新型多巴胺D3受体抑制剂的发现

根据最新解析的多巴胺D3受体的晶体结构进行活性位点分析, 建立了基于受体的药效团模型, 并对Asinex Gold Collection 数据库进行筛选, 选择7个化合物进行生物活性测试, 得到了高活性新型多巴胺受体抑制剂(04932482), 它对多巴胺D3受体抑制率达85.45%, 其Ki值为(806.75±34.58) nmol/L. 进一步对活性化合物进行结构分析, 研究了其与受体相互作用的模式, 并以此为指导提出以04932482为先导化合物进行结构改造的方向.     

多管藻中具有PTP1B酶抑制活性的新化合物

应用正相硅胶、凝胶SephadexLH20柱色谱和反相HPLC等方法,从多管藻(Polysiphoniaurceolata)中分离得到6个溴酚类化合物,通过IR、MS、1D和2DNMR等波谱技术分析鉴定,发现一个有明显的PTP1B抑制活性(IC50=4·9μg/mL)的新化合物:3-溴-4-[3-溴-4,5-二羟基苯基]甲基-5-羟甲基-1,2-二苯酚(1)。另5个已知化合物分别为3-溴-4,5-二羟基苄基乙基醚(2)、3-溴-4,5-二羟基苯甲醇(3)、3-溴-4,5-二羟基苯甲醛(4)、3,5-二溴-4-羟基苯甲醛(5)、3,5-二溴-4-羟基苯甲醇(6)。     

丝印电极法体外筛选黄嘌呤氧化酶抑制剂方法研究

建立丝印电极快速筛选黄嘌呤氧化酶(XO)抑制剂的新方法. 利用丝网印刷技术制作一次性使用的丝印碳糊电极, 采用吸附法将羧基化的多壁碳纳米管(MWNTs)修饰在电极表面, 建立测定尿酸(UA)的简单快捷的计时电流分析方法, 将该法应用于XO抑制剂的体外筛选. 在0.3 V (vs. Ag/AgCl)的工作电位下, UA在MWNTs修饰的丝印电极上产生灵敏的响应电流, 响应时间30 s, 测定UA的线性范围为2～300 μmol•L^－1, 线性方程为Y (μmol•L^－1)＝43.8240X (μA)－0.1592, r＝0.9998, 最低检出限为1 μmol•L^－1. 用该法对4种中药水提物进行了筛选, 相对空白对照组, 桑寄生500 μg•mL^－1浓度组有显著性差异. 该法简单、快速, 电极制作成本低, 所需仪器简单, 适用于体外大量筛选XO抑制剂.     

Inhibitory Activity and Mechanism Investigation of Hypericin as a Novel α-Glucosidase Inhibitor

α-glucosidase is a major enzyme that is involved in starch digestion and type 2 diabetes mellitus. In this study, the inhibition of hypericin by α-glucosidase and its mechanism were firstly investigated using enzyme kinetics analysis, real-time interaction analysis between hypericin and α-glucosidase by surface plasmon resonance (SPR), and molecular docking simulation. The results showed that hypericin was a high potential reversible and competitive α-glucosidase inhibitor, with a maximum half inhibitory concentration (IC₅₀) of 4.66 ± 0.27 mg/L. The binding affinities of hypericin with α-glucosidase were assessed using an SPR detection system, which indicated that these were strong and fast, with balances dissociation constant (KD) values of 6.56 × 10⁻⁵ M and exhibited a slow dissociation reaction. Analysis by molecular docking further revealed that hydrophobic forces are generated by interactions between hypericin and amino acid residues Arg-315 and Tyr-316. In addition, hydrogen bonding occurred between hypericin and α-glucosidase amino acid residues Lys-156, Ser-157, Gly-160, Ser-240, His-280, Asp-242, and Asp-307. The structure and micro-environment of α-glucosidase enzymes were altered, which led to a decrease in α-glucosidase activity. This research identified that hypericin, an anthracene ketone compound, could be a novel α-glucosidase inhibitor and further applied to the development of potential anti-diabetic drugs.     

Biosynthesis of Phomactin Platelet Activating Factor Antagonist Requires a Two-Enzyme Cascade

Phomactin diterpenoids possess a unique bicyclo[9.3.1]pentadecane skeleton with multiple oxidative modifications, and are good platelet-activating factor (PAF) antagonists that can inhibit PAF-induced platelet aggregation. In this study, we identified the gene cluster (phm) responsible for the biosynthesis of phomactins from a marine fungus, Phoma sp. ATCC 74077. Despite the complexity of their structures, phomactin biosynthesis only requires two enzymes: a type I diterpene cyclase PhmA and a P450 monooxygenase PhmC. PhmA was found to catalyze the formation of the phomactatriene, while PhmC sequentially catalyzes the oxidation of multiple sites, leading to the generation of structurally diverse phomactins. The rearrangement mechanism of the diterpene scaffold was investigated through isotope labeling experiments. Additionally, we obtained the crystal complex of PhmA with its substrate analogue FGGPP and elucidated the novel metal-ion-binding mode and enzymatic mechanism of PhmA through site-directed mutagenesis. This study provides the first insight into the biosynthesis of phomactins, laying the foundation for the efficient production of phomactin natural products using synthetic biology approaches.     

Discovery of a Novel Inhibitor Structure of Mycobacterium tuberculosis Isocitrate Lyase

Tuberculosis remains a global threat to public health, and dormant Mycobacterium tuberculosis leads to long-term medication that is harmful to the human body. M. tuberculosis isocitrate lyase (MtICL), which is absent in host cells, is a key rate-limiting enzyme of the glyoxylic acid cycle and is essential for the survival of dormant M. tuberculosis. The aim of this study was to evaluate natural compounds as potential MtICL inhibitors through docking and experimental verification. Screening of the TCMSP database library was done using Discovery Studio 2019 for molecular docking and interaction analysis, with the putative inhibitors of MtICL, 3-BP, and IA as reference ligands. Daphnetin (MOL005118), with a docking score of 94.8 and -CDOCKER interaction energy of 56 kcal/mol, was selected and verified on MtICL in vitro and M. smegmatis; daphnetin gave an IC₅₀ of 4.34 μg/mL for the MtICL enzyme and an MIC value of 128 μg/mL against M. smegmatis, showing enhanced potential in comparison with 3-BP and IA. The interactions and essential amino acid residues of the protein were analyzed. In summary, natural daphnetin may be a promising new skeleton for the design of inhibitors of MtICL to combat dormant M. tuberculosis.     

Engineering the Substrate Specificity of a P450 Dimerase Enables the Collective Biosynthesis of Heterodimeric Tryptophan-Containing Diketopiperazines

Heterodimeric tryptophan-containing diketopiperazines (HTDKPs) are an important class of bioactive secondary metabolites. Biosynthesis offers a practical opportunity to access their bioactive structural diversity, however, it is restricted by the limited substrate scopes of the HTDKPs-forming P450 dimerases. Herein, by genome mining and investigation of the sequence-product relationships, we unveiled three important residues (F387, F388 and E73) in these P450s that are pivotal for selecting different diketopiperazine (DKP) substrates in the upper binding pocket. Engineering these residues in Nas_F5053 significantly expanded its substrate specificity and enabled the collective biosynthesis, including 12 self-dimerized and at least 81 cross-dimerized HTDKPs. Structural and molecular dynamics analysis of F387G and E73S revealed that they control the substrate specificity via reducing steric hindrance and regulating substrate tunnels, respectively.     

Novel Dual‐Site‐Binding Neuraminidase Inhibitor from Virtual Screening by Pharmacophore and Molecular Dynamics Methods

Neuraminidase is a significant anti‐influenza target that plays crucial role in virus replication cycle. The discovery of 150‐cavity in Group‐1 neuraminidase provides us a novel mentality of designing inhibitor which can bind with both conserved site and 150‐cavity. In order to discover novel dual‐site‐binding inhibitors, a 3D chemical‐feature‐based pharmacophore model was established to cover dual‐site in neuraminidase. The dual‐site‐binding model was consistent in predicting the binding conformation of Group‐1 neuraminidase inhibitor and applied for virtual screening of Specs database. Compound 4 (ZINC05790048) that aligned well to the model was selected after multiple filtrations for molecular dynamics simulations, indicating improved binding energy with neuraminidase. It can sever as the lead compound for a novel series of inhibitors.     

合成SGLT2抑制剂埃格列净的新方法

以D-(+)-葡萄糖酸内酯为原料,经三甲硅基保护羟基后与5-溴-2-氯-4′-乙氧基二苯甲烷偶联制得(2S,3R,4S,5S,6R)-2-［4-氯-3-(4-乙氧苄基)苯基］-6-(羟甲基)-2-甲氧基四氢-2H-吡喃-3,4,5-三醇（2）; 2经羟基保护、氧化和羟醛缩合等5步反应制得(3S,4S,5R,6S)-3,4,5-三(苄氧基)-6-［4-氯-3-(4-乙氧苄基)苯基］-2-(羟甲基)-6-甲氧基四氢-2H-吡喃-2-甲醛（7）; 7经还原、脱苄同时关环制得埃格列净(1S,2S,3S,4R,5S)-5-［4-氯-3-(4-乙氧苄基)苯基］-1-(羟甲基)-6,8-二氧杂二环［3.2.1］辛烷-2,3,4-三醇,其结构经¹H NMR和LC-MS表征。     

Biosynthesis of Cosmosporasides Reveals the Assembly Line for Fungal Hybrid Terpenoid Saccharides

Fungal hybrid terpenoid saccharides constitute a new and growing family of natural products with significant biomedical and agricultural activities. One representative family is the cosmosporasides, which feature oxidized terpenoid units and saccharide moieties; however, the assembly line of these building blocks has been elusive. Herein, a cos cluster from Fusarium orthoceras was discovered for the synthesis of cosmosporaside C ( 1 ) by genome mining. A UbiA family intramembrane prenyltransferase (UbiA-type PT), a multifunctional cytochrome P450, an α,β-hydrolase, an acetyltransferase, a dimethylallyl transferase (DMAT-type PT) and a glycosyltransferase function cooperatively in the assembly of the scaffold of 1 using primary central metabolites. The absolute configuration at C4, C6 and C7 of 1 was also established. Our work clarifies the unexpected functions of UbiA-type and DMAT-type PTs and provides an example for understanding the synthetic logic of hybrid terpenoid saccharides in fungi.     

Biosynthesis of Indole Diterpene Lolitrems: Radical-Induced Cyclization of an Epoxyalcohol Affording a Characteristic Lolitremane Skeleton

Lolitrems are tremorgenic indole diterpenes that exhibit a unique 5/6 bicyclic system of the indole moiety. Although genetic analysis has indicated that the prenyltransferase LtmE and the cytochrome P450 LtmJ are involved in the construction of this unique structure, the detailed mechanism remains to be elucidated. Herein, we report the reconstitution of the biosynthetic pathway for lolitrems employing a recently established genome-editing technique for the expression host Aspergillus oryzae. Heterologous expression and bioconversion of the various intermediates revealed that LtmJ catalyzes multistep oxidation to furnish the lolitrem core. We also isolated the key reaction intermediate with an epoxyalcohol moiety. This observation allowed us to establish the mechanism of radical-induced cyclization, which was firmly supported by density functional theory calculations and a model experiment with a synthetic analogue.     

A Novel Organophosphorus Compound as a Coupling Reagent for Peptide Synthesis

3-(Diethoxyphosphoryloxy)-1,2,3-benzotriazin-4 (3H)-one 1 is a new organophosphorus compound which can be used as an efficient coupling reagent for peptide synthesis by either solution or solid phase coupling.

Standard abbreviations for amino acids and protecting groups follow the IUPAC-IUB Joint Commission on Biochemistry Nomenclature: J. Biol. Chem. 1971, 247, 977.     

Newly Designed Quinazolinone Derivatives as Novel Tyrosinase Inhibitor: Synthesis,Inhibitory Activity,and Mechanism

We synthesized a series of quinazolinone derivates as tyrosinase inhibitors and evaluated their inhibition constants. We synthesized 2-(2,6-dimethylhepta-1,5-dien-1-yl)quinazolin-4(3H)-one (Q1) from the natural citral. The concentration, which led to 50% activity loss of Q1, was 103 ± 2 μM (IC₅₀ = 103 ± 2 μM). Furthermore, we considered Q1 to be a mixed-type and reversible tyrosinase inhibitor, and determined the K_I and K_IS inhibition constants to be 117.07 μM and 423.63 μM, respectively. Our fluorescence experiment revealed that Q1 could interact with the substrates of tyrosine and L-DOPA in addition to tyrosinase. Molecular docking studies showed that the binding of Q1 to tyrosinase was driven by hydrogen bonding and hydrophobicity. Briefly, the current study confirmed a new tyrosinase inhibitor, which is expected to be developed into a novel pigmentation drug.     

新型吡咯并[2,1-f][1,2,4]三嗪酪氨酸激酶抑制剂的设计与合成

以对甲基苯磺酰甲基异腈和反-2-丁烯酸甲酯为原料,经过11步反应合成了7个4,5,6-三取代的新型吡咯并[2,1-f][1,2,4]三嗪酪氨酸激酶抑制剂,总收率3.2%～10.0%,其结构经1H NMR和MS 表征.     

Cytochrome P450 Bioconjugate as a Nanovehicle for Improved Chemotherapy Treatment

Cancer is still a growing public health problem, especially breast cancer that is one of the most important cancers in women. Chemotherapy, even though a successful treatment, is accompanied by severe side effects. Moreover, most of the drugs used for chemotherapy are administered as prodrugs and need to be transformed to the active form by cytochromes P450 (CYPs). In addition, increasing numbers of cancer tissues show lower CYP activity than the surrounding healthy tissues in which prodrugs are preferentially activated causing cytotoxicity. Here, the design of a functionalized cytochrome P450 bioconjugate is reported as nanovehicle for the enzyme direct delivery to the tumor tissue in order to improve the local drug activation. MCF‐7 breast cancer cells are treated with CYP‐polyethylene glycol bioconjugate functionalized folic acid, where it activates the prodrug tamoxifen and significantly reduces the dose of tamoxifen needed to kill the tumor cells. The CYP bioconjugate covered with polyethylene glycol shows no immunogenic activity. The advantages of increasing the site‐specific CYP activity in tumor tissues are discussed.

     

Discovery of Quinacrine as a Potent Topo II and Hsp90 Dual-Target Inhibitor,Repurposing for Cancer Therapy

Topo II and Hsp90 are promising targets. In this study, we first verified the structural similarities between Topo IIα ATPase and Hsp90α N−ATPase. Subsequently, 720 compounds from the Food and Drug Administration (FDA) drug library and kinase library were screened using the malachite green phosphate combination with the Topo II-mediated DNA relaxation and MTT assays. Subsequently, the antimalarial drug quinacrine was found to be a potential dual−target inhibitor of Topo II and Hsp90. Mechanistic studies showed that quinacrine could specifically bind to the Topo IIα ATPase domain and inhibit the activity of Topo IIα ATPase without impacting DNA cleavage. Furthermore, our study revealed that quinacrine could bind Hsp90 N−ATPase and inhibit Hsp90 activity. Significantly, quinacrine has broad antiproliferation activity and remains sensitive to the multidrug−resistant cell line MCF−7/ADR and the atypical drug−resistant tumor cell line HL−60/MX2. Our study identified quinacrine as a potential dual−target inhibitor of Topo II and Hsp90, depending on the ATP−binding domain, positioning it as a hit compound for further structural modification.     

Structure-Based Discovery and Biological Assays of a Novel PRMT5 Inhibitor for Non-Small Cell Lung Cancer

Protein arginine methyltransferase 5 (PRMT5) is a popular anticancer target that regulates histone or nonhistone methylation and is linked to the development and poor prognosis of non-small cell lung cancer. PRMT5 inhibitors have shown great promise in clinical trials as a cancer therapy. However, most inhibitors reported recently act in a SAM-competitive mode and lack structural diversity. In this paper, a novel non-SAM inhibitor, 3039-0164, was discovered by the structure-based virtual screening method. The binding mechanism of 3039-0164 to PRMT5 was revealed via molecular docking and molecular dynamics simulations. 3039-0164 inhibited PRMT5 enzymatic activity, downregulated the expression of PRMT5 downstream target genes (FGFR3 and eIF4E), and blocked the activation of the PI3K/AKT/mTOR and ERK signaling pathways. The discovery of 3039-0164 provides precise and creative hit compounds for the design optimization of PRMT5 lead compounds in non-small cell lung cancer.     

Biosynthesis of Indole Diterpene Lolitrems: Radical‐Induced Cyclization of an Epoxyalcohol Affording a Characteristic Lolitremane Skeleton

Lolitrems are tremorgenic indole diterpenes that exhibit a unique 5/6 bicyclic system of the indole moiety. Although genetic analysis has indicated that the prenyltransferase LtmE and the cytochrome P450 LtmJ are involved in the construction of this unique structure, the detailed mechanism remains to be elucidated. Herein, we report the reconstitution of the biosynthetic pathway for lolitrems employing a recently established genome‐editing technique for the expression host Aspergillus oryzae. Heterologous expression and bioconversion of the various intermediates revealed that LtmJ catalyzes multistep oxidation to furnish the lolitrem core. We also isolated the key reaction intermediate with an epoxyalcohol moiety. This observation allowed us to establish the mechanism of radical‐induced cyclization, which was firmly supported by density functional theory calculations and a model experiment with a synthetic analogue.     

A Helquat-like Compound as a Potent Inhibitor of Flaviviral and Coronaviral Polymerases

Positive-sense single-stranded RNA (+RNA) viruses have proven to be important pathogens that are able to threaten and deeply damage modern societies, as illustrated by the ongoing COVID-19 pandemic. Therefore, compounds active against most or many +RNA viruses are urgently needed. Here, we present PR673, a helquat-like compound that is able to inhibit the replication of SARS-CoV-2 and tick-borne encephalitis virus in cell culture. Using in vitro polymerase assays, we demonstrate that PR673 inhibits RNA synthesis by viral RNA-dependent RNA polymerases (RdRps). Our results illustrate that the development of broad-spectrum non-nucleoside inhibitors of RdRps is feasible.     

一种智能型阻垢剂性能快速评定装置

针对我国目前指导水处理剂阻垢性能评定的化工行业标准HG/T2160-1991——鼓泡法及其评定设备的不足,基于国外上世纪末提出的电导滴定法,研制了一种一体化智能型阻垢剂性能的快速评定装置。该装置基于微机技术应用平台,实现了水处理剂阻垢性能评定的全程自动化:建立了滴定过程中电导率——滴定液容积变化曲线突降点位置的预测算法和相应软件,奠定了阻垢效果的智能评定基础;研制了数据采样间隔自动跟踪、匹配蠕动泵转速的自动滴定装置,提高了滴定分辨率和拐点位置确定精度;自制了高灵敏度、高精密度的半导体温度传感器及其测点位置调节部件,消除了滴定过程中温度对检测评定的干扰;定量确定了测定池系统表面积垢对检测评定的影响,拟定了相应清洗程序。与现有各评定方法和设备相比,本仪器的精准度高、重复性好、评定快捷、操作简便、容重小、能耗小,能更好满足水处理剂生产企业和众多行业用户对水处理剂阻垢性能评定的需求,有望成为鼓泡法评定设备的换代产品。