Distinct Binding Modes Research Articles

Rational Design of Macrocyclic Noncovalent Inhibitors of SARS-CoV-2 Mpro from a DNA-Encoded Chemical Library Screening Hit That Demonstrate Potent Inhibition against Pan-Coronavirus Homologues and Nirmatrelvir-Resistant Variants.

The recent global COVID-19 pandemic has highlighted treatments for coronavirus infection as an unmet medical need. The main protease (Mpro) has been an important target for the development of SARS-CoV-2 direct-acting antivirals. Nirmatrelvir as a covalent Mpro inhibitor was the first such approved therapy. Although Mpro inhibitors of various chemical classes have been reported, they are generally less active against nirmatrelvir-resistant variants and have limited pan-coronavirus potential, presenting a significant human health risk upon future outbreaks. We here present a novel approach and utilized DNA-encoded chemical library screening to identify the noncovalent Mpro inhibitor 5, which demonstrated a distinct binding mode to nirmatrelvir. A macrocyclization strategy designed to lock the active conformation resulted in lactone 12 with significantly improved antiviral activity. Further optimization led to the potent lactam 26, which demonstrated exceptional potency against nirmatrelvir-resistant variants as well as against a panel of viral main proteases from other coronaviruses.

Ultra-High Adsorption Capacity of Calcium-Iron Layered Double Hydroxides for HEDP Removal through Phase Transition Processes.

Antiscalant disposal in reverse osmosis concentrate (ROC) treatment is a significant obstacle in desalination. This study investigated the adsorption performance of LDHs for removing 1-hydroxyethylidene-1,1-diphosphonic acid (HEDP). CaFe-LDH presented a specific adsorption behavior and ultrahigh adsorption capacity for HEDP, with a maximum adsorption capacity of 335.7 mg P/g (1116.5 mg HEDP/g) at pH 7.0. X-ray diffraction (XRD) demonstrated that HEDP adsorption induced a structural transformation of CaFe-LDH from a layered configuration to a highly ordered structure, leading to a noticeable phase transition. Time-of-flight secondary ion mass spectrometry (ToF-SIMS) and Raman spectroscopy further confirmed that two distinct binding modes of HEDP, relating to chelation with Ca2+ and adsorption on Fe3+ simultaneously, are connected by phosphonic acid groups (-C-PO(OH)2), forming the CaFe-HEDP complex. X-ray fluorescence (XRF) and X-ray photoelectron spectroscopy (XPS) analyses revealed that the CaFe-HEDP ternary complex exhibits a highly ordered arrangement in an oxygen-bridged framework. The construction of an oxygen-coordinated framework contributes to the incorporation of more HEDP into CaFe-LDH, leading to a well-aligned lattice in the new phase. These findings provide valuable insights into developing novel LDH-based adsorbents for removing phosphorus-containing antiscalants, establishing a sustainable approach to ROC management, and potential environmental risk reduction.

Novel non-peptide uracil-derived human gonadotropin-releasing hormone receptor antagonists

Gonadotropin-releasing hormone (GnRH) is the main regulator of the reproductive system, acting on gonadotropic cells by binding to the GnRH1 receptor (GnRH1R). Traditionally, therapies targeting this receptor have relied on peptide modulators, which required subcutaneous or intramuscular injections. Due to the limitations of the parenteral administrations, there is a growing interest in developing oral small molecule modulators of GnRH1R as more convenient therapeutic alternatives.In this study, we examined the potential of chemically modifying elagolix, the first approved non-peptide, orally active GnRH1R antagonist, to increase its atropisomeric properties by introducing new moieties. We designed and synthesized the thio-uracil (1) and cytosine (2) derivatives of elagolix, both demonstrating GnRH1R antagonistic activities, with EC50 values of 39 and 110 nM, respectively. The atropisomers of 1 and 2 were efficiently separated using silica gel chromatography, and extensive NMR investigation, supported by Density Functional Theory (DFT) calculations, allowed us to define their conformations and rotational barriers. Docking and Molecular Dynamics (MD) studies revealed that 1 and 2 bind to GnRH1R with ΔG values comparable to elagolix, but through distinct binding modes. These results highlight the potential of non-peptide modulators to effectively modulate GnRH1R activity and pave the way for developing novel modulators.

Toward Unveiling Putative Binding Sites of Interleukin-33: Insights from Mixed-Solvent Molecular Dynamics Simulations of the Interleukin-1 Family.

The interleukin (IL)-1 family is a major proinflammatory cytokine family, ranging from the well-studied IL-1s to the most recently discovered IL-33. As a new focus, IL-33 has attracted extensive research for its crucial immunoregulatory roles, leading to the development of notable monoclonal antibodies as clinical candidates. Efforts to develop small molecules disrupting IL-33/ST2 interaction remain highly desired but encounter challenges due to the shallow and featureless interfaces. The information from relative cytokines has shown that traditional binding site identification methods still struggle in mapping cryptic sites, necessitating dynamic approaches to uncover druggable pockets on IL-33. Here, we employed mixed-solvent molecular dynamics (MixMD) simulations with diverse-property probes to map the hotspots of IL-33 and identify potential binding sites. The protocol was first validated using the known binding sites of two IL-1 family members and then applied to the structure of IL-33. Our simulations revealed several binding sites and proposed side-chain rearrangements essential for the binding of a known inhibitor, aligning well with experimental NMR findings. Further microsecond-time scale simulations of this IL-33-protein complex unveiled distinct binding modes with varying occurrences. These results could facilitate future efforts in developing ligands to target challenging flexible pockets of IL-33 and IL-1 family cytokines in general.

Engineering of Phycourobilin Synthase: PubS to a Two-Electron Reductase.

Phycourobilin:ferredoxin oxidoreductase (PubS) belongs to the ferredoxin-dependent bilin reductase (FDBR) family and catalyzes the reduction of the C15=C16 double bond, followed by the C4=C5 double bond of biliverdin IXα to produce phycourobilin. Among the diverse FDBR enzymes that catalyze site-specific reduction reactions of bilins, PubS lineage is the only one that reduces the C4=C5 double bond. This family can be broadly divided into four-electron reduction enzymes, which catalyze two successive two-electron reductions, such as PubS, and two-electron reduction enzymes, which catalyze a single two-electron reduction. The crystal structures of diverse FDBRs, excluding PubS, have unraveled that there are two distinct binding modes in the substrate-binding pocket. In this study, we focused on the arginine (Arg) residues that is considered crucial for substrate-binding mode in two-electron reduction enzymes. Through sequence alignment and comparison with the predicted structure of PubS, we identified a residue in PubS that corresponds to the Arg residue in the two-electron reduction enzymes. We further introduced mutations to avoid the steric hindrance associated with changes in the binding mode. Biochemical characterization of these variants showed that we successfully modified PubS from a four-electron reduction enzyme to a two-electron reduction enzyme with the accumulation of radicals. Our results provide insight into the molecular mechanisms of the chromophore binding mode and proton donation from acidic residues.

Single-molecule imaging of SWI/SNF chromatin remodelers reveals bromodomain-mediated and cancer-mutants-specific landscape of multi-modal DNA-binding dynamics

Despite their prevalent cancer implications, the in vivo dynamics of SWI/SNF chromatin remodelers and how misregulation of such dynamics underpins cancer remain poorly understood. Using live-cell single-molecule tracking, we quantify the intranuclear diffusion and chromatin-binding of three key subunits common to all major human SWI/SNF remodeler complexes (BAF57, BAF155 and BRG1), and resolve two temporally distinct stable binding modes for the fully assembled complex. Super-resolved density mapping reveals heterogeneous, nanoscale remodeler binding “hotspots” across the nucleoplasm where multiple binding events (especially longer-lived ones) preferentially cluster. Importantly, we uncover distinct roles of the bromodomain in modulating chromatin binding/targeting in a DNA-accessibility-dependent manner, pointing to a model where successive longer-lived binding within “hotspots” leads to sustained productive remodeling. Finally, systematic comparison of six common BRG1 mutants implicated in various cancers unveils alterations in chromatin-binding dynamics unique to each mutant, shedding insight into a multi-modal landscape regulating the spatio-temporal organizational dynamics of SWI/SNF remodelers.

Structural basis of Frizzled 4 in recognition of Dishevelled 2 unveils mechanism of WNT signaling activation

WNT signaling is fundamental in development and homeostasis, but how the Frizzled receptors (FZDs) propagate signaling remains enigmatic. Here, we present the cryo-EM structure of FZD4 engaged with the DEP domain of Dishevelled 2 (DVL2), a key WNT transducer. We uncover a distinct binding mode where the DEP finger-loop inserts into the FZD4 cavity to form a hydrophobic interface. FZD4 intracellular loop 2 (ICL2) additionally anchors the complex through polar contacts. Mutagenesis validates the structural observations. The DEP interface is highly conserved in FZDs, indicating a universal mechanism by which FZDs engage with DVLs. We further reveal that DEP mimics G-protein/β-arrestin/GRK to recognize an active conformation of receptor, expanding current GPCR engagement models. Finally, we identify a distinct FZD4 dimerization interface. Our findings delineate the molecular determinants governing FZD/DVL assembly and propagation of WNT signaling, providing long-sought answers underlying WNT signal transduction.

Dopamine reuptake and inhibitory mechanisms in human dopamine transporter.

The dopamine transporter has a crucial role in regulation of dopaminergic neurotransmission by uptake of dopamine into neurons and contributes to the abuse potential of psychomotor stimulants1-3. Despite decades of study, the structure, substrate binding, conformational transitions and drug-binding poses of human dopamine transporter remain unknown. Here we report structures of the human dopamine transporter in its apo state, and in complex with the substrate dopamine, the attention deficit hyperactivity disorder drug methylphenidate, and the dopamine-uptake inhibitors GBR12909 and benztropine. The dopamine-bound structure in the occluded state precisely illustrates the binding position of dopamine and associated ions. The structures bound to drugs are captured in outward-facing or inward-facing states, illuminating distinct binding modes and conformational transitions during substrate transport. Unlike the outward-facing state, which is stabilized by cocaine, GBR12909 and benztropine stabilize the dopamine transporter in the inward-facing state, revealing previously unseen drug-binding poses and providing insights into how they counteract the effects of cocaine. This study establishes a framework for understanding the functioning of the human dopamine transporter and developing therapeutic interventions for dopamine transporter-related disorders and cocaine addiction.

Evolutionary sequence and structural basis for the distinct conformational landscapes of Tyr and Ser/Thr kinases

Protein kinases are molecular machines with rich sequence variation that distinguishes the two main evolutionary branches – tyrosine kinases (TKs) from serine/threonine kinases (STKs). Using a sequence co-variation Potts statistical energy model we previously concluded that TK catalytic domains are more likely than STKs to adopt an inactive conformation with the activation loop in an autoinhibitory folded conformation, due to intrinsic sequence effects. Here we investigate the structural basis for this phenomenon by integrating the sequence-based model with structure-based molecular dynamics (MD) to determine the effects of mutations on the free energy difference between active and inactive conformations, using a thermodynamic cycle involving many (n = 108) protein-mutation free energy perturbation (FEP) simulations in the active and inactive conformations. The sequence and structure-based results are consistent and support the hypothesis that the inactive conformation DFG-out Activation Loop Folded, is a functional regulatory state that has been stabilized in TKs relative to STKs over the course of their evolution via the accumulation of residue substitutions in the activation loop and catalytic loop that facilitate distinct substrate binding modes in trans and additional modes of regulation in cis for TKs.

Structural basis of psychedelic LSD recognition at dopamine D1 receptor

Understanding the kinetics of LSD in receptors and subsequent induced signaling is crucial for comprehending both the psychoactive and therapeutic effects of LSD. Despite extensive research on LSD's interactions with serotonin 2A and 2B receptors, its behavior on other targets, including dopamine receptors, has remained elusive. Here, we present cryo-EM structures of LSD/PF6142-bound dopamine D1 receptor (DRD1)-legobody complexes, accompanied by a β-arrestin-mimicking nanobody, NBA3, shedding light on the determinants of G protein coupling versus β-arrestin coupling. Structural analysis unveils a distinctive binding mode of LSD in DRD1, particularly with the ergoline moiety oriented toward TM4. Kinetic investigations uncover an exceptionally rapid dissociation rate of LSD in DRD1, attributed to the flexibility of extracellular loop 2 (ECL2). Moreover, G protein can stabilize ECL2 conformation, leading to a significant slowdown in ligand's dissociation rate. These findings establish a solid foundation for further exploration of G protein-coupled receptor (GPCR) dynamics and their relevance to signal transduction.

Glucoimines incorporating classical and non-classical carbonic anhydrase pharmacophores: Design, synthesis, conformational analysis, biological evaluation, and docking simulations

In this report, a series of glucoimines has been prepared by reaction of d-glucosamine and aromatic aldehydes incorporating classical and non-classical carbonic anhydrase pharmacophores. The conformational behavior of veratrole glucoimine was studied in solution and in crystalline form, by NMR and X-ray diffraction analysis, which revealed that pyranose ring adopted a 4C1 conformation. All glucoimines were tested against four isozymes of carbonic anhydrase: hCAs I and II (cytosolic, ubiquitous isozymes) and hCAs IX and XII (tumor-associated isozymes). In this study, per-O-acetylated and deprotected sulfonamide were identified as potent inhibitors of tumor-associated carbonic anhydrase isoforms. Molecular docking simulation was performed inside the active site of hCA II to evaluate the binding modes of veratrole and sulfonamide glucoimines. The last ones demonstrated the most favorable docking scores, indicating strong binding affinity towards hCAII in correlation with experimental inhibition activities. Interestingly, interaction analyses revealed distinct binding modes for ligand-hCAII complexes primarily due to the sulfonamide group.

Molecular mechanism of distinct chemokine engagement and functional divergence of the human Duffy antigen receptor

The Duffy antigen receptor is a seven-transmembrane (7TM) protein expressed primarily at the surface of red blood cells and displays strikingly promiscuous binding to multiple inflammatory and homeostatic chemokines. It serves as the basis of the Duffy blood group system in humans and also acts as the primary attachment site for malarial parasite Plasmodium vivax and pore-forming toxins secreted by Staphylococcus aureus. Here, we comprehensively profile transducer coupling of this receptor, discover potential non-canonical signaling pathways, and determine the cryoelectron microscopy (cryo-EM) structure in complex with the chemokine CCL7. The structure reveals a distinct binding mode of chemokines, as reflected by relatively superficial binding and a partially formed orthosteric binding pocket. We also observe a dramatic shortening of TM5 and 6 on the intracellular side, which precludes the formation of the docking site for canonical signal transducers, thereby providing a possible explanation for the distinct pharmacological and functional phenotype of this receptor.

Discovery of a novel exceptionally potent and orally active Nur77 ligand NB1 with a distinct binding mode for cancer therapy

The orphan nuclear receptor Nur77 is emerging as an attractive target for cancer therapy, and activating Nur77's non-genotypic anticancer function has demonstrated strong therapeutic potential. However, few Nur77 site B ligands have been identified as excellent anticancer compounds. There are no co-crystal structures of effective anticancer agents at Nur77 site B, which greatly limits the development of novel Nur77 site B ligands. Moreover, the lack of pharmaceutical ligands restricts Nur77's therapeutic proof of concept. Herein, we developed a first-in-class Nur77 site B ligand (NB1) that significantly inhibited cancer cells by mediating the Nur77/Bcl-2-related apoptotic effect at mitochondria. The X-ray crystallography suggests that NB1 is bound to the Nur77 site B with a distinct binding mode. Importantly, NB1 showed favorable pharmacokinetic profiles and safety, as evidenced by its good oral bioavailability in rats and lack of mortality, bodyweight loss, and pathological damage at the 512.0 mg/kg dose in mice. Furthermore, oral administration of NB1 demonstrated remarkable in vivo anticancer efficacy in an MDA-MB-231 xenograft model. Together, our work discovers NB1 as a new generation Nur77 ligand that activates the Nur77/Bcl-2 apoptotic pathway with a safe and effective cancer therapeutic potency.

Atomistic characterization of β2-glycoprotein I domain V interaction with anionic membranes

BackgroundInteraction of β2-glycoprotein I (β2GPI) with anionic membranes is crucial in antiphospholipid syndrome (APS), implicating the role of its membrane-binding domain, domain V (DV). The mechanism of DV binding to anionic lipids is not fully understood. ObjectivesThis study aimed to elucidate the molecular details of β2GPI DV binding to anionic membranes. MethodsWe utilized molecular dynamics simulations to investigate the structural basis of anionic lipid recognition by DV. To corroborate the membrane-binding mode identified in the highly mobile membrane mimetic simulations, we conducted additional simulations using a full membrane model. ResultsThe study identified critical regions in DV, namely the lysine-rich loop and the hydrophobic loop, which are essential for membrane association via electrostatic and hydrophobic interactions, respectively. A novel lysine pair contributing to membrane binding was also discovered, providing new insights into β2GPI’s membrane interaction. Simulations revealed 2 distinct binding modes of DV to the membrane, with mode 1 characterized by the insertion of the hydrophobic loop into the lipid bilayer, suggesting a dominant mechanism for membrane association. This interaction is pivotal for the pathogenesis of APS, as it facilitates the recognition of β2GPI by antiphospholipid antibodies. ConclusionThe study advances our understanding of the molecular interactions between β2GPI’s DV and anionic membranes, which are crucial for APS pathogenesis. It highlights the importance of specific regions in DV for membrane binding and reveals a predominant binding mode. These findings have significant implications for APS diagnostics and therapeutics, offering a deeper insight into the molecular basis of the syndrome.

Fentanyl-Type Antagonist of the μ-Opioid Receptor: Important Role of Axial Chirality in the Active Conformation.

In recent years, synthetic opioids have emerged as a predominant cause of drug-overdose-related fatalities, causing the "opioid crisis." To design safer therapeutic agents, we accidentally discovered μ-opioid receptor (MOR) antagonists based on fentanyl with a relatively uncomplicated chemical composition that potentiates structural modifications. Here, we showed the development of novel atropisomeric fentanyl analogues that exhibit more potent antagonistic activity against MOR than naloxone, a morphinan MOR antagonist. Derivatives displaying stable axial chirality were synthesized based on the amide structure of fentanyl. The aS- and aR-enantiomers exerted antagonistic and agonistic effects on the MOR, respectively, and each atropisomer interacted with the MOR by assuming a distinct binding mode through molecular docking. These findings suggest that introducing atropisomerism into fentanyl may serve as a key feature in the molecular design of future MOR antagonists to help mitigate the opioid crisis.

The Legionella collagen-like protein employs a distinct binding mechanism for the recognition of host glycosaminoglycans

Bacterial adhesion is a fundamental process which enables colonisation of niche environments and is key for infection. However, in Legionella pneumophila, the causative agent of Legionnaires’ disease, these processes are not well understood. The Legionella collagen-like protein (Lcl) is an extracellular peripheral membrane protein that recognises sulphated glycosaminoglycans on the surface of eukaryotic cells, but also stimulates bacterial aggregation in response to divalent cations. Here we report the crystal structure of the Lcl C-terminal domain (Lcl-CTD) and present a model for intact Lcl. Our data reveal that Lcl-CTD forms an unusual trimer arrangement with a positively charged external surface and negatively charged solvent exposed internal cavity. Through molecular dynamics simulations, we show how the glycosaminoglycan chondroitin-4-sulphate associates with the Lcl-CTD surface via distinct binding modes. Our findings show that Lcl homologs are present across both the Pseudomonadota and Fibrobacterota-Chlorobiota-Bacteroidota phyla and suggest that Lcl may represent a versatile carbohydrate-binding mechanism.

Three prime repair exonuclease 1 preferentially degrades the integration-incompetent HIV-1 DNA through favorable kinetics, thermodynamic, structural, and conformational properties

HIV-1 integration into the human genome is dependent on 3’-processing of the viral DNA. Recently, we reported that the cellular Three Prime Repair Exonuclease 1 (TREX1) enhances HIV-1 integration by degrading the unprocessed viral DNA, while the integration-competent 3’-processed DNA remained resistant. Here, we describe the mechanism by which the 3’-processed HIV-1 DNA resists TREX1-mediated degradation. Our kinetic studies revealed that the rate of cleavage (kcat) of the 3’-processed DNA was significantly lower (approximately 2-2.5-fold) than the unprocessed HIV-1 DNA by TREX1. The kcat values of human TREX1 for the processed U5 and U3 DNA substrates were 3.8 s-1 and 4.5 s-1, respectively. In contrast, the unprocessed U5 and U3 substrates were cleaved at 10.2 s-1 and 9.8 s-1, respectively. The efficiency of degradation (kcat/Km) of the 3’-processed DNA (U5-70.2 and U3-28.05 pM-1s-1) was also significantly lower than the unprocessed DNA (U5-103.1 and U3-65.3 pM-1s-1). Furthermore, the binding affinity (Kd) of TREX1 was markedly lower (∼ 2-fold) for the 3’-processed DNA compared to the unprocessed DNA. Molecular docking and dynamics studies revealed distinct conformational binding modes of TREX1 with the 3’-processed and unprocessed HIV-1 DNA. Particularly, the unprocessed DNA was favorably positioned in the active site with polar interactions with the catalytic residues of TREX1. Additionally, a stable complex was formed between TREX1 and the unprocessed DNA compared the 3’-processed DNA. These results pinpoint the mechanism by which TREX1 preferentially degrades the integration-incompetent HIV-1 DNA and reveal the unique structural and conformational properties of the integration-competent 3’-processed HIV-1 DNA.

Impact of Combinatorial Histone Modifications on Acetyllysine Recognition by the ATAD2 and ATAD2B Bromodomains.

The ATPase family AAA+ domain containing 2 (ATAD2) protein and its paralog ATAD2B have a C-terminal bromodomain (BRD) that functions as a reader of acetylated lysine residues on histone proteins. Using a structure-function approach, we investigated the ability of the ATAD2/B BRDs to select acetylated lysine among multiple histone post-translational modifications. The ATAD2B BRD can bind acetylated histone ligands that also contain adjacent methylation or phosphorylation marks, while the presence of these modifications significantly weakened the acetyllysine binding activity of the ATAD2 BRD. Our structural studies provide mechanistic insights into how ATAD2/B BRD-binding pocket residues coordinate the acetyllysine group in the context of adjacent post-translational modifications. Furthermore, we investigated how sequence changes in amino acids of the histone ligands impact the recognition of an adjacent acetyllysine residue. Our study highlights how the interplay between multiple combinations of histone modifications influences the reader activity of the ATAD2/B BRDs, resulting in distinct binding modes.

N-Heterocyclic Carbene Monolayers on Metal-Oxide Films: Correlations between Adsorption Mode and Surface Functionality.

N-Heterocyclic carbene (NHC) ligands have been self-assembled on various metal and semimetal surfaces, creating a covalent bond with surface metal atoms that led to high thermal and chemical stability of the self-assembled monolayer. This study explores the self-assembly of NHCs on metal-oxide films (CuOx, FeOx, and TiOx) and reveals that the properties of these metal-oxide substrates play a pivotal role in dictating the adsorption behavior of NHCs, influencing the decomposition route of the monolayer and its impact on work function values. While the attachment of NHCs onto CuOx is via coordination with surface oxygen atoms, NHCs interact with TiOx through coordination with surface metal atoms and with FeOx via coordination with both metal and oxygen surface atoms. These distinct binding modes arise due to variances in the electronic properties of the metal atoms within the investigated metal-oxide films. Contact angle and ultraviolet photoelectron spectroscopy measurements have shown a significantly higher impact of F-NHC adsorption on CuOx than on TiOx and FeOx , correlated to a preferred, averaged upright orientation of F-NHC on CuOx.

Unveiling a Novel Solvatomorphism of Anti-inflammatory Flufenamic Acid: X-ray Structure, Quantum Chemical, and In Silico Studies.

This paper delves into the polymorphism of 2-[3-(trifluoromethyl)anilino]benzoic acid, commonly referred to as flufenamic acid (FA), a pharmaceutical agent employed in treating inflammatory conditions. The central focus of the study is on a newly unearthed solvatomorphic structure of FA in methanol (FAM), and a thorough comparison is conducted with the commercially available standard structure. Employing a comprehensive approach, including X-ray crystallography, Hirshfeld surface analysis, density functional theory (DFT), molecular docking, and molecular dynamics (MD) simulations, the research aims to unravel the structural and functional implications of solvatomorphism. The X-ray crystal structure analysis brings to light notable differences between the standard FA and solvatomorphic FAM, showcasing variations in intermolecular interactions and crystal packing. Key features such as hydrogen bonding, π···π stacking, and C-H···π interactions are identified as influential factors shaping the stability and conformation of the compounds. Hirshfeld surface analysis further quantifies the nature and contribution of intermolecular interactions, providing a comprehensive perspective on molecular stability. Density functional theory offers valuable electronic structure insights, highlighting disparities in frontier molecular orbitals between FA and FAM. Molecular docking studies against prostaglandin D2 11-ketoreductase explore potential drug interactions, unveiling distinct binding modes and hydrogen bonding patterns that shed light on how the solvatomorphic structure may impact drug-target interactions. In-depth molecular dynamics simulations over 100 ns investigate the stability of the protein-ligand complex, with root mean square deviation and root mean square fluctuation analyses revealing minimal deviations and affirming the stability of FAM within the active site of the target protein.