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Inactivation of SACE_3446, a tetR family transcriptional regulator, stimulates erythromycin production in Saccharopolyspora erythraea

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Synthetic and Systems Biotechnology 2016年第1期1卷 39-46页

作者： Hang Wu Yansheng Wang Li Yuan Yongrong Mao Weiwei Wang Lin Zhu Panpan Wu Chengzhang Fu Rolf Muller david t.weaver Lixin Zhang Buchang Zhang Institute of Health Sciences School of Life SciencesAnhui UniversityHefei 230601China CAS Key Laboratory of Pathogenic Microbiology&Immunology Institute of MicrobiologyChinese Academy of SciencesBeijing 100101China Helmholtz Institute for Pharmaceutical Research Helmholtz Centre for Infection Research and Department of Pharmaceutical BiotechnologySaarland UniversityP.O.Box 1511566041 SaarbruckenGermany

Erythromycin A is a widely used antibiotic produced by Saccharopolyspora erythraea;however,its biosynthetic cluster lacks a regulatory gene,limiting the yield enhancement via regulation engineering of ***,six tetR family transcriptional regulators(tFRs)belonging to three genomic context types were individually inactivated in *** A226,and one of them,SACE_3446,was proved to play a negative role in regulating erythromycin *** and qRt-PCR analysis revealed that SACE_3446 covering intact N-terminal DNA binding domain specifically bound to the promoter regions of erythromycin biosynthetic gene eryAI,the resistant gene ermE and the adjacent gene SACE_3447(encoding a longchain fatty-acid CoA ligase),and repressed their ***,we explored the interaction relationships of SACE_3446 and previously identified tFRs(SACE_3986 and SACE_7301)associated with erythromycin *** demonstrated relatively independent regulation mode of SACE_3446 and SACE_3986 in erythromycin biosynthesis,we individually and concomitantly inactivated them in an industrial *** *** with WB,the WBΔ3446 and WBΔ3446Δ3986 mutants respectively displayed 36%and 65%yield enhancement of erythromycin A,following significantly elevated transcription of eryAI and *** cultured in a 5 L fermentor,erythromycin A ofWBΔ3446 and WBΔ3446Δ3986 successively reached 4095 mg/L and 4670 mg/L with 23%and 41%production improvement relative to *** strategy reported here will be useful to improve antibiotics production in other industrial actinomycete.

关键词： Saccharopolyspora erythraea Erythromycin tetR family SACE_3446 Regulatory network

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Capturing the target genes of BldD in Saccharopolyspora erythraea using improved genomic SELEX method

Capturing the target genes of BldD in Saccharopolyspora eryt...

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中国生物工程学会2014年学术年会暨全国生物技术大会

作者： Hang Wu Yongrong Mao Meng Chen Hui Pan Hao Wu Jiali Li Zhongdong Xu Hualing Yuan david t.weaver Lixin Zhang Buchang Zhang Institute of Health Sciences School of Life Sciences Anhui University School of Life Sciences Hefei Normal University CAS Key Laboratory of Pathogenic Microbiology & Immunology Institute of Microbiology Chinese Academy of Sciences

BldD(SACE077), a key developmental regulator in actinomycetes, is the first identified transcriptional factor in Saccharopolyspora erythraea positively regulating erythromycin production and morphological differentiation. Although the Bld D of S. erythraea binds to the promoters of erythromycin biosynthetic genes, the interaction affinities are relatively low, implying the existence of its other target genes in S. erythraea. through the genomic SELEX method that we herein improved, four DNA sequences of S. erythraea A226, corresponding to the promoter regions of SACE306(beta-galactosidase), SACE811(50 S ribosomal protein L25), SACE410(fumarylacetoacetate hydrolase) and SACE014(aldehyde dehydrogenase), were captured with all three BldD concentrations of 0.5μM, 1μM and 2μM, while the previously identified intergenic regions of ery BIV-ery AI and ermE-eryCI plus the promoter region of SACE115, the amfC homolog for aerial mycelium formation, could be captured only when the BldD’s concentration reached 2μM. EMSA analysis indicated that BldD specifically bound to above seven DNA sequences, and qRt-PCR assay showed that the transcriptional levels of above-mentioned target genes decreased when bldD was disrupted in A226. Furthermore, SACE115 and SACE306 in A226 were individually inactivated, showing that SACE115 was predominantly involved in aerial mycelium formation, while SACE306 mainly controlled erythromycin production. this study provides valuable information for better understanding of the pleiotropic regulator BldD in ***, and the improved method may be useful for uncovering regulatory networks of other transcription factors.

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红霉素生物合成的独特调控机制解析及其遗传改造

红霉素生物合成的独特调控机制解析及其遗传改造

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中国生物工程学会2014年学术年会暨全国生物技术大会

作者：吴杭陈萌毛永荣李伟伟刘敬涛黄训端周英叶邦策张立新 david t.weaver 张部昌安徽大学生命科学学院健康科学研究院华东理工大学生物反应器工程国家重点实验室中国科学院微生物研究所病原微生物与免疫学重点实验室

红霉素A是人类医药中普遍使用的一种大环内酯类抗生素,通常是由糖多孢红霉菌进行工业化生产。然而,红霉素生物合成基因簇中不存在调控基因,这限制了我们通过调控基因的改造来提高红霉素产量。tetR家族调控基因广泛存在于原核微生物中,其... 详细信息

红霉素A是人类医药中普遍使用的一种大环内酯类抗生素,通常是由糖多孢红霉菌进行工业化生产。然而,红霉素生物合成基因簇中不存在调控基因,这限制了我们通过调控基因的改造来提高红霉素产量。tetR家族调控基因广泛存在于原核微生物中,其N和C端分别带有DNA结合和配体结合结构域,可调控抗生素合成、形态分化等重要生命过程。糖多孢红霉素基因组中共编码了101个tetR转录因子,通过本实验室此前发展一种线形DNA片段同源重组的染色体基因快速失活技术,我们在糖多孢红霉菌中获得了多个tetR家族调控基因的敲除突变株,并鉴定了其中一个基因SACE301可正调控红霉素的生物合成。实时荧光定量PCR(qRt-PCR)与凝胶迁移实验(EMSA)表明SACE301可激活红霉素合成基因ery AI和抗性基因erm E的转录,并结合它们的启动子区域,然而结合的亲和力较低。为此,我们设计了一种过表达方法,在整合型表达载体p SEt152中串联表达3个拷贝的SACE301,并导入到糖多孢红霉菌中,使其分别额外表达1,2和3个拷贝的SACE301。结果发现在过表达菌株中,随着SACE301,eryAI和erm E的转录水平的提高,红霉素A产量成比例地增加,且对红霉素的抗性也相应增强。同样地,在红霉素高产工业菌株WB中过表达SACE301,相比出发菌株,过表达菌株WB/7301, WB/2×7301 and WB/3×7301摇瓶发酵的产量分别提高了17%、29%和42%。在5 L发酵罐中进一步测试产量最高的WB/3×7301菌株,红霉素A产量达到4.23克/升,比出发菌株WB(3.32克/升)提高了约27%。本研究极大增加了我们对红霉素生物合成独特调控机制的认识,并设计了一种新的策略过表达调控基因,可显著提高红霉素的产量,也为遗传改造调控基因提高其它抗生素产量提供借鉴。

关键词：糖多孢红霉菌红霉素 tetR家族调控基因过表达

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c-Ab1以控制蛋白酶体亚基PSMA7通过泛素蛋白酶体降解调节蛋白酶体丰度

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科学新闻 2016年第1期 111-111页

作者： Dapei Li Qincai Dong Qingping tao Jing Gu Yan Cui Xuefeng Jiang Jing Yuan Weihua Li Rao Xu Yanwen Jin Ping Li david t.weaver Qingjun Ma 刘萱曹诚军事医学科学院生物工程研究所军事医学科学院病原微生物生物安全国家重点实验室中国生物工程学会

文章简介泛素—蛋白酶体是维持细胞稳态所必须的蛋白降解途径,但人们对蛋白酶体亚基自身降解的研究甚少。该课题组研究表明,20S蛋白酶体PSMA7亚基可被泛素化并通过蛋白酶体途径降解,非受体酪氨酸激酶c-Abl可通过磷酸化PSMA7的Y 106位氨... 详细信息

文章简介泛素—蛋白酶体是维持细胞稳态所必须的蛋白降解途径,但人们对蛋白酶体亚基自身降解的研究甚少。该课题组研究表明,20S蛋白酶体PSMA7亚基可被泛素化并通过蛋白酶体途径降解,非受体酪氨酸激酶c-Abl可通过磷酸化PSMA7的Y 106位氨基酸残基抑制PSMA7通过泛素一蛋白酶体降解。BRCA1是PSMA7泛素化特异的E3泛素连接酶。c-Abl/Arg通过调节细胞内PSMA7的供应调控蛋

关键词：蛋白酶体亚基泛素化 c-Ab1 PSMA7 丰度

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Functional diversity of small-mammal postcrania is linked to both substrate preference and body size

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Current Zoology 2020年第5期66卷 539-553页

作者： Lucas N.weaver david M.Grossnickle Department of Biology Life Sciences BuildingUniversity of WashingtonSeattleWA 98195USA

Selective pressures favor morphologies that are adapted to distinct ecologies,resulting in trait partition!ng among ***,the effects of these selective pressures vary across taxa,especially because morphology is also influenced by factors such as phylogeny,body size,and functional *** this study,we examine how these factors impact functional diversification in *** has been proposed that trait partitioning among mammalian ecomorphotypes is less pronoun ced at small body sizes due to biomecha nical,energetic,and environ mental factors that favor a"generalist"body plan,whereas larger taxa exhibit more substantial functional *** title this the Diverge nee Hypothesis(DH)because it predicts greater morphological divergence among ecomorphotypes at larger body *** test DH by using phylogenetic comparative methods to examine the postcranial skeletons of 129 species of taxonomically diverse,small-tomedium-sized(tegorize as either"tree-dwellers"or"ground-dwellers."In some analyses,the morphologies of ground-dwellers and tree-dwellers suggest greater between-group differentiation at larger sizes,providing some evidence for ***,this trend is n either particularly strong nor supported by all an *** stead,a more pronoun ced patter n emerges that is distinct from the predictions of DH:within-group phenotypic disparity increases with body size in both ground-dwellers and tree-dwellers,driven by morphological outliers among"medium'-sized ***,evolutionary increases in body size are more closely linked to increases in within-locomotor-group disparity tha n to in creases in betwee n-group *** discuss biomechanical and ecological factors that may drive these evolutionary patter ns,and we emphasize the significant evolutionary influences of ecology and body size on phenotypic diversity.

关键词： adaptive landscapes ecomorphology morphometries phenotypic diversity phylogenetic comparative methods trait partitioning

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Late Ordovician Mass Extinction: Earth, fire and ice

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National Science Review 2024年第1期11卷 51-65页

作者： david A.t.Harper Palaeoecosystems Group Department of Earth SciencesDurham University

the Late Ordovician Mass Extinction was the earliest of the ‘big’ five extinction events and the earliest to affect the trajectory of metazoan life. two phases have been identified near the start of the Hirnantian period and in the middle. It was a massive taxonomic extinction, a weak phylogenetic extinction and a relatively benign ecological extinction. A rapid cooling, triggering a major ice age that reduced the temperature of surface waters, prompted a drop in sea level of some 100 m and introduced toxic bottom waters onto the shelves. these symptoms of more fundamental planetary processes have been associated with a range of factors with an underlying driver identified as volcanicity. Volcanic eruptions, and other products, may have extended back in time to at least the Sandbian and early Katian, suggesting the extinctions were more protracted and influential than hitherto documented.

关键词： Ordovician LOME paleoecology palaeobiogeography biotas

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Combining innovative science and policy to improve air quality in cities with refining and chemicals manufacturing： the case study of Houston, texas, USA

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Frontiers of Chemical Science and Engineering 2017年第3期11卷 293-304页

作者： david t. Allen University of texas Austin Department of Chemical Engineering and Center for Energy and Environmental Resources Austin TX 78759 USA

In Houston, a combination of urban emissions from a city of 4 million people, coupled with emissions from extensive petroleum refining and chemical manufacturing, leads to conditions for photochemistry that are unique in the United States, and historically, the city had experienced some of the highest ozone concentrations recorded in the United States. Large air quality field studies （the texas Air Quality Studies or texAQS I and II） were conducted to determine root causes of the high ozone concentrations. Hundreds of air quality investigators, from around the world, deployed instruments on aircraft, on ships, and at fixed ground sites to make extensive air quality measurements; detailed photochemical modeling was used to interpret and assess the implications of the measurements. the texas Air Quality Studies revealed that both continuous and episodic emissions of light alkenes, which came to be called highly reactive volatile organic compounds, played a critical role in the formation of ozone and other photochemical oxidants in the region. Under- standing and quantifying the role of these emissions in regional air quality required innovations in characterizing emissions and in photochemical modeling. Reducing emissions required innovative policy approaches. these coupled scientific and policy innovations are described, and the result, substantially cleaner air for Houston, is documented. the lessons learned from the Houston air quality experience are relevant to cities with similar population and industrial profiles around the world.

关键词： ozone air quality highly reactive volatile organic compounds Houston

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Influence of incubation temperature on sea turtle hatchling quality

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Integrative Zoology 2017年第5期12卷 352-360页

作者： david t.BOOtH School of Biological Sciences The University of QueenslandSt LuciaQueenslandAustralia

Since the 1980s it has been known that incubation temperature influences the sex ratio of sea turtle hatchlings emerging from their nests,and there has been much speculation on how global climate change might threaten sea turtle populations by raising nest temperatures and causing highly female-biased hatchling sex *** recently,studies have indicated that incubation temperature can also influence the size and locomotor performance of sea turtle *** I review recent studies that have explored the influence of incubation temperature on sea turtle hatchling quality in terms of hatchling morphology and locomotor performance.I also discuss the likely underlying mechanisms responsible for incubation temperature-induced differences in hatchling locomotor performance,and how an increase in nest temperature associated with global warming might affect recruitment of sea turtle hatchlings.

关键词： locomotor performance nest temperature phenotype sea turtle hatchling

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Public Risk Perception Attitude and Information-Seeking Efficacy on Floods:A Formative Study for Disaster Preparation Campaigns and Policies

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International Journal of Disaster Risk Science 2020年第5期11卷 592-601页

作者： Do Kyun david Kim t.Phillip Madison Department of Communication University of Louisiana at LafayetteLafayette.LA 70503USA

Floods are among the most frequent and devastating natural hazards and disasters in many southern states in the United *** study examined the relationship and reciprocal predictability between two theoretical constructs-risk perception attitude(RPA)and informationseeking efficacy(ISE)——in regard to pluvial *** addition,this study extended these theoretical constructs to investigate differences in RPA and ISE among potential audience segments,providing practitioners with applicable insights for designing effective flood prevention and risk management *** of data from 716 residents in south Louisiana revealed a statistically strong relationship between RPA and *** research also identified specific audience segments that would benefit from an increase in RPA and ISE concerning *** meaningful findings infonn a discussion of the theoretical and practical implications of the relationship between RPA and ISE and guide future disaster preparation campaigns and policies.

关键词： Communication campaign Disaster preparation Flood victims Information-seeking efficacy Pluvial flood Risk perception attitude

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Biobanking and omics

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Frontiers in Biology 2018年第4期13卷 287-292页

作者： david t. Harris Executive Director Biorepository University of Arizona PO Box 245221. AHSC 6122 Tucson AZ 85724 USA

BACKGROUND： the ＂Era of Big Data＂ and ＂Precision Medicine＂ is now upon us. that is, interrogation of large data sets obtained from groups of similar patients or from the patient themselves over time will now hypothetically permit therapies to be designed to provide maximal efficacy with minimal side effects. However, such discoveries depend upon recruitment of very large numbers of subjects （tens of thousands） along with their associated biospecimens and medical records. When considering the establishment of a biobank or the refocusing of an existing repository for the purpose of＂omics＂ research （i.e., genomics, metabolomics, proteomics, microbiomics, etc.） and/or precision medicine, there are a number of considerations to ponder. Each of these facets is discussed. OBJECtIVE： the objective of this review is to describe best practices for the establishment and operations ofa biobank that will be used for omics （genomics, proteomics, metabolomics, microbiomics） analyses based on published literature and our own practical experiences. MEtHODS： We describe the most commonly described approaches to a variety of biobanking issues, including our own practical experiences over the past 5 years. RESULtS： Based on the particular biobanking situation and downstream application, we have described best practices based on the literature and own experience, taking into consideration ease of application and costs. CONCLUSIONS： the banking of various types of clinical biospecimens has many valuable uses but often depends on overall costs versus sample utility. In addition, specimen flexibility is important but is influenced by the ease or difficulty of the application. It is always preferable to collect and stored a biospecimen in a format that allows for multiple types of downstream analyses, but that often requires additional expertise, equipment and reagents that can increase overall costs. We have described the methodologies most successfully applied to many situations.

关键词： biobanking genomics metabolomics microbiomics methodology precision medicine big data

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