사상설 진균 Aspergillus nidulans의 Phospholipase B 유전자(plb A)의 클로링

Molecular Cloning of a Putative Gene Encoding Phospholipase B (plbA) from Aspergillus nidulans

  • 홍사현 (국립보건연구원 감염병센터 장내세균팀) ;
  • 조은민 (한국기초과학지원연구원 서울센터 메타볼륨분석연구팀) ;
  • 송승은 (한국기초과학지원연구원 서울센터 메타볼륨분석연구팀) ;
  • 엄치용 (한국기초과학지원연구원 서울센터 메타볼륨분석연구팀)
  • Hong, Sa-Hyun (Division of Enteric Bacterial Infections, Center for Infectious Diseases, Korean National Institute of Health) ;
  • Cho, Eun-Min (Metabolome Analysis Team, Seoul Center, Korea Basic Science Institute) ;
  • Song, Seung-Eun (Metabolome Analysis Team, Seoul Center, Korea Basic Science Institute) ;
  • Eom, Chi-Yong (Metabolome Analysis Team, Seoul Center, Korea Basic Science Institute)
  • 발행 : 2008.09.28

초록

Phospholipase B(PLB) families는 phospholipase(PL), lysophospholipase(LPL) 그리고 lysophospholipase-transacylase(LHA)의 활성을 공유하고 있는 효소이다. 본 연구에서는, 사상성 진균인 Aspergillus nidulans에서 최초로 lipase motifs를 보유하고 있는 단백질 Phospholipase B를 인코딩하는 유전자(plb A)를 클로닝하였다. plb A는 다양한 PLB 효소들의 lipase 보존영역들의 염기서열 정보에 근거하여 제작한 probe를 이용하여 A. nidulans genomic DNA library로 부터 분리하였다. Phospholipase B 유전자의 염기서열을 결정한 결과 626아미노산으로 구성된 단백질을 코드하고 있었다. PlbA는 Penicillium notatum의 PLB와는 72%의 높은 상동성을 나타내었으나, 다른 생물유래의 PLA와는 낮은 상동성을 나타내었다.

The phospholipase B (PLB) families are enzymes sharing phospholipase (PL), lysophospholipase (LPL) and lysophospholipase-transacylase (LPTA) activities. In this study, we report the putative gene encoding phospholipase B (plbA) containing lipase motifs was cloned for the first time from the filamentous fungus, Aspergillus nidulans. plbA was isolated from A. nidulans genomic DNA library using a PCR-amplified probe, which is designed on the basis of sequence information derived from the conserved lipase regions of various PLBs. The deduced product of plbA is of 626 amino acids. From the assigned sequence, PlbA showed 72% identity with Penicillium notatum PLB but have low similarity with phospholipase A of other organisms.

키워드

참고문헌

  1. Chen, S. C., L. C. Wright, J. C. Golding, and T. C. Sorrell. 2000. Purification and characterization of secretory phospholipase B, lysophospholipase and lysophospholipase/transacylase from a virulent strain of the pathogenic fungus Cryptococcus neoformans. Biochem. J. 347: 431-439 https://doi.org/10.1042/0264-6021:3470431
  2. Cox, G. M., H. C. McDade, S. C. Chen, S. C. Tucker, M. Gottfredsson, L. C. Wright, T. C. Sorrell, S. D. Leidich, A. Casadevall, M. A. Ghannoum, and J. R. Perfect. 2001. Extracellular phospholipase activity is a virulence factor for Cryptococcus neoformans. Mol. Microbiol. 39: 166-175 https://doi.org/10.1046/j.1365-2958.2001.02236.x
  3. Fyrst, H., B. Oskouian, F. A. Kuypers, and J. D. Saba. 1999. The PLB2 gene of Saccharomyces cerevisiae confers resistance to lysophosphatidylcholine and encodes a phospholipase B/lysophospholipase. Biochem. 38: 5864-5871 https://doi.org/10.1021/bi9824590
  4. Ghannoum, M. A. 2000. Potential role of phospholipases in virulence and fungal pathogenesis. Clin. Microbiol. Rev. 13: 122-143 https://doi.org/10.1128/CMR.13.1.122-143.2000
  5. Hanel, A. and M. Gelb. 1995. Multiple enzymatic activities of the human cytosolic 85 kDa phospholipase $A_{2}$: hydrolytic reactions and acyl transfer to glycerol. Biochem. 34: 7807-7818 https://doi.org/10.1021/bi00024a004
  6. Ibrahim, A. S., F. Mirbod, S. G. Filler, Y. Banno, G. T. Cole, Y. Kitajima, J. E. Edwards Jr, Y. Nozawa, and M. A. Ghannoum 1995. Evidence implicating phospholipase as a virulence factor of Candida albicans. Infect. Immun. 63: 1993-1998
  7. Johnson, D. R., R. S. Bhatnagar, L. J. Knoll, and J. I. Gordon. 1994. Genetic and biochemical studies of protein N-myristoylation. Annu. Rev. Biochem. 63: 869-914 https://doi.org/10.1146/annurev.bi.63.070194.004253
  8. Lee, K. S., J. L. Patton, M. Fido, L. K. Hines, S. D. Kohlwein, F. Paltauf, S. A. Henry, and D. E. Levin. 1994. The Saccharomyces cerevisiae PLB1 gene encodes a protein required for lysophospholipase and phospholipase B activity. J Biol. Chem. 269: 19725-19730
  9. Leidich, S. D., A. S. Ibrahim, Y. Fu, A. Koul, C. Jessup, J. Vitullo, W. Fonzi, F. Mirbod, S. Nakashima, Y. Nozawa, and M. A. Ghannoum. 1998. Cloning and disruption of CaPLB1, a phospholipase B gene involved in the pathogenicity of Candida albicans. J. Biol. Chem. 273: 26078-26086 https://doi.org/10.1074/jbc.273.40.26078
  10. Loo, R. W., K. Conde-Frieboes, L. J. Reynolds, and E. A. Dennis. 1997. Activation, inhibition, and regiospecificity of the lysophospholipase activity of the 85-kDa group IV cytosolic phospholipase $A_{2}$. J. Biol. Chem. 272: 19214-19219 https://doi.org/10.1074/jbc.272.31.19214
  11. Masuda, N., N. Kitamura, and K. Saito. 1991. Primary structure of protein moiety of Penicillium notatum phospholipase B deduced from the cDNA. Eur. J. Biochem. 202: 783-787 https://doi.org/10.1111/j.1432-1033.1991.tb16433.x
  12. Merkel, O., M. Fido, J. A. Mayr, H. Pruger, F. Raab, G. Zandonella, S. D. Kohlwein, and F. Paltauf. 1999. Characterization and function in vivo of two novel phospholipases B/lysophospholipases from Saccharomyces cerevisiae. J. Biol. Chem. 274: 28121-28127 https://doi.org/10.1074/jbc.274.40.28121
  13. Reynolds, L., L. Hughes, A. Louis, R. Kramer, and E. Dennis 1993. Metal ion and salt effects on the phospholipase $A_{2}$, lysophospholipase, and transacylase activities of human cytosolic phospholipase$A_{2}$. Biochim. Biophys. Acta. 1167: 272-280 https://doi.org/10.1016/0005-2760(93)90229-3
  14. Rowlands, R. T. and G. Turner. 1973. Nuclear and extranuclear inheritance of oligomycin resistance in Aspergillus nidulans. Mol. Gen. Genet. 126: 201-16 https://doi.org/10.1007/BF00267531
  15. Saito, K., J. Sugatani, and T. Okumura. 1991. Phospholipase B from Penicillium notatum. Methods Enzymol. 197: 446-456 https://doi.org/10.1016/0076-6879(91)97170-4
  16. Schultz, J., F. Milpetz, P. Bork, and C. P. Ponting. 1998. SMART, a simple modular architecture research tool: identification of signaling domains. Proc. Natl. Acad. Sci. USA. 95: 5857-64
  17. Shen, D. K., A. D. Noodeh, A. Kazemi, R. Grillot, G. Robson, and J. F. Brugere. 2004. Characterisation and expression of phospholipases B from the opportunistic fungus Aspergillus fumigatus. FEMS Microbiol. Lett. 239: 87-93 https://doi.org/10.1016/j.femsle.2004.08.019
  18. Tamura, M., T. Ajayi, L. R. Allmond, K. Moriyama, J. P. Wiener-Kronish, and T. Sawa. 2004. Lysophospholipase A activity of Pseudomonas aeruginosa type III secretory toxin ExoU. Biochem. Biophys. Res. Commun. 316: 323-331 https://doi.org/10.1016/j.bbrc.2004.02.050
  19. Takemori, H., F. N. Zolotaryov, L. Ting, T. Urbain, T. Komatsubara, O. Hatano, M. Okamoto, and H. Tojo. 1998. Identification of functional domains of rat intestinal phospholipase B/lipase. Its cDNA cloning, expression, and tissue distribution. J. Biol. Chem. 273: 2222-2231 https://doi.org/10.1074/jbc.273.4.2222
  20. Wang, A. and E. A. Dennis. 1999. Mammalian lysophospholipases. Biochim. Biophys. Acta. 1439: 1-16 https://doi.org/10.1016/S1388-1981(99)00063-3