产生芳香环聚酮类天然产物放线菌的分子筛选研究

目的建立一套简便、快速、高效的芳香环聚酮类化合物产生菌基因筛选体系，为聚酮类药物筛选搭建一个新的技术平台；认识PKS—Ⅱ基因在不同环境放线菌群中的分布规律，为新药筛选中微生物资源的定向分离提供依据。方法通过对芳香环聚酮类化合物生物合成途径所用的Ⅱ型聚酮合酶氨基酸和核苷酸序列的同源性比较分析，设计一对简并引物且以微波炉法提取的基因组DNA为模板扩增KS／CLF功能域约0．8kb目的基因片段，依据放线菌基因组中Ⅱ型PKS合酶基因的存在与否标定可能的芳香环聚酮类天然产物产生菌。结果利用建立的芳香环聚酮类化合物产生菌基因筛选体系对99株嗜碱放线菌、100株链霉菌、47株嗜盐放线菌和776株一般放线菌进行了筛选，研究表明链霉菌、嗜碱放线菌、嗜盐放线菌和一般放线菌Ⅱ型聚酮合酶基因的阳性率分别为54％、29．3％、25．5％和24．1％。结论组合放线菌基因组DNA快速提取技术和PKSⅡ基因简并引物PCR扩增技术建立了快速、简便、高效的芳香环聚酮类化合物产生菌基因筛选体系并对不同环境条件的放线菌菌群的PKSⅡ基因分布进行了研究。结果表明放线菌PKSⅡ聚酮合酶分布具有广泛性；而且同时表明链霉菌是芳香环聚酮类化合物的最丰富来源；在极端环境放线菌中，嗜盐放线菌的PKSⅡ基因在中度嗜盐放线菌分布的频率远高于嗜盐放线菌，嗜碱和中度嗜盐放线菌也是芳香环聚酮类化合物潜在的丰富资源。本研究为新药筛选中新的有效菌源的确定提供了可靠依据。     

产生大环聚酮类天然产物放线菌的分子筛选研究

目的 建立一套简便、快速、高效的大环聚酮类化合物产生菌基因筛选体系，为聚酮类药物筛选搭建一个新的技术平台；认识PKS—I基因在不同环境放线菌群中的分布规律，为新药筛选中微生物资源的定向分离提供依据。方法 通过对大环聚酮类化合物生物合成途径所用的I型聚酮合成酶氨基酸和核苷酸序列的同源性比较分析，设计了一对引物用于扩增KS／AT功能域约1．6kb目的基因片段，依据放线菌基因组中I型PKS合成酶基因的存在与否标定可能的大环聚酮类天然产物产生菌。结果 利用建立的大环聚酮类化合物产生菌基因筛选体系对98株嗜碱放线菌、99株链霉菌、46株嗜盐放线菌和757株稀有放线菌进行了筛选，研究表明嗜碱放线菌、链霉菌、嗜盐放线菌和稀有放线菌I型聚酮合成酶基因的阳性率分别为26．5％、20．4％、15．2％和9．35％。结论 利用组合放线菌基因组DNA快速提取技术和PKS—I基因兼并引物PCR扩增技术建立了快速、简便、高效的大环聚酮类化合物产生菌基因筛选体系；并对不同环境条件的放线菌菌群的PKS—I基因分布进行了研究，结果 不仅表明了放线菌PKS I聚酮合成酶分布的广泛性，而且表明极端环境放线菌，尤其是嗜碱放线菌是大环聚酮类化合物潜在的丰富资源，为新药筛选有效菌源的确定提供了可靠依据。     

真菌Drechslera catenaria聚酮类代谢产物及聚酮合成酶基因的初步研究

目的 分离分析内脐蠕孢真菌(Drchslera catenaria)的次级聚酮类代谢产物,初步分析其非还原型Ⅰ型聚酮合成酶基因组成,为该菌株中大黄酚生物合成机制的阐明奠定理论基础.方法 应用Czapek Dox培养基发酵培养,酸化乙酸乙酯提取,硅胶柱层析及薄层制备等方法,对该菌株产生的聚酮类代谢产物进行分离纯化;利用紫外(UV),液相-质谱联用(LC-MS)和核磁共振谱(1H-NMR)鉴定化学结构;基于真菌聚酮合成酶保守序列设计引物,克隆与代谢产物合成相关的聚酮合成酶基因.结果与结论 从试验菌株菌丝体及发酵液中分离获得4个芳香聚酮类化合物,其结构分别为大黄酚(chrysophanol),长蠕孢素(helminthosporin),冰岛青霉素(islandicin),链蠕孢素(catenarin),其中后3个为首次从该真菌中得到.从基因组DNA中得到一段非还原性聚酮合成酶(PKS)基因序列(约5.3kb),与Pyrenophora tritici-repentis Pt-1C-BFP中黄色色素合成相关的基因片段有较高类似度,很有可能与内脐蠕孢真菌中大黄酚的生物合成有关;在距离该基因2.5kb处克隆到一段基因序列(0.79kb),与P.triticirepentis Pt-1C-BFP中β-酮酯酰基-ACP-还原酶(KR)基因完全相似,其很可能负责大黄素的6位羰基经还原形成大黄酚.     

药用微生物代谢产物筛选的新进展

微生物生长繁殖过程中产生的代谢产物种类繁多,结构各异,是寻找生物活性物质的重要天然宝库之一。自Fleming发现青霉素之后,人们不断从微生物代谢产物中筛选到有价值的抗菌药物,为人类抗感染疾病的治疗作出了极大贡献。随着抗生素数量的增加,随机筛选抗生素的重复机率高,鉴别工作量大。以致人们一度对从微生物代谢物中寻找抗生素丧失信心。近年来,生物学各相关学科的发展及各种新技术的应用,基于酶抑制剂,抗原-抗体,受体-配基相互作用原理和应用生物技术建立了许多各具特色的筛选方法,新发现抗生素的种类和数量不断增加。从微生物代谢物中寻找抗生素仍不失为新抗生素研究的重要途径之一。     

聚酮合酶与药物筛选的研究进展

由于结构多样化和生物活性多样性,聚酮化合物已经成为新药的重要来源.聚酮合酶(polyketide synthase,PKS)是合成聚酮类物质的酶复合体.本文介绍了聚酮合酶的分类、作用机制及相关药物筛选的研究进展.并对通过宏基因组技术,从自然环境中获得新聚酮化合物的基因筛选方法的研究进展进行了总结.     

小单孢菌及其产生的次级生物活性代谢产物

原核微生物小单孢菌产生了740多个生物活性物质，化学结构类型多样，生物活性各异，主要为抗生素和一些人体酶抑制剂。小单孢菌不但能产生链霉菌产生的抗生素化学类型而且还有其独到之处。最受人重视的抗生素是庆大霉素、最主要的特征是产生氨基糖苷类抗生素和大环内酯类抗生素。福建省微生物研究所自20世纪60年代成功研发庆大霉素之后不问断地从我国各地水生环境中分离到多种抗生素、几种酶抑制剂和其他生物活性物质。本文还阐述近年来国外在小单孢菌中发现的一些有实用价值和有前途的微生物新药物，特别是烯二炔类抗肿瘤抗生素。     

聚酮化合物及其组合生物合成

聚酮化合物的组合生物合成是当前国际化学与生物学交叉学科研究的热点之一，也正在发展成为药物创新超常规的重要手段。本文对近十年来聚酮化合物，特别是Ⅰ型聚酮化合物的组合生物合成的常用技术和方法学发展进行了回顾和展望。     

部分黄酮类化合物的Ⅱ相代谢产物及其药理活性研究进展

黄酮类化合物为一大类广泛存在于食物和药物中的天然产物,具有多种药理作用。近来研究表明黄酮类化合物在体内主要以苷元的Ⅱ相代谢产物形式存在,包括不同取代位点的一系列苷元的葡糖醛酸苷、硫酸酯及混合结合物。其中一些Ⅱ相代谢产物已被证明具有抗炎、抗氧化、抗肿瘤、影响代谢酶和转运体等方面的药理活性。文中对文献报道的黄酮、异黄酮、黄酮醇、二氢黄酮、黄烷-3-醇和花色素等黄酮类化合物的Ⅱ相代谢产物及其药理作用进行了综述。     

植物培养细胞次级代谢产物商产细胞系的筛选

对植物培养细胞中高产次级代谢产物细胞系筛选的方法与研究作了概述;包括高产变异系和突变体的产生和筛选等。同时对高产细胞系的稳定性及保存方法和当前研究的进展进行了讨论。     

化合物结构与代谢激活的毒性分析

药物开发的不同阶段，人们通过系列的体外试验研究、动物实验以及随后人体应用的结果试图揭示药物的毒副作用，目的是为确保人体用药的安全性。其中代谢转化诱导的毒性占据药物毒性的很大一方面，尤其反应性代谢产物引发的毒性近几年越来越引起重视。本文综述了几类代表性化合物结构产生反应性代谢产物的情况，包括苯醌亚胺、噻吩环、氮翁离子、环氧化物、硫脲等，试图从代谢角度揭示药物分子结构与药物毒性之间的关系，为药物的设计与开发提供参考。     

Discovery of angucyclinone polyketides from marine actinomycetes with a genomic DNA-based PCR assay targeting type II polyketide synthase

Angucyclinones are aromatic polyketides produced by type II polyketide synthases (PKS) and are mainly found in terrestrial actinomycetes. To discover more angucyclinones from marine actinomycetes, a genomic DNA-based PCR assay targeting type II polyketide synthases was performed. Among the 167 marine actinomycetes strains screened, twelve strains were identified as the “positive” strains possessing type II PKS-encoding genes based on the sequencing of PCR products. One of the 12 “positive” strains, Streptomyces sp. PKU-MA00218 was selected for the large-scale fermentation based on the HPLC and TLC analysis. Four angucyclinones, 6-deoxy-8-O-methylrabelomycin (1), 8-O-methylrabelomycin (2), 8-O-methyltetrangulol (3), C-ring cleavage product of angucyclinone C (4), were isolated and their structures were elucidated based on spectroscopic analyses. The isolation of angucyclinones 1–4 highlights the power of genome mining technologies based on biosynthetic knowledge in natural products discovery.     

基于网络药理学和分子对接技术解读川皮苷改善代谢综合征的作用机制（英文）

采用网络药理学方法探讨川皮苷改善代谢综合征的作用机制。首先利用TCMSP、TCMIP、TCMID、ETCM、HERB、NPASS和NPACT等数据库获取川皮苷作用靶点;在Dis Ge NET、Drug Bank等6个数据库中获取代谢综合征的相关靶点,筛选出与川皮苷作用靶点的共同部分构建PPI网络,并利用R语言对交集靶点进行GO和KEGG通路富集分析;最后对川皮苷和关键疾病靶点进行分子对接验证。结果收集到川皮苷作用靶点105个,代谢综合征相关靶点1975个。上述靶点取交集,获得了60个川皮苷改善代谢综合征的潜在靶点。PPI分析发现,川皮苷改善代谢综合征的关键靶点为TP53、MAPK8、AKT1、GSK3B、HSP90AA1、CTNNB1、JUN、AR、ESR1、CCND1、HRAS、TNF和PPARA。功能富集分析发现,脂质和动脉粥样硬化通路及糖尿病并发症中的AGE-RAGE信号通路在川皮苷改善代谢综合征过程中发挥重要作用。分子对接结果显示,川皮苷与上述13个核心基因具有很强的亲和力。综上所述,推测川皮苷通过脂质和动脉粥样硬化通路及糖尿病并发症中的AGE-RAGE信号通路发挥改善代谢综合...     

Mutagenicity of aromatic amines and amides with chemical models for cytochrome P450 in Ames assay

Chemical models for cytochrome P450, consisting of water-insoluble or water-soluble iron porphyrin plus an oxidant, have been used to detect the mutagenicity of promutagens in genotoxicity assays. The procedure for using chemical models for cytochrome P450 as substitutes for the S9 mix in the Ames assay have been already established. Aromatic amines and amides require metabolic activation by cytochrome P450 when they exert their mutagenicity in Salmonella typhimurium strains. In this study, we optimized the conditions of the assay using a water-soluble chemical model, 5,10,15,20-tetrakis(1-methylpyridinium-4-yl)porphyrinatoiron(III) pentachloride (4-MPy), plus tert-butyl hydroperoxide (t-BuOOH), magnesium monoperoxyphthalate, or iodosylbenzene, by comparing the mutagenicity of 2-aminofluorene (AF) in the Ames test. The model with 4-MPy/t-BuOOH showed the highest AF mutagenic potency. The chemical model activated 2-naphthylamine, 4-aminobiphenyl, and benzidine in S. typhimurium TA98. In aromatic amides, the model with 4-MPy/t-BuOOH weakly activated 2-acetylaminofluorene (AAF). To detect higher mutagenicity of aromatic amides, we used a higher concentration of 4-MPy/t-BuOOH by a factor of 5 over that used for aromatic amines, and then detected the mutagenicity of AAF, 2-acetylaminoanthracene, and 2-acetylamino-9-fluorenone. Furthermore, we concluded that the AAF mutagenicity in the presence of 4-MPy/t-BuOOH is derived from N-hydroxylacetylamino compounds.     

Biomonitoring of aromatic amines V: acetylation and deacetylation in the metabolic activation of aromatic amines as determined by haemoglobin binding

Aromatic amines are metabolically activated by N-oxidation of either the amine or the acetamide as a first step and esterification of the resulting N-hydroxyl derivatives as a second step. Both pathways may lead to DNA-adducts and subsequently to DNA lesions and mutations. Since the accumulation of non-acetylated adducts has been associated with tumour initiating properties, the balance between acetylation and deacetylation may greatly influence the biological effect. Hydrolysable haemoglobin adducts representing the bioavailability of N-hydroxylamines and the corresponding nitroso-derivatives were analysed following oral administration to female Wistar rats of two arylamine-acetamide couples: 4-aminobiphenyl and 2-aminofluorene, and two arylamine-acetamide-diacetamide triples: benzidine and 3,3′-dichlorobenzidine. The results show that the mono-acetamides are readily deacetylated in vivo whereas the diacetamides are not. A dynamic equilibrium is indicated to exist between acetylation and deacetylation, which depends on substrate specificity, and the role of deacetylation is emphasised. In addition, acetylation polymorphism was studied with 4-chloroaniline and 3,3′-dichlorobenzidine in slow acetylating A/J and rapid acetylating C57BL/6J mice. The slow acetylator genotype was associated with significantly higher haemoglobin-adduct levels for both arylamines. The results provide additional support for the use of haemoglobin adducts in biomonitoring as a dosimeter for the biologically active dose of arylamines/arylacetamides. Moreover, biomonitoring of haemoglobin adducts may provide information about an individual's susceptibility to the toxic and carcinogenic effects of these chemicals. Received: 19 January 1998 / Accepted: 7 April 1998     

以非洲猪瘟病毒DNA聚合酶X为靶点的海洋药物计算机虚拟筛选

摘 要：目的 从天然产物中高效快速的筛选出一批有效针对非洲猪瘟病毒的药物苗头化合物,为寻找能抑制猪瘟蔓延的生物饲料提供线索。方法 利用计算机辅助药物设计技术,以非洲猪瘟病毒。DNA聚合酶X的DNA结合区为靶点,进行虚拟筛选,在陆生和海洋天然产物中,化合物通过文献搜索和数据库查询明确部分天然产物的来源。结果 从陆生天然产物库中筛选出16个苗头化合物,其中有5个源自中国;在海洋化合物数据库MarineChem3D中得到了59个苗头化合物,其中有8个源自中国。结论 我国境内分布有13种潜在的具抑制非洲猪瘟效果的天然产物,其中大部分来源于海洋生物。     

Methemoglobin formation and binding to blood constituents as indicators for the formation,availability and reactivity of activated metabolites derived from trans-4-aminostilbene and related aromatic amines

trans-4-Aminostilbene derivatives exhibit higher acute and chronic toxicity than 4-aminobibenzyl derivatives. Yet, trans-4-aminostilbene produced less methemoglobin in female Wistar rats than 4-aminobibenzyl. This cannot be explained by differences in N-oxidation since trans-4-nitrosostilbene was also less efficient than 4-nitrosobibenzyl.The fate of intravenously injected, highly and specifically ³H-labeled trans-4- aminostilbene, cis-4-aminostilbene, 4-aminobibenzyl, trans-4-nitrosostilbene and 4-nitrosobibenzyl was investigated. The results indicate that trans-4-aminostilbene and 4-aminobibenzyl are N-oxidized to a similar extent and primary activation products of trans-4-aminostilbene appear even faster in the blood. However, intermediates originating during methemoglobin formation are more reactive and covalently bind to hemoglobin 2–3 times as much with trans-stilbene as compared to bibenzyl derivatives. As a consequence the availability of these intermediates in the cyclic process and thus methemoglobin formation is reduced.Therefore, binding to hemoglobin rather than levels of methemoglobin appears to be an indicator for the availability and reactivity of some activated aromatic amine metabolites.A preliminary account of part of this work has been published (Neumann and Wieland, 1973)     

电视时间与代谢综合征的关系研究

目的:探讨社区人群看电视时间与代谢综合征(metabolic syndrome,MS)的关系。方法:采用横断面调查方法 ,调查社区人群看电视时间并检测其主要代谢指标。结果:MS组与非代谢综合征(NMS)组相比,看电视的时间更长。与每日看电视时间少于60min的亚组相比,其余各组血压及各项主要代谢指标显著升高;每日看电视时间120～180 min及超过180 min的亚组均显著增高MS的风险[OR:1.16(1.10,1.23);1.35(1.22,1.50);P<0.01]。结论:看电视时间超过120 min/d显著增加MS风险。     

贵州地区体检人群高尿酸血症流行病学研究

目的探讨贵州地区20岁以上人群的高尿酸血症的发病情况。方法对我院2009年到2011年期间到我院进行体检的健康人群2 616例,于空腹静脉抽血查血糖、血脂、血尿酸、肾功能等,并作身高、体重、血压等测定。结果2 616例体检人群,男性1 929例,女性686例。高尿酸血症共551例,占总抽样人群21.06%,其中女性74例,占10.79%,男性477例,占24.73%。男女两组组间比较差异具有显著性(χ2=59.38,P<0.05)。男性血尿酸均值显著高于女性(t=30.23,P<0.05),女性在50岁以后血尿酸水平逐渐升高接近男性。男性高尿酸血症组年龄显著高于女性组(t=3.773,P<0.05)。高尿酸人群中患高血糖、高血压、超重、高脂血症的患病率显著高于总体人群(P<0.05)。结论贵州地区体检人群高尿酸血症较高。男性的高尿酸血症发病率高于女性,女性于更年期后血尿酸发病率逐渐接近男性。体重指数超标、高血压、高血糖、高脂血症是高尿酸血症的重要相关因素。     

The compromise of virtual screening and its impact on drug discovery

Introduction: Docking and structure-based virtual screening (VS) have been standard approaches in structure-based design for over two decades. However, our understanding of the limitations, potential, and strength of these techniques has enhanced, raising expectations.

Areas covered: Based on a survey of reports in the past five years, we assess whether VS: (1) predicts binding poses in agreement with crystallographic data (when available); (2) is a superior screening tool, as often claimed; (3) is successful in identifying chemical scaffolds that can be starting points for subsequent lead optimization cycles. Data shows that knowledge of the target and its chemotypes in postprocessing lead to viable hits in early drug discovery endeavors.

Expert opinion: VS is capable of accurate placements in the pocket for the most part, but does not consistently score screening collections accurately. What matters is capitalization on available resources to get closer to a viable lead or optimizable series. Integration of approaches, subjective hit selection guided by knowledge of the receptor or endogenous ligand, libraries driven by experimental guides, validation studies to identify the best docking/scoring that reproduces experimental findings, constraints regarding receptor–ligand interactions, thoroughly designed methodologies, and predefined cutoff scoring criteria strengthen VS’s position in pharmaceutical research.