Porin Associates with Tom22 to Regulate the Mitochondrial Protein Gate Assembly.

Mitochondria import nearly all of their resident proteins from the cytosol, and the TOM complex functions as their entry gate. The TOM complex undergoes a dynamic conversion between the majority population of a three-channel gateway ("trimer") and the minor population that lacks Tom22 and has only two Tom40 channels ("dimer"). Here, we found that the porin Por1 acts as a sink to bind newly imported Tom22. This Por1 association thereby modulates Tom22 integration into the TOM complex, guaranteeing formation of the functional trimeric TOM complex. Por1 sequestration of Tom22 dissociated from the trimeric TOM complex also enhances the dimeric TOM complex, which is preferable for the import of TIM40/MIA-dependent proteins into mitochondria. Furthermore, Por1 appears to contribute to cell-cycle-dependent variation of the functional trimeric TOM complex by chaperoning monomeric Tom22, which arises from the cell-cycle-controlled variation of phosphorylated Tom6.

Attenuation of SARS-CoV-2 replication and associated inflammation by concomitant targeting of viral and host cap 2'-O-ribose methyltransferases.

The SARS-CoV-2 infection cycle is a multistage process that relies on functional interactions between the host and the pathogen. Here, we repurposed antiviral drugs against both viral and host enzymes to pharmaceutically block methylation of the viral RNA 2'-O-ribose cap needed for viral immune escape. We find that the host cap 2'-O-ribose methyltransferase MTr1 can compensate for loss of viral NSP16 methyltransferase in facilitating virus replication. Concomitant inhibition of MTr1 and NSP16 efficiently suppresses SARS-CoV-2 replication. Using in silico target-based drug screening, we identify a bispecific MTr1/NSP16 inhibitor with anti-SARS-CoV-2 activity in vitro and in vivo but with unfavorable side effects. We further show antiviral activity of inhibitors that target independent stages of the host SAM cycle providing the methyltransferase co-substrate. In particular, the adenosylhomocysteinase (AHCY) inhibitor DZNep is antiviral in in vitro, in ex vivo, and in a mouse infection model and synergizes with existing COVID-19 treatments. Moreover, DZNep exhibits a strong immunomodulatory effect curbing infection-induced hyperinflammation and reduces lung fibrosis markers ex vivo. Thus, multispecific and metabolic MTase inhibitors constitute yet unexplored treatment options against COVID-19.

Hexestrol, an estrogen receptor agonist, inhibits Lassa virus entry.

Lassa virus (LASV) is the causative agent of human Lassa fever which in severe cases manifests as hemorrhagic fever leading to thousands of deaths annually. However, no approved vaccines or antiviral drugs are currently available. Recently, we screened approximately 2,500 compounds using a recombinant vesicular stomatitis virus (VSV) expressing LASV glycoprotein GP (VSV-LASVGP) and identified a P-glycoprotein inhibitor as a potential LASV entry inhibitor. Here, we show that another identified candidate, hexestrol (HES), an estrogen receptor agonist, is also a LASV entry inhibitor. HES inhibited VSV-LASVGP replication with a 50% inhibitory concentration (IC50) of 0.63 µM. Importantly, HES also inhibited authentic LASV replication with IC50 values of 0.31 µM-0.61 µM. Time-of-addition and cell-based membrane fusion assays suggested that HES inhibits the membrane fusion step during virus entry. Alternative estrogen receptor agonists did not inhibit VSV-LASVGP replication, suggesting that the estrogen receptor itself is unlikely to be involved in the antiviral activity of HES. Generation of a HES-resistant mutant revealed that the phenylalanine at amino acid position 446 (F446) of LASVGP, which is located in the transmembrane region, conferred resistance to HES. Although mutation of F446 enhanced the membrane fusion activity of LASVGP, it exhibited reduced VSV-LASVGP replication, most likely due to the instability of the pre-fusion state of LASVGP. Collectively, our results demonstrated that HES is a promising anti-LASV drug that acts by inhibiting the membrane fusion step of LASV entry. This study also highlights the importance of the LASVGP transmembrane region as a target for anti-LASV drugs.IMPORTANCELassa virus (LASV), the causative agent of Lassa fever, is the most devastating mammarenavirus with respect to its impact on public health in West Africa. However, no approved antiviral drugs or vaccines are currently available. Here, we identified hexestrol (HES), an estrogen receptor agonist, as the potential antiviral candidate drug. We showed that the estrogen receptor itself is not involved in the antiviral activity. HES directly bound to LASVGP and blocked membrane fusion, thereby inhibiting LASV infection. Through the generation of a HES-resistant virus, we found that phenylalanine at position 446 (F446) within the LASVGP transmembrane region plays a crucial role in the antiviral activity of HES. The mutation at F446 caused reduced virus replication, likely due to the instability of the pre-fusion state of LASVGP. These findings highlight the potential of HES as a promising candidate for the development of antiviral compounds targeting LASV.

Temperature-Sensitive Substrate and Product Binding Underlie Temperature-Compensated Phosphorylation in the Clock.

Temperature compensation is a striking feature of the circadian clock. Here we investigate biochemical mechanisms underlying temperature-compensated, CKIδ-dependent multi-site phosphorylation in mammals. We identify two mechanisms for temperature-insensitive phosphorylation at higher temperature: lower substrate affinity to CKIδ-ATP complex and higher product affinity to CKIδ-ADP complex. Inhibitor screening of ADP-dependent phosphatase activity of CKIδ identified aurintricarboxylic acid (ATA) as a temperature-sensitive kinase activator. Docking simulation of ATA and mutagenesis experiment revealed K224D/K224E mutations in CKIδ that impaired product binding and temperature-compensated primed phosphorylation. Importantly, K224D mutation shortens behavioral circadian rhythms and changes the temperature dependency of SCN's circadian period. Interestingly, temperature-compensated phosphorylation was evolutionary conserved in yeast. Molecular dynamics simulation and X-ray crystallography demonstrate that an evolutionally conserved CKI-specific domain around K224 can provide a structural basis for temperature-sensitive substrate and product binding. Surprisingly, this domain can confer temperature compensation on a temperature-sensitive TTBK1. These findings suggest the temperature-sensitive substrate- and product-binding mechanisms underlie temperature compensation.

Pocket to concavity: a tool for the refinement of protein-ligand binding site shape from alpha spheres.

SUMMARY: Understanding the binding site of the target protein is essential for rational drug design. Pocket detection software predicts the ligand binding site of the target protein; however, the predicted protein pockets are often excessively estimated in comparison with the actual volume of the bound ligands. This study proposes a refinement tool for the pockets predicted by an alpha sphere-based approach, Pocket to Concavity (P2C). P2C is divided into two modes: Ligand-Free (LF) and Ligand-Bound (LB) modes. The LF mode provides the shape of the deep and druggable concavity where the core scaffold can bind. The LB mode searches the deep concavity around the bound ligand. Thus, P2C is useful for identifying and designing desirable compounds in Structure-Based Drug Design (SBDD). AVAILABILITY AND IMPLEMENTATION: Pocket to Concavity is freely available at https://github.com/genki-kudo/Pocket-to-Concavity. This tool is implemented in Python3 and Fpocket2.

Biological Evaluation of Isosteric Applicability of 1,3-Substituted Cuneanes as m-Substituted Benzenes Enabled by Selective Isomerization of 1,4-Substituted Cubanes.

We herein evaluate a biological applicability of 1,3-substituted cuneanes as an isostere of m-substituted benzenes based on its structural similarity. An investigation of a method to obtain 1,3-substituted cuneanes by selective isomerization of 1,4-substituted cubanes enables this attempt by giving a key synthetic step to obtain a cuneane analogs of pharmaceuticals having m-substituted benzene moiety. Biological evaluation of the synthesized analogs and in silico study of the obtained result revealed a potential usage of cuneane skeleton in medicinal chemistry.

Lipid bilayer membrane permeability mechanism of the K-Ras(G12D)-inhibitory bicyclic peptide KS-58 elucidated by molecular dynamics simulations.

Peptides are mid-size molecules (700-2000 g/mol) and have attracted particular interest as therapeutic modalities as they are superior in controlling protein-protein interactions, a process that is a typical drug target category, compared with small molecules (<500 g/mol). In 2020, we identified KS-58 (1333 g/mol) as a K-Ras(G12D)-inhibitory bicyclic peptide and suggested its cell membrane permeability. However, the membrane permeability mechanism had not been elucidated. In this study, we aim to clarify the mechanism by molecular dynamics (MD) simulations. Initially, we simulated the molecular conformations of KS-58 in water (a polar solvent) and in chloroform (a non-polar solvent). The identified stable conformations were significantly different in each solvent. KS-58 behaves as a chameleon-like molecule as it alters its polar surface area (PSA) depending on the solvent environment. It was also discovered that orientation of Asp's side chain is a critical energy barrier for KS-58 altering its conformation from hydrophilic to lipophilic. Taking these properties into consideration, we simulated its lipid bilayer membrane permeability. KS-58 shifted toward the inside of the lipid bilayer membrane with altering its conformations to lipophilic. When the simulation condition was set in deionized form of that carboxy group of Asp, KS-58 traveled deeper inside the cell membrane. PSA and the depth of the membrane penetration correlated. In vitro data suggested that cell membrane permeability of KS-58 is improved in weakly acidic conditions leading to partial deionization of the carboxy group. Our data provide an example of the molecular properties of mid-size peptides with membrane accessibility and propose an effective metadynamics approach to elucidate such molecular mechanisms by MD simulations.

Site Identification and Next Choice Protocol for Hit-to-Lead Optimization.

Time efficiency and cost savings are major challenges in drug discovery and development. In this process, the hit-to-lead stage is expected to improve efficiency because it primarily exploits the trial-and-error approach of medicinal chemists. This study proposes a site identification and next choice (SINCHO) protocol to improve the hit-to-lead efficiency. This protocol selects an anchor atom and growth site pair, which is desirable for a hit-to-lead strategy starting from a 3D complex structure. We developed and fine-tuned the protocol using a training data set and assessed it using a test data set of the preceding hit-to-lead strategy. The protocol was tested for experimentally determined structures and molecular dynamics (MD) ensembles. The protocol had a high prediction accuracy for applying MD ensembles, owing to the consideration of protein flexibility. The SINCHO protocol enables medicinal chemists to visualize and modify functional groups in a hit-to-lead manner.

Structural and Energetic Basis for Differential Binding of Ebola and Marburg Virus Glycoproteins to a Bat-Derived Niemann-Pick C1 Protein.

BACKGROUND: Our previous study demonstrated that the fruit bat (Yaeyama flying fox)-derived cell line FBKT1 showed preferential susceptibility to Ebola virus (EBOV), whereas the human cell line HEK293T was similarly susceptible to EBOV and Marburg virus (MARV). This was due to 3 amino acid differences of the endosomal receptor Niemann-Pick C1 (NPC1) between FBKT1 and HEK293T (ie, TET and SGA, respectively, at positions 425-427), as well as 2 amino acid differences at positions 87 and 142 of the viral glycoprotein (GP) between EBOV and MARV. METHODS/RESULTS: To understand the contribution of these amino acid differences to interactions between NPC1 and GP, we performed molecular dynamics simulations and binding free energy calculations. The average binding free energies of human NPC1 (hNPC1) and its mutant having TET at positions 425-427 (hNPC1/TET) were similar for the interaction with EBOV GP. In contrast, hNPC1/TET had a weaker interaction with MARV GP than wild-type hNPC1. As expected, substitutions of amino acid residues at 87 or 142 in EBOV and MARV GPs converted the binding affinity to hNPC1/TET. CONCLUSIONS: Our data provide structural and energetic insights for understanding potential differences in the GP-NPC1 interaction, which could influence the host tropism of EBOV and MARV.

Lysoglycosphingolipids have the ability to induce cell death through direct PI3K inhibition.

Sphingolipidoses are inherited metabolic disorders associated with glycosphingolipids accumulation, neurodegeneration, and neuroinflammation leading to severe neurological symptoms. Lysoglycosphingolipids (lysoGSLs), also known to accumulate in the tissues of sphingolipidosis patients, exhibit cytotoxicity. LysoGSLs are the possible pathogenic cause, but the mechanisms are still unknown in detail. Here, we first show that lysoGSLs are potential inhibitors of phosphoinositide 3-kinase (PI3K) to reduce cell survival signaling. We found that phosphorylated Akt was commonly reduced in fibroblasts from patients with sphingolipidoses, including GM1/GM2 gangliosidoses and Gaucher's disease, suggesting the contribution of lysoGSLs to the pathogenesis. LysoGSLs caused cell death and decreased the level of phosphorylated Akt as in the patient fibroblasts. Extracellularly administered lysoGM1 permeated the cell membrane to diffusely distribute in the cytoplasm. LysoGM1 and lysoGM2 also inhibited the production of phosphatidylinositol-(3,4,5)-triphosphate and the translocation of Akt from the cytoplasm to the plasma membrane. We also predicted that lysoGSLs could directly bind to the catalytic domain of PI3K by in silico docking study, suggesting that lysoGSLs could inhibit PI3K by directly interacting with PI3K in the cytoplasm. Furthermore, we revealed that the increment of lysoGSLs amounts in the brain of sphingolipidosis model mice correlated with the neurodegenerative progression. Our findings suggest that the down-regulation of PI3K/Akt signaling by direct interaction of lysoGSLs with PI3K in the brains is a neurodegenerative mechanism in sphingolipidoses. Moreover, we could propose the intracellular PI3K activation or inhibition of lysoGSLs biosynthesis as novel therapeutic approaches for sphingolipidoses because lysoGSLs should be cell death mediators by directly inhibiting PI3K, especially in neurons.

Identification of Chemicals That Abrogate Folate-Dependent Inhibition of Starch Accumulation in Non-Photosynthetic Plastids of Arabidopsis.

Folate, also known as vitamin B9, is an essential cofactor for a variety of enzymes and plays a crucial role in many biological processes. We previously reported that plastidial folate prevents starch biosynthesis triggered by the influx of sugar into non-starch-accumulating plastids, such as etioplasts, and chloroplasts under darkness; hence the loss of plastidial folate induces the accumulation of starch in plastids. To understand the molecular mechanism underlying this phenomenon, we screened our in-house chemical library and searched their derivatives to identify chemicals capable of inducing starch accumulation in etioplasts. The results revealed four chemicals, compounds #120 and #375 and their derivatives, compounds #120d and #375d, respectively. The derivative compounds induced starch accumulation in etioplasts and suppressed hypocotyl elongation in dark-grown Arabidopsis seedlings. They also inhibited the post-germinative growth of seedlings under illumination. All four chemicals contained the sulfonamide group as a consensus structure. The sulfonamide group is also found in sulfa drugs, which exhibit antifolate activity, and in sulfonylurea herbicides. Further analyses revealed that compound #375d induces starch accumulation by inhibiting folate biosynthesis. By contrast, compound #120d neither inhibited folate biosynthesis nor exhibited the herbicide activity. Protein and metabolite analyses suggest that compound #120d abrogates folate-dependent inhibition of starch accumulation in etioplasts by enhancing starch biosynthesis.

Unfolding is the driving force for mitochondrial import and degradation of the Parkinson's disease-related protein DJ-1.

Diverse genes associated with familial Parkinson's disease (familial Parkinsonism) have been implicated in mitochondrial quality control. One such gene, PARK7 encodes the protein DJ-1, pathogenic mutations of which trigger its translocation from the cytosol to the mitochondrial matrix. The translocation of steady-state cytosolic proteins like DJ-1 to the mitochondrial matrix upon missense mutations is rare, and the underlying mechanism remains to be elucidated. Here, we show that the protein unfolding associated with various DJ-1 mutations drives its import into the mitochondrial matrix. Increasing the structural stability of these DJ-1 mutants restores cytosolic localization. Mechanistically, we show that a reduction in the structural stability of DJ-1 exposes a cryptic N-terminal mitochondrial-targeting signal (MTS), including Leu10, which promotes DJ-1 import into the mitochondrial matrix for subsequent degradation. Our work describes a novel cellular mechanism for targeting a destabilized cytosolic protein to the mitochondria for degradation.

AAp-MSMD: Amino Acid Preference Mapping on Protein-Protein Interaction Surfaces Using Mixed-Solvent Molecular Dynamics.

Peptides have attracted much attention recently owing to their well-balanced properties as drugs against protein-protein interaction (PPI) surfaces. Molecular simulation-based predictions of binding sites and amino acid residues with high affinity to PPI surfaces are expected to accelerate the design of peptide drugs. Mixed-solvent molecular dynamics (MSMD), which adds probe molecules or fragments of functional groups as solutes to the hydration model, detects the binding hotspots and cryptic sites induced by small molecules. The detection results vary depending on the type of probe molecule; thus, they provide important information for drug design. For rational peptide drug design using MSMD, we proposed MSMD with amino acid residue probes, named amino acid probe-based MSMD (AAp-MSMD), to detect hotspots and identify favorable amino acid types on protein surfaces to which peptide drugs bind. We assessed our method in terms of hotspot detection at the amino acid probe level and binding free energy prediction with amino acid probes at the PPI site for the complex structure that formed the PPI. In hotspot detection, the max-spatial probability distribution map (max-PMAP) obtained from AAp-MSMD detected the PPI site, to which each type of amino acid can bind favorably. In the binding free energy prediction using amino acid probes, ΔGFE obtained from AAp-MSMD roughly estimated the experimental binding affinities from the structure-activity relationship. AAp-MSMD, with amino acid probes, provides estimated binding sites and favorable amino acid types at the PPI site of a target protein.

Modeling the structure of the transmembrane domain of T1R3, a subunit of the sweet taste receptor, with neohesperidin dihydrochalcone using molecular dynamics simulation.

Neohesperidin dihydrochalcone (NHDC) is a sweetener, which interacts with the transmembrane domain (TMD) of the T1R3 subunit of the human sweet taste receptor. Although NHDC and a sweet taste inhibitor lactisole share similar structural motifs, they have opposite effects on the receptor. This study involved the creation of an NHDC-docked model of T1R3 TMD through mutational analyses followed by in silico simulations. When certain NHDC derivatives were docked to the model, His7345.44 was demonstrated to play a crucial role in activating T1R3 TMD. The NHDC-docked model was then compared with a lactisole-docked inactive form, several residues were characterized as important for the recognition of NHDC; however, most of them were distinct from those of lactisole. Residues such as His6413.33 and Gln7947.38 were found to be oriented differently. This study provides useful information that will facilitate the design of sweeteners and inhibitors that interact with T1R3 TMD.

Molecular action of larvicidal flavonoids on ecdysteroidogenic glutathione S-transferase Noppera-bo in Aedes aegypti.

BACKGROUND: Mosquito control is a crucial global issue for protecting the human community from mosquito-borne diseases. There is an urgent need for the development of selective and safe reagents for mosquito control. Flavonoids, a group of chemical substances with variable phenolic structures, such as daidzein, have been suggested as potential mosquito larvicides with less risk to the environment. However, the mode of mosquito larvicidal action of flavonoids has not been elucidated. RESULTS: Here, we report that several flavonoids, including daidzein, inhibit the activity of glutathione S-transferase Noppera-bo (Nobo), an enzyme used for the biosynthesis of the insect steroid hormone ecdysone, in the yellow fever mosquito Aedes aegypti. The crystal structure of the Nobo protein of Ae. aegypti (AeNobo) complexed with the flavonoids and its molecular dynamics simulation revealed that Glu113 forms a hydrogen bond with the flavonoid inhibitors. Consistent with this observation, substitution of Glu113 with Ala drastically reduced the inhibitory activity of the flavonoids against AeNobo. Among the identified flavonoid-type inhibitors, desmethylglycitein (4',6,7-trihydroxyisoflavone) exhibited the highest inhibitory activity in vitro. Moreover, the inhibitory activities of the flavonoids correlated with the larvicidal activity, as desmethylglycitein suppressed Ae. aegypti larval development more efficiently than daidzein. CONCLUSION: Our study demonstrates the mode of action of flavonoids on the Ae. aegypti Nobo protein at the atomic, enzymatic, and organismal levels.

Distinct Profiles of Desensitization of µ-Opioid Receptors Caused by Remifentanil or Fentanyl: In Vitro Assay with Cells and Three-Dimensional Structural Analyses.

Remifentanil (REM) and fentanyl (FEN) are commonly used analgesics that act by activating a µ-opioid receptor (MOR). Although optimal concentrations of REM can be easily maintained during surgery, it is sometimes switched to FEN for optimal pain regulation. However, standards for this switching protocol remain unclear. Opioid anesthetic efficacy is decided in part by MOR desensitization; thus, in this study, we investigated the desensitization profiles of REM and FEN to MOR. The efficacy and potency during the 1st administration of REM or FEN in activating the MOR were almost equal. Similarly, in ß arrestin recruitment, which determines desensitization processes, they showed no significant differences. In contrast, the 2nd administration of FEN resulted in a stronger MOR desensitization potency than that of REM, whereas REM showed a higher internalization potency than FEN. These results suggest that different ß arrestin-mediated signaling caused by FEN or REM led to their distinct desensitization and internalization processes. Our three-dimensional analysis, with in silico binding of REM and FEN to MOR models, highlighted that REM and FEN bound to similar but distinct sites of MOR and led to distinct ß arrestin-mediated profiles, suggesting that distinct binding profiles to MOR may alter ß arrestin activity, which accounts for MOR desensitization and internalization.

AlphaFold version 2.0 elucidates the binding mechanism between VIPR2 and KS-133, and reveals an S-S bond (Cys25-Cys192) formation of functional significance for VIPR2.

The vasoactive intestinal peptide receptor 2 (VIPR2) has attracted attention as a drug target for the treatment of mental disorders, cancer, and immune diseases. In 2021, we identified the peptide KS-133 as a VIPR2-selective antagonist. In this study, we aimed to elucidate the binding mechanism between VIPR2 and KS-133. To this end, VIPR2/KS-133 and VIPR2/vasoactive intestinal peptide (VIP) complex models were constructed through AlphaFold version 2.0 and molecular dynamic simulations. Our models revealed that: (i) both KS-133 and VIP have helical structures, (ii) the interaction residues on VIPR2 for both peptides are similar, and (iii) the orientation of their helices upon their binding to VIPR2 are different by ∼45°. Interestingly, in the process of constructing the aforementioned models, an S-S bond formation between Cys25 and Cys192 of the human VIPR2 was identified. Although these two Cys residues are highly conserved among species (i.e., corresponding to Cys24 and Cys191 in the mouse), no previous reports regarding this S-S bond formation exist. In order to clarify the potential role of this S-S bond in the VIPR2 has functional consequences, a cell line expressing the mouse VIPR2(Cys24Ala, Cys191Ala) was generated. During the VIP stimulation of this cell line, the phosphorylation of AKT (a downstream signal marker of VIPR2) was found to be significantly attenuated, thereby suggesting that the S-S bond has a functional significance for VIPR2. Our study not only elucidates the VIPR2-binding mechanism of KS-133 for the first time, but also provides new insights into the structural biology of VIPR2.

MDContactCom: a tool to identify differences of protein molecular dynamics from two MD simulation trajectories in terms of interresidue contacts.

SUMMARY: Comparing results from multiple MD simulations performed under different conditions is essential during the initial stages of analysis. We propose a tool called MD Contact Comparison (MDContactCom) that compares residue-residue contact fluctuations of two MD trajectories, quantifies the differences, identifies sites that exhibit large differences and visualizes those sites on the protein structure. Using this method, it is possible to identify sites affected by varying simulation conditions and reveal the path of propagation of the effect even when differences between the 3D structure of the molecule and the fluctuation RMSF of each residue is unclear. MDContactCom can monitor differences in complex protein dynamics between two MD trajectories and identify candidate sites to be analyzed in more detail. As such, MDContactCom is a versatile software package for analyzing most MD simulations. AVAILABILITY AND IMPLEMENTATION: MDContactCom is freely available for download on GitLab. The software is implemented in Python3. https://gitlab.com/chiemotono/mdcontactcom. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Inverse Mixed-Solvent Molecular Dynamics for Visualization of the Residue Interaction Profile of Molecular Probes.

To ensure efficiency in discovery and development, the application of computational technology is essential. Although virtual screening techniques are widely applied in the early stages of drug discovery research, the computational methods used in lead optimization to improve activity and reduce the toxicity of compounds are still evolving. In this study, we propose a method to construct the residue interaction profile of the chemical structure used in the lead optimization by performing "inverse" mixed-solvent molecular dynamics (MSMD) simulation. Contrary to constructing a protein-based, atom interaction profile, we constructed a probe-based, protein residue interaction profile using MSMD trajectories. It provides us the profile of the preferred protein environments of probes without co-crystallized structures. We assessed the method using three probes: benzamidine, catechol, and benzene. As a result, the residue interaction profile of each probe obtained by MSMD was a reasonable physicochemical description of the general non-covalent interaction. Moreover, comparison with the X-ray structure containing each probe as a ligand shows that the map of the interaction profile matches the arrangement of amino acid residues in the X-ray structure.

[In-Silico Drug Discovery Support-Current Situation and Challenges].

In the drug discovery field, the current problems are sluggish drug development due to the depletion of target molecules and other factors, and rising development costs. In silico drug discovery is expected to be a drug discovery support technology that will lead to the discovery of novel drug target molecules, active sites, lead compounds to more efficient development processes. In silico drug discovery can be broadly classified into methods directed by ligand information(ligand-based drug design: LBDD)and methods based on the 3-dimensional structure of target proteins(structure-based drug design: SBDD). LBDD method is based on similar structural and physicochemical properties in overall structure, or substructures and pharmacophore, using known ligands information, and has the advantage that it can be applied even when the 3-dimensional structure of the target protein is unknown. On the other hand, SBDD is a method to discovery and design compounds directed to the 3-dimensional structure of the target protein based on the'lock and key'theory, in which the target protein selects and binds to specific ligands, and has the advantage of leading to the discovery of diversity compounds. This paper outlines the basics of LBDD and SBDD, and the latest topics using AI and large-scale simulations. Furthermore, as an example of in-silico drug discovery support, in-silico drug discovery support research for the discovery of protein-protein interaction inhibitors targeting early-stage lung adenocarcinoma is also introduced.