Skip to main content
SpringerLink
Log in

Cell specificity and molecular mechanism of antibacterial and antitumor activities of carboxyl-terminal RWL-tagged antimicrobial peptides

  • Original Article
  • Published:
Amino Acids Aims and scope Submit manuscript

Abstract

Antimicrobial peptides (AMPs) constitute a diverse class of naturally occurring or synthetic antimicrobial molecules that have potential for use in the treatment of drug-resistant infections. Several undesirable properties of AMPs, however, may ultimately hinder their development as antimicrobial agents. Thus, new synthetic strategies, including primarily the de novo design of AMPs, urgently need to be developed. In this study, a series of peptides, H-(RWL) n (n = 1, 2, 3, 4, or 5), were designed. H represents GLRPKYS from the C-terminal sequence of AvBD-4. Our results showed that these RWL-tagged peptides can kill not only bacteria but also human hepatocellular carcinoma HepG2 cells. However, the peptide tagged with two repeats of RWL (GW13) showed less affinity to human embryonic lung fibroblast MRC-5 cells or human red blood cells (hRBCs) than HepG2 cells. These results demonstrated that GW13, with high amphiphilicity, exerted great selectivity toward bacteria and cancer cells, sparing host mammalian cells. The mechanism of action against bacteria was elucidated through combined studies of scanning electron microscopy (SEM) and fluorescence assays, showing that the peptide possessed membrane-lytic activities against microbial cells. The fluorescence assays illustrated that GW13 induced apoptosis in HepG2 cells. The cell morphology of HepG2 cells, observed by SEM, further illustrated that GW13 causes cell death by damaging the cell membrane. Our results indicate that GW13 has considerable potential for future development as an antimicrobial and antitumor agent.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9
Fig. 10
Fig. 11

Similar content being viewed by others

Abbreviations

AMP:

Antimicrobial peptide

MIC:

Minimum inhibitory concentration

MHC:

Minimal hemolytic concentration

CD:

Circular dichroism

SDS:

Sodium dodecyl sulfate

PC:

Phosphatidylcholine

PE:

Phosphatidylethanolamine

PG:

Phosphatidylglycerol

MH:

Mueller–Hinton

PI:

Propidium iodide

CCCP:

Carbonyl cyanide m-chlorophenylhydrazone

References

  • Banki K, Hutter E, Gonchoroff NJ, Perl A (1999) Elevation of mitochondrial transmembrane potential and reactive oxygen intermediate levels are early events and occur independently from activation of caspases in Fas signaling. J Immunol 162(3):1466–1479

    CAS  PubMed Central  PubMed  Google Scholar 

  • Bell A (2011) Antimalarial peptides: the long and the short of it. Curr Pharm Des 17(25):2719–2731

    Article  CAS  PubMed  Google Scholar 

  • Bell G, Gouyon PH (2003) Arming the enemy: the evolution of resistance to self-proteins. Microbiology 149(Pt 6):1367–1375

    CAS  PubMed  Google Scholar 

  • Beven L, Castano S, Dufourcq J, Wieslander A, Wroblewski H (2003) The antibiotic activity of cationic linear amphipathic peptides: lessons from the action of leucine/lysine copolymers on bacteria of the class Mollicutes. Eur J Biochem 270(10):2207–2217

    CAS  PubMed  Google Scholar 

  • Burkert U, Allinger NL (1982) Molecular mechanics, vol 177. American Chemical Society, Washington DC

    Google Scholar 

  • Chen Y, Guarnieri MT, Vasil AI, Vasil ML, Mant CT, Hodges RS (2007) Role of peptide hydrophobicity in the mechanism of action of alpha-helical antimicrobial peptides. Antimicrob Agents Chemother 51(4):1398–1406

    CAS  PubMed Central  PubMed  Google Scholar 

  • Chen C, Pan F, Zhang S, Hu J, Cao M, Wang J, Xu H, Zhao X, Lu JR (2010) Antibacterial activities of short designer peptides: a link between propensity for nanostructuring and capacity for membrane destabilization. Biomacromolecules 11(2):402–411

    CAS  PubMed  Google Scholar 

  • Chen C, Hu J, Zhang S, Zhou P, Zhao X, Xu H, Zhao X, Yaseen M, Lu JR (2012) Molecular mechanisms of antibacterial and antitumor actions of designed surfactant-like peptides. Biomaterials 33(2):592–603

    CAS  PubMed  Google Scholar 

  • Cruciani RA, Barker JL, Zasloff M, Chen H-C, Colamonici O (1991) Antibiotic magainins exert cytolytic activity against transformed cell lines through channel formation. Proc Natl Acad Sci USA 88(9):3792–3796

    CAS  PubMed Central  PubMed  Google Scholar 

  • Dathe M, Nikolenko H, Klose J, Bienert M (2004) Cyclization increases the antimicrobial activity and selectivity of arginine- and tryptophan-containing hexapeptides. Biochemistry 43(28):9140–9150

    CAS  PubMed  Google Scholar 

  • Deslouches B, Phadke SM, Lazarevic V, Cascio M, Islam K, Montelaro RC, Mietzner TA (2005) De novo generation of cationic antimicrobial peptides: influence of length and tryptophan substitution on antimicrobial activity. Antimicrob Agents Chemother 49(1):316–322

    CAS  PubMed Central  PubMed  Google Scholar 

  • Dong N, Ma Q–Q, Shan A-S, Lv Y-F, Hu W, Gu Y, Li Y-Z (2012a) Novel design of short antimicrobial peptides derived from the bactericidal domain of avian-defensin-4. Protein Pept Lett 19(11):1212–1219

    CAS  PubMed  Google Scholar 

  • Dong N, Ma Q–Q, Shan A-S, Wang L, Sun W-Y, Li Y-Z (2012b) Influence of truncation of avian-defensin-4 on biological activity and peptide-membrane interaction. Protein Pept Lett 19(4):430–438

    CAS  PubMed  Google Scholar 

  • Dong N, Ma Q, Shan A, Lv Y, Hu W, Gu Y, Li Y (2012c) Strand length-dependent antimicrobial activity and membrane-active mechanism of arginine- and valine-rich beta-hairpin-like antimicrobial peptides. Antimicrob Agents Chemother 56(6):2994–3003

    CAS  PubMed Central  PubMed  Google Scholar 

  • Guillen Schlippe YV, Hartman MC, Josephson K, Szostak JW (2012) In vitro selection of highly modified cyclic peptides that act as tight binding inhibitors. J Am Chem Soc 134(25):10469–10477

    CAS  PubMed Central  Google Scholar 

  • Hancock RE, Lehrer R (1998) Cationic peptides: a new source of antibiotics. Trends Biotechnol 16(2):82–88

    CAS  PubMed  Google Scholar 

  • Hancock RE, Sahl HG (2006) Antimicrobial and host-defense peptides as new anti-infective therapeutic strategies. Nat Biotechnol 24(12):1551–1557

    CAS  PubMed  Google Scholar 

  • Haney EF, Hunter HN, Matsuzaki K, Vogel HJ (2009) Solution NMR studies of amphibian antimicrobial peptides: linking structure to function? Biochim Biophys Acta 1788(8):1639–1655

    CAS  PubMed  Google Scholar 

  • Hoskin DW, Ramamoorthy A (2008) Studies on anticancer activities of antimicrobial peptides. Biochim Biophys Acta 1778(2):357–375

    CAS  PubMed Central  PubMed  Google Scholar 

  • Hu H, Xue J, Swarts BM, Wang Q, Wu Q, Guo Z (2009) Synthesis and antibacterial activities of N-glycosylated derivatives of tyrocidine A, a macrocyclic peptide antibiotic. J Med Chem 52(7):2052–2059

    CAS  PubMed  Google Scholar 

  • Hu J, Chen C, Zhang S, Zhao X, Xu H, Zhao X, Lu JR (2011) Designed antimicrobial and antitumor peptides with high selectivity. Biomacromolecules 12(11):3839–3843

    CAS  PubMed  Google Scholar 

  • Ladokhin AS, White SH (1999) Folding of amphipathic alpha-helices on membranes: energetics of helix formation by melittin. J Mol Biol 285(4):1363–1369

    CAS  PubMed  Google Scholar 

  • Lee KH, Lee DG, Park Y, Kang DI, Shin SY, Hahm KS, Kim Y (2006) Interactions between the plasma membrane and the antimicrobial peptide HP (2-20) and its analogues derived from Helicobacter pylori. Biochem J 394(Pt 1):105–114. doi:10.1042/Bj20051574

    CAS  PubMed Central  PubMed  Google Scholar 

  • Liu Z, Brady A, Young A, Rasimick B, Chen K, Zhou C, Kallenbach NR (2007) Length effects in antimicrobial peptides of the (RW)n series. Antimicrob Agents Chemother 51(2):597–603

    CAS  PubMed Central  PubMed  Google Scholar 

  • Liu Y, Xia X, Xu L, Wang Y (2013) Design of hybrid beta-hairpin peptides with enhanced cell specificity and potent anti-inflammatory activity. Biomaterials 34(1):237–250

    CAS  PubMed  Google Scholar 

  • Niidome T, Matsuyama N, Kunihara M, Hatakeyama T, Aoyagi H (2005) Effect of chain length of cationic model peptides on antibacterial activity. Bull Chem Soc Jpn 78(3):473–476

    CAS  Google Scholar 

  • Rice LB (2003) Do we really need new anti-infective drugs? Curr Opin Pharmacol 3(5):459–463

    CAS  PubMed  Google Scholar 

  • Riedl S, Zweytick D, Lohner K (2011) Membrane-active host defense peptides–challenges and perspectives for the development of novel anticancer drugs. Chem Phys Lipids 164(8):766–781

    CAS  PubMed Central  PubMed  Google Scholar 

  • Rubinchik E, Dugourd D, Algara T, Pasetka C, Friedland HD (2009) Antimicrobial and antifungal activities of a novel cationic antimicrobial peptide, omiganan, in experimental skin colonisation models. Int J Antimicrob Agents 34(5):457–461

    CAS  PubMed  Google Scholar 

  • Sato H, Feix JB (2006) Peptide-membrane interactions and mechanisms of membrane destruction by amphipathic alpha-helical antimicrobial peptides. Biochim Biophys Acta 1758(9):1245–1256

    CAS  PubMed  Google Scholar 

  • Selsted ME, Ouellette AJ (2005) Mammalian defensins in the antimicrobial immune response. Nat Immunol 6(6):551–557

    CAS  PubMed  Google Scholar 

  • Selsted ME, Novotny MJ, Morris WL, Tang YQ, Smith W, Cullor JS (1992) Indolicidin, a novel bactericidal tridecapeptide amide from neutrophils. J Biol Chem 267(7):4292–4295

    CAS  PubMed  Google Scholar 

  • Shafer WM, Hubalek F, Huang M, Pohl J (1996) Bactericidal activity of a synthetic peptide (CG 117-136) of human lysosomal cathepsin G is dependent on arginine content. Infect Immun 64(11):4842–4845

    CAS  PubMed Central  PubMed  Google Scholar 

  • Sitaram N, Nagaraj R (1999) Interaction of antimicrobial peptides with biological and model membranes: structural and charge requirements for activity. Biochim Biophys Acta 1462(1–2):29–54

    CAS  PubMed  Google Scholar 

  • Standley SM, Toft DJ, Cheng H, Soukasene S, Chen J, Raja SM, Band V, Band H, Cryns VL, Stupp SI (2010) Induction of cancer cell death by self-assembling nanostructures incorporating a cytotoxic peptide. Cancer Res 70(8):3020–3026

    CAS  PubMed Central  PubMed  Google Scholar 

  • Strom MB, Haug BE, Skar ML, Stensen W, Stiberg T, Svendsen JS (2003) The pharmacophore of short cationic antibacterial peptides. J Med Chem 46(9):1567–1570

    CAS  PubMed  Google Scholar 

  • Subbalakshmi C, Sitaram N (1998) Mechanism of antimicrobial action of indolicidin. FEMS Microbiol Lett 160(1):91–96

    CAS  PubMed  Google Scholar 

  • Subbalakshmi C, Bikshapathy E, Sitaram N, Nagaraj R (2000) Antibacterial and hemolytic activities of single tryptophan analogs of indolicidin. Biochem Biophys Res Commun 274(3):714–716

    CAS  PubMed  Google Scholar 

  • Tam JP, Lu YA, Yang JL (2002) Antimicrobial dendrimeric peptides. Eur J Biochem 269(3):923–932

    CAS  PubMed  Google Scholar 

  • Wang H, Xu K, Liu L, Tan JP, Chen Y, Li Y, Fan W, Wei Z, Sheng J, Yang Y–Y (2010) The efficacy of self-assembled cationic antimicrobial peptide nanoparticles against Cryptococcus neoformans for the treatment of meningitis. Biomaterials 31(10):2874–2881

    CAS  PubMed  Google Scholar 

  • Wang J, Wong ES, Whitley JC, Li J, Stringer JM, Short KR, Renfree MB, Belov K, Cocks BG (2011) Ancient antimicrobial peptides kill antibiotic-resistant pathogens: australian mammals provide new options. PLoS One 6(8):e24030

    CAS  PubMed Central  PubMed  Google Scholar 

  • Wieprecht T, Apostolov O, Beyermann M, Seelig J (1999) Thermodynamics of the alpha-helix-coil transition of amphipathic peptides in a membrane environment: implications for the peptide-membrane binding equilibrium. J Mol Biol 294(3):785–794

    CAS  PubMed  Google Scholar 

  • Wiradharma N, Khan M, Yong LK, Hauser CAE, Seow SV, Zhang SG, Yang YY (2011a) The effect of thiol functional group incorporation into cationic helical peptides on antimicrobial activities and spectra. Biomaterials 32(34):9100–9108

    CAS  PubMed  Google Scholar 

  • Wiradharma N, Khoe U, Hauser CA, Seow SV, Zhang S, Yang YY (2011b) Synthetic cationic amphiphilic α-helical peptides as antimicrobial agents. Biomaterials 32(8):2204–2212

    CAS  PubMed  Google Scholar 

  • Yang L, Harroun TA, Weiss TM, Ding L, Huang HW (2001) Barrel-stave model or toroidal model? A case study on melittin pores. Biophys J 81(3):1475–1485

    PubMed Central  PubMed  Google Scholar 

  • Yeung AT, Gellatly SL, Hancock RE (2011) Multifunctional cationic host defence peptides and their clinical applications. Cell Mol Life Sci 68(13):2161–2176

    CAS  PubMed  Google Scholar 

  • Yin LM, Edwards MA, Li J, Yip CM, Deber CM (2012) Roles of hydrophobicity and charge distribution of cationic antimicrobial peptides in peptide-membrane interactions. J Biol Chem 287(10):7738–7745

    CAS  PubMed Central  PubMed  Google Scholar 

  • Yoon WH, Park HD, Lim K, Hwang BD (1996) Effect of O-glycosylated mucin on invasion and metastasis of HM7 human colon cancer cells. Biochem Biophys Res Commun 222(3):694–699

    CAS  PubMed  Google Scholar 

  • Zasloff M (2002) Antimicrobial peptides of multicellular organisms. Nature 415(6870):389–395. doi:10.1038/415389a

    CAS  PubMed  Google Scholar 

  • Zelezetsky I, Pag U, Sahl H-G, Tossi A (2005) Tuning the biological properties of amphipathic α-helical antimicrobial peptides: rational use of minimal amino acid substitutions. Peptides 26(12):2368–2376

    CAS  PubMed  Google Scholar 

  • Zheng L, McQuaw CM, Ewing AG, Winograd N (2007) Sphingomyelin/phosphatidylcholine and cholesterol interactions studied by imaging mass spectrometry. J Am Chem Soc 129(51):15730–15731

    CAS  PubMed Central  PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported financially by the National Basic Research Program (2012CB124703), the National Natural Science Foundation of China (31272453), the China Agriculture Research System (CARS-36), the Postgraduate Innovative Research Projects of Heilongjiang Province (YJSCX2012-004HLJ), and the Open Projects of Key Laboratory of Feed Science, College of Heilongjiang Province (yy-2012-03).

Conflict of interest

We have no proprietary, financial, professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the position presented in, or the review of, the manuscript entitled: “Cell specificity and molecular mechanism of antibacterial and antitumor activities of carboxyl-terminal RWL-tagged antimicrobial peptides”.

Author information

Authors and Affiliations

  1. Laboratory of Molecular Nutrition and Immunity, Institute of Animal Nutrition, Northeast Agricultural University, 59 Mucai Street, Harbin, 150030, China

    N. Dong, X. Zhu, Y. F. Lv, Q. Q. Ma & A. S. Shan

  2. State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, 5625 Ren Min Street, Changchun, 130022, China

    J. G. Jiang

Authors
  1. N. Dong
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. X. Zhu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Y. F. Lv
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Q. Q. Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. J. G. Jiang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. A. S. Shan
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to A. S. Shan.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Dong, N., Zhu, X., Lv, Y.F. et al. Cell specificity and molecular mechanism of antibacterial and antitumor activities of carboxyl-terminal RWL-tagged antimicrobial peptides. Amino Acids 46, 2137–2154 (2014). https://doi.org/10.1007/s00726-014-1761-8

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00726-014-1761-8

Keywords

Search

Navigation