Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Expression and Purification of a Cathelicidin-Derived Antimicrobial Peptide, CRAMP

  • Park Eu-Jin (Department of Advanced Fusion Technology and Bio/Molecular Informatics Center, Konkuk University) ;
  • Chae Young-Kee (Department of Chemistry and Recombinant Protein Expression Center, Sejong University) ;
  • Lee Jee-Young (Department of Advanced Fusion Technology and Bio/Molecular Informatics Center, Konkuk University) ;
  • Lee Byoung-Jae (Laboratory of Molecular Genetics, School of Biological Sciences, Institute of Molecular Biology and Genetics) ;
  • Kim Yang-Mee (Department of Advanced Fusion Technology and Bio/Molecular Informatics Center, Konkuk University)
  • Published : 2006.09.01
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Abstract

Application of recombinant protein production and particularly their isotopic enrichment has stimulated development of a range of novel multidimensional heteronuclear NMR techniques. Peptides in most cases are amenable to assignment and structure determination without the need for isotopic labeling. However, there are many cases where the availability of $^{15}N$ and/or $^{13}C$ labeled peptides is useful to study the structure of peptides with more than 30 residues and the interaction between peptides and membrane. CRAMP (Cathelicidin-Related AntiMicrobial Peptide) was identified from a cDNA clone derived from mouse femoral marrow cells as a member of cathelicidin-derived antimicrobial peptides. CRAMP was successfully expressed as a GST-fused form in E. coli and purified using affinity chromatography and reverse-phase chromatography. The yield of the CRAMP was 1.5 mg/l 1. According to CD spectra, CRAMP adopted ${\alpha}$-helical conformation in membrane-mimetic environments. Isotope labeling of CRAMP is expected to make it possible to study the structure and dynamic properties of CRAMP in various membrane systems.

Keywords

References

  1. Bajenova, O. V., R. Zimmer, E. Stolper, J. Salisbury-Rowswell, A. Nanji, and P. Thomas. 2001. Heterogeneous RNA-binding protein M4 is a receptor for carcinoembryonic antigen in Kupffer cells. J. Biol. Chem. 276: 31067-31073 https://doi.org/10.1074/jbc.M104093200
  2. Baneyx, F. 1999. Recombinant protein expression in Escherichia coli. Curr. Opin. Biotechnol. 10: 411-421 https://doi.org/10.1016/S0958-1669(99)00003-8
  3. Brogden, K. A. 2005. Antimicrobial peptides: Pore formers or metabolic inhibitors in bacteria. Nature Rev. Microbiol. 3: 238-250 https://doi.org/10.1038/nrmicro1098
  4. Cipakova, I., J. Gasperik, and E. Hostinova. 2006. Expression and purification of human antimicrobial peptide, dermcidin, in Escherichia coli. Protein Expr. Purif. 45: 269-274 https://doi.org/10.1016/j.pep.2005.07.002
  5. Cornells, P. 2000. Expressing genes in different Escherichia coli. compartments. Curr. Opin. Biotechnol. 11: 450-454 https://doi.org/10.1016/S0958-1669(00)00131-2
  6. Delaglio, F., S. Frzesiek, G. W. Vuister, G. Zhu, J. Pfeifer, and A. Bax. 1995. NMRPipe: A multidimensional spectral processing system based on UNIX pipes. J. Biomol. NMR 6: 277-293
  7. Gallo, R. L., K. J. Kim, M. Bernfield, C. A. Kozak, M. Zanetti, L. Merluzzi, and R. Gennaro. 1997. Identification of CRAMP, a cathelin-related antimicrobial peptide expressed in the embryonic and adult mouse. J. Biol. Chem. 272: 13088-13093 https://doi.org/10.1074/jbc.272.20.13088
  8. Goddard, T. D. and D. G. Kneller. 2004. Sparky 3 (3.110ed.). University of California, San Francisco
  9. Halbhuber, Z., Z. Petrmichlova, K. Alexciev, E. Thulin, and D. Stys. 2003. Overexpression and purification of recombinant membrane PsbH protein in Escherichia coli. Protein Expr. Purif. 32: 18-27 https://doi.org/10.1016/S1046-5928(03)00188-8
  10. Hanning, G. and S. C. Markrides. 1998. Strategies for optimizing heterologous protein expression in Escherichia coli. Trends Biotechnol. 16: 54-60 https://doi.org/10.1016/S0167-7799(97)01155-4
  11. Kim, Y., K. Park, S. W. Kang, S. Y. Shin, K. S. Hahtn, and Y. Kim. 2002. Solution structure of a cathelicidin-derived antimicrobial peptide, CRAMP, as determined by NMR spectroscopy. J. Pept. Res. 60: 1-9 https://doi.org/10.1034/j.1399-3011.2002.01968.x
  12. Lee, B. Y. 2005. Expression of human interleukin-11 and granulocyte-macrophage colony-stimulating factor in transgenic plants. J. Microbiol. Biotechnol. 15: 1304-1309
  13. Liu, S. and X. Zhang. 2004. Expression and purification of a novel rice (Oryza sativa L.) mitochondrial ATP synthase small subunit in Escherichia coli. Protein Expr. Purif. 37: 306-310 https://doi.org/10.1016/j.pep.2004.06.010
  14. Lin, Y. W., D. X. Zhao, Z. H. Wang, W. H. Yu, and Z. X. Huang. 2006. Expression of lipase-solubilized bovine liver microsomal cytochrome b (5) in Escherichia coli as a glutathione S-transferase fusion protein (GST-cyt b (5)). Protein Expr. Purif. 45: 352-358 https://doi.org/10.1016/j.pep.2005.06.006
  15. Oh, J. S. and S. C. Yoon. 2005. In vivo $^{13}C$-NMR spectroscopic study of polyhydroxyalkanoic acid degradation kinetics in bacteria. J. Microbiol. Biotechnol. 15: 1330-1336
  16. Ohk, S. H. and D. H. Bai. 2004. Purification and characterization of cell wall hydrolase from alkalophilic Bacillus mutanolyticus YU5215. J. Microbiol. Biotechnol. 14: 1142-1149
  17. Park, K., D. Oh, S. Y. Shin, K. S. Hahm, and Y. Kim. 2002. Structural studies of porcine myeloid antibacterial peptide PMAP-23 and its analogues in DPC micelles by NMR spectroscopy. Biochem. Biophys. Res. Commun. 290: 204-212 https://doi.org/10.1006/bbrc.2001.6173
  18. Pestonjamasp, V. K., K. H. Huttner, and R. L. Gallo. 2001. Processing site and gene structure for the marine antimicrobial peptide CRAMP. Peptides 22: 1643-1650 https://doi.org/10.1016/S0196-9781(01)00499-5
  19. Piccinini, M., A. Merighi, R. Bruno, P. Cascio, M. Curto, S. Mioletti, C. Ceruti, and M. T. Rinaudo. 1996. Affinity purification and characterization of protein gene product 9.5 (PGP9.5) from retina. Biochem. J. 318: 711-716 https://doi.org/10.1042/bj3180711
  20. Shi, W. J., H. J. Xu, J. A. Cheng, and C. X. Zhang. 2004. Expression of the melittin gene of Apis cerana cerana in Escherichia coli. Protein Expr. Purif. 37: 213-219 https://doi.org/10.1016/j.pep.2004.05.010
  21. Shin, H. J., S. K. Lee, J. J. Choi, and S. H. Koh. 2005. Cloning, expression, and characterization of a family B-type DNA polymerase from the hyperthermophillic crenarchaeon Pyrobaculum arsenaticum and its application to PCR. J. Microbiol. Biotechnol. 15: 1359-1367
  22. Ueki, N. and Y. Ochiai. 2005. Structural stabilities of recombinant Scombridae fish myoglobins. Biosci. Biotechnol. Biochem. 69: 1935-1943 https://doi.org/10.1271/bbb.69.1935
  23. Xiao, Y, Y Cai, Y. R. Bommineni, S. C. Fernando, O. Prakash, S. E. Gilliland, and G. Zhang. 2006. Identification and functional characterization of three chicken cathelicidins with potent antimicrobial activity. J. Biol. Chem. 281: 2858-2867 https://doi.org/10.1074/jbc.M507180200
  24. Zhang, L., T. Falla, M. Wu, S. Fidai, J. Burian, W. Kay, and R. E. Hancock. 1998. Determinants of recombinant production of antimicrobial cationic peptides and creation of peptide variants in bacteria. Biochem. Biophys. Res. Commun. 247: 674-680 https://doi.org/10.1006/bbrc.1998.8848