The Human Knockout Gene CLYBL Connects Itaconate to Vitamin B12.

CLYBL encodes a ubiquitously expressed mitochondrial enzyme, conserved across all vertebrates, whose cellular activity and pathway assignment are unknown. Its homozygous loss is tolerated in seemingly healthy individuals, with reduced circulating B12 levels being the only and consistent phenotype reported to date. Here, by combining enzymology, structural biology, and activity-based metabolomics, we report that CLYBL operates as a citramalyl-CoA lyase in mammalian cells. Cells lacking CLYBL accumulate citramalyl-CoA, an intermediate in the C5-dicarboxylate metabolic pathway that includes itaconate, a recently identified human anti-microbial metabolite and immunomodulator. We report that CLYBL loss leads to a cell-autonomous defect in the mitochondrial B12 metabolism and that itaconyl-CoA is a cofactor-inactivating, substrate-analog inhibitor of the mitochondrial B12-dependent methylmalonyl-CoA mutase (MUT). Our work de-orphans the function of human CLYBL and reveals that a consequence of exposure to the immunomodulatory metabolite itaconate is B12 inactivation.

Discovery and characterization of natural product luteolin as an effective inhibitor of human pyridoxal kinase.

Pyridoxal kinase (PDXK) is an essential enzyme in the synthesis of pyridoxal 5-phosphate (PLP), the active form of vitamin B6, which plays a pivotal role in maintaining the enzyme activity necessary for cell metabolism. Thus, PDXK has garnered attention as a potential target for metabolism regulation and tumor therapy. Despite this interest, existing PDXK inhibitors have faced limitations, including weak suppressive activity, unclear mechanisms of action, and associated toxic side effects. In this study, we present the discovery of a novel PDXK inhibitor, luteolin, through a high-throughput screening approach based on enzyme activity. Luteolin, a natural product, exhibits micromolar-level affinity for PDXK and effectively inhibits the enzyme's activity in vitro. Our crystal structures reveal that luteolin occupies the ATP binding pocket through hydrophobic interactions and a weak hydrogen bonding pattern, displaying reversible characteristics as confirmed by biochemical assays. Moreover, luteolin disrupts vitamin B6 metabolism by targeting PDXK, thereby inhibiting the proliferation of leukemia cells. This research introduces a novel screening method for identifying high-affinity and potent PDXK inhibitors and sheds light on clarification of the structural mechanism of PDXK-luteolin for subsequent structure optimization of inhibitors.

Sphondin efficiently blocks HBsAg production and cccDNA transcription through promoting HBx degradation.

Hepatitis B surface antigen (HBsAg) loss and seroconversion, which is considered as functional cure of chronic Hepatitis B virus (HBV) infection, is rarely achieved even after long-term antiviral treatments. Therefore, new antiviral strategies interfering with other HBV replication steps are required, especially those that could efficiently inhibit HBsAg production. Here, we identified novel anti-HBV compounds that could potently block HBsAg expression from cccDNA by screening a natural compound library derived from Chinese traditional medical plants by a novel screening strategy. The combination of ELISA assay detecting the HBsAg and real-time PCR detecting HBV RNAs as indicator for cccDNA transcriptional activity were used. The antiviral activity of a candidate compound and underlying mechanism were evaluated in HBV-infected cells and a humanized liver mouse model. Herein, we selected a highly effective low-cytotoxic compound sphondin, which could effectively inhibit both intracellular HBsAg production and HBV RNAs levels. Moreover, we found that sphondin markedly inhibited cccDNA transcriptional activity without affecting cccDNA level. Mechanistic study found sphondin preferentially bound to HBx protein by residue Arg72, which led to increased 26S proteasome-mediated degradation of HBx. Sphondin treatment significantly reduced the recruitment of HBx to cccDNA, which subsequently led to inhibition of cccDNA transcription and HBsAg expression. The absence of HBx or R72A mutation potently abrogated the antiviral effect induced by sphondin in HBV-infected cells. Collectively, sphondin may be considered as a novel and natural antiviral agent directly targeting HBx protein, which effectively inhibited cccDNA transcription and HBsAg expression.

Inhibition of Autophagy by a Small Molecule through Covalent Modification of the LC3 Protein.

The autophagic ubiquitin-like protein LC3 functions through interactions with LC3-interaction regions (LIRs) of other autophagy proteins, including autophagy receptors, which stands out as a promising protein-protein interaction (PPI) target for the intervention of autophagy. Post-translational modifications like acetylation of Lys49 on the LIR-interacting surface could disrupt the interaction, offering an opportunity to design covalent small molecules interfering with the interface. Through screening covalent compounds, we discovered a small molecule modulator of LC3A/B that covalently modifies LC3A/B protein at Lys49. Activity-based protein profiling (ABPP) based evaluations reveal that a derivative molecule DC-LC3in-D5 exhibits a potent covalent reactivity and selectivity to LC3A/B in HeLa cells. DC-LC3in-D5 compromises LC3B lipidation in vitro and in HeLa cells, leading to deficiency in the formation of autophagic structures and autophagic substrate degradation. DC-LC3in-D5 could serve as a powerful tool for autophagy research as well as for therapeutic interventions.

YTHDF2 Binds to 5-Methylcytosine in RNA and Modulates the Maturation of Ribosomal RNA.

5-Methylcytosine is found in both DNA and RNA; although its functions in DNA are well established, the exact role of 5-methylcytidine (m5C) in RNA remains poorly defined. Here we identified, by employing a quantitative proteomics method, multiple candidate recognition proteins of m5C in RNA, including several YTH domain-containing family (YTHDF) proteins. We showed that YTHDF2 could bind directly to m5C in RNA, albeit at a lower affinity than that toward N6-methyladenosine (m6A) in RNA, and this binding involves Trp432, a conserved residue located in the hydrophobic pocket of YTHDF2 that is also required for m6A recognition. RNA bisulfite sequencing results revealed that, after CRISPR-Cas9-mediated knockout of the YTHDF2 gene, the majority of m5C sites in rRNA (rRNA) exhibited substantially augmented levels of methylation. Moreover, we found that YTHDF2 is involved in pre-rRNA processing in cells. Together, our data expanded the functions of the YTHDF2 protein in post-transcriptional regulations of RNA and provided novel insights into the functions of m5C in RNA biology.

Interaction assessments of the first S-adenosylmethionine competitive inhibitor and the essential interacting partner methylosome protein 50 with protein arginine methyltransferase 5 by combined computational methods.

Protein arginine methyltransferase 5 (PRMT5) is the most promising anticancer target in PRMT family. In this study, based on the first S-adenosylmethionine (SAM) competitive small molecule inhibitor (17, compound number is from original paper) of PRMT5 reported in our recent paper, we determined the molecular mechanism of 17 interacting with PRMT5 by computational methods. Previously reported CMP5 was also thought of as a SAM competitive inhibitor of PRMT5, but the direct inhibition activity against PRMT5 at enzymatic level was not provided. Therefore, we tested the half-maximal inhibitory concentration (IC50) of CMP5 against PRMT5 at enzymatic level for the purpose of summarizing the interaction characteristics of SAM binding site inhibitors with PRMT5. Additionally, as the essential interacting partner of PRMT5, the binding attributes of the WD-repeat-containing protein MEP50 (methylosome protein 50) was investigated, and nine key residues that contribute most to PRMT5:MEP50 interaction were identified. These results could be helpful in discovering new potent and specific inhibitors of PRMT5, as well as in designing mutant residue assay to modulate the catalytic activity of PRMT5.

Deciphering the role of dimer interface in intrinsic dynamics and allosteric pathways underlying the functional transformation of DNMT3A.

BACKGROUND: DNMT3A, as de novo DNA methyltransferase, is essential for regulating gene expression through cellular development and differentiation. The functions of DNMT3A rely on its oligomeric states and allosteric regulations between its catalytic domain and binding partners. Despite recent resolution of autoinhibitory and active DNMT3A/3L crystal structures, the mechanism of their functional motions and interdomain allostery in regulating the activity remains to be established. METHODS: The hybrid approach, comprising Elastic Network Models coupled with information theory, Protein Structure Network, and sequence evolution analysis was employed to investigate intrinsic dynamics and allosteric properties of DNMT3A resolved in autoinhibitory and active states. RESULTS: The conformational transition between two states is characterized by global motions, and the homo-dimer displays the similar dynamic properties as tetramer, acting as the basic functional unit. The hinge residues with restricted fluctuations are clustered at the dimer interface, which are predicted to enjoy remarkably efficient signal transduction properties. The allosteric pathways through the dimer interface are achieved by a cascade of interactions predominantly involving conserved and co-evolved residues. CONCLUSIONS: Our results suggest that structural topology coupled with global motions indicates the structural origin of the functional transformation of DNMT3A. The comprehensive analysis further highlights the pivotal role of the dimer interface of DNMT3A both in defining the quaternary structure dynamics and establishing interdomain communications. GENERAL SIGNIFICANCE: Understanding the global motions of DNMT3As not only provides mechanical insights into the functions of such molecular machines, but also reveals the mediators that determine their allosteric regulations.

Development of a high-throughput fluorescence polarization assay for the discovery of EZH2-EED interaction inhibitors.

Aberrant activity of enhancer of zeste homolog 2 (EZH2) is associated with a wide range of human cancers. The interaction of EZH2 with embryonic ectoderm development (EED) is required for EZH2's catalytic activity. Inhibition of the EZH2-EED complex thus represents a novel strategy for interfering with the oncogenic potentials of EZH2 by targeting both its catalytic and non-catalytic functions. To date, there have been no reported high-throughput screening (HTS) assays for inhibitors acting at the EZH2-EED interface. In this study, we developed a fluorescence polarization (FP)-based HTS system for the discovery of EZH2-EED interaction inhibitors. The tracer peptide sequences, positions of fluorescein labeling, and a variety of physicochemical conditions were optimized. The high Z' factors (>0.9) at a variety of DMSO concentrations suggested that this system is robust and suitable for HTS. The minimal sequence requirement for the EZH2-EED interaction was determined by using this system. A pilot screening of an in-house compound library containing 1600 FDA-approved drugs identified four compounds (apomorphine hydrochloride, oxyphenbutazone, nifedipine and ergonovine maleate) as potential EZH2-EED interaction inhibitors.

Identification of small molecule inhibitors targeting the SMARCA2 bromodomain from a high-throughput screening assay.

SMARCA2 is a critical catalytic subunit of the switch/sucrose non-fermenting (SWI/SNF) chromatin remodeling complexes. Dysregulation of SMARCA2 is associated with several diseases, including some cancers. SMARCA2 is multi-domain protein containing a bromodomain (BRD) that specifically recognizes acetylated lysine residues in histone tails, thus playing an important role in chromatin remodeling. Many potent and specific inhibitors targeting other BRDs have recently been discovered and have been widely used for cancer treatments and biological research. However, hit discovery targeting SMARCA2-BRD is particularly lacking. To date, there is a paucity of reported high-throughput screening (HTS) assays targeting the SMARCA2-BRD interface. In this study, we developed an AlphaScreen HTS system for the discovery of SMARCA2-BRD inhibitors and optimized the physicochemical conditions including pH, salt concentrations and detergent levels. Through an established AlphaScreen-based high-throughput screening assay against an in-house compound library, DCSM06 was identified as a novel SMARCA2-BRD inhibitor with an IC50 value of 39.9±3.0 µmol/L. Surface plasmon resonance demonstrated the binding between SMARCA2-BRD and DCSM06 (Kd=38.6 µmol/L). A similarity-based analog search led to identification of DCSM06-05 with an IC50 value of 9.0±1.4 µmol/L. Molecular docking was performed to predict the binding mode of DCSM06-05 and to decipher the structural basis of the infiuence of chemical modifications on inhibitor potency. DCSM06-05 may be used as a starting point for further medicinal chemistry optimization and could function as a chemical tool for SMARCA2-related functional studies.

Conformation and dynamics of the C-terminal region in human phosphoglycerate mutase 1.

Phosphoglycerate mutase 1 (PGAM1), an important enzyme in glycolysis, is overexpressed in a number of human cancers, thus has been proposed as a promising metabolic target for cancer treatments. The C-terminal portion of the available crystal structures of PGAM1 and its homologous proteins is partially disordered, as evidenced by weak electron density. In this study, we identified the conformational behavior of the C-terminal region of PGAM1 as well as its role during the catalytic cycle. Using the PONDR-FIT server, we demonstrated that the C-terminal region was intrinsically disordered. We applied the Monte Carlo (MC) method to explore the conformational space of the C-terminus and conducted a series of explicit-solvent molecular dynamics (MD) simulations, and revealed that the C-terminal region is inherently dynamic; large-scale conformational changes in the C-terminal segment led to the structural transition of PGAM1 from the closed state to the open state. Furthermore, the C-terminal segment influenced 2,3-bisphosphoglycerate (2,3-BPG) binding. The proposed swing model illustrated a critical role of the C-terminus in the catalytic cycle through the conformational changes. In conclusion, the C-terminal region induces large movements of PGAM1 from the closed state to the open state and influences cofactor binding during the catalytic cycle. This report describes the dynamic features of the C-terminal region in detail and should aid in design of novel and efficient inhibitors of PGAM1. A swing mechanism of the C-terminal region is proposed, to facilitate further studies of the catalytic mechanism and the physiological functions of its homologues.

Identification of novel EZH2 inhibitors through pharmacophore-based virtual screening and biological assays.

Polycomb repressive complex 2 (PRC2) acts as a primary writer for di- and tri-methylation of histone H3 at lysine 27. This protein plays an essential role in silencing gene expression. Enhancer of zeste 2 (EZH2), the catalytic subunit of PRC2, is considered as a promising therapeutic target for cancer. GSK126, a specific inhibitor of EZH2, is undergoing phase I trials for hypermethylation-related cancers. In addition, many derivatives of GSK126 are also commonly used in laboratory investigations. However, studies on the mechanism and drug development of EZH2 are limited by the absence of structural diversity of these inhibitors because they share similar SAM-like scaffolds. In this study, we generated a pharmacophore model based on reported EZH2 inhibitors and performed in silico screenings. Experimental validations led to the identification of two novel EZH2 inhibitors, DCE_42 and DCE_254, with IC50 values of 23 and 11µM, respectively. They also displayed significant anti-proliferation activity against lymphoma cell lines. Thus, we discovered potent EZH2 inhibitors with novel scaffold using combined in silico screening and experimental study. Results from this study can also guide further development of novel specific EZH2 inhibitors.

Identification of Novel Disruptor of Telomeric Silencing 1-like (DOT1L) Inhibitors through Structure-Based Virtual Screening and Biological Assays.

Histone methyltransferases are involved in many important biological processes, and abnormalities in these enzymes are associated with tumorigenesis and progression. Disruptor of telomeric silencing 1-like (DOT1L), a key hub in histone lysine methyltransferases, has been reported to play an important role in the processes of mixed-lineage leukemia (MLL)-rearranged leukemias and validated to be a potential therapeutic target. In this study, we identified a novel DOT1L inhibitor, DC_L115 (CAS no. 1163729-79-0), by combining structure-based virtual screening with biochemical analyses. This potent inhibitor DC_L115 shows high inhibitory activity toward DOT1L (IC50 = 1.5 µM). Through a process of surface plasmon resonance-based binding assays, DC_L115 was founded to bind to DOT1L with a binding affinity of 0.6 µM in vitro. Moreover, this compound selectively inhibits MLL-rearranged cell proliferation with an IC50 value of 37.1 µM. We further predicted the binding modes of DC_L115 through molecular docking analysis and found that the inhibitor competitively occupies the binding site of S-adenosylmethionine. Overall, this study demonstrates the development of potent DOT1L inhibitors with novel scaffolds.

P4HA2 hydroxylates SUFU to regulate the paracrine Hedgehog signaling and promote B-cell lymphoma progression.

Aberrations in the Hedgehog (Hh) signaling pathway are significantly prevailed in various cancers, including B-cell lymphoma. A critical facet of Hh signal transduction involves the dynamic regulation of the suppressor of fused homolog (SUFU)-glioma-associated oncogene homolog (GLI) complex within the kinesin family member 7 (KIF7)-supported ciliary tip compartment. However, the specific post-translational modifications of SUFU-GLI complex within this context have remained largely unexplored. Our study reveals a novel regulatory mechanism involving prolyl 4-hydroxylase 2 (P4HA2), which forms a complex with KIF7 and is essential for signal transduction of Hh pathway. We demonstrate that, upon Hh pathway activation, P4HA2 relocates alongside KIF7 to the ciliary tip. Here, it hydroxylates SUFU to inhibit its function, thus amplifying the Hh signaling. Moreover, the absence of P4HA2 significantly impedes B lymphoma progression. This effect can be attributed to the suppression of Hh signaling in stromal fibroblasts, resulting in decreased growth factors essential for malignant proliferation of B lymphoma cells. Our findings highlight the role of P4HA2-mediated hydroxylation in modulating Hh signaling and propose a novel stromal-targeted therapeutic strategy for B-cell lymphoma.

Adenanthin exhibits anti-inflammatory effects by covalently targeting the p65 subunit in the NF-κB signaling pathway.

Adenanthin is a structurally unique ent-kaurane diterpenoid isolated from Rabdosia adenantha, a traditional Chinese medicinal plant with potent anti-cancer and anti-inflammatory activities. However, its anti-inflammatory molecular mechanism remains largely elusive to date. Here, we developed an affinity-based label-free protein profiling (ALFPP) to identify potential covalent targets of electrophilic natural products with ketone or aldehyde groups. Using ALFPP, we identified 27 potential covalent targets of adenanthin, among which p65 (RelA) has been associated with its anti-inflammatory activities. Through a series of experiments, including LC-MS/MS, molecular docking, electrophoretic mobility shift assays (EMSA), and genome editing, we demonstrated that adenanthin could covalently modify the Cys38 residue of p65 to affect the binding of DNA to p65, thereby inhibiting the NF-κB signaling pathway. ALFPP will facilitate the target identification of electrophilic carbonylated natural products, especially those containing α, ß-unsaturated keto groups. Furthermore, the elucidation of the molecular mechanism of adenanthin will contribute to new drug development of adenanthin to treat inflammations and cancers, enhancing the possibility for its clinical application.

m6A-driven NAT10 translation facilitates fatty acid metabolic rewiring to suppress ferroptosis and promote ovarian tumorigenesis through enhancing ACOT7 mRNA acetylation.

RNA epigenetic modifications have been implicated in cancer progression. However, the interplay between distinct RNA modifications and its role in cancer metabolism remain largely unexplored. Our study demonstrates that N-acetyltransferase 10 (NAT10) is notably upregulated in ovarian cancer (OC), correlating with poor patient prognosis. IGF2BP1 enhances the translation of NAT10 mRNA in an m6A-dependent manner in OC cells. NAT10 drives tumorigenesis by mediating N4-acetylcytidine (ac4C) modification of ACOT7 mRNA, thereby augmenting its stability and translation. This NAT10-ACOT7 axis modulates fatty acid metabolism in cancer cells and promotes tumor progression by suppressing ferroptosis. Additionally, our research identifies fludarabine as a small molecule inhibitor targeting NAT10, inhibits the ac4C modification and expression of ACOT7 mRNA. By using cell derived xenograft model and patient derived organoid model, we show that fludarabine effectively suppresses ovarian tumorigenesis. Overall, our study highlights the pivotal role of the NAT10-ACOT7 axis in the malignant cancer progression, underscoring the potential of targeting NAT10-mediated ac4C modification as a viable therapeutic strategy for this disease.

Itaconate protects ferroptotic neurons by alkylating GPx4 post stroke.

Neuronal ferroptosis plays a key role in neurologic deficits post intracerebral hemorrhage (ICH). However, the endogenous regulation of rescuing ferroptotic neurons is largely unexplored. Here, we analyzed the integrated alteration of metabolomic landscape after ICH using LC-MS and MALDI-TOF/TOF MS, and demonstrated that aconitate decarboxylase 1 (Irg1) and its product itaconate, a derivative of the tricarboxylic acid cycle, were protectively upregulated. Deficiency of Irg1 or depletion of neuronal Irg1 in striatal neurons was shown to exaggerate neuronal loss and behavioral dysfunction in an ICH mouse model using transgenic mice. Administration of 4-Octyl itaconate (4-OI), a cell-permeable itaconate derivative, and neuronal Irg1 overexpression protected neurons in vivo. In addition, itaconate inhibited ferroptosis in cortical neurons derived from mouse and human induced pluripotent stem cells in vitro. Mechanistically, we demonstrated that itaconate alkylated glutathione peroxidase 4 (GPx4) on its cysteine 66 and the modification allosterically enhanced GPx4's enzymatic activity by using a bioorthogonal probe, itaconate-alkyne (ITalk), and a GPx4 activity assay using phosphatidylcholine hydroperoxide. Altogether, our research suggested that Irg1/itaconate-GPx4 axis may be a future therapeutic strategy for protecting neurons from ferroptosis post ICH.

Artemisinins ameliorate polycystic ovarian syndrome by mediating LONP1-CYP11A1 interaction.

Polycystic ovary syndrome (PCOS), a prevalent reproductive disorder in women of reproductive age, features androgen excess, ovulatory dysfunction, and polycystic ovaries. Despite its high prevalence, specific pharmacologic intervention for PCOS is challenging. In this study, we identified artemisinins as anti-PCOS agents. Our finding demonstrated the efficacy of artemisinin derivatives in alleviating PCOS symptoms in both rodent models and human patients, curbing hyperandrogenemia through suppression of ovarian androgen synthesis. Artemisinins promoted cytochrome P450 family 11 subfamily A member 1 (CYP11A1) protein degradation to block androgen overproduction. Mechanistically, artemisinins directly targeted lon peptidase 1 (LONP1), enhanced LONP1-CYP11A1 interaction, and facilitated LONP1-catalyzed CYP11A1 degradation. Overexpression of LONP1 replicated the androgen-lowering effect of artemisinins. Our data suggest that artemisinin application is a promising approach for treating PCOS and highlight the crucial role of the LONP1-CYP11A1 interaction in controlling hyperandrogenism and PCOS occurrence.

A novel in vitro prognostic model of bladder cancer based on urine-derived living tumor cells.

Bladder cancer (BLCA) remains a difficult malignancy to manage because of its high recurrence, intense follow-up, and invasive diagnostic and treatment techniques. Immune checkpoint inhibitors (ICIs) have forged a new direction for the treatment of BLCA, but it is currently challenging to predict whether an individual patient will be sensitive to ICIs. We collected 43 urine/tumor samples from BLCA patients for primary bladder cancer cells (BCCs) culturing using our previously reported BCC culture platform. We used flow cytometry (FCM) to measure the expression levels of Programmed Death-Ligand 1 (PD-L1) on BCCs before and after interferon-gamma (IFN-γ) treatment and found that PD-L1 expression and the sensitivities to IFN-γ varied among patients. RNA-sequencing, western blotting, and programmed death-1 (PD-1) binding assays confirmed that the BCC FCM-based PD-L1 detection platform (BC-PD-L1) was reliable and was not hindered by the  glycosylation of PD-L1. In the subsequent retrospective study, we found that IFN-γ-stimulated PD-L1 (sPD-L1) expression on BCCs detected by BC-PD-L1 could predict the prognosis of BLCA patients. Importantly, the prognostic value was similar or even better in urine-derived BC-PD-L1 (UBC-PD-L1). Transcriptome analysis showed that BCCs with high sPD-L1 tended to enrich genes associated with the collagen-containing extracellular matrix, cell-cell adhesion, and positive regulation of the immune system. In addition, the UBC-PD-L1 also exhibited predictive value for ICI response in BLCA patients. In conclusion, as a novel personalized urine-detection method, UBC-PD-L1 may provide a rapid, accurate, and non-invasive tool for monitoring tumor progression, predicting therapeutic responses, and helping improve BLCA clinical treatment in future.

Natural product P57 induces hypothermia through targeting pyridoxal kinase.

Induction of hypothermia during hibernation/torpor enables certain mammals to survive under extreme environmental conditions. However, pharmacological induction of hypothermia in most mammals remains a huge challenge. Here we show that a natural product P57 promptly induces hypothermia and decreases energy expenditure in mice. Mechanistically, P57 inhibits the kinase activity of pyridoxal kinase (PDXK), a key metabolic enzyme of vitamin B6 catalyzing phosphorylation of pyridoxal (PL), resulting in the accumulation of PL in hypothalamus to cause hypothermia. The hypothermia induced by P57 is significantly blunted in the mice with knockout of PDXK in the preoptic area (POA) of hypothalamus. We further found that P57 and PL have consistent effects on gene expression regulation in hypothalamus, and they may activate medial preoptic area (MPA) neurons in POA to induce hypothermia. Taken together, our findings demonstrate that P57 has a potential application in therapeutic hypothermia through regulation of vitamin B6 metabolism and PDXK serves as a previously unknown target of P57 in thermoregulation. In addition, P57 may serve as a chemical probe for exploring the neuron circuitry related to hypothermia state in mice.

An optimization algorithm for conformer generation based on the bond contribution ranking.

Many works in computational drug discovery require the conformer generation of small molecules. Most existing tools aim to generate diverse conformers and deal with all of the rotatable bonds without distinction. There are some problems in existing approaches, such as the combinatorial explosion of conformers along with the number of rotatable bonds increasing and the incomplete sampling of the conformational space. Here, we present an optimized conformer generation algorithm based on the bond contribution ranking (ABCR) to find the optimal conformer under any specified scoring function. Compared with existing methods, our method can improve molecular conformational searching and protein-ligand docking performance. Meanwhile, our method has the same or broader coverage of conformational space in the global conformer sampling. Our research shows it can achieve the optima with small numbers of generated conformers and small numbers of iterations.