain polar group. In Cu(aa)2, two coordinating groups per amino acid bind to copper(II) in-plane, while the third group can bind apically, which yields many possibilities for axial and planar bonds, that is, for bidentate and tridentate binding. So far, the experimental studies of physiological Cu(aa)2 compounds in solutions have not specified their complete geometries. This paper provides a brief review of my group's research on structural properties of physiological Cu(aa)2 calculated using the density functional theory (DFT) to locate low-energy conformers that can coexist in aqueous solutions. These DFT investigations have revealed high conformational flexibility of ternary Cu(aa)2 compounds for tridentate or bidentate chelation, which may explain copper(II) exchange reactions in the plasma and inform the development of small multifunctional copper(II)-binding drugs with several possible copper(II)-binding groups. Furthermore, our prediction of metal ion affinities for Cu2+ binding with amino-acid ligands in low-energy conformers with different coordination modes of five physiological Cu(aa)2 in aqueous solution supports the findings of their abundance in human plasma obtained with chemical speciation modelling.

Design and performance of sulfur and selenium-substituted triarylboron D3-A TADF emitters for OLED applications

This study presents the design, synthesis, and comprehensive theoretical and photophysical analysis of two new D3-A type thermally activated delayed fluorescence (TADF) emitters for organic light-emitting diode (OLED) applications. Utilizing a triarylboron core as the electron-accepting group and phenothiazine (PTZ) or phenoselenazine (PSZ) as electron-donating units, the molecules BTP-S and BTP-Se were developed. The D3-A structure supports the separation of frontier molecular orbitals (FMOs), leading to minimized singlet-triplet energy gaps (ΔEST), which are crucial for the TADF mechanism. Density functional theory (DFT) calculations presented that BTP-S and BTP-Se exhibit band gaps (Eg) of 2.52 and 3.23 eV, respectively, with BTP-S showing an ΔEST value as low as 0.007 eV for the S1-T1 transition at the lowest energy conformation. Photophysical studies revealed high photoluminescence quantum yields (PLQYs) for both compounds, with BTP-S achieving up to 85 % in mCP films and BTP-Se up to 59 %. In vacuum-processed OLEDs, BTP-S achieved a maximum external quantum efficiency (EQE) of 25.3 %, a current efficiency (ηc) of 195.8 cd/A, and a maximum luminance (Lmax) of 17356 cd/m2, while BTP-Se reached an EQE of 7.5 %, an ηc of 132.19 cd/A, and an Lmax of 16826 cd/m2 likely limited by the contributions of a folded-donor conformer enabled by the Se substitution. These findings underscore the impact of donor unit selection and conformation on the TADF characteristics, and provide valuable insights for designing high-performance OLED materials.

Experiments and Calculation on New N,N-bis-Tetrahydroacridines.

Tetrahydroacridines arouse particular interest due to the potential possibilities of application in the medical field and protection against corrosion. Bis-tetrahydroacridines were newly synthesized by Pfitzinger condensation of 5,5'-(ethane-1,2-diyl) diindoline-2,3-dione with several cyclanones. NMR, MS, and FT-IR were used to prove their molecular structure. In addition, a computer-aided study was performed for the lowest energy conformers of each structure, in vacuum conditions, at ground state using DFT models to assess their electronic properties. UV-Vis and voltammetric methods (cyclic voltammetry, differential pulse voltammetry, and rotating disk electrode voltammetry) were used to investigate their optical and electrochemical properties. The results obtained for these π-conjugated heteroaromatic compounds lead to the conclusion that they have real potential in applications in different fields such as pharmaceuticals and especially as corrosion inhibitors.

Novel Insecticidal Pyridylhydrazono Derivatives Identified via Scaffold Hopping and Conformation Regulation Strategies.

Insect transient receptor potential vanilloid (TRPV) channels are critical targets for insecticides. In this study, various scaffold-hopping strategies were employed in the rational design of pyridylhydrazono derivatives as potential insect TRPV channels modulators. Insecticidal bioassay demonstrated that the initial target compounds exhibited lower insecticidal activity compared to pymetrozine, with the optimal compound B3 exhibiting a mortality rate of 53.3 % against Aphis craccivora at 400 mg L-1. Conformation analysis indicated that the high energy barrier required for the transition from the lowest-energy conformation to the active conformation may be a key factor contributing to the reduced insecticidal activities of the target compounds. Further structural optimizations aimed at reducing this energy barrier through binding mode-based conformation regulation led to the identification of optimal target 4-(3'-pyridylhydrazono)pyrazol-5-one derivatives C1 and C2. These compounds exhibited reduced transition energy barriers and improved insecticidal activity, with moderate mortality rate of 66.3 % and 75.7 % against A. craccivora at 400 mg L-1, respectively. These findings provide valuable insights for future research on the discovery of insect TRPV modulators and have significant implications for the development of more effective agricultural insecticides.

AlphaFold predictions of fold-switched conformations are driven by structure memorization

Recent work suggests that AlphaFold (AF)–a deep learning-based model that can accurately infer protein structure from sequence–may discern important features of folded protein energy landscapes, defined by the diversity and frequency of different conformations in the folded state. Here, we test the limits of its predictive power on fold-switching proteins, which assume two structures with regions of distinct secondary and/or tertiary structure. We find that (1) AF is a weak predictor of fold switching and (2) some of its successes result from memorization of training-set structures rather than learned protein energetics. Combining >280,000 models from several implementations of AF2 and AF3, a 35% success rate was achieved for fold switchers likely in AF’s training sets. AF2’s confidence metrics selected against models consistent with experimentally determined fold-switching structures and failed to discriminate between low and high energy conformations. Further, AF captured only one out of seven experimentally confirmed fold switchers outside of its training sets despite extensive sampling of an additional ~280,000 models. Several observations indicate that AF2 has memorized structural information during training, and AF3 misassigns coevolutionary restraints. These limitations constrain the scope of successful predictions, highlighting the need for physically based methods that readily predict multiple protein conformations.

Characterization and conformational analysis of a novel hydrazonoyltetrazole: Crystallographic and theoretical investigations of an unexpected reaction product

A new hydrazonoyltetrazole was isolated as a byproduct from a previously reported reaction, and characterized by 1H and 13C NMR spectroscopy, infrared spectroscopy, single-crystal X-ray diffraction, and high-resolution mass spectroscopy. Intermolecular interactions in the crystal state were examined by Hirshfeld surface analysis (HSA), 2D fingerprint plots, and noncovalent interaction analysis (NCI). Geometry optimization in the gaseous phase was executed at the ωB97X-D4/def2-TZVPP level of theory, and subsequent maps of molecular electrostatic potential (MEP) were generated and analyzed. The calculated MEP was in good agreement with the HSA and NCI results. Conformational analysis was performed using a CREST/CENSO protocol. Two low-energy solution conformers were obtained. To confirm the presence of the second conformer, 1H NMR spectra were calculated, the transition state connecting the two conformers was found, and their energies were obtained with the DLPNOCCSD(T) scheme. A barrier of only 4 kcal mol-1 was found between them, and their MEP maps were also created. The dispersion energies between fragments of the three structures were calculated using the local energy decomposition (LED) scheme.

Multiple Parameter Replica Exchange Gaussian Accelerated Molecular Dynamics for Enhanced Sampling and Free Energy Calculation of Biomolecular Systems.

This study introduces a novel method named multiple parameter replica exchange Gaussian accelerated molecular dynamics (MP-Rex-GaMD), building on the Gaussian accelerated molecular dynamics (GaMD) algorithm. GaMD enhances sampling and retrieves free energy information for biomolecular systems by adding a harmonic boost potential to smooth the potential energy surface without the need for predefined reaction coordinates. Our innovative approach advances the acceleration power and energetic reweighting accuracy of GaMD by incorporating a replica exchange algorithm that enables the exchange of multiple parameters, including the GaMD boost parameters of force constant and energy threshold, as well as temperature. Applying MP-Rex-GaMD to the three model systems of dialanine, chignolin, and HIV protease, we demonstrate its superior capability over conventional molecular dynamics and GaMD simulations in exploring protein conformations and effectively navigating various biomolecular states across energy barriers. MP-Rex-GaMD allows users to accurately map free energy landscapes through energetic reweighting, capturing the ensemble of biomolecular states from low-energy conformations to rare high-energy transitions within practical computational time scales.