Methanotrophs을 이용한 메탄 저감 기술 최신 동향

조경숙 ( Kyung-suk Cho ),정혜경 ( Hyekyeng Jung ) 한국미생물생명공학회(구 한국산업미생물학회) 2017 한국미생물·생명공학회지 Vol.45 No.3

메탄은 자연적인 발생원과 인위적인 발생원에 의해 배출되며 지구온난화를 야기하는 대표적인 온실가스이다. 메탄을 탄소원과 에너지원으로 이용하는 메탄산화세균은 메탄의 생물학적 산화에 중요한 역할을 한다. 메탄산화세균의 서식지는 매우 다양하며 메탄산화반응의 핵심 효소인 methane monooxygenases (MMOs)는 메탄뿐 아니라 다른 기질을 산화할 수 있는 기질특이성을 가지고 있다. 이러한 메탄산화세균의 특성으로 인해 생물학적 메탄 저감 기술과 생물정화기술 분야에서 메탄산화세균의 활용에 대한 연구가 활발히 진행되고 있다. 본 총설 논문에서는 메탄산화세균의 종류, MMOs의 특성과 메탄산화세균의 고농도 배양 기술에 관한 최근 정보를 정리하였다. 또한 메탄산화세균을 이용한 생물학적 메탄 저감 관련 실험실 규모와 매립지 현장에서의 기술 개발 현황 및 적용 결과를 소개하였다. 이러한 생물학적 메탄 저감 시스템에서 메탄산화세균의 군집 거동 특성도 고찰하였다. 마지막으로, 메탄산화세균을 활용한 생물공학기술의 혁신을 위해 필요한 과제로 대사활성이 우수하거나 신규대사능력을 가진 메탄산화세균의 지속적인 탐색 연구, 고농도 세포 대량배양기술 개발 및 미생물 컨소시움(메탄산화세균과 비메탄산화세균의 컨소시움) 디자인 및 관리 기술 등이 필요함을 제안하였다. Methane, which is emitted from natural and anthropogenic sources, is a representative greenhouse gas for global warming. Methanotrophs are widespread in the environment and play an important role in the biological oxidation of methane via methane monooxygenases (MMOs), key enzymes for methane oxidation with broad substrate specificity. Methanotrophs have attracted attention as multifunctional bacteria with promising applications in biological methane mitigation technology and environmental bioremediation. In this review, we have summarized current knowledge regarding the biodiversity of methanotrophs, catalytic properties of MMOs, and high-cell density cultivation technology. In addition, we have reviewed the recent advances in biological methane mitigation technologies using methanotrophs in field-scale systems as well as in lab-scale bioreactors. We have also surveyed information on the dynamics of the methanotrophic community in biological systems and discussed the various challenges pertaining to methanotroph- related biotechnological innovation, such as identification of suitable methanotrophic strains with better and/or novel metabolic activity, development of high-cell density mass cultivation technology, and the microbial consortium (methanotrophs and non-methanotrophs consortium) design and control technology.

Characterization of tobermolite as a bed material for selective growth of methanotrophs in biofiltration

Kim, T.G.,Jeong, S.Y.,Cho, K.S. Elsevier Science Publishers 2014 Journal of biotechnology Vol.173 No.-

Tobermolite was characterized as a bed material for methanotrophic biofiltration. A lab-scale biofilter packed with tobermolite was operated for different operation times under identical conditions. The three different runs showed similar acclimation patterns of methane oxidation, with methane removal efficiency increasing rapidly for the first few days and peaking within three weeks, after which the efficiency remained stable. The mean methane removal capacities ranged from 766gm<SUP>-3</SUP>d<SUP>-1</SUP> to 974gm<SUP>-3</SUP>d<SUP>-1</SUP> after acclimation. Pyrosequencing indicated that the methanotrophic proportion (methanotroph/bacteria) increased to 71-94% within three weeks. Type I methanotrophs Methylocaldum and Methylosarcina were dominant during the initial growth period, then Methylocaldum alone dominated the methanotrophic community. A community comparison showed that total bacterial and methanotrophic communities were temporally stable after the initial growth period. Quantitative PCR showed that methanotrophic density increased during the first 3-4 weeks, then remained stable over 120 days. Tobermolite can provide a special habitat for the selective growth of methanotrophs, resulting in rapid acclimation. Tobermolite also allows the microbial community and methanotrophic density to remain stable, resulting in stable methane biofiltration.

Microbial Community Analysis of a Methane-Oxidizing Biofilm Using Ribosomal Tag Pyrosequencing

( Tae Gwan Kim ),( Eun Hee Lee ),( Kyung Suk Cho ) 한국미생물 · 생명공학회 2012 Journal of microbiology and biotechnology Vol.22 No.3

Current ecological knowledge of methanotrophic biofilms is incomplete, although they have been broadly studied in biotechnological processes. Four individual DNA samples were prepared from a methanotrophic biofilm, and a multiplex 16S rDNA pyrosequencing was performed. A complete library (before being de-multiplexed) contained 33,639 sequences (average length, 415 nt). Interestingly, methanotrophs were not dominant, only making up 23% of the community. Methylosinus, Methylomonas, and Methylosarcina were the dominant methanotrophs. Type II methanotrophs were more abundant than type I (56 vs. 44%), but less richer and diverse. Dominant non-methanotrophic genera included Hydrogenophaga, Flavobacterium, and Hyphomicrobium. The library was de-multiplexed into four libraries, with different sequencing efforts (3,915-20,133 sequences). Sørrenson abundance similarity results showed that the four libraries were almost identical (indices > 0.97), and phylogenetic comparisons using UniFrac test and P-test revealed the same results. It was demonstrated that the pyrosequencing was highly reproducible. These survey results can provide an insight into the management and/or manipulation of methanotrophic biofilms.

Bioconversion of methane to chemicals using metabolic engineered methanotroph

이은열 한국공업화학회 2018 한국공업화학회 연구논문 초록집 Vol.2018 No.0

Methane is considered as next-generation carbon feedstock for industrial biotechnology in production of value-added chemicals and fuels due to low price and abundance (1, 2). Methane can be biologically converted to value-added chemicals using methanotrophs as a biocatalyst. In order to produce chemicals using methanotrophs, biosynthetic pathways for target products should be reconstructed in connection with C1 assimilation pathway in methanotrophs. In this study, recent progresses on bioconversion of methane to chemicals using metabolic engineered methanotrophs will be presented and discussed.

Metabolic engineering of methanotrophs and its application to production of chemicals and biofuels from methane

Lee, Ok Kyung,Hur, Dong Hoon,Nguyen, Diep Thi Ngoc,Lee, Eun Yeol John WileySons, Ltd 2016 Biofuels, Bioproducts and Biorefining Vol.10 No.6

<P>Methane-assimilating bacteria, methanotrophs, can play an important role in producing various value-added chemicals and biofuels from methane, which is considered a next-generation carbon feedstock. The capability to engineer the metabolic pathway of methanotrophs is a key success factor for enhancing methane-to-product conversion efficiency. Recently, OMICS studies on several model methanotrophs have been conducted and provided strategies to engineer methanotrophs. Here, we present a review on the current progresses and future prospects of metabolic engineering of methanotrophs and its application to chemical and biofuel production from methane. (c) 2016 Society of Chemical Industry and John Wiley & Sons, Ltd</P>

Microbial consortia including methanotrophs: some benefits of living together

Rajendra Singh,류재원,Si Wouk Kim 한국미생물학회 2019 The journal of microbiology Vol.57 No.11

With the progress of biotechnological research and improvements made in bioprocessing with pure cultures, microbial consortia have gained recognition for accomplishing biological processes with improved effectiveness. Microbes are indispensable tool in developing bioprocesses for the production of bioenergy and biochemicals while utilizing renewable resources due to technical, economic and environmental advantages. They communicate with specific cohorts in close proximity to promote metabolic cooperation. Use of positive microbial associations has been recognized widely, especially in food industries and bioremediation of toxic compounds and waste materials. Role of microbial associations in developing sustainable energy sources and substitutes for conventional fuels is highly promising with many commercial prospects. Detoxification of chemical contaminants sourced from domestic, agricultural and industrial wastes has also been achieved through microbial catalysis in pure and co-culture systems. Methanotrophs, the sole biological sink of greenhouse gas methane, catalyze the methane monooxygenasemediated oxidation of methane to methanol, a high energy density liquid and key platform chemical to produce commodity chemical compounds and their derivatives. Constructed microbial consortia have positive effects, such as improved biomass, biocatalytic potential, stability etc. In a methanotroph- heterotroph consortium, non-methanotrophs provide key nutrient factors and alleviate the toxicity from the culture. Non-methanotrophic organisms biologically stimulate the growth and activity of methanotrophs via production of growth stimulators. However, methanotrophs in association with cocultured microorganisms are in need of further exploration and thorough investigation to study their interaction mode and application with improved effectiveness.

Effect of Ammonium Chloride on the Mixed Methanotrophs Species Composition and Methanol Metabolism

김이태,윤영한 한국수처리학회 2018 한국수처리학회지 Vol.26 No.6

This study analyzed the utility of ammonium chloride (NH4Cl) as a nitrogen source for methanotroph communities. When cultured in nitrate mineral salt (NMS) medium, the methanotroph community we identified four families, seven genera, and 16 type I and type II species of methanotrophs. Among species in the Methylobacter genus, Methylobacter marinus could be actively cultured in NMS medium without NaCl addition. Following the addition of 25 mM NH4Cl, the numbers of the type I genera Methylomonas, Methylococcus, and Methylobacter were increased, whereas the numbers of the type II genera Methylocystis and Methylosinus were decreased after 5 days. In methanotroph communities, certain concentrations of NH4Cl affected methane consumption and growth of methanotrophs at the community level. NH4Cl caused a considerable decrease in the methane consumption rate and the expression of soluble methane monooxygenases (sMMOs) but did not inhibit the growth of Methylomonas methanica expressing sMMO. These results could be attributed to competitive antagonism of MMOs due to their direct involvement in ammonia oxidation.

메탄산화균의 메탄 산화속도에 미치는 암모니아의 영향

이수연,류희욱,조경숙 한국냄새환경학회 2012 실내환경 및 냄새 학회지 Vol.11 No.1

Rhizosphere and non-rhizosphere soils were sampled from landfill area, riparian wetland, and rice paddy. The consortia were obtained by methane enrichment culture using the soils. The effects of ammonia on methane oxidation in the consortia were evaluated. Compared with methane oxidation rates without ammonia, the rates with ammonia of 1mg-N/bottle were similar or slightly lower. However, their methane oxidation rates were significantly reduced with 2~4mg-N ammonia/bottles. The effect of ammonia on the methanotrophic abundance was estimated by using a quantitative real-time PCR method targeting particulate methane monooxygenase gene. Ammonia didn’t negatively influence on the methanotrophic abundance although it inhibited the methane oxidation activity by methanotrophs. 매립지, 습지 및 논으로부터 근권 및 비근권 토양을 채취하여 메탄으로 농화 배양한 후, 농화배양액의 메탄 산화에 미치는 암모니아 농도의 영향을 정량적으로 분석하였다. 순수 메탄만을 첨가한 조건에서의 메탄 산화속도와비교하여, 암모니아 첨가량이 1mg-N/bottle까지는 메탄 산화속도는 거의 유사하거나 약간 감소하는 경향을 보였다. 그러나, 암모니아 첨가량이 2~4mg-N/bottles인 조건에서는 메탄 산화속도가 급격하게 감소하였다. Particulate methane monooxygenase gene을 이용한 quantitative real-time PCR 기법을 활용하여 메탄산화균에 미치는 암모니아 영향을 정량적으로 분석한 결과, 메탄산화균의 군집 밀도는 암모니아 첨가에 의해 감소되지 않았다.

메탄가스 전환 미생물촉매 개량을 위한 플라스미드 복제 시작점 예측

김민식(Min-Sik Kim) 한국신재생에너지학회 2023 신재생에너지 Vol.19 No.4

Methane is the second most emitted greenhouse gas after carbon dioxide. Despite lower emissions than those of carbon dioxide, methane receives significant attention owing to its more than 20-fold higher global warming potential. Consequently, the importance of research on methanotrophic bacteria, microorganisms capable of converting methane gas into high-value materials, is increasingly emphasized. In the case of methanotrophic bacteria, knowledge on episomal plasmids that can be used for genetic engineering remains lacking, which poses significant challenges to the engineering process. The replication origin sequences of natural plasmids within methanotrophic bacteria have been predicted through in silico methods. The basic characteristics of the replication origin, such as a high A/T ratio, repetitive sequences, and proximity to proteins related to replication, have been used as criteria for identifying the replication origin. As a result, a region with a sequence of 18 base pairs repeated eight times could be identified. The putative replication origin sequence thus identified generally takes the form of iterons, but it also possesses unique features such as the length of the gap between iterons and the repetition of identical iteron sequences. This information can be valuable for future design of episomal plasmids applicable to methanotrophs.

Biofuel upgrade reactions via phase-transfer catalysis of methanotrophs

박예림,김동호,최규환,김용우,이은열,박범준 한국공업화학회 2021 Journal of Industrial and Engineering Chemistry Vol.95 No.-

Methane, one of the six major greenhouse gases, poses a serious environmental problem with thepotential for global warming 20 times that of CO2. Although a large amount of methane is generatedworldwide, its recovery and utilization are very low. One of the environmentally friendly ways to usemethane is the biological gas-to-liquids (Bio-GTL) process, in which methane is biologically converted touseful products by microorganisms. Methanotrophs are strains that can convert methane to methanol atambient conditions using methane monooxygenase (MMO). Here, we report an efficient phase-transfercatalysis system for methane-to-methanol conversion using methanotrophs. The methanotroph used inthe work is Methylomicrobium alcaliphilum 20Z of which the methanol dehydrogenase (MDH) enzyme isremoved to enhance methanol accumulation. The phase-transfer catalysis system does not require anyseparation processes and facilitates the mass transfer of methane gas, thereby increasing the methanolproductivity and lowering the production cost. The methanol productivity is 0.717 g/L/h, which issuperior to the results reported to date. In addition, the use of a cellulose-membrane reactor systemenables multiple biocatalytic reactions without a significant decrease in productivity.