생명과학회지 (Journal of Life Science)

  • 제19권5호
  • /
  • Pages.566-572
  • /
  • 2009
  • /
  • 1225-9918(pISSN)
  • /
  • 2287-3406(eISSN)
한국생명과학회 (Korean Society of Life Science)
권호 표지
DOI QR코드

DOI QR Code

Serratia marcescens KCTC 2172로부터 pst operon의 클로닝 및 해석

Molecular Cloning and Analysis of Phosphate Specific Transport (pst) Operon from Serratia marcescens KCTC 2172

  • Lee, Seung-Jin (Department of Biotechnology, College of Natural Resources and Life Science) ;
  • Lee, Yong-Seok (Department of Biotechnology, College of Natural Resources and Life Science) ;
  • Lee, Sang-Cheol (Department of Biotechnology, College of Natural Resources and Life Science) ;
  • Park, In-Hye (Department of Biotechnology, College of Natural Resources and Life Science) ;
  • Ahn, Soon-Cheol (School of Medicine, College of Medicine, Pusan National University) ;
  • Choi, Yong-Lark (Department of Biotechnology, College of Natural Resources and Life Science)
  • 발행 : 2009.05.30
PDF 다운로드
⟨ 이전 논문 다음 논문 ⟩

초록

S. marcescens KCTC 2172로부터 유전자 은행을 작성하여 재조합 클론 pDH3를 얻었으며, pDH3 유래의 서브클론을 작성하였다. 플라스미드 pPH4의 전염기서열 5,137 bp 영역을 결정한 결과 3개의 ORF가 있음을 확인하였다. 이들은 pst 오페론의 pstC, pstA, 및 pstB, 세 유전자를 동일 전사방향으로 코드하고 있었다. 타 세균의 유전자와 비교한 결과 S. marcescens의 pst 오페론은 pstS와 phoU가 결손되어 있다. 조절영역에는 CRP 결합영역과 pho box 서열이 존재하였다. 보고된 유전자와 상동성 조사결과, PstC 단백질은 Yersinia sp., Vibrio sp. 및 Pseudomonas sp.와는 49, 37, 33%의 상동성을, PstA 단백질은 Yersinia sp., Vibrio sp. 및 Pseudomonas sp.와 64, 51, 47%의 상동성을, PstB 단백질은 Methanocaldococcus sp., E. coli 및 Mycoplasma sp.와 60, 50, 48%의 상동성을 나타내었다. Pst 유전자들은 조절영역의 cAMP-CRP 복합체에 의해 in vivo에서 양성적으로 발현됨을 확인하였다. Pst 오페론을 포함하는 플라스미드를 도입한 대장균은 인산운송에 관여하는 능력을 확인하였다.

A recombinant plasmid, pDH3, was obtained from the genomic library of Serattia marcescens KCTC 2172, and several recombinant subclones constructed from pDH3. The nucleotide sequence of a 5,137 bp segment, pPH4, was determined and three open reading frames were detected. The three ORFs encoded the phosphate specific transport (pst) operon, which was pstC, pstA, and pstB, with the same direction of transcription. Comparison of the pst operon of S. marcescens with that of other organisms revealed that the genes for pstS and phoU were missing. A potential CRP bonding site and pho box sequence was found in the upstream of the putative promoter at the regulatory region. Analysis of the nucleotide sequence showed that homology in amino acid sequences between the PstC protein and Yersinia sp., Vibrio sp., and Pseudomonas sp. were 49, 37 and 33%, respectively. The PstA protein and Yersinia sp., Vibrio sp., and Pseudomonas sp. showed homologies of 64, 51, and 47%, respectively. PstB protein and Methanocaldococcus sp., E. coli, and Mycoplasma sp. showed homologies of 60, 50, and 48%, respectively. The pst genes could be expressed in vivo and positively regulated by cAMP-CRP. The E. coli strain harboring plasmid pPH7, with pst genes, increased with the transport of phosphate.

키워드

참고문헌

  1. Allenby, N. E. E., N. O’Connor, Z. Pragai, N. M. Carter, M. Miethke, S. Engelmann, M. Hecker, A. Wipat, A. C. Ward, and C. R. Harwood. 2004. Post-transcriptional regulation of the Bacillus subtilis pst operon encoding a phosphate-specific ABC transporter. Microbiology 150, 2619-2628 https://doi.org/10.1099/mic.0.27126-0
  2. Atalla, A. and A. Schumann. 2003. The pst operon of Bacillus subtilis is specially induced by alkali stress. J. Bacteriol. 185, 5019-5022 https://doi.org/10.1128/JB.185.16.5019-5022.2003
  3. Braibant, M., P. Lefevre, L. Wit, P. Peirs, J. Ooms, K. Huhgen, A. B.. Anderson, and J. A. Content. 1996. Mycobacterium tuberculosis gene cluster encoding proteins of a phosphate transport homologous to the Escherichia coli Pst system. Gene 176, 171-176 https://doi.org/10.1016/0378-1119(96)00242-9
  4. Ficsher, R. J., S. Oehmcke, U. Meyer, M. Mix, K. Schwarz, T. Fielder, and H. Barl. 2006. Transcription of the pst operon of Clostridium acetobutyricum is dependent on phosphate concentration and pH. J. Bacteriol. 188, 5469-5478 https://doi.org/10.1128/JB.00491-06
  5. Gal, S. W., Y. J. Choi, C. Y. Kim, Y. H. Cheong, J. D. Bahk, and M. J. Cho. 1998. Cloning of the 52-kDa chitinase gene from Serratia marcescens KCTC 2172 and its proteolytic cleavage into an active 35-kDa enzyme. FEMS Microbiol. Lett. 160, 151-158 https://doi.org/10.1111/j.1574-6968.1998.tb12905.x
  6. Hadimann, A., L. L. Daniels, and B. L. Wanner. 1998. Use of new methods for construction of tightly regulated arabinose and rhamnose promoter fusions in studies of the Escherichia coli phosphate regulon. J. Bacteriol. 180, 1277-1286
  7. Harris, R. M., D. C. Webb, S. M. Howitt, and G. B. Cox. 2001. Characterization of PitA and PitB from Escherichia coli. J. Bacteriol. 183, 5008-5014 https://doi.org/10.1128/JB.183.17.5008-5014.2001
  8. Jenkins, D. and V. Tandoi. 1991. The applied microbiology of enhanced biological phosphate removal-accomplishments and needs. Water Res. 25, 1471-1478 https://doi.org/10.1016/0043-1354(91)90177-R
  9. Kato, J., K. Yamada, A. Muramatsu, and H. Ohtake. 1993. Genetic improvement of Escherichia coli for enhanced biological removal of phosphate from wastewater. Appl. Environ. Microbiol. 59, 3744-3749
  10. Lee, S. J., O. R. Song, Y. C. Lee, Y. C. and Y. L. Choi. 2003. Molecular characterization of polyphosphate kinase (ppk) gene from Serratia marcescens. Biotechnol. Lett. 25, 191-197 https://doi.org/10.1023/A:1022386514487
  11. Lee, S.J., Y. S. Lee, Y. C. Lee, and Y. L. Choi. 2006. Molecular characterization of polyphosphate (PolyP) operon from Serratia marcescens. J. Basic Microbiol. 46, 108-115 https://doi.org/10.1002/jobm.200510038
  12. Magota, K., N. Otsuji, T. Miki, T. Horiuchi, S. Tsunasawa, J. Kondo, F. Sakiyama, M. Amemura, T. Morita, H. Shinagawa, and A. Nakata. 1984. Nucleotide sequence of the phoS gene, the structural gene for the phosphate-binding protein of Escherichia coli. J. Bacteriol. 157, 909-917
  13. Mathew, J. A., Y. P. Tan, P. S. Srinivasa Rao, T. M. Lim, and K. Y. Leung. 2001. Edwardsiella tarda mutant defective in siderophore production, motility, serum resistance and catalase activity. Microbiology 147, 449-457
  14. Monds, R. D., M. W. Silby, and H. K. Mahanty. 2001. Expression of the Pho regulon negatively regulates biofilm formation by Pseudomonas aureofaciens PAI47.2. Mol. Microbiol. 42, 415-426 https://doi.org/10.1046/j.1365-2958.2001.02641.x
  15. Nakata, T., Y. Sakai, K. Shibata, J. Kato, A. Kuroda, and H. Ohtake. 1996. Molecular analysis of the phosphate-specific transport operon of Pseudomonas aeruginosa. Mol. Genet. Genomomics 250, 692-698
  16. Novak, R., A. Cauwels, E. Charpentier, and E. Tuomanen. 1999. Identification of a Streptococcus pneumoniae gene locus encoding proteins of an ABC phosphate transporter and a two-component regulatory system. J. Bacteriol. 181, 1126-1133
  17. Qi, Y., Y. Kobayashi, and F. M. Hulett. 1997. The pst operon of Bacillus subtilis has a phosphate-related promoter and is involved in phosphate transport but not in regulation of the pho regulon. J. Bacteriol. 179, 2534-2539
  18. Sambrook, J. and D. W. Russel. 2001. Molecular cloning: A Laboratory Manual, 3rd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York., USA
  19. Stephen, J., V. Dien, S.. Keyhani, C. Yang, and J. D. Keasling. 1997. Manipulation of independent synthesis and degradation of phosphate in Escherichia coli for phosphate secretion from the cell. Appl. Environ. Microbiol. 63, 1689-1695
  20. Wu, H., H. Kosaka, J. Kato, A. Kuroda, T. Ikeda, N. Kakiguchi, and H. Ohtake. 1999. Cloning and characterization of Pseudomonas putida genes encoding the phosphate-specific transport system. J. Biosci. Bioeng. 87, 273-279 https://doi.org/10.1016/S1389-1723(99)80031-0
  21. Yoo, J. S., H. S. Kim, S. Y. Chung, and Y. L. Choi. 2000. Characterization of crp, the cyclic AMP receptor protein gene of Serratia marcescens KCTC 2172. J. Microbiol. Biotechnol. 10, 670-676