Microbiology and Biotechnology Letters (한국미생물·생명공학회지)

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
권호 표지

Cloning and Nucleotide Sequence Analysis of xylC Gene Encoding 5C-2HMS Dehydrogenase from Pseudomonas sp. S-47.

Pseudomonas sp. S-47로부터 5-Chloro-2-Hydroxymuconic Semialdehyde Dehydrogenase를 암호화하는 xylG 유전자의 클로닝 및 염기서열 분석

  • Park, Song-Yi (Department of Microbiology, Chungbuk National University) ;
  • Lee, Dong-Hoon (Department of Microbiology, Chungbuk National University) ;
  • Kim, Young-Soo (Department of Pharmacy,Chungbuk National University) ;
  • Lee, Kyung (Department of Microbiology, Changwon National University) ;
  • Kim, Chi-Kyung (Department of Microbiology, Chungbuk National University)
  • 박송이 (충북대학교 미생물학과 및 유전공학연구소) ;
  • 이동훈 (충북대학교 미생물학과 및 유전공학연구소) ;
  • 김영수 (충북대학교 약학과) ;
  • 이경 (창원대학교 미생물학과) ;
  • 김치경 (충북대학교 미생물학과 및 유전공학연구소)
  • Published : 2002.03.01
Download PDF
⟨ Previous Next ⟩

Abstract

Pseudomonas sp. S-47 is capable of degrading 4-chlorobenzoate to produce 5-chloro-2-hydroxymuconic semialdehyde (5C-2HMS) by the enzymes encoding by xylXYZLTE cluster. In this study, the resulting 5C-2HMS was confirmed to be transformed to 5-chloro-2-hydroxymuconic acid (5C-2HMA) by 5C-2HMS dehydrogenase. The xylG gene encoding 5C-2HMS dehydrogenase was cloned from the chromosomal DNA of strain S-47. The nucleotide sequence of xylG showed to be composed of 1,600 base pairs with ATG initiation and TGA termination codons. A deduced amino acid sequence of the 5C-2HMS dehydrogenase (XylG) exhibited 98％, 93％, and 89％ identity with those of the dehydrogenases from P. putida mt-2, P. putida G7, and Pseudomonas sp. CF600, respectively.

Pseudomonas sp. S-47은 xylXYZLTE 유전자에 의하여 암호화되는 효소군에 의하여 4CBA를 분해하여 5-chloro-2-hydroxymuconic semialdehyde(5C-2HMS)를 생성하는데, 본 연구에서는 이 5C-2HMS의 다음 분해과정을 확인하였다. xylXYZLTE 유전자와 5-chloro-2-hydroxymuconic semialdehyde dehydrogenase(5C-2HMSD)를 암호화하고 xylG 유전자를 포함하는 재조합 균주인 pCSS202로부터, xylG 유전자를 포함하는 재조합 플라스미드 pENV5를 만들었다. 이 플라스미드는 2-hydroxymuconic semialdehyde, 3-chloro-muconate, 2-hydroxy-6-oxohepta-2,4-dienoate, 2-hydroxy-5-methylmuconic semialdehyde와 같은 aromatic compound 에서 분해능을 나타냈으며, 그 중 5C-2HMS에서 가장 높은 분해능을 나타내었다. 또한 5C-2HMSD를 암호화하는 유전자인 xylC의 염기서열을 분석한 결과, 약 1,600 bp의 염기와 486개의 amino acid residue를 갖고있는 것을 확인하였다. P. sp. S-47의 xylG 유전자를 비교 분석한 결과 P. putida CF600, P. putida G7과 P. putida mt-2 등의 5C-2HMS dehydro-genase와 85％ 이상의 amino acid homology를 보여주었다.

Keywords

References

  1. Arendorf, J. J. and D. D. Focht. 1995.A meta-cleavage pathway for 4-chlorobenzoate, an intermediate in the metabolism of 4-chlorobiphenyl by Pseudomonas cepacia P166. Appl. Environ. Microbiol. 61: 443-447.
  2. Brinkmann, U. and W. Reineke. 1992. Degradation of chlorotoluenes by in vivo constructed hybrid strains: problems of enzyme specificity, induction and prevention of meta-path way. FEMS Microbial. Lett. 96: 81-88. https://doi.org/10.1111/j.1574-6968.1992.tb05397.x
  3. Cerdan, P., A. Wasserfallen, M. Rekik, K. N. Timmis, and S. Harayama. 1994. Substrate specificity of catechol 2,3-dioxygenases encoded by TOL plasmid pWWO of Pseudomonas putida and its relationship to cell growth. J. Bacterial. 176: 6074-6081,
  4. Harayama, S., M. Rekik, A. Wasserfallen, and A. Bairoch. 1987. Evolutionary relationships between catabolic pathways for aromatics: conservation of gene order and nucleotide sequence of catechol oxidation genes of pWWO and NAH7 plasmids. Mol. cen. Genet. 210: 241-247.
  5. Harayama, S. and R. Monique. 1989. Bacterial aromatic ring-cleavage enzymes are classified into two different gene familes.J. BioI. Chem. 264: 15328-15333.
  6. Harayama, S and M. Rekik. 1990. The meta cleavage operon of TOL degradative plasmid pWWO comprises 13 genes. Mol. Gen. Genet, 221: 113-120.
  7. Harayama, S., M. Kok, and E. L. Neidle. 1992. Functional and evolutionary relationships among diverse oxygenases. Annu, Rev. Microbial. 46: 565-601.
  8. Hom, J. M., S. Harayama, and K. N. Timmis. 1991. DNA sequence determination of the TOL plasmid (PWWO) xylGFJ genes of Pseudomonas putida. Mol, Microbial. 5: 2459-2474.
  9. Hughes, E. L., R. C. Bayly, and R. A. Skurray, 1984, Evidence for isofunctional enzymes in the degradation of phenol, m- and p-toluate, and p-cresol via catechol metacleavage pathways in Alcaligenes eutrophus. J. Bacterial. 158: 79-83.
  10. Inoue, J., J. P. Shaw, M. Rekik, and S. Harayama. 1995. Overlapping substrate specificities of benzaldehyde dehydrogenase (the xylC gene product) and 2-hydroxymuconic semialdehyde dehydrogenase (the xylG gene product) encoded by TOL plasmid pWWO of Pseudomonas putida. J. Bacterial. 177: 1196-1201.
  11. Keil, H., S. Keil, R. W Pickup, and P. A. Wiliams. 1985. Evolutionary conservation of genes coding for meta pathway enzymes within TOL plasmids pWWO and pWW53. J. Bacterial. 164: 887-895.
  12. Kim, K. P., D. J. Seo, K. H. Min, J. O. Ka,Y K. Park, and C. K. Kim. 1997. Characteristics of catechol 2,3-dioxygenase produced by 4-chlorobenzoate degrading Pseudomonas sp S47. J. Microbial. 35: 295-299.
  13. Kim, K. P., D. J. Seo, D. H. Lee, Y Kim and C. K. Kim. 1998. Cloning and expression in E. coli of the genes responsible for degradation of 4-chlorobenzoate and 4-chlorocatechol from Pseudomonas sp. S-47. J. Microbiol. 36: 99-105.
  14. Kim, Y, B. Choi, J. Lee, H. Chang, and K. R. Min. 1992. Characterization of catechol 2,3-dioxygenase. Biochem. BioPhys. Res. Comm. 183: 77-82. https://doi.org/10.1016/0006-291X(92)91611-S
  15. Noh, S. J., Y Kim, K. H. Min, T. B. Karegoudar, and C. K. Kim. 2000. Cloning and nucleotide sequence analysis of xylE gene responsible for meta-Cleavage of 4-chlorocatechol from Pseudomonas sp. S-47. Mol. Cells. 10: 475--479.
  16. Park, D. W, Y Kim, S. M. Lee, J. O. Ka, and C. K. Kim. 2000. Cloning and nucleotide sequence analysis of xylL gene responsible for 4CBA-dihydrodiol dehydrogenase from Pseudomonas sp. S-47. J. Microbial. 38: 275-280.
  17. Sambrook, J., E. F. Fritisch, and T. Maniatis. 1989. Molecular cloning: A laboratory mannual. 2nd. ed. Cold Spring Harbour, NY
  18. Sentchilo, V S., A. N. Perebituk, A. J. B. Zehnder, and J. Roelof van der Meer. 2000. Molecular diversity of plasmids bearing genes that encode toluene and xylene metabolism in Pseudomonas strains Isolated from different contaminated sites in belarus. Appl. Environ. Microbiol. 66: 2842-2852.
  19. Seo, D. I., J. C. Chae, K. P. Kim, Y S. Kim, K. S. Lee, and C. K. Kim. 1998. A pathway for 4-chlorobenzoate degradation by Pseudomonas sp. S-47. J. Microbial. Biotech. 8: 96100.
  20. Zaitsev, G. M., T. V Tsoi, V G. Grishenkov, E. G. Plotnikova and A. M. Boronin. 1991. Genetic control of degradation of chlorinated benzoic acids in Arthrobacter globiformis, Corynebacterium sepedonicum and Pseudomonas cepacia strains. FEMS Microbiol. Lett. 81: 171-176.