[논문]Sensitive and Rapid Phenotyping of Microbes With Soluble Methane Monooxygenase Using a Droplet-Based Assay

Lee, Hyewon; Baek, Ji In; Kim, Su Jin; Kwon, Kil Koang; Rha, Eugene; Yeom, Soo-Jin; Kim, Haseong; Lee, Dae-Hee; Kim, Dong-Myung; Lee, Seung-Goo

doi:10.3389/fbioe.2020.00358

[해외논문] Sensitive and Rapid Phenotyping of Microbes With Soluble Methane Monooxygenase Using a Droplet-Based Assay 원문보기

Frontiers in bioengineering and biotechnology, v.8, 2020년, pp.358 -

Lee, Hyewon (Synthetic Biology and Bioengineering Research Center, Korea Research Institute of Bioscience and Biotechnology , Daejeon , South Korea) , Baek, Ji In (Synthetic Biology and Bioengineering Research Center, Korea Research Institute of Bioscience and Biotechnology , Daejeon , South Korea) , Kim, Su Jin (Synthetic Biology and Bioengineering Research Center, Korea Research Institute of Bioscience and Biotechnology , Daejeon , South Korea) , Kwon, Kil Koang (Synthetic Biology and Bioengineering Research Center, Korea Research Institute of Bioscience and Biotechnology , Daejeon , South Korea) , Rha, Eugene (Synthetic Biology and Bioengineering Research Center, Korea Research Institute of Bioscience and Biotechnology , Daejeon , South Korea) , Yeom, Soo-Jin (Synthetic Biology and Bioengineering Research Center, Korea Research Institute of Bioscience and Biotechnology , Daejeon , South Korea South Kore) , Kim, Haseong , Lee, Dae-Hee , Kim, Dong-Myung , Lee, Seung-Goo

Abstract ▼ AI-Helper

Methanotrophs with soluble methane monooxygenase (sMMO) show high potential for various ecological and biotechnological applications. Here, we developed a high throughput method to identify sMMO-producing microbes by integrating droplet microfluidics and a genetic circuit-based biosensor system. sMMO-producers and sensor cells were encapsulated in monodispersed droplets with benzene as the substrate and incubated for 5 h. The sensor cells were analyzed as the reporter for phenol-sensitive transcription activation of fluorescence. Various combinations of methanotrophs and biosensor cells were investigated to optimize the performance of our droplet-integrated transcriptional factor biosensor system. As a result, the conditions to ensure sMMO activity to convert the starting material, benzene, into phenol, were determined. The biosensor signals were sensitive and quantitative under optimal conditions, showing that phenol is metabolically stable within both cell species and accumulates in picoliter-sized droplets, and the biosensor cells are healthy enough to respond quantitatively to the phenol produced. These results show that our system would be useful for rapid evaluation of phenotypes of methanotrophs showing sMMO activity, while minimizing the necessity of time-consuming cultivation and enzyme preparation, which are required for conventional analysis of sMMO activity.

Keyword

참고문헌 (56)

Agresti J. J. Antipov E. Abate A. R. Ahn K. Rowat A. C. Baret J. C. ( 2010 ). Ultrahigh-throughput screening in drop-based microfluidics for directed evolution. Proc. Natl. Acad. Sci. U.S.A. 107 4004 ？ 4009 . 10.1073/pnas.0910781107 20142500

상세보기
Borodina E. Nichol T. Dumont M. G. Smith T. J. Murrell J. C. ( 2007 ). Mutagenesis of the “leucine gate” to explore the basis of catalytic versatility in soluble methane monooxygenase. Appl. Environ. Microbiol. 73 6460 ？ 6467 . 10.1128/aem.00823-07 17704278

상세보기
Chistoserdova L. ( 2018 ). Applications of methylotrophs: can single carbon be harnessed for biotechnology? Curr. Opin. Biotechnol. 50 189 ？ 194 . 10.1016/j.copbio.2018.01.012 29414059

상세보기
Chistoserdova L. Kalyuzhnaya M. G. ( 2018 ). Current trends in methylotrophy. Trends Microbiol. 26 703 ？ 714 . 10.1016/j.tim.2018.01.011 29471983

상세보기
Choi S. L. Rha E. Lee S. J. Kim H. Kwon K. Jeong Y. S. ( 2014 ). Toward a generalized and high-throughput enzyme screening system based on artificial genetic circuits. ACS Synth. Biol. 3 163 ？ 171 . 10.1021/sb400112u 24295047

상세보기
Collins M. L. Buchholz L. A. Remsen C. C. ( 1991 ). Effect of copper on Methylomonas albus BG8. Appl. Environ. Microbiol. 57 1261 ？ 1264 . 10.1128/aem.57.4.1261-1264.1991 16348466

상세보기
Conrado R. J. Gonzalez R. ( 2014 ). Chemistry. Envisioning the bioconversion of methane to liquid fuels. Science 343 621 ？ 623 . 10.1126/science.1246929 24503844

상세보기
Crombie A. Murrell J. C. ( 2011 ). Development of a system for genetic manipulation of the facultative methanotroph Methylocella silvestris BL2. Methods Enzymol. 495 119 ？ 133 . 10.1016/B978-0-12-386905-0.00008-5 21419918

상세보기
Dassama L. M. Kenney G. E. Rosenzweig A. C. ( 2017 ). Methanobactins: from genome to function. Metallomics 9 7 ？ 20 . 10.1039/c6mt00208k 27905614

상세보기
de Almeida J. M. Alnoch R. C. De Souza E. M. Mitchell D. A. Krieger N. ( 2019 ). Metagenomics: is it a powerful tool to obtain lipases for application in biocatalysis? Biochim. Biophys. Acta Proteins Proteom. 1868 : 140320 . 10.1016/j.bbapap.2019.140320 31756433

상세보기
Duarte J. M. Barbier I. Schaerli Y. ( 2017 ). Bacterial microcolonies in gel beads for high-throughput screening of libraries in synthetic biology. ACS Synth. Biol. 6 1988 ？ 1995 . 10.1021/acssynbio.7b00111 28803463

상세보기
Giuffrida M. C. Cigliana G. Spoto G. ( 2018 ). Ultrasensitive detection of lysozyme in droplet-based microfluidic devices. Biosens Bioelectron . 104 8 ？ 14 . 10.1016/j.bios.2017.12.042 29294408

상세보기
Graham D. W. Korich D. G. Leblanc R. P. Sinclair N. A. Arnold R. G. ( 1992 ). Applications of a colorimetric plate assay for soluble methane monooxygenase activity. Appl. Environ. Microbiol. 58 2231 ？ 2236 . 10.1128/aem.58.7.2231-2236.1992 1637160

상세보기
Grosse S. Laramee L. Wendlandt K. D. Mcdonald I. R. Miguez C. B. Kleber H. P. ( 1999 ). Purification and characterization of the soluble methane monooxygenase of the type II methanotrophic Bacterium Methylocysitis sp. strain WI 14. Appl. Environ. Microbiol. 65 3929 ？ 3935 . 10.1128/aem.65.9.3929-3935.1999 10473397

상세보기
Guo M. T. Rotem A. Heyman J. A. Weitz D. A. ( 2012 ). Droplet microfluidics for high-throughput biological assays. Lab Chip 12 2146 ？ 2155 . 10.1039/c2lc21147e 22318506

상세보기
Hosokawa M. Hoshino Y. Nishikawa Y. Hirose T. Yoon D. H. Mori T. ( 2015 ). Droplet-based microfluidics for high-throughput screening of a metagenomic library for isolation of microbial enzymes. Biosens Bioelectron. 67 379 ？ 385 . 10.1016/j.bios.2014.08.059 25194237

상세보기
Joensson H. N. Andersson Svahn H. ( 2012 ). Droplet microfluidics？a tool for single-cell analysis. Angew Chem. Int. Ed. Engl. 51 12176 ？ 12192 . 10.1002/anie.201200460 23180509

상세보기
Kalyuzhnaya M. G. Yang S. Rozova O. N. Smalley N. E. Clubb J. Lamb A. ( 2013 ). Highly efficient methane biocatalysis revealed in a methanotrophic bacterium. Nat. Commun. 4 : 2785 . 10.1038/ncomms3785 24302011

상세보기
Kaushik A. M. Hsieh K. Chen L. Shin D. J. Liao J. C. Wang T. H. ( 2017 ). Accelerating bacterial growth detection and antimicrobial susceptibility assessment in integrated picoliter droplet platform. Biosens Bioelectron. 97 260 ？ 266 . 10.1016/j.bios.2017.06.006 28609716

상세보기
Kim S. K. Kim S. H. Subhadra B. Woo S. G. Rha E. Kim S. W. ( 2018 ). A genetically encoded biosensor for monitoring isoprene production in engineered Escherichia coli . ACS Synth. Biol. 7 2379 ？ 2390 . 10.1021/acssynbio.8b00164 30261142

상세보기
Kim H. Kwon K. K. Seong W. Lee S. G. ( 2016a ). Multi-enzyme screening using a high-throughput genetic enzyme screening system. J. Vis. Exp. e54059, 1？7. 10.3791/54059 27584951

상세보기
Kim H. Rha E. Seong W. Yeom S. J. Lee D. H. Lee S. G. ( 2016b ). A cell-cell communication-based screening system for novel microbes with target enzyme activities. ACS Synth. Biol. 5 1231 ？ 1238 . 10.1021/acssynbio.5b00287 27452868

상세보기
Lawton T. J. Rosenzweig A. C. ( 2016 ). Methane-oxidizing enzymes: an upstream problem in biological gas-to-liquids conversion. J. Am. Chem. Soc. 138 9327 ？ 9340 . 10.1021/jacs.6b04568 27366961

상세보기
Lee J. Y. Sung B. H. Oh S. H. Kwon K. K. Lee H. Kim H. ( 2019 ). C1 compound biosensors: design, functional study, and applications. Int. J. Mol. Sci. 20 : 2253 . 10.3390/ijms20092253 31067766

상세보기
Leemhuis H. Kelly R. M. Dijkhuizen L. ( 2009 ). Directed evolution of enzymes: library screening strategies. IUBMB Life 61 222 ？ 228 . 10.1002/iub.165 19180668

상세보기
Lim H. G. Jang S. Jang S. Seo S. W. Jung G. Y. ( 2018 ). Design and optimization of genetically encoded biosensors for high-throughput screening of chemicals. Curr. Opin. Biotechnol. 54 18 ？ 25 . 10.1016/j.copbio.2018.01.011 29413747

상세보기
Lloyd J. S. Finch R. Dalton H. Murrell J. C. ( 1999 ). Homologous expression of soluble methane monooxygenase genes in Methylosinus trichosporium OB3b. Microbiology 145(Pt 2) , 461 ？ 470 . 10.1099/13500872-145-2-461 10075428

상세보기
Meyer A. Pellaux R. Potot S. Becker K. Hohmann H. P. Panke S. ( 2015 ). Optimization of a whole-cell biocatalyst by employing genetically encoded product sensors inside nanolitre reactors. Nat. Chem. 7 673 ？ 678 . 10.1038/nchem.2301 26201745

상세보기
Miller A. R. Keener W. K. Watwood M. E. Roberto F. F. ( 2002 ). A rapid fluorescence-based assay for detecting soluble methane monooxygenase. Appl. Microbiol. Biotechnol. 58 183 ？ 188 . 10.1007/s00253-001-0885-4 11876411

상세보기
Murrell J. C. Mcdonald I. R. Gilbert B. ( 2000 ). Regulation of expression of methane monooxygenases by copper ions. Trends Microbiol. 8 221 ？ 225 . 10.1016/s0966-842x(00)01739-x 10785638

상세보기
Myung J. Kim M. Pan M. Criddle C. S. Tang S. K. ( 2016 ). Low energy emulsion-based fermentation enabling accelerated methane mass transfer and growth of poly(3-hydroxybutyrate)-accumulating methanotrophs. Bioresour. Technol. 207 302 ？ 307 . 10.1016/j.biortech.2016.02.029 26896714

상세보기
Ngara T. R. Zhang H. ( 2018 ). Recent advances in function-based metagenomic screening. Genomics Proteomics Bioinformatics. 6 405 ？ 415 . 10.1016/j.gpb.2018.01.002 30597257

상세보기
Nielsen A. K. Gerdes K. Degn H. Murrell J. C. ( 1996 ). Regulation of bacterial methane oxidation: transcription of the soluble methane mono-oxygenase operon of Methylococcus capsulatus (Bath) is repressed by copper ions. Microbiology 142(Pt 5) , 1289 ？ 1296 . 10.1099/13500872-142-5-1289 8704968

상세보기
Puri A. W. Owen S. Chu F. Chavkin T. Beck D. A. Kalyuzhnaya M. G. ( 2015 ). Genetic tools for the industrially promising methanotroph Methylomicrobium buryatense . Appl. Environ. Microbiol. 81 1775 ？ 1781 . 10.1128/AEM.03795-14 25548049

상세보기
Rohlhill J. Sandoval N. R. Papoutsakis E. T. ( 2017 ). Sort-Seq approach to engineering a formaldehyde-inducible promoter for dynamically regulated Escherichia coli growth on methanol. ACS Synth. Biol. 6 1584 ？ 1595 . 10.1021/acssynbio.7b00114 28463494

상세보기
Ross M. O. Rosenzweig A. C. ( 2017 ). A tale of two methane monooxygenases. J. Biol. Inorg Chem. 22 307 ？ 319 . 10.1007/s00775-016-1419-y 27878395

상세보기
Schneider C. A. Rasband W. S. Eliceiri K. W. ( 2012 ). NIH image to imageJ: 25 years of image analysis. Nat. Methods 9 671 ？ 675 . 10.1038/nmeth.2089 22930834

상세보기
Selvamani V. Ganesh I. Maruthamuthu M. K. Eom G. T. Hong S. H. ( 2017 ). Engineering chimeric two-component system into Escherichia coli from Paracoccus denitrificans to sense methanol. Biotechnol. Bioproce. Eng. 22 225 ？ 230 . 10.4014/jmb.1611.11070 28372037

원문보기 상세보기
Semrau J. D. ( 2011 ). Bioremediation via methanotrophy: overview of recent findings and suggestions for future research. Front. Microbiol. 2 : 209 . 10.3389/fmicb.2011.00209 22016748

상세보기
Semrau J. D. Dispirito A. A. Yoon S. ( 2010 ). Methanotrophs and copper. FEMS Microbiol. Rev. 34 496 ？ 531 . 10.1111/j.1574-6976.2010.00212.x 20236329

상세보기
Shah N. N. Hanna M. L. Jackson K. J. Taylor R. T. ( 1995 ). Batch cultivation of Methylosinus trichosporium OB3B: IV. Production of hydrogen-driven soluble or particulate methane monooxygenase activity. Biotechnol. Bioeng. 45 229 ？ 238 . 10.1002/bit.260450307 18623142

상세보기
Shaofeng H. Shuben L. Jiayin X. Jianzhong N. Chungu X. Haidong T. ( 2007 ). Purification and biochemical characterization of soluble methane monooxygenase hydroxylase from Methylosinus trichosporium IMV 3011. Biosci. Biotechnol. Biochem. 71 122 ？ 129 . 17213640

상세보기
Shindell D. Kuylenstierna J. C. Vignati E. Van Dingenen R. Amann M. Klimont Z. ( 2012 ). Simultaneously mitigating near-term climate change and improving human health and food security. Science 335 183 ？ 189 . 10.1126/science.1210026 22246768

상세보기
Siedler S. Khatri N. K. Zsohar A. Kjaerbolling I. Vogt M. Hammar P. ( 2017 ). Development of a bacterial biosensor for rapid screening of yeast p-coumaric acid production. ACS Synth. Biol. 6 1860 ？ 1869 . 10.1021/acssynbio.7b00009 28532147

상세보기
Sirajuddin S. Rosenzweig A. C. ( 2015 ). Enzymatic oxidation of methane. Biochemistry 54 2283 ？ 2294 . 10.1021/acs.biochem.5b00198 25806595

상세보기
Smith T. J. Murrell J. C. ( 2011 ). Mutagenesis of soluble methane monooxygenase. Methods Enzymol. 495 135 ？ 147 . 10.1016/B978-0-12-386905-0.00009-7 21419919

상세보기
Strong P. J. Xie S. Clarke W. P. ( 2015 ). Methane as a resource: can the methanotrophs add value? Environ. Sci. Technol. 49 4001 ？ 4018 . 10.1021/es504242n 25723373

상세보기
Tapscott T. Guarnieri M. T. Henard C. A. ( 2019 ). Development of a CRISPR/Cas9 system for Methylococcus capsulatus in vivo gene editing. Appl. Environ. Microbiol. 85 : AEM.00340-19 . 10.1128/AEM.00340-19 30926729

상세보기
Tavormina P. L. Kellermann M. Y. Antony C. P. Tocheva E. I. Dalleska N. F. Jensen A. J. ( 2017 ). Starvation and recovery in the deep-sea methanotroph Methyloprofundus sedimenti . Mol. Microbiol. 103 242 ？ 252 . 10.1111/mmi.13553 27741568

상세보기
Thorsen T. Roberts R. W. Arnold F. H. Quake S. R. ( 2001 ). Dynamic pattern formation in a vesicle-generating microfluidic device. Phys. Rev. Lett. 86 4163 ？ 4166 . 10.1103/physrevlett.86.4163 11328121

상세보기
van der Helm E. Genee H. J. Sommer M. O. A. ( 2018 ). The evolving interface between synthetic biology and functional metagenomics. Nat. Chem. Biol. 14 752 ？ 759 . 10.1038/s41589-018-0100-x 30013060

상세보기
Wang Y. H. Wei K. Y. Smolke C. D. ( 2013 ). Synthetic biology: advancing the design of diverse genetic systems. Annu. Rev. Chem. Biomol. Eng. 4 69 ？ 102 . 10.1146/annurev-chembioeng-061312-103351 23413816

상세보기
West C. A. Salmond G. P. Dalton H. Murrell J. C. ( 1992 ). Functional expression in Escherichia coli of proteins B and C from soluble methane monooxygenase of Methylococcus capsulatus (Bath). J. Gen. Microbiol. 138 1301 ？ 1307 . 10.1099/00221287-138-7-1301 1512560

상세보기
Whittenbury R. Phillips K. C. Wilkinson J. F. ( 1970 ). Enrichment, isolation and some properties of methane-utilizing bacteria. J. Gen. Microbiol. 61 205 ？ 218 . 10.1099/00221287-61-2-205 5476891

상세보기
Yeom S. J. Kim M. Kwon K. K. Fu Y. Rha E. Park S. H. ( 2018 ). A synthetic microbial biosensor for high-throughput screening of lactam biocatalysts. Nat. Commun. 9 5053 . 10.1038/s41467-018-07488-0 30498220

상세보기
Yu Z. Chistoserdova L. ( 2017 ). Communal metabolism of methane and the rare Earth element switch. J. Bacteriol. 199 : e0328-17 . 10.1128/JB.00328-17 28630125

상세보기

LOADING...

활용도 분석정보

상세보기

다운로드

내보내기

활용도 Top5 논문

해당 논문의 주제분야에서 활용도가 높은 상위 5개 콘텐츠를 보여줍니다.
더보기 버튼을 클릭하시면 더 많은 관련자료를 살펴볼 수 있습니다.

표제어: PCR

동의어: Packet Collision Rate

용어 설명 출처 목록 (6)

용어 설명: PCR은 세균 특이성이 있는 primer를 이용하여 적은 수의 세균이 있을지라도 쉽게 검출할 수 있는 유용한 방법이며, 이를 이용하여 구강 내 치면세균막이나 타액에서 직접 세균을 검출할 수 있게 되었다[8].

내보내기 구분	파일저장 인쇄 메일전송
구성항목	기본정보 상세정보 관리번호, 논문명, 저널/프로시딩명, 저자 , 발행년, 권, 호, 시작페이지, 끝페이지, 발행기관 관리번호, 논문명, 대등논문명, 저자 , 저널/프로시딩명, 발행기관, 발행년, 발행언어, 권, 호, 시작페이지, 끝페이지, ISBN, ISSN, 주제분야, 키워드, 초록(한글), 초록(영문), 저자(소속기관)
저장형식	Text(ASCII format) Excel format RefWorks Direct Export RIS format (for Reference Manager, ProCite, EndNote), Scholar's Aids, Mendeley
메일정보	받는사람 (필수) @ 보내는사람 (선택) @ 제목 내용 KISTI 검색결과 이메일 서비스
안내	총 건의 자료가 검색되었습니다. 다운받으실 자료의 인덱스를 입력하세요. (1-10,000) 검색결과의 순서대로 최대 10,000건 까지 다운로드가 가능합니다. 데이타가 많을 경우 속도가 느려질 수 있습니다.(최대 2~3분 소요) 다운로드 파일은 UTF-8 형태로 저장됩니다. 파일의 내용이 제대로 보이지 않을실 때는 웹브라우저 상단의 보기 -> 인코딩 -> 자동선택 여부를 확인하십시오. ~ Text(ASCII format) Excel format

연합인증