Molecular cloning of a novel cecropin-like peptide gene from the swallowtail butterfly, Papilio xuthus

Kim, Seong-Ryul;Choi, Kwang-Ho;Kim, Sung-Wan;Hwang, Jae-Sam;Goo, Tae-Won;Kim, Iksoo

  • Received : 2015.10.30
  • Accepted : 2015.11.03
  • Published : 2015.12.31


A new cecropin-like antimicrobial peptide (Px-CLP) gene was isolated from the immunechallenged larvae of the swallowtail butterfly, Papilio xuthus, by employing annealing control primer (ACP)-based GeneFishing PCR. The full-length cDNA of Px-CLP is 310 nucleotides encoding a 70 amino acid precursor that contains a putative 22-residue signal peptide, a 4-residue propeptide, a presumed 37-residue mature peptide, and an uncommon 7-residue acidic pro-region at the C-terminus. The deduced amino acid sequence of Px-CLP showed significant identities with other Lepidopteran cecropin D type peptides. RT-PCR revealed that the Px-CLP transcript was detected at significant level after injection with bacterial lipopolysaccharide (LPS). The peptides with or without C-terminal acidic sequence region were synthesized on-solid phage and submitted to antibacterial activity assay. The synthetic 37-mer peptide (Px-CLPa), which removed C-terminal acidic sequence region, was showed exclusively antibacterial activity against E. coli ML35; meanwhile, a 44-mer peptide (Px-CLPb) with C-terminal acidic peptide region was not active. This result suggests that Px-CLP is produced as a larger precursor containing a C-terminal pro-region that is subsequently removed by C-terminal modification.


Papilio xuthus;antimicrobial peptide;RT-PCR;antibacterial activity


  1. Pillai A, Ueno S, Zhang H, Lee JM, Kato Y (2005) Cecropin P1 and novel nematode cecropins: a bacteria-inducible antimicrobial peptide family in the nematode Ascaris suum. Biochem J 390, 207-214.
  2. Saito A, Ueda K, Imamura M, Atsumi S, Tabunoki H, Miura N et al. (2005) Purification and cDNA cloning of a cecropin from the longicorn beetle, Acalolepta luxuriosa. Comp Biochem Physiol B 142, 317-323.
  3. Steiner H, Hultmark D, Engstrom A, Bennich H, Boman HG (1981) Sequence and specificity of two antibacterial proteins involved in insect immunity. Nature 292, 246-248.
  4. Vizioli J, Bulet P, Charlet M, Lowenberger C, Blass C, Muller HM et al. (2000) Cloning and analysis of a cecropin gene from the malaria vector mosquito, Anopheles gambiae. Insect Mol Biol 9, 75-84.
  5. Zhao C, Liaw L, Lee IH, Lehrer RI (1997) cDNA cloning of three cecropin-like antimicrobial peptides (Styelins) from the tunicate, Styela clava. FEBS Lett 412, 144-148.
  6. Gudmundsson GH, Lidholm DA, Asling B, Gan R, Boman HG (1991) The cecropin locus. Cloning and expression of a gene cluster encoding three antibacterial peptides in Hyalophora cecropia. J Biol Chem 266, 11510-11517.
  7. Hara S, Taniai K, Kato Y, Yamakawa M (1994) Isolation and α-amidation of the non-amidated form of cecropin D from larvae of Bombyx mori. Comp Biochem Physiol B 108, 303-308.
  8. Hoffman JA, Kafatos FC, Janeway CA, Ezekowitz RA (1999) Phylogenetic perspectives in innate immunity. Science 284, 1313-1318.
  9. Jenssen H, Hamill P, Hancock RE (2006) Peptide antimicrobial agents, Clin Microbiol Rev 19, 491-511.
  10. Kim SR, Hong MY, Park SW, Choi KH, Yun EY, Goo TW, et al. (2010) Characterization and cDNA cloning of cecropin-like antimicrobial peptide, Papiliocin from the swallowtail butterfly, Papilio xuthus. Mol Cells 29, 419-423.
  11. Kylsten P, Samakovlis C, Hultmark D (1990). The cecropin locus in Drosophila; a compact gene cluster involved in the response to infection. EMBO J 9, 217-224.
  12. Li W, Li Z, Du C, Chen W, Pang Y (2007). Characterization and expression of a cecropin-like gene from Helicoverpa armigera. Comp Biochem Physiol B 148, 417-425.
  13. Liang Y, Wang JX, Zhao XF, Du XJ, Xue JF (2006) Molecular cloning and characterization of cecropin from the housefly (Musca domestica), and its expression in Escherichia coli. Dev Comp Immunol 30, 249-257.
  14. Morishima I, Suginaka S, Ueno T, Hirano H (1990) Isolation and structure of cecropins, inducible antibacterial peptides, from the silkworm, Bombyx mori. Comp Biochem Physiol B 95, 551-554.
  15. Boman HG, Boman IA, Andreu D, Li ZQ, Merrifield RB, Schlenstedt G et al. (1989). Chemical synthesis and enzymic processing of precursor forms of cecropins A and B. J Biol Chem 264, 5852-5860.
  16. Boman HG (1995) Peptide antibiotics and their role in innate immunity. Annu Rev Immunol 13, 61-92.
  17. Bulet P, Hetru C, Dimarcq J, Hoffmann D (1999) Antimicrobial peptides in insects; structure and function. Dev Comp Immunol 23, 329-344.
  18. Chalk R, Townson H, Ham PJ (1995) Brugia pahangi: the effects of cecropins on microfilariae in vitro and in Aedes Aegypti. Exp Parasitol 80, 401-406.
  19. Callaway JE, Lai J, Haselbeck B, Baltaian M, Bonnesen SP, Weickmann J et al. (1993) Modification of the C terminus of cecropin is essential for broad-spectrum antimicrobial activity. Antimicrob Agents Chemother 37, 1614-1619.
  20. Cociancich S, Bulet P, Hetru C, Hoffmann JA (1994) The inducible antibacterial peptides of insects. Parasitol Today, 132-139.
  21. DeLucca AJ, Bland JM, Jacks TJ, Grimm C, Cleveland TE, Walsh TJ (1997) Fungicidal activity of cecropin A. Antimicrob Agents Chemother 41, 481-483.


Grant : 누에 생체공장화 시스템 이용 천연항생제 생산 기술