미생물학회지 (Korean Journal of Microbiology)

  • 제41권1호
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  • Pages.1-7
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  • 2005
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  • 0440-2413(pISSN)
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  • 2383-9902(eISSN)
한국미생물학회 (The Microbiological Society of Korea)
권호 표지

Methylophaga aminosulfidovorans SKI bfmo 오페론의 클로닝 및 구조 분석

Cloning and Structural Analysis of bfmo Operon in Methylophaga aminosulfidovorans SK1

  • Lim Hyun Sook (Graduate School of Education, Chosun University) ;
  • Goo Jae Whan (Department of Environmental engineering, Chosun University) ;
  • Kim Lee Hyun (Graduate School of Education, Chosun University) ;
  • Kim Si Wouk (Department of Environmental engineering, Chosun University) ;
  • Cho Eun Hee (Department of Biology Education, Chosun University)
  • 발행 : 2005.03.01
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초록

Methylophaga aminosulfidovorans SK1 (KCTC 10323 BP)은 단일 탄소원, 질소원 그리고 에너지원으로 난분해성 화합물인 트리메틸아민을 이용할 수 있다. M. aminosulfidovorans SK1는 진핵세포의 flavin-containing monooxygenase와 유사한 유전자(bFMO)를 지니고 있으며 대장균에서 발현된 재조합 단백질은 강력한 트리메틸아민 산화활성을 보인다. 본 연구에서는 bEMO의 기능과 조절 메커니즘을 연구하기 위하여 bfmo의 상단부 및 하단부 유전자의 염기서열을 결정하였다. bfmo 상단부의 세 개의 열린해독틀은 잘 보존된 nitrate/nitrite response regulators와 methyl accepting protein 유사단백질을 암호화하였다. 하단부의 두 개의 작은 열린해독틀은 기능은 알려져 있지 않지만 진정세균계에서 잘 보존된 단백질의 일종으로 나타났다. 역전사효소 중합효소증폭반응을 통하여 여섯 개의 유전자는 세 개의 독립된 오페론으로 구성되어 있음을 확인하였다. bfmo의 상단부에 위치하는 세 개의 조절유전자는 두 개의 프로모터에서 전사되었다. 그리고 이와 독립적으로 bfmo와 두 개의 하단부 유전자가 하나의 전사단위를 이루고 있다.

Methylophaga aminosulfidovorans SK1 (KCTC 10323 BP) can utilize trimethylamine as a sole carbon, nitrogen, and energy source. The bacterial flavin-containing monooxygenase (bFMO) gene was identified in the strain and the recombinant enzyme expressed in E. coli oxidized trimethylamine. To study the function and regulation of the bfmo, over 8,000 nucleotide sequences of the neighboring regions including the bfmo were determined. Three open reading frames proceeding to the bfmo gene encoded analogues to highly conserved nitrate/nitrite sensing two-component system regulators and a methyl accepting protein. Two small open reading frames just downstream of the bfmo gene showed no similar proteins of known functions but the sequences were conserved among other bacteria. Reverse transcription-polymerase chain reaction analysis showed that the six putative genes consisted of three transcription units. The three regulatory genes located upstream of the bfmo gene formed two separate transcription units. The bfmo and the two downstream genes were transcribed from a single promoter.

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참고문헌

  1.  Altschul, S.F., T.L. Madden, A.A. Schaffer, J. Zhang, Z. Zhang, W. Miller, and D.J. Lipman. 1997. Gapped BLAST and PSI- BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25, 3389-3402 https://doi.org/10.1093/nar/25.17.3389
  2. Anthony, C. 1982. The Biochemistry of methylotrophs. Academic press, New York
  3. Baikalov, I., I. Schroder, M. Kaczor-Grzeskowiak, K. Grzeskowiak, R.P. Gunsalus and R.E. Dickerson. 1996. Structure of the Escherichia coli response regulator NarL. Biochemistry. 35, 11053-1161 https://doi.org/10.1021/bi960919o
  4. Barrett, E.L. and H.S. Kwan. 1985. Bacterial reduction of trimethylamine oxide. Annu Rev Microbiol. 39, 131-149 https://doi.org/10.1146/annurev.mi.39.100185.001023
  5. Choi, H.S., J.K. Kim, E.H. Cho, Y.C. Kim, J.I. Kim, and S.W. Kim. 2003. A novel flavin-containing monooxygenase from Methylophaga sp strain SK1 and its indigo synthesis in Escherichia coli. Biochem Biophys Res. Commun. 306(4), 930-936 https://doi.org/10.1016/S0006-291X(03)01087-8
  6. Dahl, J.S., Mehta, R.J. and Hoare, D.S. 1972. New obligate methylotroph. J. Bacteriol. 109(2), 916-921
  7. Doronina, N.V., T.D. Darmaeva, and Y.A. Trotsenko. 2003. Methylophaga alcalica sp. nov., a novel alkaliphilic and moderately halophilic, obligately methylotrophic bacterium from the East Mongolian saline soda lake. Int. J. Syst. Evol. Microbiol. 53, 223-229 https://doi.org/10.1099/ijs.0.02267-0
  8.  Doronina, N.V., T.D. Darmaeva, and Y.A. Trotsenko. 2003. Methylophaga natronica sp. nov., a new alkaliphilic and moderately halophilic, restricted-facultatively methylotrophic bacterium from Soda Lake of the Southern Transbaikal Region. Syst. Appl. Microbiol. 26, 382-389 https://doi.org/10.1078/072320203322497419
  9. Doronina, N.V., T.D. Li, E.G. Ivanova, O.V. Rodionova, and Y.A. Trotsenko, 2004. Methylophaga murata sp. nov. - haloalcalotolerant aerobic methylotroph from destroyed marble. Unpublished (as of 20 August 2004)
  10.  Eldridge, A.M., H.S. Kang, E. Johnson, R. Gunsalus, F.W. Dahlquist, 2002. Effect of phosphorylation on the interdomain interaction of the response regulator, NarL. Biochemistry. 41, 15173-15180 https://doi.org/10.1021/bi026254+
  11. Guest, I. and D.R. Varma, 1992. Teratogenic and macromolecular synthesis inhibitory effects of trimethylamine on mouse embryos in culture. J. Toxicol Environ Health. 36, 27-41 https://doi.org/10.1080/15287399209531621
  12.  Hartig, E., U. Schiek, K.U. Vollack, and W.G. Zumft, 1999. Nitrate and nitrite control of respiratory nitrate reduction in denitrifying Pseudomonas stutzeri by a two-component regulatory system homologous to NarXL of Escherichia coli. J. Bacteriol. 181, 3658-65
  13. Janvier, M. and P.A.D. Grimont, 1995. The genus Methylophaga, a new line of descent within phylogenetic branch $\gamma$ of Proteobacteria. Microbial Paris. 146, 543-550
  14. Janvier, M., C. Frehel, F. Grimont, and F. Gasser, 1985. Methylophaga marina gen. nov., sp. nov. and Methylophaga thalassica sp. nov., marine methylotrophs. Int. J. Syst. Bacteriol. 35, 131-139 https://doi.org/10.1099/00207713-35-2-131
  15.  Kim, S.G., H.S. Bae, and S.T. Lee, 2001. A novel denitrifying bacterial isolate that degrades trimethylamine both aerobically and anaerobically via two different pathways. Arch Microbiol. 176, 271-277 https://doi.org/10.1007/s002030100319
  16. Koh, M., C.S. Kim, Y.A. Kim, H.S. Choi, E.H. Cho, E. Kim, Y.M. Kim and S.W. Kim, 2002. Properties of electron carriers in the process of methanol oxidation in a new restricted facultative marine methylotrophic bacterium, Methylophaga sp. MP. J. Microbiol. Biotechnol. 12, 476-482
  17. Lee, A.I., A. Delgado, and R.P. Gunsalus, 1999. Signal-dependent phosphorylation of the membrane-bound NarX two-component sensor-transmitter protein of Escherichia coli: nitrate elicits a superior anion ligand response compared to nitrite. J. Bacteriol. 181, 5309-5316
  18. Maris, A.E., M.R. Sawaya, M. Kaczor-Grzeskowiak, M.R. Jarvis, S.M. Bearson, M.L. Kopka, I. Schroder, R.P. Gunsalus, and R.E. Dickerson, 2002. Dimerization allows DNA target site recognition by the NarL response regulator. Nat. Struct .Biol. 9, 771-8 https://doi.org/10.1038/nsb845
  19. Moune, S., N. Manac'h, A. Hirschler, P. Caumette, J.C. Willison, and R. Matheron, 1999. Haloanaerobacter salinarius sp. nov., a novel halophilic fermentative bacterium that reduces glycine-betaine to trimethylamine with hydrogen or serine as electron donors; emendation of the genus Haloanaerobacter. Int. J. Syst. Bacteriol. 49 Pt 1, 103-112 https://doi.org/10.1099/00207713-49-1-103
  20. Schroder, I., R. Cavicchioli, and R.P. Gunsalus, 1994. Phosphorylation and dephosphorylation of the NarQ, NarX, and NarL proteins of the nitrate-dependent two-component regulatory system of Escherichia coli. J. Bacteriol. 176, 4985-92 https://doi.org/10.1128/jb.176.16.4985-4992.1994
  21.  Stackebrandt, E., R.G.E. Murray, and H.G. Trtiper, 1988. Proteobacteria classis nov., a name for the phylogenetic taxon that includes the 'purple bacteria and their relatives'. Int. J. Syst. Bacteriol. 38, 321-325 https://doi.org/10.1099/00207713-38-3-321
  22.  Thompson, J.D., D.G. Higgins, and T.J. Gibson, 1994. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 22:4673-4680 https://doi.org/10.1093/nar/22.22.4673
  23. Tsuji, K., H.C. Tsien, R.S. Hanson, S.R. DePalma, R. Sehoitz, and S. LaRoche, 1990. 16S ribosomal RNA sequence analysis for determination of phylogenetic relationship among methylotrophs. J. Gen. Microbiol. 136, 1-10 https://doi.org/10.1099/00221287-136-1-1
  24.  Urakami, T. and K. Komagata, 1987. Characterization of species of marine methylotrophs of the genus Methylophagn. Int. J. Syst. Bacteriol. 37, 402-406 https://doi.org/10.1099/00207713-37-4-402
  25.  Vedenina, I. and N.I. Govorukhina, 1988. Formation of a methylotrophic denitrifying biocenosis in a system of sewage treatment for nitrates. Mikrobiologia. 57, 320-328
  26.  Yancey, P.H., W.R. Blake, and J. Conley, 2002. Unusual organic osmolytes in deep-sea animals: adaptations to hydrostatic pressure and other perturbants. Comp. Biochem. Physiol. A Mol. Integr. Physiol. 133, 667-76 https://doi.org/10.1016/S1095-6433(02)00182-4
  27.  Zhao, Y., S.K. Christensen, C. Fankhauser, J.R. Cashman, J.D. Cohen, D. Weigel, and J. Chory, 2001. A role for flavin monooxygenase- like enzymes in auxin biosynthesis. Science. 291, 306-309 https://doi.org/10.1126/science.291.5502.306
  28.  Ziegler, D.M. 1988. Flavin-containing monooxygenases: catalytic mechanism and substrate specificities. Drug Metab. Rev. 19, 1-32 https://doi.org/10.3109/03602538809049617
  29. de Zwart, J.M.M., P.N. Nelisse, and J.G. Kuenen, 1996. Isolation and characterization of Methylophaga sulfidovorans sp. nov.: an obligately methylotrophic, aerobic, dimethylsulfide oxidizing bacterium from microbial mat. FEMS Microbiol. Ecol. 20, 261-270