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. 1999 Dec 15;344(Pt 3):845–849.

Identification of an anti-mycobacterial domain in NK-lysin and granulysin.

D Andreu 1, C Carreño 1, C Linde 1, H G Boman 1, M Andersson 1
PMCID: PMC1220707  PMID: 10585872

Abstract

NK-lysin and granulysin are homologous cationic anti-bacterial peptides produced by pig and human cytolytic lymphocytes, respectively. The solution structure of NK-lysin comprises five amphipathic alpha-helices. To investigate the properties of a helix-loop-helix region postulated to be a membrane-docking part of NK-lysin, we synthesized 22- and 29-residue peptides reproducing this region for both NK-lysin and granulysin. CD spectroscopy of the synthetic peptides in a liposomal solution showed spectra typical of alpha-helical peptides. The peptides were active against Gram-positive and Gram-negative bacteria, with the two NK-lysin peptides showing higher anti-bacterial activities than the two from granulysin. One NK-lysin peptide was active against Pseudomonas aeruginosa and Staphylococcus aureus, two organisms against which NK-lysin is inactive. Granulysin peptides were inactive against these bacteria, in contrast with granulysin, which is known to be active against them. Both NK-lysin and all synthetic analogues killed Mycobacterium tuberculosis and K562 tumour cells, but did not display haemolytic activity. These results identify a potent anti-mycobacterial domain in NK-lysin and granulysin consisting of a 22-residue (helix 3) sequence plus a disulphide-constrained loop.

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Selected References

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  1. Andersson M., Curstedt T., Jörnvall H., Johansson J. An amphipathic helical motif common to tumourolytic polypeptide NK-lysin and pulmonary surfactant polypeptide SP-B. FEBS Lett. 1995 Apr 10;362(3):328–332. doi: 10.1016/0014-5793(95)00268-e. [DOI] [PubMed] [Google Scholar]
  2. Andersson M., Girard R., Cazenave P. Interaction of NK lysin, a peptide produced by cytolytic lymphocytes, with endotoxin. Infect Immun. 1999 Jan;67(1):201–205. doi: 10.1128/iai.67.1.201-205.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Andersson M., Gunne H., Agerberth B., Boman A., Bergman T., Sillard R., Jörnvall H., Mutt V., Olsson B., Wigzell H. NK-lysin, a novel effector peptide of cytotoxic T and NK cells. Structure and cDNA cloning of the porcine form, induction by interleukin 2, antibacterial and antitumour activity. EMBO J. 1995 Apr 18;14(8):1615–1625. doi: 10.1002/j.1460-2075.1995.tb07150.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Andersson M., Holmgren A., Spyrou G. NK-lysin, a disulfide-containing effector peptide of T-lymphocytes, is reduced and inactivated by human thioredoxin reductase. Implication for a protective mechanism against NK-lysin cytotoxicity. J Biol Chem. 1996 Apr 26;271(17):10116–10120. doi: 10.1074/jbc.271.17.10116. [DOI] [PubMed] [Google Scholar]
  5. Andreu D., Albericio F., Solé N. A., Munson M. C., Ferrer M., Barany G. Formation of disulfide bonds in synthetic peptides and proteins. Methods Mol Biol. 1994;35:91–169. doi: 10.1385/0-89603-273-6:91. [DOI] [PubMed] [Google Scholar]
  6. Andreu D., Rivas L. Animal antimicrobial peptides: an overview. Biopolymers. 1998;47(6):415–433. doi: 10.1002/(SICI)1097-0282(1998)47:6<415::AID-BIP2>3.0.CO;2-D. [DOI] [PubMed] [Google Scholar]
  7. Bellamy W., Takase M., Yamauchi K., Wakabayashi H., Kawase K., Tomita M. Identification of the bactericidal domain of lactoferrin. Biochim Biophys Acta. 1992 May 22;1121(1-2):130–136. doi: 10.1016/0167-4838(92)90346-f. [DOI] [PubMed] [Google Scholar]
  8. Boman H. G. Peptide antibiotics and their role in innate immunity. Annu Rev Immunol. 1995;13:61–92. doi: 10.1146/annurev.iy.13.040195.000425. [DOI] [PubMed] [Google Scholar]
  9. Fahrner R. L., Dieckmann T., Harwig S. S., Lehrer R. I., Eisenberg D., Feigon J. Solution structure of protegrin-1, a broad-spectrum antimicrobial peptide from porcine leukocytes. Chem Biol. 1996 Jul;3(7):543–550. doi: 10.1016/s1074-5521(96)90145-3. [DOI] [PubMed] [Google Scholar]
  10. Greenfield N., Fasman G. D. Computed circular dichroism spectra for the evaluation of protein conformation. Biochemistry. 1969 Oct;8(10):4108–4116. doi: 10.1021/bi00838a031. [DOI] [PubMed] [Google Scholar]
  11. Hill C. P., Yee J., Selsted M. E., Eisenberg D. Crystal structure of defensin HNP-3, an amphiphilic dimer: mechanisms of membrane permeabilization. Science. 1991 Mar 22;251(5000):1481–1485. doi: 10.1126/science.2006422. [DOI] [PubMed] [Google Scholar]
  12. Hultmark D., Engström A., Andersson K., Steiner H., Bennich H., Boman H. G. Insect immunity. Attacins, a family of antibacterial proteins from Hyalophora cecropia. EMBO J. 1983;2(4):571–576. doi: 10.1002/j.1460-2075.1983.tb01465.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Iwanaga S., Muta T., Shigenaga T., Seki N., Kawano K., Katsu T., Kawabata S. Structure-function relationships of tachyplesins and their analogues. Ciba Found Symp. 1994;186:160–175. doi: 10.1002/9780470514658.ch10. [DOI] [PubMed] [Google Scholar]
  14. Johansson J., Gudmundsson G. H., Rottenberg M. E., Berndt K. D., Agerberth B. Conformation-dependent antibacterial activity of the naturally occurring human peptide LL-37. J Biol Chem. 1998 Feb 6;273(6):3718–3724. doi: 10.1074/jbc.273.6.3718. [DOI] [PubMed] [Google Scholar]
  15. Kokryakov V. N., Harwig S. S., Panyutich E. A., Shevchenko A. A., Aleshina G. M., Shamova O. V., Korneva H. A., Lehrer R. I. Protegrins: leukocyte antimicrobial peptides that combine features of corticostatic defensins and tachyplesins. FEBS Lett. 1993 Jul 26;327(2):231–236. doi: 10.1016/0014-5793(93)80175-t. [DOI] [PubMed] [Google Scholar]
  16. Kreil G. Antimicrobial peptides from amphibian skin: an overview. Ciba Found Symp. 1994;186:77–90. doi: 10.1002/9780470514658.ch5. [DOI] [PubMed] [Google Scholar]
  17. Lehrer R. I., Ganz T., Selsted M. E. Defensins: endogenous antibiotic peptides of animal cells. Cell. 1991 Jan 25;64(2):229–230. doi: 10.1016/0092-8674(91)90632-9. [DOI] [PubMed] [Google Scholar]
  18. Liepinsh E., Andersson M., Ruysschaert J. M., Otting G. Saposin fold revealed by the NMR structure of NK-lysin. Nat Struct Biol. 1997 Oct;4(10):793–795. doi: 10.1038/nsb1097-793. [DOI] [PubMed] [Google Scholar]
  19. Middlebrook G., Reggiardo Z., Tigertt W. D. Automatable radiometric detection of growth of Mycobacterium tuberculosis in selective media. Am Rev Respir Dis. 1977 Jun;115(6):1066–1069. doi: 10.1164/arrd.1977.115.6.1066. [DOI] [PubMed] [Google Scholar]
  20. Miyakawa Y., Ratnakar P., Rao A. G., Costello M. L., Mathieu-Costello O., Lehrer R. I., Catanzaro A. In vitro activity of the antimicrobial peptides human and rabbit defensins and porcine leukocyte protegrin against Mycobacterium tuberculosis. Infect Immun. 1996 Mar;64(3):926–932. doi: 10.1128/iai.64.3.926-932.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Morrisett J. D., David J. S., Pownall H. J., Gotto A. M., Jr Interaction of an apolipoprotein (apoLP-alanine) with phosphatidylcholine. Biochemistry. 1973 Mar 27;12(7):1290–1299. doi: 10.1021/bi00731a008. [DOI] [PubMed] [Google Scholar]
  22. Peña S. V., Hanson D. A., Carr B. A., Goralski T. J., Krensky A. M. Processing, subcellular localization, and function of 519 (granulysin), a human late T cell activation molecule with homology to small, lytic, granule proteins. J Immunol. 1997 Mar 15;158(6):2680–2688. [PubMed] [Google Scholar]
  23. Romeo D., Skerlavaj B., Bolognesi M., Gennaro R. Structure and bactericidal activity of an antibiotic dodecapeptide purified from bovine neutrophils. J Biol Chem. 1988 Jul 15;263(20):9573–9575. [PubMed] [Google Scholar]
  24. Ruysschaert J. M., Goormaghtigh E., Homblé F., Andersson M., Liepinsh E., Otting G. Lipid membrane binding of NK-lysin. FEBS Lett. 1998 Mar 27;425(2):341–344. doi: 10.1016/s0014-5793(98)00261-0. [DOI] [PubMed] [Google Scholar]
  25. Stenger S., Hanson D. A., Teitelbaum R., Dewan P., Niazi K. R., Froelich C. J., Ganz T., Thoma-Uszynski S., Melián A., Bogdan C. An antimicrobial activity of cytolytic T cells mediated by granulysin. Science. 1998 Oct 2;282(5386):121–125. doi: 10.1126/science.282.5386.121. [DOI] [PubMed] [Google Scholar]
  26. Thennarasu S., Nagaraj R. Synthetic peptides corresponding to the beta-hairpin loop of rabbit defensin NP-2 show antimicrobial activity. Biochem Biophys Res Commun. 1999 Jan 19;254(2):281–283. doi: 10.1006/bbrc.1998.9933. [DOI] [PubMed] [Google Scholar]
  27. Tschopp J., Hofmann K. Cytotoxic T cells: more weapons for new targets? Trends Microbiol. 1996 Mar;4(3):91–94. doi: 10.1016/0966-842X(96)81522-8. [DOI] [PubMed] [Google Scholar]
  28. Zimmermann G. R., Legault P., Selsted M. E., Pardi A. Solution structure of bovine neutrophil beta-defensin-12: the peptide fold of the beta-defensins is identical to that of the classical defensins. Biochemistry. 1995 Oct 17;34(41):13663–13671. doi: 10.1021/bi00041a048. [DOI] [PubMed] [Google Scholar]

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