Complex Binding Mechanism Research Articles

Structural and Functional Characterization of a Fish Type I Subgroup d IFN Reveals Its Binding to Receptors.

Teleost fish type I IFNs and the associated receptors from the cytokine receptor family B (CRFB) are characterized by remarkable diversity and complexity. How the fish type I IFNs bind to their receptors is still not fully understood. In this study, we demonstrate that CRFB1 and CRFB5 constitute the receptor pair through which type I subgroup d IFN (IFNd) from large yellow croaker, Larimichthys crocea, activates the conserved JAK-STAT signaling pathway as a part of the antiviral response. Our data suggest that L. crocea IFNd (LcIFNd) has a higher binding affinity with L. crocea CRFB5 (LcCRFB5) than with LcCRFB1. Furthermore, we report the crystal structure of LcIFNd at a 1.49-Å resolution and construct structural models of LcIFNd in binary complexes with predicted structures of extracellular regions of LcCRFB1 and LcCRFB5, respectively. Despite striking similarities in overall architectures of LcIFNd and its ortholog human IFN-ω, the receptor binding patterns between LcIFNd and its receptors show that teleost and mammalian type I IFNs may have differentially selected helices that bind to their homologous receptors. Correspondingly, key residues mediating binding of LcIFNd to LcCRFB1 and LcCRFB5 are largely distinct from the receptor-interacting residues in other fish and mammalian type I IFNs. Our findings reveal a ligand/receptor complex binding mechanism of IFNd in teleost fish, thus providing new insights into the function and evolution of type I IFNs.

Determining epitope specificity of T-cell receptors with transformers.

T-cell receptors (TCRs) on T cells recognize and bind to epitopes presented by the major histocompatibility complex in case of an infection or cancer. However, the high diversity of TCRs, as well as their unique and complex binding mechanisms underlying epitope recognition, make it difficult to predict the binding between TCRs and epitopes. Here, we present the utility of transformers, a deep learning strategy that incorporates an attention mechanism that learns the informative features, and show that these models pre-trained on a large set of protein sequences outperform current strategies. We compared three pre-trained auto-encoder transformer models (ProtBERT, ProtAlbert, and ProtElectra) and one pre-trained auto-regressive transformer model (ProtXLNet) to predict the binding specificity of TCRs to 25 epitopes from the VDJdb database (human and murine). Two additional modifications were performed to incorporate gene usage of the TCRs in the four transformer models. Of all 12 transformer implementations (four models with three different modifications), a modified version of the ProtXLNet model could predict TCR-epitope pairs with the highest accuracy (weighted F1 score 0.55 simultaneously considering all 25 epitopes). The modification included additional features representing the gene names for the TCRs. We also showed that the basic implementation of transformers outperformed the previously available methods, i.e. TCRGP, TCRdist, and DeepTCR, developed for the same biological problem, especially for the hard-to-classify labels. We show that the proficiency of transformers in attention learning can be made operational in a complex biological setting like TCR binding prediction. Further ingenuity in utilizing the full potential of transformers, either through attention head visualization or introducing additional features, can extend T-cell research avenues. Data and code are available on https://github.com/InduKhatri/tcrformer.

Elucidation of binding mechanism, affinity, and complex structure between mWT1 tumor-associated antigen peptide and HLA-A*24:02.

We have applied our advanced computational and experimental methodologies to investigate the complex structure and binding mechanism of a modified Wilms' Tumor 1 (mWT1) protein epitope to the understudied Asian-dominant allele HLA-A*24:02 (HLA-A24) in aqueous solution. We have applied our developed multicanonical molecular dynamics (McMD)-based dynamic docking method to analyze the binding pathway and mechanism, which we verified by comparing the highest probability structures from simulation with our experimentally solved x-ray crystal structure. Subsequent path sampling MD simulations elucidated the atomic details of the binding process and indicated that first an encounter complex is formed between the N-terminal's positive charge of the 9-residue mWT1 fragment peptide and a cluster of negative residues on the surface of HLA-A24, with the major histocompatibility complex (MHC) molecule preferring a predominantly closed conformation. The peptide first binds to this closed MHC conformation, forming an encounter complex, after which the binding site opens due to increased entropy of the binding site, allowing the peptide to bind to form the native complex structure. Further sequence and structure analyses also suggest that although the peptide loading complex would help with stabilizing the MHC molecule, the binding depends in a large part on the intrinsic affinity between the MHC molecule and the antigen peptide. Finally, our computational tools and analyses can be of great benefit to study the binding mechanism of different MHC types to their antigens, where it could also be useful in the development of higher affinity variant peptides and for personalized medicine.

Charge transfer interaction between 2, 3-Diamino-5-bromopyridine and 2, 4-Dinitrophenol: Synthesis, spectroscopic characterization, DNA binding analysis, and density functional studies

The e-donor 2, 3-Diamino-5-bromopyridine (5BPY) and the e-acceptor 2, 4-Dinitrophenol (DNP) were both involved in the synthesis of a new charge transfer complex, which was then characterized experimentally as well as theoretically. Several spectroscopic techniques were used to analyze the produced solid compound. 1H NMR, FT-IR, P-XRD and SEM-EDX analyses all confirmed the presence of charge and proton transfers in the resulting complex. Analysis of the complex DNA binding ability was carried out using electron absorption spectroscopy; the CT complex binding mechanism was determined to be intercalative, and the intrinsic binding constant (Kb) value was determined to be 7.2 × 106 M−1. To corroborate the results of the experiments, theoretical calculations were performed using DFT with a basis set of CAM-B3LYP/6-31 G (d, p). In accordance with the experimental findings, we calculated and analyzed the molecular electrostatic potential maps (MEPs), geometrical parameters and Mullikan atomic charges. The presence of an H-bond also affects the stability of the complex, in addition to e-transfer. The experimental findings are in agreement with the DFT calculations.

Selenium Status: Its Interactions with Dietary Mercury Exposure and Implications in Human Health.

Selenium is an essential trace element in humans and animals and its role in selenoprotein and enzyme antioxidant activity is well documented. Food is the principal source of selenium, and it is important that selenium status in the body is adequately maintained for physiological functions. There has been increasing attention on the role of selenium in mitigating the toxic effects of mercury exposure from dietary intake in humans. In contrast, mercury is a neurotoxin, and its continuous exposure can cause adverse health effects in humans. The interactions of selenium and mercury are multi-factorial and involve complex binding mechanisms between these elements at a molecular level. Further insights and understanding in this area may help to evaluate the health implications of dietary mercury exposure and selenium status. This review aims to summarise current information on the interplay of the interactions between selenium and mercury in the body and the protective effect of selenium on at-risk groups in a population who may experience long-term mercury exposure.

Synthesis, spectroscopic characterization, DNA binding and DFT/PCM calculations of new Hydrogen-bonded charge transfer complex between 4-dimethylaminopyridine and Chloranilic acid

A charge transfer hydrogen bonded complex between the electron donor 4-dimethylaminopyridine (4-DMAP) and the electron acceptor Chloranilic acid (CLA) has been synthesized. The complex was studied experimentally and theoretically. A chemical reaction developed due to strong H-bonding, followed by proton transfer from the CLA to the 4-DMAP in acetonitrile medium (ACN). The straight line approach was employed to verify the 1:1 M ratio of the synthesised CT complex. The Benesi-Hildebrand equation was employed to compute the association constant (KCT), molar absorptivity (εCT), and other physical parameters. The synthesized complex was characterized spectroscopically. The presence of proton and charge transfers in the resultant complex was supported by 1H NMR, FT-IR and SEM-EDX investigations. The complex DNA binding ability of complex was investigated using electron absorption spectroscopy to reveal that the CT complex binding mechanism is intercalative and the intrinsic binding constant (Kb) value is 6.7 × 104 M−1. Theoretical calculations using DFT at the basis set CAM-B3LYP/6–31 G (d, p) were used to validate the experimental outcomes. The Mullikan atomic charges, geometrical parameters, and molecular electrostatic potential maps (MEPs) were computed and interpreted in conformity with the experimental results. In addition to e-transfer, the presence of an H-bond influences complex stability. There is good agreement between experimental results and DFT calculations.

The Charge Transfer Complex between 2, 3-diamino-5-bromopyridine and Chloranilic acid: Preparation, Spectroscopic Characterization, DNA binding, and DFT/PCM analysis

A charge transfer hydrogen bonded complex was prepared and experimentally explored in an acetonitrile (ACN) medium between the proton acceptor (electron donor) 2, 3-Diamino-5-bromopyridine and the proton donor (electron acceptor) chloranilic acid. The stoichiometry of the charge transfer complex is 1:1. The Benesi-Hildebrand equation is used to calculate the molar absorptivity (εCT), association constant (KCT) and other spectroscopic physical characteristics. The solid compound was synthesized and studied using several spectroscopic methods. The presence of charge and proton transfers in the resultant complex was supported by 1H NMR, FT-IR and SEM-EDX investigations. The complex DNA binding ability was investigated using electron absorption spectroscopy, and the CT complex binding mechanism is intercalative. The intrinsic binding constant (Kb) value is 5.2 × 106M−1. The good binding affinity of the CT complex makes it potentially suitable for usage as a pharmaceutical in the future. Molecular docking calculations have been performed between CT complex and DNA (ID = 1BNA) to study the CT-DNA interaction theoretically. To corroborate the experimental findings, calculations based on DFT were carried out in the gas and PCM analysis where the existence of charge and hydrogen transfers. Finally, good agreement between experimental and theoretical computations was observed confirming that the basis set used is appropriate for the system under examination.

Uncovering the Relationship between Tissue-Specific TF-DNA Binding and Chromatin Features through a Transformer-Based Model.

Chromatin features can reveal tissue-specific TF-DNA binding, which leads to a better understanding of many critical physiological processes. Accurately identifying TF-DNA bindings and constructing their relationships with chromatin features is a long-standing goal in the bioinformatic field. However, this has remained elusive due to the complex binding mechanisms and heterogeneity among inputs. Here, we have developed the GHTNet (General Hybrid Transformer Network), a transformer-based model to predict TF-DNA binding specificity. The GHTNet decodes the relationship between tissue-specific TF-DNA binding and chromatin features via a specific input scheme of alternative inputs and reveals important gene regions and tissue-specific motifs. Our experiments show that the GHTNet has excellent performance, achieving about a 5% absolute improvement over existing methods. The TF-DNA binding mechanism analysis shows that the importance of TF-DNA binding features varies across tissues. The best predictor is based on the DNA sequence, followed by epigenomics and shape. In addition, cross-species studies address the limited data, thus providing new ideas in this case. Moreover, the GHTNet is applied to interpret the relationship among TFs, chromatin features, and diseases associated with AD46 tissue. This paper demonstrates that the GHTNet is an accurate and robust framework for deciphering tissue-specific TF-DNA binding and interpreting non-coding regions.

Competition in drug binding and … the race to equilibrium.

Binding kinetics has become a popular topic in pharmacology due to its potential contribution to the selectivity and duration of drug action. Yet, the overall kinetic aspects of complex binding mechanisms are still merely described in terms of elaborate algebraic equations. Interestingly, it has been recommended some 10 years ago to examine such mechanisms in terms of binding fluxes instead of the conventional rate constants. Alike the velocity of product formation in enzymology, those fluxes refer to the velocity by which one target species converts into another one. Novel binding flux-based approaches are utilized to get a better visual insight into the "competition" between two drugs/ligands for a single target as well as between induced fit- and conformational selection pathways for a single ligand within a thermodynamic cycle. The present data were obtained by differential equation-based simulations. Early on, the ligand-binding steps "race" to equilibrium (i.e., when their forward and reverse fluxes are equal) at their individual pace. The overall/global equilibrium is only reached later on. For the competition association assays, this parting might produce a transient "overshoot" of one of the bound target species. A similar overshoot may also show up within a thermodynamic cycle and, at first glance, suggest that the induced fit pathway dominates. Yet, present findings show that under certain circumstances, it could rather be the other way round. Novel binding flux-based approaches offer visually attractive insights into crucial aspects of "complex" binding mechanisms under non-equilibrium conditions.

Discovery of Complex Binding and Reaction Mechanisms from Ternary Gases in Rare Earth Metal-Organic Frameworks.

Understanding the selectivity of metal-organic frameworks (MOFs) to complex acid gas streams will enable their use in industrial applications. Herein, ab initio molecular dynamic simulations (AIMD) were used to simulate ternary gas mixtures (H2 O-NO2 -SO2 )in rare earth 2,5-dihydroxyterephthalic acid (RE-DOBDC) MOFs. Stronger H2 O gas-metal binding arose from thermal vibrations in the MOF sterically hindering access of SO2 and NO2 molecules to the metal sites. Gas-gas and gas-linker interactions within the MOF framework resulted in the formation of multiple secondary gas species including HONO, HNO2 , NOSO, and HNO3 - . Four gas adsorption sites were identified along with a new de-protonation reaction mechanism not observable through experiment. This study not only provides valuable information on competitive gas binding energies in the MOF, it also provides important chemical insights into transient chemical reactions and mechanisms.

A novel peptide-based fluorescent probe for highly selective detection of mercury (II) ions in real water samples and living cells based on aggregation-induced emission effect

A new fluorescent probe TPE-GHK was synthesized containing a tetrastyrene (TPE) derivative as fluorophore and classical tripeptide (Gly-His-Lys-NH2) as a receptor based on the aggregation-induced emission (AIE) mechanism. TPE-GHK displayed high selectivity and rapid fluorescent "turn-on" response to Hg2+ among other competitive metal ions. The 2:1 complex binding mechanism of TPE-GHK toward Hg2+ was verified by fluorometric titration, Job's plots, and ESI-HRMS spectra. The fluorescent emission showed a good linear response in the range of 0-1.0 μM with the low detection limit of 28.6 nM. Meanwhile, TPE-GHK exhibited the excellent biocompatibility and low toxicity and was successfully applied in monitoring Hg2+ in living CAKI 2 cells, which demonstrated its potential application in environment and biological science. More importantly, TPE-GHK could be used to detect Hg2+ in two real water samples and also was successfully designed as test strips.

Towards an Extrinsic Formalization of Featherweight Java in Agda

Featherweight Java is one of the most popular calculi which specify object-oriented programming features. It has been used as the basis for investigating novel language functionalities, as well as to specify and understand the formal properties of existing features for languages in this paradigm. However, when considering mechanized formalization, it is hard to find an implementation for languages with complex structures and binding mechanisms as Featherweight Java. In this paper we formalize Featherweight Java, implementing the static and dynamic semantics in Agda, and proving the main safety properties for this calculus.

Recent Advancements for the Recognization of Nitroaromatic Explosives Using Calixarene Based Fluorescent Probes.

In this era, explosives are easily available compared to the early days. Thus, more effective detection of explosives has become the main concern of homeland security. In the past decades, a large number of sensing materials have been developed for the detection of explosives in solid, vapor, and solution states through fluorescence methods. In recent years, great efforts have been devoted to developing new fluorescent materials with various sensing mechanisms for detecting explosives in order to achieve super-sensitivity, ultra-selectivity, as well as fast response time. Modified calixarenes have high potentials to detect nitroaromatic compounds (NACs) due to their favorable structural properties. It summarizes the detection of NACs by the modified calixarene system formed by the complex. Various methodologies responsible for complex formation and binding mechanisms (PET, FRET, EE, etc.) are the centerpiece of this review. Finally, conclusions and future outlook are presented and discussed.

Unraveling binding mechanism and kinetics of macrocyclic Gαq protein inhibitors

G proteins represent intracellular switches that transduce signals relayed from G protein-coupled receptors. The structurally related macrocyclic depsipeptides FR900359 (FR) and YM-254890 (YM) are potent, selective inhibitors of the Gαq protein family. We recently discovered that radiolabeled FR and YM display strongly divergent residence times, which translates into significantly longer antiasthmatic effects of FR. The present study is aimed at investigating the molecular basis for this observed disparity. Based on docking studies, we mutated amino acid residues of the Gαq protein predicted to interact with FR or YM, and recombinantly expressed the mutated Gαq proteins in cells in which the native Gαq proteins had been knocked out by CRISPR-Cas9. Both radioligands showed similar association kinetics, and their binding followed a conformational selection mechanism, which was rationalized by molecular dynamics simulation studies. Several mutations of amino acid residues near the putative binding site of the “lipophilic anchors” of FR, especially those predicted to interact with the isopropyl group present in FR but not in YM, led to dramatically accelerated dissociation kinetics. Our data indicate that the long residence time of FR depends on lipophilic interactions within its binding site. The observed structure-kinetic relationships point to a complex binding mechanism of FR, which likely involves snap-lock- or dowel-like conformational changes of either ligand or protein, or both. These experimental data will be useful for the design of compounds with a desired residence time, a parameter that has now been recognized to be of utmost importance in drug development.

Discovery of Small Molecule Modulators targeting Secretin Receptors

Class B G protein-coupled receptors (GPCRs) are a highly important group of therapeutic targets with strong physiologic implications in cardiovascular, metabolic, or gastrointestinal systems. Endogenously activated by moderate-length peptide hormones, class B GPCRs entertain a complex binding mechanism to induce stimulatory signaling processes. The secretin receptor (SCTR) is a prototypical member of this receptor family, having clinical relevance in multiple diseases including heart failure, digestive disorders, diabetes and obesity. [1-4] However, its endogenous ligand secretin and peptide-based analogs are the only ligands known to interact with SCTRs, likely due to challenges in class B GPCR drug discovery [5]. To find orally available therapeutic leads targeting this “undruggable” receptor, we focused on identifying positive allosteric modulators (PAMs), which act via structurally distinct binding pockets and potentiate hormonal signaling of native agonists. Thus, we developed a testing funnel tailored to discover SCTR small molecule modulators. After screening a 12,000-compound library with a unique primary assay that used a mixture of stimulators, we identified three structurally related scaffolds (A-C) enhancing peptidic SCTR activation [5]. Applying a comprehensive toolbox with binding and functional assays, we studied structure-activity relationships of a well-considered set of commercially available analogs incorporating these three scaffolds. Our investigations afforded an optimized analog B2, which showed substantial positive cooperativity to low-potent peptide agonists and significantly decelerated secretin dissociation. Due to the innate electrophilicity of scaffold set A, we recognized a structural similarity to PAMs acting on glucagon-like peptide-1 receptor (GLP-1R), a prominent member of class B GPCR family. We further demonstrated that a subset of SCTR PAMs was able to augment not only SCTR signaling but also GLP-1R functions. A recent study described that dual activation of SCTRs and GLP-1Rs was benefiting glucose homeostasis and appetite regulation while avoiding proliferative effects on pancreas mass [6]. Determining cooperativity between PAMs and selected orthosteric peptides, we established distinct selectivity and probe dependency profiles. To decipher reversible and covalent mechanisms-of-action as well as potential binding sites of novel PAMs, we conducted glutathione reactivity, cAMP-washout and receptor mutational studies. [7] To conclude, we discovered a unique class of SCTR modulators providing useful pharmacologic tools to study SCTR- or GLP-1R-specific activation pathways and explore novel allosteric binding sites with potential to instigate the development of first-in-class therapeutics for treating comorbid conditions obesity and diabetes. [7] References [1] Annu Rev Pharmacol Toxicol, 2020, 60, 89-107; [2] Trends in Biochemical Sciences, 2017, 12, 946-960; [3] Experimental Physiology, 2013, 5, 973-987; [4] Current Neuropharmacology, 2007, 3, 168-179; [5] SLAS Discovery, 2021, 26, 1-16; [6] Journal of Peptide Science, 2017, 12, 845-854; [7] submitted to Biochemical Pharmacology 2021.

Submolecular probing of the complement C5a receptor\u2013ligand binding reveals a cooperative two-site binding mechanism

A current challenge to produce effective therapeutics is to accurately determine the location of the ligand-biding site and to characterize its properties. So far, the mechanisms underlying the functional activation of cell surface receptors by ligands with a complex binding mechanism remain poorly understood due to a lack of suitable nanoscopic methods to study them in their native environment. Here, we elucidated the ligand-binding mechanism of the human G protein-coupled C5a receptor (C5aR). We discovered for the first time a cooperativity between the two orthosteric binding sites. We found that the N-terminus C5aR serves as a kinetic trap, while the transmembrane domain acts as the functional site and both contributes to the overall high-affinity interaction. In particular, Asp282 plays a key role in ligand binding thermodynamics, as revealed by atomic force microscopy and steered molecular dynamics simulation. Our findings provide a new structural basis for the functional and mechanistic understanding of the GPCR family that binds large macromolecular ligands.

Donepezil Inhibits Acetylcholinesterase via Multiple Binding Modes at Room Temperature.

Donepezil is a second generation acetylcholinesterase (AChE) inhibitor for treatment of Alzheimer's disease (AD). AChE is important for neurotransmission at neuromuscular junctions and cholinergic brain synapses by hydrolyzing acetylcholine into acetate and choline. In vitro data support that donepezil is a reversible, mixed competitive and noncompetitive inhibitor of AChE. The experimental fact then suggests a more complex binding mechanism beyond the molecular view in X-ray models resolved at cryogenic temperatures that show a unique binding mode of donepezil in the active site of the enzyme. Aiming at clarifying the mechanism behind that mixed competitive and noncompetitive nature of the inhibitor, we have applied molecular dynamics (MD) simulations and docking and free-energy calculations to investigate microscopic details and energetics of donepezil association for conditions of substrate-free and -bound states of the enzyme. Liquid-phase MD simulation at room temperature shows AChE transits between "open" and "closed" conformations to control accessibility to the active site and ligand binding. As shown by docking and free-energy calculations, association of donepezil involves its reversible axial displacement and reorientation in the active site of the enzyme, assisted by water molecules. Donepezil binds equally well the main-door anionic binding site PAS, the acyl pocket, and the catalytic site CAS by respectively adopting outward-inward-inward orientations regardless of substrate occupancy-the overall stability of that reaction process depends however on co-occupancy of the enzyme being preferential for its substrate-free state. All together, our findings support a physiologically relevant mechanism of AChE inhibition by donepezil involving multistable interactions modes at the molecular origin of the inhibitor's activity.

Specific Conformational Dynamics and Expansion Underpin a Multi-Step Mechanism for Specific Binding of p27 with Cdk2/Cyclin A

The protein p27, a prominent regulatory protein in eukaryotes and an intrinsically disordered protein (IDP), regulates cell division by causing cell cycle arrest when bound in ternary complex with cyclin-dependent kinase (Cdk2) and cyclins (e.g., Cdk2/Cyclin A). We present an integrative study of p27 and its binding to Cdk2/Cyclin A complex by performing single-molecule multiparameter fluorescence spectroscopy, stopped-flow experiments, and molecular dynamics simulations. Our results suggest that unbound p27 adopts a compact conformation and undergoes conformational dynamics across several orders of magnitude in time (nano-to milliseconds), reflecting a multi-step mechanism for binding Cdk2/Cyclin A. Mutagenesis studies reveal that the region D1 in p27 plays a significant role in mediating the association kinetics, undergoing conformational rearrangement upon initial binding. Additionally, FRET experiments indicate an expansion of p27 throughout binding. The detected local and long-range structural dynamics suggest that p27 exhibits a limited binding surface in the unbound form, and stochastic conformational changes in D1 facilitate initial binding to Cdk2/Cyclin A complex. Furthermore, the post-kinase inhibitory domain (post-KID) region of p27 exchanges between distinct conformational ensembles: an extended regime exhibiting worm-like chain behavior, and a compact ensemble, which may protect p27 against nonspecific interactions. In summary, the binding interaction involves three steps: (i) D1 initiates binding, (ii) p27 wraps around Cdk2/Cyclin A and D2 binds, and (iii) the fully-formed fuzzy ternary complex is formed concomitantly with an extension of the post-KID region. An understanding of how the IDP nature of p27 underpins its functional interactions with Cdk2/Cyclin A provides insight into the complex binding mechanisms of IDPs and their regulatory mechanisms.

Synthesis, crystal structure, photophysical properties, density functional theory calculations and molecular docking studies on Cd(II) complex of 4,4′-dimethyl-2,2′-dipyridyl

Abstract A Cd(II) complex containing 4,4′-dimethyl-2,2′-dipyridyl (dmbpy) and nitrate (NO3−) was synthesized, and its crystal structure was determined by single crystal X-ray diffraction method. A detailed investigation on the electronic absorption spectrum was performed by the utilizing of UV–Vis spectroscopy and TD-B3LYP/LanL2DZ level. The highest occupied and lowest unoccupied molecular orbitals as well as the other molecular orbitals which are active in electronic spectrum were calculated for the Cd(II) complex. Natural bond orbital analysis was performed to provide evidences about the charge transfer interactions occurred in the Cd(II) complex. Total static dipole moment (μ), mean polarizibility (α) and mean first-order hyperpolarizibility (β) parameters were calculated to evaluate nonlinear optical properties of the Cd(II) complex. Molecular docking study for the Cd(II) complex to the target DNA was performed to understand complex binding mechanisms to a therapeutic target.

A Multiscale Simulation Approach to Modeling Drug-Protein Binding Kinetics.

Drug-target binding kinetics has recently emerged as a sometimes critical determinant of in vivo efficacy and toxicity. Its rational optimization to improve potency or reduce side effects of drugs is, however, extremely difficult. Molecular simulations can play a crucial role in identifying features and properties of small ligands and their protein targets affecting the binding kinetics, but significant challenges include the long time scales involved in (un)binding events and the limited accuracy of empirical atomistic force fields (lacking, e.g., changes in electronic polarization). In an effort to overcome these hurdles, we propose a method that combines state-of-the-art enhanced sampling simulations and quantum mechanics/molecular mechanics (QM/MM) calculations at the BLYP/VDZ level to compute association free energy profiles and characterize the binding kinetics in terms of structure and dynamics of the transition state ensemble. We test our combined approach on the binding of the anticancer drug Imatinib to Src kinase, a well-characterized target for cancer therapy with a complex binding mechanism involving significant conformational changes. The results indicate significant changes in polarization along the binding pathways, which affect the predicted binding kinetics. This is likely to be of widespread importance in binding of ligands to protein targets.