1887

Abstract

Bacterial cell wall hydrolases are essential for peptidoglycan remodelling in regard to bacterial cell growth and division. In this study, peptidoglycan hydrolases (PGHs) of different strains were investigated. First, the genome sequence of CD034 and NRRL B-30929 was analysed for the presence of PGHs. Of 23 putative PGHs with different predicted hydrolytic specificities, the glycosyl hydrolase family 25 domain-containing homologues GH25B and GH25N from CD034 and NRRL B-30929, respectively, were selected and characterized in detail. Zymogram analysis confirmed hydrolysing activity on bacterial cell walls for both enzymes. Subsequent reversed-phase HPLC and MALDI-TOF MS analysis of the peptidoglycan breakdown products from strains CD034 and NRRL B-30929, and from GG, which served as a reference, revealed that GH25B and GH25N have -acetylmuramidase activity. Both enzymes were identified as cell wall-associated proteins by means of immunofluorescence microscopy and cellular fractionation, as well as by the ability of purified recombinant GH25B and GH25N to bind to cell walls . Moreover, similar secondary structures mainly composed of β-sheets and nearly identical thermal stabilities with values around 49 °C were found for the two -acetylmuramidases by far-UV circular dichroism spectroscopy. The functional and structural data obtained are discussed and compared to related PGHs. In this study, a major -acetylmuramidase from was characterized in detail for the first time.

Funding
This study was supported by the:
  • Austrian Science Fund (Award P24305-B20 and P21954-B20)
  • Austrian Science Fund (Award W1224)
  • Hochschuljubiläumsstiftung der Stadt Wien (Award H-2442/2012)
  • INRA
  • Région Ile de France
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.078162-0
2014-08-01
2024-12-12
Loading full text...

Full text loading...

/deliver/fulltext/micro/160/8/1807.html?itemId=/content/journal/micro/10.1099/mic.0.078162-0&mimeType=html&fmt=ahah

References

  1. Anzengruber J., Pabst M., Neumann L., Sekot G., Heinl S., Grabherr R., Altmann F., Messner P., Schäffer C. ( 2014). Protein O-glucosylation in Lactobacillus buchneri. Glycoconj J 31:117–131 [View Article][PubMed]
    [Google Scholar]
  2. Atrih A., Bacher G., Allmaier G., Williamson M. P., Foster S. J. ( 1999). Analysis of peptidoglycan structure from vegetative cells of Bacillus subtilis 168 and role of PBP 5 in peptidoglycan maturation. J Bacteriol 181:3956–3966[PubMed]
    [Google Scholar]
  3. Barrett J. F., Dolinger D. L., Schramm V. L., Shockman G. D. ( 1984). The mechanism of soluble peptidoglycan hydrolysis by an autolytic muramidase. A processive exodisaccharidase. J Biol Chem 259:11818–11827[PubMed]
    [Google Scholar]
  4. Blättel V., Wirth K., Claus H., Schlott B., Pfeiffer P., König H. ( 2009). A lytic enzyme cocktail from Streptomyces sp. B578 for the control of lactic and acetic acid bacteria in wine. Appl Microbiol Biotechnol 83:839–848 [View Article][PubMed]
    [Google Scholar]
  5. Bustamante N., Rico-Lastres P., García E., García P., Menéndez M., Menéndez M. ( 2012). Thermal stability of Cpl-7 endolysin from the Streptococcus pneumoniae bacteriophage Cp-7; cell wall-targeting of its CW_7 motifs. PLoS ONE 7:e46654 [View Article][PubMed]
    [Google Scholar]
  6. Callewaert L., Walmagh M., Michiels C. W., Lavigne R. ( 2011). Food applications of bacterial cell wall hydrolases. Curr Opin Biotechnol 22:164–171 [View Article][PubMed]
    [Google Scholar]
  7. Claes I. J. J., Schoofs G., Regulski K., Courtin P., Chapot-Chartier M.-P., Rolain T., Hols P., von Ossowski I., Reunanen J. & other authors ( 2012). Genetic and biochemical characterization of the cell wall hydrolase activity of the major secreted protein of Lactobacillus rhamnosus GG. PLoS ONE 7:e31588 [View Article][PubMed]
    [Google Scholar]
  8. Clarke C. A., Scheurwater E. M., Clarke A. J. ( 2010). The vertebrate lysozyme inhibitor Ivy functions to inhibit the activity of lytic transglycosylase. J Biol Chem 285:14843–14847 [View Article][PubMed]
    [Google Scholar]
  9. Courtin P., Miranda G., Guillot A., Wessner F., Mézange C., Domakova E., Kulakauskas S., Chapot-Chartier M.-P. ( 2006). Peptidoglycan structure analysis of Lactococcus lactis reveals the presence of an l,d-carboxypeptidase involved in peptidoglycan maturation. J Bacteriol 188:5293–5298 [View Article][PubMed]
    [Google Scholar]
  10. Danner H., Holzer M., Mayrhuber E., Braun R. ( 2003). Acetic acid increases stability of silage under aerobic conditions. Appl Environ Microbiol 69:562–567 [View Article][PubMed]
    [Google Scholar]
  11. De Man J. C., Rogosa M., Sharpe M. E. ( 1960). A medium for the cultivation of lactobacilli. J Appl Microbiol 23:130–135
    [Google Scholar]
  12. Delcour J., Ferain T., Deghorain M., Palumbo E., Hols P. ( 1999). The biosynthesis and functionality of the cell-wall of lactic acid bacteria. Antonie van Leeuwenhoek 76:159–184 [View Article][PubMed]
    [Google Scholar]
  13. Dezélée P., Bricas E. ( 1970). Structure of the peptidoglycan in Escherichia coli B and Bacillus megaterium KM. Stereospecific synthesis of two meso-diaminopimelic acid peptides with the tetrapeptide subunit of bacterial cell wall peptidoglycan. Biochemistry 9:823–831 [View Article][PubMed]
    [Google Scholar]
  14. Eikmeyer F. G., Köfinger P., Poschenel A., Jünemann S., Zakrzewski M., Heinl S., Mayrhuber E., Grabherr R., Pühler A. & other authors ( 2013). Metagenome analyses reveal the influence of the inoculant Lactobacillus buchneri CD034 on the microbial community involved in grass ensiling. J Biotechnol 167:334–343 [View Article][PubMed]
    [Google Scholar]
  15. García P., Paz González M., García E., García J. L., López R. ( 1999). The molecular characterization of the first autolytic lysozyme of Streptococcus pneumoniae reveals evolutionary mobile domains. Mol Microbiol 33:128–138 [View Article][PubMed]
    [Google Scholar]
  16. Heinl S., Wibberg D., Eikmeyer F., Szczepanowski R., Blom J., Linke B., Goesmann A., Grabherr R., Schwab H. & other authors ( 2012). Insights into the completely annotated genome of Lactobacillus buchneri CD034, a strain isolated from stable grass silage. J Biotechnol 161:153–166 [View Article][PubMed]
    [Google Scholar]
  17. Höltje J.-V. ( 1996a). Molecular interplay of murein synthases and murein hydrolases in Escherichia coli.. Microb Drug Resist 2:99–103 [View Article][PubMed]
    [Google Scholar]
  18. Höltje J.-V. ( 1996b). A hypothetical holoenzyme involved in the replication of the murein sacculus of Escherichia coli.. Microbiology 142:1911–1918 [View Article][PubMed]
    [Google Scholar]
  19. Holzer M., Mayrhuber E., Danner H., Braun R. ( 2003). The role of Lactobacillus buchneri in forage preservation. Trends Biotechnol 21:282–287 [View Article][PubMed]
    [Google Scholar]
  20. Janesch B., Koerdt A., Messner P., Schäffer C. ( 2013). The S-layer homology domain-containing protein SlhA from Paenibacillus alvei CCM 2051(T) is important for swarming and biofilm formation. PLoS ONE 8:e76566 [View Article][PubMed]
    [Google Scholar]
  21. Kawamura T., Shockman G. D. ( 1983). Purification and some properties of the endogenous, autolytic N-acetylmuramoylhydrolase of Streptococcus faecium, a bacterial glycoenzyme. J Biol Chem 258:9514–9521[PubMed]
    [Google Scholar]
  22. Kawata S., Takemura T., Yokogawa K. ( 1983). Characterization of two N-acetylmuramidases from Streptomyces globisporus 1829. Agric Biol Chem 47:1501–1508 [View Article]
    [Google Scholar]
  23. Kelly S. M., Jess T. J., Price N. C. ( 2005). How to study proteins by circular dichroism. Biochim Biophys Acta 1751:119–139 [View Article][PubMed]
    [Google Scholar]
  24. Laemmli U. K. ( 1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685 [View Article][PubMed]
    [Google Scholar]
  25. Layec S., Decaris B., Leblond-Bourget N. ( 2008). Diversity of Firmicutes peptidoglycan hydrolases and specificities of those involved in daughter cell separation. Res Microbiol 159:507–515 [View Article][PubMed]
    [Google Scholar]
  26. Leclerc D., Asselin A. ( 1989). Detection of bacterial cell wall hydrolases after denaturing polyacrylamide gel electrophoresis. Can J Microbiol 35:749–753 [View Article][PubMed]
    [Google Scholar]
  27. Lepeuple A.-S., Van Gemert E., Chapot-Chartier M.-P. ( 1998). Analysis of the bacteriolytic enzymes of the autolytic Lactococcus lactis subsp. cremoris strain AM2 by renaturing polyacrylamide gel electrophoresis: identification of a prophage-encoded enzyme. Appl Environ Microbiol 64:4142–4148[PubMed]
    [Google Scholar]
  28. Liu S., Leathers T. D., Copeland A., Chertkov O., Goodwin L., Mills D. A. ( 2011). Complete genome sequence of Lactobacillus buchneri NRRL B-30929, a novel strain from a commercial ethanol plant. J Bacteriol 193:4019–4020 [View Article][PubMed]
    [Google Scholar]
  29. Martinez-Fleites C., Korczynska J. E., Davies G. J., Cope M. J., Turkenburg J. P., Taylor E. J. ( 2009). The crystal structure of a family GH25 lysozyme from Bacillus anthracis implies a neighboring-group catalytic mechanism with retention of anomeric configuration. Carbohydr Res 344:1753–1757 [View Article][PubMed]
    [Google Scholar]
  30. Möschl A., Schäffer C., Sleytr U. B., Messner P., Christian R., Schulz G. ( 1993). Characterization of the S-layer glycoproteins of two lactobacilli. Advances in Bacterial Paracrystalline Surface Layers vol. 252281–284 Beveridge T. J., Koval S. F. New York: Plenum Press; [View Article]
    [Google Scholar]
  31. Niu S., Luo M., Tang J., Zhou H., Zhang Y., Min X., Cai X., Zhang W., Xu W. & other authors ( 2013). Structural basis of the novel S. pneumoniae virulence factor, GHIP, a glycosyl hydrolase 25 participating in host-cell invasion. PLoS ONE 8:e68647 [View Article][PubMed]
    [Google Scholar]
  32. Oude Elferink S. J. W. H., Krooneman J., Gottschal J. C., Spoelstra S. F., Faber F., Driehuis F. ( 2001). Anaerobic conversion of lactic acid to acetic acid and 1, 2-propanediol by Lactobacillus buchneri. Appl Environ Microbiol 67:125–132 [View Article][PubMed]
    [Google Scholar]
  33. Parisien A., Allain B., Zhang J., Mandeville R., Lan C. Q. ( 2008). Novel alternatives to antibiotics: bacteriophages, bacterial cell wall hydrolases, and antimicrobial peptides. J Appl Microbiol 104:1–13[PubMed]
    [Google Scholar]
  34. Pérez-Dorado I., González A., Morales M., Sanles R., Striker W., Vollmer W., Mobashery S., García J. L., Martínez-Ripoll M. & other authors ( 2010). Insights into pneumococcal fratricide from the crystal structures of the modular killing factor LytC. Nat Struct Mol Biol 17:576–581 [View Article][PubMed]
    [Google Scholar]
  35. Rau A., Hogg T., Marquardt R., Hilgenfeld R. ( 2001). A new lysozyme fold. Crystal structure of the muramidase from Streptomyces coelicolor at 1.65 Å resolution. J Biol Chem 276:31994–31999 [View Article][PubMed]
    [Google Scholar]
  36. Regulski K., Courtin P., Meyrand M., Claes I. J. J., Lebeer S., Vanderleyden J., Hols P., Guillot A., Chapot-Chartier M. P. ( 2012). Analysis of the peptidoglycan hydrolase complement of Lactobacillus casei and characterization of the major γ-d-glutamyl-l-lysyl-endopeptidase. PLoS ONE 7:e32301 [View Article][PubMed]
    [Google Scholar]
  37. Rolain T., Bernard E., Courtin P., Bron P. A., Kleerebezem M., Chapot-Chartier M.-P., Hols P. ( 2012). Identification of key peptidoglycan hydrolases for morphogenesis, autolysis, and peptidoglycan composition of Lactobacillus plantarum WCFS1. Microb Cell Fact 11:11 [View Article][PubMed]
    [Google Scholar]
  38. Rolain T., Bernard E., Beaussart A., Degand H., Courtin P., Egge-Jacobsen W., Bron P. A., Morsomme P., Kleerebezem M. & other authors ( 2013). O-glycosylation as a novel control mechanism of peptidoglycan hydrolase activity. J Biol Chem 288:22233–22247 [View Article][PubMed]
    [Google Scholar]
  39. Schäffer C., Kählig H., Christian R., Schulz G., Zayni S., Messner P. ( 1999). The diacetamidodideoxyuronic-acid-containing glycan chain of Bacillus stearothermophilus NRS 2004/3a represents the secondary cell-wall polymer of wild-type B. stearothermophilus strains. Microbiology 145:1575–1583 [View Article][PubMed]
    [Google Scholar]
  40. Schäffer C., Müller N., Mandal P. K., Christian R., Zayni S., Messner P. ( 2000). A pyrophosphate bridge links the pyruvate-containing secondary cell wall polymer of Paenibacillus alvei CCM 2051 to muramic acid. Glycoconj J 17:681–690 [View Article][PubMed]
    [Google Scholar]
  41. Scheurwater E., Reid C. W., Clarke A. J. ( 2008). Lytic transglycosylases: bacterial space-making autolysins. Int J Biochem Cell Biol 40:586–591 [View Article][PubMed]
    [Google Scholar]
  42. Schleifer K. H., Kandler O. ( 1972). Peptidoglycan types of bacterial cell walls and their taxonomic implications. Bacteriol Rev 36:407–477[PubMed]
    [Google Scholar]
  43. Spath K., Heinl S., Egger E., Grabherr R. ( 2012). Lactobacillus plantarum and Lactobacillus buchneri as expression systems: evaluation of different origins of replication for the design of suitable shuttle vectors. Mol Biotechnol 52:40–48 [View Article][PubMed]
    [Google Scholar]
  44. Typas A., Banzhaf M., Gross C. A., Vollmer W. ( 2012). From the regulation of peptidoglycan synthesis to bacterial growth and morphology. Nat Rev Microbiol 10:123–136[PubMed]
    [Google Scholar]
  45. Vollmer W. ( 2012). Bacterial growth does require peptidoglycan hydrolases. Mol Microbiol 86:1031–1035 [View Article][PubMed]
    [Google Scholar]
  46. Vollmer W., Pilsl H., Hantke K., Höltje J.-V., Braun V. ( 1997). Pesticin displays muramidase activity. J Bacteriol 179:1580–1583[PubMed]
    [Google Scholar]
  47. Vollmer W., Blanot D., de Pedro M. A. ( 2008a). Peptidoglycan structure and architecture. FEMS Microbiol Rev 32:149–167 [View Article][PubMed]
    [Google Scholar]
  48. Vollmer W., Joris B., Charlier P., Foster S. ( 2008b). Bacterial peptidoglycan (murein) hydrolases. FEMS Microbiol Rev 32:259–286 [View Article][PubMed]
    [Google Scholar]
  49. Whisstock J. C., Lesk A. M. ( 1999). SH3 domains in prokaryotes. Trends Biochem Sci 24:132–133 [View Article][PubMed]
    [Google Scholar]
/content/journal/micro/10.1099/mic.0.078162-0
Loading
/content/journal/micro/10.1099/mic.0.078162-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error