利用“移树养苗”策略挖掘粤蓝链霉菌隐性活性次级代谢产物

doi:10.13560/j.cnki.biotech.bull.1985.2017-0276

生物技术通报 ›› 2017, Vol. 33 ›› Issue (9): 145-152.doi: 10.13560/j.cnki.biotech.bull.1985.2017-0276

利用“移树养苗”策略挖掘粤蓝链霉菌隐性活性次级代谢产物

林海州¹,²,³ 陈洲琴² 王燕⁴ 郭俊² 朱红惠² 邓名荣²

1. 中国科学院南海海洋研究所,广州 510301；
2. 广东省微生物研究所省部共建华南应用微生物国家重点实验室广东省菌种保藏与应用重点实验室广东省微生物应用新技术公共实验室,广州 510070；
3. 中国科学院大学,北京 100049；
4. 广东省科技图书馆,广州 510070

收稿日期:2017-04-07 出版日期:2017-09-01 发布日期:2017-09-15
作者简介:林海州,男,硕士研究生,研究方向：天然产物化学；E-mail：linhaizhou2016@163.com
基金资助:
国家自然科学基金项目（31470188）,广东省科技计划项目（2013B010102017,2015A020211022,2016A010105013）,广州市科技计划项目（201504010035）,广东省科学院科研平台环境与能力建设专项资金项目（2016GDASPT-0302）

Mining the Cryptic Bioactive Secondary Metabolites from Streptomyces vietnamensis Using a‘Tree-Removal’Strategy

LIN Hai-zhou¹,²,³ CHEN Zhou-qin² WANG Yan⁴ GUO Jun² ZHU Hong-hui² DENG Ming-rong²

1. South China Sea Institute of Oceanology,the Chinese Academy of Sciences,Guangzhou 510301；
2. State Key Laboratory of Applied Microbiology Southern China,Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application,Guangdong Open Laboratory of Applied Microbiology,Guangdong Institute of Microbiology,Guangzhou 510070；
3. University of Chinese Academy of Sciences,Beijing 100049；
4. Guangdong Science and Technology Library,Guangzhou 510070

Received:2017-04-07 Published:2017-09-01 Online:2017-09-15

摘要/Abstract

摘要：

利用适当的策略激活粤蓝链霉菌中潜在的隐性次级代谢途径,以期获得新的活性物质。以粤蓝链霉菌突变株DMR1为研究对象,该突变株的主导产物榴菌素的关键生物合成基因gra-orf1-3被敲除,不能产生榴菌素。通过改变营养条件、添加诱导物和热激等方法处理菌株,利用琼脂糖扩散法及TLC-生物显影等方法检测发酵粗提物的抑菌活性。结果显示,突变株DMR1的高氏一号和ISP3培养基发酵粗提物对枯草芽孢杆菌ATCC6633、金黄色葡萄球菌ATCC6538以及耐药的金黄色葡萄球菌ATCC43300和206具有显著抑菌活性；添加3% DMSO诱导后,高氏一号和ISP3培养基的发酵粗提物不仅对上述菌株的抑制活性增强,还具有抑制大肠杆菌ATCC8739、白色念珠菌ATCC10231以及多株耐药菌大肠杆菌和沙门氏菌的活性。粤蓝链霉菌中新发现的活性物质对革兰氏阳性菌、革兰氏阴性菌以及酵母菌具有广泛的抗菌活性,且部分发酵粗提物对所有测试的耐药菌均具有显著活性,有可能是新型的抗生素。结果也表明“移树养苗”策略能够提高隐性活性次级代谢产物的挖掘效率。

关键词: 隐性次级代谢产物, “移树养苗”策略, 激活, 多重耐药菌, 粤蓝链霉菌

Abstract:

This study aims to discover novel bioactive substances by activating the cryptic biosynthetic gene clusters of secondary metabolites from Streptomyces vietnamensis with appropriate strategies. The study was carried out in a S. vietnamensis mutant DMR1. In this mutant,the key biosynthetic genes,gra-orf1-3,of the dominant product were knocked out,resulting in deficiency in granaticin production. The strain was subject to nutrient condition changing,inducer adding and heat shocking. The antimicrobial activities of the fermentation crude extracts were tested using agarose diffusion method and TLC-bioautography. Results showed that the fermentation crude extracts of Gause No.1 and ISP3 presented significant inhibitory effects against Bacillus subtilis ATCC6633,Staphylococcus aureus ATCC6538 as well as multidrug resistant Staphylococcus aureus strains ATCC43300 and 206. With adding 3% DMSO to the media,not only did the activities of the fermentation crude extracts of Gause No.1 and ISP3 against the afore said strains increase,but also the spectrum broadened into against Escherichia coli ATCC8739,Candida albicans ATCC10231 and multidrug resistant E. coli and Salmonella sp. strains. The newly discovered bioactive substances from S. vietnamensis showed extensive activities against both gram-positive and gram-negative bacteria as well as yeast. More importantly,some of the crude extracts were significantly active against all the tested drug-resistant strains. These results indicate a possibility of a novel type of antibiotic. The results presented in this paper illustrate that the so-called ‘tree-removal’ strategy can improve the mining efficiency of the cryptic bioactive secondary metabolites.

Key words: cryptic secondary metabolites, ‘tree-removal&rsquo, strategy, activation, multidrug resistant bacterium, Streptomyces vietnamensis

林海州, 陈洲琴王燕郭俊朱红惠邓名荣. 利用“移树养苗”策略挖掘粤蓝链霉菌隐性活性次级代谢产物[J]. 生物技术通报, 2017, 33(9): 145-152.

LIN Hai-zhou, CHEN Zhou-qin WANG Yan GUO Jun ZHU Hong-hui DENG Ming-rong. Mining the Cryptic Bioactive Secondary Metabolites from Streptomyces vietnamensis Using a‘Tree-Removal’Strategy[J]. Biotechnology Bulletin, 2017, 33(9): 145-152.

参考文献

[1] Butler MS, Robertson AAB, Cooper MA. Natural product and natural product derived drugs in clinical trials[J] . Natural Product Reports, 2014, 31（11）：1612-1661.
[2] Newman DJ, Cragg GM. Natural products as sources of new drugs over the 30 years from 1981 to 2010[J] . Journal of Natural Products, 2012, 75（3）：311-335.
[3] Li JWH, Vederas JC. Drug discovery and natural products：End of an era or an endless frontier？[J] . Science, 2009, 325（5937）：161-165.
[4] Bentley SD, Chater KF, Cerdeno-Tarraga AM, et al. Complete genome sequence of the model actinomycete Streptomyces coelicolor A3（2）[J] . Nature, 2002, 417（6885）：141-147.
[5] Mardis ER. The impact of next-generation sequencing technology on genetics[J] . Trends in Genetics, 2008, 24（3）：133-141.
[6] Shen B. A new golden age of natural products drug discovery[J] . Cell, 2015, 163（6）：1297-1300.
[7] Bode HB, Bethe B, H?fs R, et al. Big effects from small changes：Possible ways to explore nature’s chemical diversity[J] . Chembiochem, 2002, 3（7）：619-627.
[8] Mearns-Spragg A, Bregu M, Boyd KG, et al. Cross-species induction and enhancement of antimicrobial activity produced by epibiotic bacteria from marine algae and invertebrates, after exposure to terrestrial bacteria[J] . Letters in Applied Microbiology, 1998, 27（3）：142-146.
[9] Shima J, Hesketh A, Okamoto S, et al. Induction of actinorhodin production by rpsL（encoding ribosomal protein S12）mutations that confer streptomycin resistance in Streptomyces lividans and Streptomyces coelicolor A3（2）[J] . Journal of Bacteriology, 1996, 178（24）：7276-7284.
[10] Zarins-Tutt JS, Barberi TT, Gao H, et al. Prospecting for new bacterial metabolites：a glossary of approaches for inducing, activating and upregulating the biosynthesis of bacterial cryptic or silent natural products[J] . Natural Product Reports, 2016, 33（1）：54-72.
[11] Rutledge PJ, Challis GL. Discovery of microbial natural products by activation of silent biosynthetic gene clusters[J] . Nature Reviews Microbiology, 2015, 13（8）：509-523.
[12] 齐志, 孙东昌, 裘娟萍. 微生物沉默基因簇激活方法的研究进展[J] . 科技通报, 2016, 32（2）：82-86.
[13] Gomez-Escribano JP, Song L, Fox DJ, et al. Structure and biosynthesis of the unusual polyketide alkaloid coelimycin P1, a metabolic product of the cpk gene cluster of Streptomyces coelicolor M145[J] . Chemical Science, 2012, 3（9）：2716-2720.
[14] Zhu HH, Guo J, Yao Q, et al. Streptomyces vietnamensis sp. nov., a streptomycete with violet blue diffusible pigment isolated from soil in Vietnam[J] . International Journal of Systematic and Evolutionary Microbiology, 2007, 57（8）：1770-1774.
[15] 邓名荣, 朱春华, 郭俊, 等. 粤蓝链霉菌中紫罗兰蓝色素成分的分离与鉴定[J] . 华南理工大学学报：自然科学版, 2011, 39（5）：132-137.
[16] Deng MR, Guo J, Ma LY, et al. Complete genome sequence of Streptomyces vietnamensis GIMV4. 0001T, a genetically manipulable producer of the benzoisochromanequinone antibiotic granaticin[J] . Journal of Biotechnology, 2015, 200：6-7.
[17] Deng MR, Guo J, Zhu HH. Streptomyces vietnamensis GIMV4. 0001：a granaticin-producing strain that can be readily genetically manipulated[J] . Journal of Antibiotics, 2011, 64（4）：345-347.
[18] 邓名荣, 郭俊, 朱红惠. 粤蓝链霉菌代谢产物的抗菌抗肿瘤活性及相关基因的初步研究[J] . 天然产物研究与开发, 2010, 22（3）：367-372.
[19] Romero-Rodriguez A, Robledo-Casados I, Sanchez S. An overview on transcriptional regulators in Streptomyces[J] . Biochimica Et Biophysica Acta-Gene Regulatory Mechanisms, 2015, 1849（8）：1017-1039.
[20] Gottelt M, Kol S, Gomezescribano JP, et al. Deletion of a regulatory gene within the cpk gene cluster reveals novel antibacterial activity in Streptomyces coelicolor A3（2）[J] . Microbiology, 2010, 156（8）：2343-2353.
[21] Butler MS, Blaskovich MAT, Owen JG, et al. Old dogs and new tricks in antimicrobial discovery[J] . Current Opinion in Microbiology, 2016, 33：25-34.
[22] Gavrish E, Sit Clarissa S, CaoS, et al. Lassomycin, a ribosomally synthesized cyclic peptide, kills Mycobacterium tuberculosis by targeting the ATP-dependent protease ClpC1P1P2[J] . Chemistry & Biology, 2014, 21（4）：509-518.
[23] Yan X, Ge H, Huang T, et al. Strain prioritization and genome mining for enediyne natural products[J] . mBio, 2016, 7（6）：e02104-16.

利用“移树养苗”策略挖掘粤蓝链霉菌隐性活性次级代谢产物

Mining the Cryptic Bioactive Secondary Metabolites from Streptomyces vietnamensis Using a‘Tree-Removal’Strategy

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价

[1]	吕秋谕, 孙培媛, 冉彬, 王佳蕊, 陈庆富, 李洪有. 苦荞转录因子基因FtbHLH3的克隆、亚细胞定位及表达分析[J]. 生物技术通报, 2023, 39(8): 194-203.
[2]	刘奎, 李兴芬, 杨沛欣, 仲昭晨, 曹一博, 张凌云. 青杄转录共激活因子PwMBF1c的功能研究与验证[J]. 生物技术通报, 2023, 39(5): 205-216.
[3]	刘金升, 陈振娅, 霍毅欣, 郭淑元. FACS技术在酶定向进化中的应用[J]. 生物技术通报, 2023, 39(10): 93-106.
[4]	程英, 靳明辉, 萧玉涛. 鳞翅目昆虫基因编辑技术研究进展[J]. 生物技术通报, 2020, 36(3): 18-28.
[5]	叶洲杰, 王心睿. CRISPR系统转化医学研究进展[J]. 生物技术通报, 2020, 36(11): 188-197.
[6]	侯成林, 杨艳坤, 陈嘉荔, 白仲虎. Mxr1磷酸化水平受Ptp调控机理的初步研究[J]. 生物技术通报, 2019, 35(7): 108-113.
[7]	徐杰, 黄建忠, 李力. 基因组挖掘技术及其在真菌中的研究进展[J]. 生物技术通报, 2019, 35(11): 201-207.
[8]	杨先友, 黄有军, 张通, 黄春颖. 拟南芥转录激活因子AtWRI1研究进展[J]. 生物技术通报, 2016, 32(6): 13-18.
[9]	高敬, 魏迪, 池振奋, 张癸荣, 张万明, 聂凌云. TALENs新型模块组装法及活性鉴定方法[J]. 生物技术通报, 2016, 32(1): 220-228.
[10]	裘天休, 李长红, 陈炯. 大黄鱼Wap65-2基因成熟肽区酵母双杂交诱饵载体的构建及鉴定[J]. 生物技术通报, 2015, 31(8): 108-113.
[11]	陆睿,宋绍征,祁正强,葛欣,邵宾,成勇. 重组人纤溶酶原激活剂转基因兔的制备及其表达产物的检测[J]. 生物技术通报, 2015, 31(10): 216-221.
[12]	宿明星,孙颖颢,施鹤,李秋莉. 植物非生物胁迫相关转录因子研究方法[J]. 生物技术通报, 2015, 31(1): 51-60.
[13]	周莲洁, 张富春, 王艳. 盐穗木转录因子HcSCL13转录激活的体外鉴定及植物表达载体的构建[J]. 生物技术通报, 2014, 0(5): 76-82.
[14]	曾凡才, 顾洪, 王轲, 周红. 针对ALK4基因的TALEN质粒构建与活性鉴定[J]. 生物技术通报, 2014, 0(5): 210-216.
[15]	李鑫, 王正明, 薛伟, 初明光. 棉花中一个新型转录因子GhWRI1的表达特征与功能分析[J]. 生物技术通报, 2013, 0(6): 80-86.