전체메뉴
검색
Article Search

JMB Journal of Microbiolog and Biotechnology

QR Code QR Code

Research article

References

  1. Athmaram TN, Saraswat S, Singh AK, Rao MK, Gopalan N, Suryanarayana VVS, Rao PVL. 2012. Influence of copy number on the expression levels of pandemic influenza hemagglutinin recombinant protein in methylotrophic yeast Pichia pastoris. Virus Genes 45: 440-451.
    Pubmed CrossRef
  2. Barton MD. 2000. Antibiotic use in animal feed and its impact on human health. Nutr. Res. Rev. 13: 279-299.
    Pubmed CrossRef
  3. Brinch KS, Sandberg A, Baudoux P, Van Bambeke F, Tulkens PM, Frimodt-Møller N, et al. 2009. Plectasin shows intracellular activity against Staphylococcus aureus in human THP-1 monocytes and in a mouse peritonitis model. Antimicrob. Agents Chemother. 53: 4801-4808.
    Pubmed PMC CrossRef
  4. Buckholz RG, Gleeson MA. 1991. Yeast systems for the commercial production of heterologous proteins. Nat. Biotechnol. 9: 1067-1072.
    CrossRef
  5. Cereghino JL, Cregg JM. 2000. Heterologous protein expression in the methylotrophic yeast Pichia pastoris. FEMS Microbiol. Rev. 24: 45-66.
    Pubmed CrossRef
  6. Dhople V, Krukemeyer A, Ramamoorthy A. 2006. The human beta-defensin-3, an antibacterial peptide with multiple biological functions. BBA Biomembr. 1758: 1499-1512.
    Pubmed CrossRef
  7. Hancock REW, Patrzykat A. 2002. Clinical development of cationic antimicrobial peptides: from natural to novel antibiotics. Curr. Drug Targets 2: 79-83.
    CrossRef
  8. Hancock REW, Sahl HG. 2006. Antimicrobial and hostdefense peptides as new anti-infective therapeutic strategies. Nat. Biotechnol. 24: 1551-1557.
    Pubmed CrossRef
  9. Hara S, Mukae H, Sakamoto N, Ishimoto H, Amenomori M, Fujita H, et al. 2008. Plectasin has antibacterial activity and no affect on cell viability or IL-8 production. Biochem. Biophys. Res. Commun. 374: 709-713.
    Pubmed CrossRef
  10. Hunt I. 2005. From gene to protein: a review of new and enabling technologies for multi-parallel protein expression. Protein Expr. Purif. 40: 1-22.
    Pubmed CrossRef
  11. Iwatani S, Zendo T, Yoneyama F, Nakayama J, Sonomoto K. 2007. Characterization and structure analysis of a novel bacteriocin, lacticin Z, produced by Lactococcus lactis QU 14. Biosci. Biotechnol. Biochem. 71: 1984-1992.
    Pubmed CrossRef
  12. Jing XL, Luo XG, Tian WJ, Lv LH, Jiang Y, Wang N, Zhang TC. 2010. High-level expression of the antimicrobial peptide plectasin in Escherichia coli. Curr. Microbiol. 61: 197-202.
    Pubmed CrossRef
  13. Jonasson P, Nygren PÅ, Jörnvall H, Johansson BL, Wahren J, Uhlén M, Ståhl S. 2000. Integrated bioprocess for production of human proinsulin C-peptide via heat release of an intracellular heptameric fusion protein. J. Biotechnol. 76: 215-226.
    CrossRef
  14. Kim H K, Chun DS, Kim J S, Yun CH, Lee JH, Hong SK, Kang DK. 2006. Expression of the cationic antimicrobial peptide lactoferricin fused with the anionic peptide in Escherichia coli. Appl. Microbiol. Biotechnol. 72: 330-338.
    Pubmed CrossRef
  15. Kim JM, Jang SA, Yu BJ, Sung BH, Cho JH, Kim SC. 2008. High-level expression of an antimicrobial peptide histonin as a natural form by multimerization and furin-mediated cleavage. Appl. Microbiol. Biotechnol. 78: 123-130.
    Pubmed CrossRef
  16. Laemmli UK. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680-685.
    Pubmed CrossRef
  17. Lee SJ, Lee JH, Jin HJ, Lee JH, Ryu HY, Kim Y, et al. 2000. A novel technique for the effective production of short peptide analogs from concatameric short peptide multimers. Mol. Cells 10: 236-240.
    Pubmed CrossRef
  18. Lewis RV, Hinman M, Kothakota S, Fournier MJ. 1996. Expression and purification of a spider silk protein: a new strategy for producing repetitive proteins. Protein Expr. Purif. 7: 400-406.
    Pubmed CrossRef
  19. Liu YF, Luo JC, Xu CY, Ren FC, Peng C, Wu GY, Zhao JD. 2000. Purification, characterization, and molecular cloning of the gene of a seed-specific antimicrobial protein from pokeweed. Plant Physiol. 122: 1015-1024.
    Pubmed PMC CrossRef
  20. Luo HY, Huang HQ, Yang PL, Wang YR, Yuan TZ, Wu NF, et al. 2007. A novel phytase appA from Citrobacter amalonaticus CGMCC 1696: gene cloning and overexpression in Pichia pastoris. Curr. Microbiol. 55: 185-192.
    Pubmed CrossRef
  21. Maemoto A, Qu X, Rosengren KJ, Tanabe H, HenschenEdman A, Craik DJ, Ouellette AJ. 2004. Functional analysis of the α-defensin disulfide array in mouse cryptdin-4. J. Biol. Chem. 279: 44188-44196.
    Pubmed CrossRef
  22. Monroe S, Polk R. 2000. Antimicrobial use and bacterial resistance. Curr. Opin. Microbiol. 3: 496-501.
    CrossRef
  23. Murphey ED, Sherwood ER. 2008. Pretreatment with the gram-positive bacterial cell wall molecule peptidoglycan improves bacterial clearance and decreases inflammation and mortality in mice challenged with Pseudomonas aeruginosa. Microbes Infect. 10: 1244-1250.
    Pubmed PMC CrossRef
  24. Mygind PH, Fischer RL, Schnorr KM, Hansen MT, Sönksen CP, Ludvigsen S, et al. 2005. Plectasin is a peptide antibiotic with therapeutic potential from a saprophytic fungus. Nature 437: 975-980.
    Pubmed CrossRef
  25. Romanos MA, Scorer CA, Clare JJ. 1992. Foreign gene expression in yeast: a review. Yeast 8: 423-488.
    Pubmed CrossRef
  26. Schneider T, Kruse T, Wimmer R, Wiedemann I, Sass V, Pag U, et al. 2010. Plectasin, a fungal defensin, targets the bacterial cell wall precursor Lipid II. Science 328: 1168-1172.
    Pubmed CrossRef
  27. Schnorr KM, Hansen MT, Mygind PH, Segura DR, Hoegenh Kristensen HH, et al. 2011. Antimicrobial polypeptides from Pseudoplectania nigrella. US Patent 7,972,814.
  28. Schwarz S, Kehrenberg C, Walsh T R. 2 001. U se o f antimicrobial agents in veterinary medicine and food animal production. Int. J. Antimicrob. Agents 17: 431-437.
    CrossRef
  29. Singh M P, Greenstein M. 2 000. A ntibacterial l eads from microbial natural products discovery. Curr. Opin. Drug Discov. Devel. 3: 167-176.
    Pubmed
  30. Smith DL, Harris AD, Johnson JA, Silbergeld EK, Morris JG. 2002. Animal antibiotic use has an early but important impact on the emergence of antibiotic resistance in human commensal bacteria. Proc. Natl. Acad. Sci. USA 99: 6434-6439.
    Pubmed PMC CrossRef
  31. Tanabe H, Ayabe T, Maemoto A, Ishikawa C, Inaba Y, Sato R, et al. 2007. Denatured human α-defensin attenuates the bactericidal activity and the stability against enzymatic digestion. Biochem. Biophys. Res. Commun. 358: 349-355.
    Pubmed CrossRef
  32. Tian ZG, Dong TT, Yang YL, Teng D, Wang JH. 2009. Expression of antimicrobial peptide LH multimers in Escherichia coli C43 (DE3). Appl. Microbiol. Biotechnol. 83: 143149.
    Pubmed CrossRef
  33. Tian ZG, Teng D, Yang YL, Luo J, Feng XJ, Fan Y, et al. 2007. Multimerization and fusion expression of bovine lactoferricin derivative LfcinB15-W4,10 in Escherichia coli. Appl. Microbiol. Biotechnol. 75: 117-124.
    Pubmed CrossRef
  34. Vizioli J, Salzet M. 2002. Antimicrobial peptides versus parasitic infections? Trends Parasitol. 18: 475-476.
    CrossRef
  35. Wang XQ, Yang F, Liu C, Zhou HJ, Wu GY, Qiao SY, et al. 2012. Dietary supplementation with the probiotic Lactobacillus fermentum I5007 and the antibiotic aureomycin differentially affects the small intestinal proteomes of weanling piglets. J. Nutr. 142: 7-13.
    Pubmed CrossRef
  36. Yamada O, Sakamoto K, Tominaga M, Nakayama T, Koseki T, Fujita A, Akita O. 2005. Cloning and heterologous expression of the antibiotic peptide (ABP) genes from Rhizopus oligosporus NBRC 8631. Biosci. Biotechnol. Biochem. 69: 477-482.
    Pubmed CrossRef
  37. Zasloff M. 2002. Antimicrobial peptides of multicellular organisms. Nature 415: 389-395.
    Pubmed CrossRef
  38. Zhang J, Yang YL, Teng D, Tian ZG, Wang SR, Wang JH. 2011. Expression of plectasin in Pichia pastoris and its characterization as a new antimicrobial peptide against Staphyloccocus and Streptococcus. Protein Expr. Purif. 78: 189196.
    Pubmed CrossRef

Related articles in JMB

More Related Articles

Article

Research article

J. Microbiol. Biotechnol. 2016; 26(3): 461-468

Published online March 28, 2016 https://doi.org/10.4014/jmb.1508.08091

Copyright © The Korean Society for Microbiology and Biotechnology.

Expression of a Tandemly Arrayed Plectasin Gene from Pseudoplectania nigrella in Pichia pastoris and its Antimicrobial Activity

Jin Wan 1, 2, Yan Li 1, 2, Daiwen Chen 1, 2, Bing Yu 1, 2, Ping Zheng 1, 2, Xiangbing Mao 1, 2, Jie Yu 1, 2 and Jun He 1, 2*

1Institute of Animal Nutrition, Sichuan Agricultural University, Ya’an, Sichuan 625014, P.R. China, 2Key Laboratory of Animal Disease-Resistance Nutrition, Ministry of Education, Beijing 100029, P.R. China

Received: August 31, 2015; Accepted: December 3, 2015

Abstract

In recent years, various naturally occurring defence peptides such as plectasin have attracted
considerable research interest because they could serve as alternatives to antibiotics. However,
the production of plectasin from natural microorganisms is still not commercially feasible
because of its low expression levels and weak stability. A tandemly arrayed plectasin gene
(1,002 bp) from Pseudoplectania nigrella was generated using the isoschizomer construction
method, and was inserted into the pPICZαA vector and expressed in Pichia pastoris. The
selected P. pastoris strain yielded 143 μg/ml recombinant plectasin (Ple) under the control of
the methanol-inducible alcohol oxidase 1 (AOX1) promoter. Ple was estimated by SDS-PAGE
to be 41 kDa. In vitro studies have shown that Ple efficiently inhibited the growth of several
gram-positive bacteria such as Streptococcus suis and Staphylococcus aureus. S. suis is the most
sensitive bacterial species to Ple, with a minimum inhibitory concentration (MIC) of 4 μg/ml.
Importantly, Ple exhibited resistance to pepsin but it was quite sensitive to trypsin and
maintained antimicrobial activity over a wide pH range (pH 2.0 to 10.0). P. pastoris offers an
attractive system for the cost-effective production of Ple. The antimicrobial activity of Ple
suggested that it could be a potential alternative to antibiotics against S. suis and S. aureus
infections.

Keywords: Plectasin, Expression, Purification, Pichia pastoris, Antimicrobial activity

References

  1. Athmaram TN, Saraswat S, Singh AK, Rao MK, Gopalan N, Suryanarayana VVS, Rao PVL. 2012. Influence of copy number on the expression levels of pandemic influenza hemagglutinin recombinant protein in methylotrophic yeast Pichia pastoris. Virus Genes 45: 440-451.
    Pubmed CrossRef
  2. Barton MD. 2000. Antibiotic use in animal feed and its impact on human health. Nutr. Res. Rev. 13: 279-299.
    Pubmed CrossRef
  3. Brinch KS, Sandberg A, Baudoux P, Van Bambeke F, Tulkens PM, Frimodt-Møller N, et al. 2009. Plectasin shows intracellular activity against Staphylococcus aureus in human THP-1 monocytes and in a mouse peritonitis model. Antimicrob. Agents Chemother. 53: 4801-4808.
    Pubmed KoreaMed CrossRef
  4. Buckholz RG, Gleeson MA. 1991. Yeast systems for the commercial production of heterologous proteins. Nat. Biotechnol. 9: 1067-1072.
    CrossRef
  5. Cereghino JL, Cregg JM. 2000. Heterologous protein expression in the methylotrophic yeast Pichia pastoris. FEMS Microbiol. Rev. 24: 45-66.
    Pubmed CrossRef
  6. Dhople V, Krukemeyer A, Ramamoorthy A. 2006. The human beta-defensin-3, an antibacterial peptide with multiple biological functions. BBA Biomembr. 1758: 1499-1512.
    Pubmed CrossRef
  7. Hancock REW, Patrzykat A. 2002. Clinical development of cationic antimicrobial peptides: from natural to novel antibiotics. Curr. Drug Targets 2: 79-83.
    CrossRef
  8. Hancock REW, Sahl HG. 2006. Antimicrobial and hostdefense peptides as new anti-infective therapeutic strategies. Nat. Biotechnol. 24: 1551-1557.
    Pubmed CrossRef
  9. Hara S, Mukae H, Sakamoto N, Ishimoto H, Amenomori M, Fujita H, et al. 2008. Plectasin has antibacterial activity and no affect on cell viability or IL-8 production. Biochem. Biophys. Res. Commun. 374: 709-713.
    Pubmed CrossRef
  10. Hunt I. 2005. From gene to protein: a review of new and enabling technologies for multi-parallel protein expression. Protein Expr. Purif. 40: 1-22.
    Pubmed CrossRef
  11. Iwatani S, Zendo T, Yoneyama F, Nakayama J, Sonomoto K. 2007. Characterization and structure analysis of a novel bacteriocin, lacticin Z, produced by Lactococcus lactis QU 14. Biosci. Biotechnol. Biochem. 71: 1984-1992.
    Pubmed CrossRef
  12. Jing XL, Luo XG, Tian WJ, Lv LH, Jiang Y, Wang N, Zhang TC. 2010. High-level expression of the antimicrobial peptide plectasin in Escherichia coli. Curr. Microbiol. 61: 197-202.
    Pubmed CrossRef
  13. Jonasson P, Nygren PÅ, Jörnvall H, Johansson BL, Wahren J, Uhlén M, Ståhl S. 2000. Integrated bioprocess for production of human proinsulin C-peptide via heat release of an intracellular heptameric fusion protein. J. Biotechnol. 76: 215-226.
    CrossRef
  14. Kim H K, Chun DS, Kim J S, Yun CH, Lee JH, Hong SK, Kang DK. 2006. Expression of the cationic antimicrobial peptide lactoferricin fused with the anionic peptide in Escherichia coli. Appl. Microbiol. Biotechnol. 72: 330-338.
    Pubmed CrossRef
  15. Kim JM, Jang SA, Yu BJ, Sung BH, Cho JH, Kim SC. 2008. High-level expression of an antimicrobial peptide histonin as a natural form by multimerization and furin-mediated cleavage. Appl. Microbiol. Biotechnol. 78: 123-130.
    Pubmed CrossRef
  16. Laemmli UK. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680-685.
    Pubmed CrossRef
  17. Lee SJ, Lee JH, Jin HJ, Lee JH, Ryu HY, Kim Y, et al. 2000. A novel technique for the effective production of short peptide analogs from concatameric short peptide multimers. Mol. Cells 10: 236-240.
    Pubmed CrossRef
  18. Lewis RV, Hinman M, Kothakota S, Fournier MJ. 1996. Expression and purification of a spider silk protein: a new strategy for producing repetitive proteins. Protein Expr. Purif. 7: 400-406.
    Pubmed CrossRef
  19. Liu YF, Luo JC, Xu CY, Ren FC, Peng C, Wu GY, Zhao JD. 2000. Purification, characterization, and molecular cloning of the gene of a seed-specific antimicrobial protein from pokeweed. Plant Physiol. 122: 1015-1024.
    Pubmed KoreaMed CrossRef
  20. Luo HY, Huang HQ, Yang PL, Wang YR, Yuan TZ, Wu NF, et al. 2007. A novel phytase appA from Citrobacter amalonaticus CGMCC 1696: gene cloning and overexpression in Pichia pastoris. Curr. Microbiol. 55: 185-192.
    Pubmed CrossRef
  21. Maemoto A, Qu X, Rosengren KJ, Tanabe H, HenschenEdman A, Craik DJ, Ouellette AJ. 2004. Functional analysis of the α-defensin disulfide array in mouse cryptdin-4. J. Biol. Chem. 279: 44188-44196.
    Pubmed CrossRef
  22. Monroe S, Polk R. 2000. Antimicrobial use and bacterial resistance. Curr. Opin. Microbiol. 3: 496-501.
    CrossRef
  23. Murphey ED, Sherwood ER. 2008. Pretreatment with the gram-positive bacterial cell wall molecule peptidoglycan improves bacterial clearance and decreases inflammation and mortality in mice challenged with Pseudomonas aeruginosa. Microbes Infect. 10: 1244-1250.
    Pubmed KoreaMed CrossRef
  24. Mygind PH, Fischer RL, Schnorr KM, Hansen MT, Sönksen CP, Ludvigsen S, et al. 2005. Plectasin is a peptide antibiotic with therapeutic potential from a saprophytic fungus. Nature 437: 975-980.
    Pubmed CrossRef
  25. Romanos MA, Scorer CA, Clare JJ. 1992. Foreign gene expression in yeast: a review. Yeast 8: 423-488.
    Pubmed CrossRef
  26. Schneider T, Kruse T, Wimmer R, Wiedemann I, Sass V, Pag U, et al. 2010. Plectasin, a fungal defensin, targets the bacterial cell wall precursor Lipid II. Science 328: 1168-1172.
    Pubmed CrossRef
  27. Schnorr KM, Hansen MT, Mygind PH, Segura DR, Hoegenh Kristensen HH, et al. 2011. Antimicrobial polypeptides from Pseudoplectania nigrella. US Patent 7,972,814.
  28. Schwarz S, Kehrenberg C, Walsh T R. 2 001. U se o f antimicrobial agents in veterinary medicine and food animal production. Int. J. Antimicrob. Agents 17: 431-437.
    CrossRef
  29. Singh M P, Greenstein M. 2 000. A ntibacterial l eads from microbial natural products discovery. Curr. Opin. Drug Discov. Devel. 3: 167-176.
    Pubmed
  30. Smith DL, Harris AD, Johnson JA, Silbergeld EK, Morris JG. 2002. Animal antibiotic use has an early but important impact on the emergence of antibiotic resistance in human commensal bacteria. Proc. Natl. Acad. Sci. USA 99: 6434-6439.
    Pubmed KoreaMed CrossRef
  31. Tanabe H, Ayabe T, Maemoto A, Ishikawa C, Inaba Y, Sato R, et al. 2007. Denatured human α-defensin attenuates the bactericidal activity and the stability against enzymatic digestion. Biochem. Biophys. Res. Commun. 358: 349-355.
    Pubmed CrossRef
  32. Tian ZG, Dong TT, Yang YL, Teng D, Wang JH. 2009. Expression of antimicrobial peptide LH multimers in Escherichia coli C43 (DE3). Appl. Microbiol. Biotechnol. 83: 143149.
    Pubmed CrossRef
  33. Tian ZG, Teng D, Yang YL, Luo J, Feng XJ, Fan Y, et al. 2007. Multimerization and fusion expression of bovine lactoferricin derivative LfcinB15-W4,10 in Escherichia coli. Appl. Microbiol. Biotechnol. 75: 117-124.
    Pubmed CrossRef
  34. Vizioli J, Salzet M. 2002. Antimicrobial peptides versus parasitic infections? Trends Parasitol. 18: 475-476.
    CrossRef
  35. Wang XQ, Yang F, Liu C, Zhou HJ, Wu GY, Qiao SY, et al. 2012. Dietary supplementation with the probiotic Lactobacillus fermentum I5007 and the antibiotic aureomycin differentially affects the small intestinal proteomes of weanling piglets. J. Nutr. 142: 7-13.
    Pubmed CrossRef
  36. Yamada O, Sakamoto K, Tominaga M, Nakayama T, Koseki T, Fujita A, Akita O. 2005. Cloning and heterologous expression of the antibiotic peptide (ABP) genes from Rhizopus oligosporus NBRC 8631. Biosci. Biotechnol. Biochem. 69: 477-482.
    Pubmed CrossRef
  37. Zasloff M. 2002. Antimicrobial peptides of multicellular organisms. Nature 415: 389-395.
    Pubmed CrossRef
  38. Zhang J, Yang YL, Teng D, Tian ZG, Wang SR, Wang JH. 2011. Expression of plectasin in Pichia pastoris and its characterization as a new antimicrobial peptide against Staphyloccocus and Streptococcus. Protein Expr. Purif. 78: 189196.
    Pubmed CrossRef