Skip to main content
SpringerLink
Log in

Lysozyme inhibitor conferring bacterial tolerance to invertebrate type lysozyme

  • Research Article
  • Published:
Cellular and Molecular Life Sciences Aims and scope Submit manuscript

Abstract

Invertebrate (I-) type lysozymes, like all other known lysozymes, are dedicated to the hydrolysis of peptidoglycan, the major bacterial cell wall polymer, thereby contributing to the innate immune system and/or digestive system of invertebrate organisms. Bacteria on the other hand have developed several protective strategies against lysozymes, including the production of periplasmic and/or membrane-bound lysozyme inhibitors. The latter have until now only been described for chicken (C-) type lysozymes. We here report the discovery, purification, identification and characterization of the first bacterial specific I-type lysozyme inhibitor from Aeromonas hydrophila, which we designate PliI (periplasmic lysozyme inhibitor of the I-type lysozyme). PliI has homologs in several proteobacterial genera and contributes to I-type lysozyme tolerance in A. hydrophila in the presence of an outer membrane permeabilizer. These and previous findings on C-type lysozyme inhibitors suggest that bacterial lysozyme inhibitors may have an important function, for example, in bacteria-host interactions.

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

Similar content being viewed by others

References

  1. Jollès J, Jollès P (1975) The lysozyme from Asterias rubens. Eur J Biochem 54:19–23

    Article  PubMed  Google Scholar 

  2. Ito Y, Yoshikawa A, Hotani T, Fukuda S, Sugimura K, Imoto T (1999) Amino acid sequences of lysozymes newly purified from invertebrates imply wide distribution of a novel class in the lysozyme family. Eur J Biochem 259:456–461

    Article  CAS  PubMed  Google Scholar 

  3. Goto T, Abe Y, Kakuta Y, Takeshita K, Imoto T, Ueda T (2007) Crystal structure of Tapes japonica lysozyme with substrate analogue—structural basis of the catalytic mechanism and manifestation of its chitinase activity accompanied by quaternary structural change. J Biol Chem 282:27459–27467

    Article  CAS  PubMed  Google Scholar 

  4. Bachali S, Jager M, Hassanin A, Schoentgen F, Jolles P, Fiala-Medioni A, Deutsch JS (2002) Phylogenetic analysis of invertebrate lysozymes and the evolution of lysozyme function. J Mol Evol 54:652–664

    Article  CAS  PubMed  Google Scholar 

  5. Nilsen IW, Myrnes B (2001) The gene of chlamysin, a marine invertebrate-type lysozyme, is organized similar to vertebrate but different from invertebrate chicken-type lysozyme genes. Gene 269:27–32

    Article  CAS  PubMed  Google Scholar 

  6. Cloud-Hansen KA, Peterson SB, Stabb EV, Goldman WE, McFall-Ngai MJ, Handelsman J (2006) Breaching the great wall: peptidoglycan and microbial interactions. Nat Rev Microbiol 4:710–716

    Article  CAS  PubMed  Google Scholar 

  7. Dziarski R (2003) Recognition of bacterial peptidoglycan by the innate immune system. Cell Mol Life Sci 60:1793–1804

    Article  CAS  PubMed  Google Scholar 

  8. Park JW, Kim CH, Kim JH, Je BR, Roh KB, Kim SJ, Lee HH, Ryu JH, Lim JH, Oh BH, Lee WJ, Ha NC, Lee BL (2007) Clustering of peptidoglycan recognition protein-SA is required for sensing lysine-type peptidoglycan in insects. Proc Natl Acad Sci USA 104:6602–6607

    Article  CAS  PubMed  Google Scholar 

  9. Monchois V, Abergel C, Sturgis J, Jeudy S, Claverie J (2001) Escherichia coli ykfE ORFan gene encodes a potent inhibitor of C-type lysozyme. J Biol Chem 276:18437–18441

    Article  CAS  PubMed  Google Scholar 

  10. Callewaert L, Aertsen A, Deckers D, Vanoirbeek KGA, Vanderkelen L, Van Herreweghe JM, Masschalck B, Nakimbugwe D, Robben J, Michiels CW (2008) A new family of lysozyme inhibitors contributing to lysozyme tolerance in Gram-negative bacteria. PLoS Pathog 4(3):e1000019

    Google Scholar 

  11. Deckers D, Masschalck B, Aertsen A, Callewaert L, Van Tiggelen CGM, Atanassova M, Michiels CW (2004) Periplasmic lysozyme inhibitor contributes to lysozyme resistance in Escherichia coli. Cell Mol Life Sci 61:1229–1237

    Article  CAS  PubMed  Google Scholar 

  12. Callewaert L, Vanoirbeek K, Lurquin I, Michiels C, Aertsen A (2009) The Rcs two-component system regulates expression of lysozyme inhibitors and is induced by exposure to lysozyme. J Bacteriol 191:1979–1981

    Article  CAS  PubMed  Google Scholar 

  13. Daigle F, Graham JE, Curtiss R (2001) Identification of Salmonella typhi genes expressed within macrophages by selective capture of transcribed sequences (SCOTS). Mol Microbiol 41:1211–1222

    Article  CAS  PubMed  Google Scholar 

  14. Deckers D, Vanlint D, Callewaert L, Aertsen A, Michiels CW (2008) Role of the lysozyme inhibitor Ivy in growth or survival of Escherichia coli and Pseudomonas aeruginosa bacteria in hen egg white and in human saliva and breast milk. Appl Environ Microbiol 74:4434–4439

    Article  CAS  PubMed  Google Scholar 

  15. Abergel C, Monchois V, Byrne D, Chenivesse S, Lembo F, Lazzaroni JC, Claverie JM (2007) Structure and evolution of the Ivy protein family, unexpected lysozyme inhibitors in Gram-negative bacteria. Proc Natl Acad Sci USA 104:6394–6399

    Article  CAS  PubMed  Google Scholar 

  16. Yum S, Kim MJ, Xu Y, Jin XL, Yoo HY, Park JW, Gong JH, Choe KM, Lee BL, Ha NC (2009) Structural basis for the recognition of lysozyme by MliC, a periplasmic lysozyme inhibitor in Gram-negative bacteria. Biochem Biophys Res Commun 378:244–248

    Article  CAS  PubMed  Google Scholar 

  17. Takeshita K, Hashimoto Y, Thujihata Y, So T, Ueda T, Iomoto T (2004) Determination of the complete cDNA sequence, construction of expression systems, and elucidation of fibrinolytic activity for Tapes japonica lysozyme. Protein Expr Purif 36:254–262

    Article  CAS  PubMed  Google Scholar 

  18. Callewaert L, Masschalck B, Deckers D, Nakimbugwe D, Atanassova M, Aertsen A, Michiels CW (2005) Purification of Ivy, a lysozyme inhibitor from Escherichia coli, and characterisation of its specificity for various lysozymes. Enzyme Microb Technol 37:205–211

    Article  CAS  Google Scholar 

  19. Shevchenko A, Tomas H, Havlis J, Olsen JV, Mann M (2006) In-gel digestion for mass spectrometric characterization of proteins and proteomes. Nat Protoc 1:2856–2860

    Article  CAS  PubMed  Google Scholar 

  20. Dumont D, Noben JP, Raus J, Stinissen P, Robben J (2004) Proteomic analysis of cerebrospinal fluid from multiple sclerosis patients. Proteomics 4:2117–2124

    Article  CAS  PubMed  Google Scholar 

  21. Chain PS, Carniel E, Larimer FW, Lamerdin J, Stoutland PO, Regala WM, Georgescu AM, Vergez LM, Land ML, Motin VL, Brubaker RR, Fowler J, Hinnebusch J, Marceau M, Medigue C, Simonet M, Chenal-Francisque V, Souza B, Dacheux D, Elliott JM, Derbise A, Hauser LJ, Garcia E (2004) Insights into the evolution of Yersinia pestis through whole-genome comparison with Yersinia pseudotuberculosis. Proc Natl Acad Sci USA 101:13826–13831

    Article  CAS  PubMed  Google Scholar 

  22. Kyomuhendo P, Myrnes B, Nilsen I (2007) A cold-active salmon goose-type lysozyme with high heat tolerance. Cell Mol Life Sci 64:2841–2847

    Article  CAS  PubMed  Google Scholar 

  23. Erova T, Pillai L, Fadl A, Sha J, Wang S, Galindo C, Chopra A (2006) DNA adenine methyltransferase influences the virulence of Aeromonas hydrophila. Infect Immun 74:410–424

    Article  CAS  PubMed  Google Scholar 

  24. Seshadri R, Joseph SW, Chopra AK, Sha J, Shaw J, Graf J, Haft D, Wu M, Ren Q, Rosovitz MJ, Madupu R, Tallon L, Kim M, Jin S, Vuong H, Stine OC, Ali A, Horneman AJ, Heidelberg JF (2006) Genome sequence of Aeromonas hydrophila ATCC 7966T: jack of all trades. J Bacteriol 188:8272–8282

    Article  CAS  PubMed  Google Scholar 

  25. Bendtsen JD, Nielsen H, von Heijne G, Brunak S (2004) Improved prediction of signal peptides: SignalP 3.0. J Mol Biol 340:783–795

    Article  PubMed  Google Scholar 

  26. Rice P, Longden I, Bleasby A (2000) EMBOSS: the European molecular biology open software suite. Trends Genet 16:276–277

    Article  CAS  PubMed  Google Scholar 

  27. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215:403–410

    CAS  PubMed  Google Scholar 

  28. Bulet P, Stöcklin R, Menin L (2004) Anti-microbial peptides: from invertebrates to vertebrates. Immunol Rev 198:169–184

    Article  CAS  PubMed  Google Scholar 

  29. Gonzalez M, Gueguen Y, Destoumieux-Garzón D, Romestand B, Fievet J, Pugnière M, Roquet F, Escoubas JM, Vandenbulcke F, Levy O, Sauné L, Bulet P, Bachère E (2007) Evidence of a bactericidal permeability increasing protein in an invertebrate, the Crassostrea gigas Cg-BPI. Proc Natl Acad Sci USA 104:17759–17764

    Article  CAS  PubMed  Google Scholar 

  30. Ottaviani D, Santarelli S, Bacchiocchi S, Masini L, Ghittino C, Bacchiocchi I (2006) Occurrence and characterization of Aeromonas spp. in mussels from the Adriatic Sea. Food Microbiol 23:418–422

    Article  CAS  PubMed  Google Scholar 

  31. Evangelista-Barreto N, Vieira R, Carvalho F, Torres R, Sant’Anna E, Rodrigues D, Reis C (2006) Aeromonas spp. isolated from oysters (Crassostrea rhizophorea) from a natural oyster bed, Ceará, Brazil. Rev Inst Med Trop Sao Paulo 48:129–133

    PubMed  Google Scholar 

  32. Ristori C, Iaria S, Gelli D, Rivera I (2007) Pathogenic bacteria associated with oysters (Crassostrea brasiliana) and estuarine water along the south coast of Brazil. Int J Environ Health Res 17:259–269

    Article  CAS  PubMed  Google Scholar 

  33. Tsai G, Chen T (1996) Incidence and toxigenicity of Aeromonas hydrophila in seafood. Int J Food Microbiol 31:121–131

    Article  CAS  PubMed  Google Scholar 

  34. Miñana-Galbis D, Farfán M, Fusté M, Lorén J (2007) Aeromonas bivalvium sp. nov., isolated from bivalve molluscs. Int J Syst Evol Microbiol 57:582–587

    Article  PubMed  Google Scholar 

  35. Miñana-Galbis D, Farfán M, Fusté M, Lorén J (2004) Aeromonas molluscorum sp. nov., isolated from bivalve molluscs. Int J Syst Evol Microbiol 54:2073–2078

    Article  PubMed  Google Scholar 

  36. Graf J, Kikuchi Y, Rio R (2006) Leeches and their microbiota: naturally simple symbiosis models. Trends Microbiol 14:365–371

    Article  CAS  PubMed  Google Scholar 

  37. Zavalova LL, Baskova IP, Lukyanov SA, Sass AV, Snezhkov EV, Akopov SB, Artamonova II, Archipova VS, Nesmeyanov VA, Kozlov DG, Benevolensky SV, Kiseleva VI, Poverenny AM, Sverdlov ED (2000) Destabilase from the medicinal leech is a representative of a novel family of lysozymes. Biochim Biophys Acta Protein Struct Mol Enzymol 1478:69–77

    Article  CAS  Google Scholar 

  38. Kikuchi Y, Meng X, Fukatsu T (2005) Gut symbiotic bacteria of the genus Burkholderia in the broad-headed bugs Riptortus clavatus and Leptocorisa chinensis (Heteroptera: Alydidae). Appl Environ Microbiol 71:4035–4043

    Article  CAS  PubMed  Google Scholar 

  39. Kikuchi Y, Hosokawa T, Fukatsu T (2007) Insect-microbe mutualism without vertical transmission: a stinkbug acquires a beneficial gut symbiont from the environment every generation. Appl Environ Microbiol 73:4308–4316

    Article  CAS  PubMed  Google Scholar 

  40. Hinnebusch BJ (2005) The evolution of flea-borne transmission in Yersinia pestis. Current Issues in Molecular Biology 7:197–212

    CAS  PubMed  Google Scholar 

  41. Tokuda G, Yamada A, Nakano K, Arita NO, Yamasaki H (2008) Colonization of Sulfurovum sp on the gill surfaces of Alvinocaris longirostris, a deep-sea hydrothermal vent shrimp. Mar Ecol Evol Perspect 29:106–114

    CAS  Google Scholar 

  42. Frith MC, Saunders NFW, Kobe B, Bailey TL (2008) Discovering sequence motifs with arbitrary insertions and deletions. PLoS Comput Biol 4(4):e1000071

    Google Scholar 

  43. Liu M, Zhang SC, Liu ZH, Li HY, Xu AL (2006) Characterization, organization and expression of AmphiLysC, an acidic c-type lysozyme gene in amphioxus Branchiostoma belcheri tsingtauense. Gene 367:110–117

    Article  CAS  PubMed  Google Scholar 

  44. Simon R, Priefer U, Puhler A (1983) Broad host range mobilization system for in vivo genetic engineering: transposon mutagenesis in Gram negative bacteria. Biotechnology 1:784–791

    Article  CAS  Google Scholar 

  45. Dombrecht B, Vanderleyden J, Michiels J (2001) Stable RK2-derived cloning vectors for the analysis of gene expression and gene function in Gram-negative bacteria. Mol Plant Microbe Interact 14:426–430

    Article  CAS  PubMed  Google Scholar 

  46. Philippe N, Alcaraz JP, Coursange E, Geiselmann J, Schneider D (2004) Improvement of pCVD442, a suicide plasmid for gene allele exchange in bacteria. Plasmid 51:246–255

    Article  CAS  PubMed  Google Scholar 

  47. Datsenko KA, Wanner BL (2000) One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci USA 97:6640–6645

    Article  CAS  PubMed  Google Scholar 

  48. Crooks G, Hon G, Chandonia J, Brenner S (2004) WebLogo: a sequence logo generator. Genome Res 14:1188–1190

    Article  CAS  PubMed  Google Scholar 

  49. Babu MM, Sankaran K (2002) DOLOP-database of bacterial lipoproteins. Bioinformatics 18:641–643

    Article  CAS  Google Scholar 

  50. Babu MM, Priya ML, Selvan AT, Madera M, Gough J, Aravind L, Sankaran K (2006) A database of bacterial lipoproteins (DOLOP) with functional assignments to predicted lipoproteins. J Bacteriol 188:2761–2773

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

J.M.V.H. holds a doctoral, A.A. and L.C. a postdoctoral fellowship from the Research Foundation-Flanders (F.W.O.-Vlaanderen). L.V. was supported by a doctoral fellowship from the Flemish Institute for the Promotion of Scientific Technological Research (IWT). This work was further financially supported by Research Grants from FWO-Vlaanderen (G.0308.05 and G.0363.08) and by the Research Fund K.U.Leuven (research project GOA/03/10). The authors wish to thank: T. Iomoto (Sojo University, Ikeda Kumamoto, Japan) for Pichia pastoris YJT46, Elisabeth Carniel (Insitut Pasteur, Paris Cedex, France) for Yersinia pseudotuberculosis IP32953, Inge W. Nilsen (NOFIMA, Tromsø, Norway) for Salmon goose type lysozyme (SalG), Dominique Schneider (Université Joseph FOURIER, Saint Martin d’Heres, France) for plasmid pDS132, Joerg Graf (Department of Molecular and Cellular Biology, University of Connecticut, USA) for Aeromonas veronii biovar Sobria 391 Hm 21, and J. Robben (Department of Chemistry, K.U. Leuven) for the mass spectrometry analysis.

Author information

Authors and Affiliations

  1. Laboratory of Food Microbiology, Leuven Food Science and Nutrition Research Centre (LFoRCe), Katholieke Universiteit Leuven, Kasteelpark Arenberg 22, 3001, Leuven, Belgium

    J. M. Van Herreweghe, L. Vanderkelen, L. Callewaert, A. Aertsen & C. W. Michiels

  2. Laboratory for Pharmaceutical Biology, Katholieke Universiteit Leuven, O&N II Herestraat 49, 3000, Leuven, Belgium

    G. Compernolle & P. J. Declerck

Authors
  1. J. M. Van Herreweghe
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. L. Vanderkelen
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. L. Callewaert
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. A. Aertsen
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. G. Compernolle
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. P. J. Declerck
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. C. W. Michiels
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to C. W. Michiels.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Van Herreweghe, J.M., Vanderkelen, L., Callewaert, L. et al. Lysozyme inhibitor conferring bacterial tolerance to invertebrate type lysozyme. Cell. Mol. Life Sci. 67, 1177–1188 (2010). https://doi.org/10.1007/s00018-009-0241-x

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00018-009-0241-x

Keywords

Search

Navigation