Studies on Two Exoenzymes Which Lyse Wine-Spoiling Bacteria

Patrick Sebastian, Harald Claus, Helmut König

Institute for Microbiology and Wine Research, Johannes Gutenberg University of Mainz, Mainz, Germany.

DOI: 10.4236/aim.2014.49059                

Abstract

Microorganisms play an important role in the conversion of grape juice into wine. Different species of yeast are mainly responsible for the production of ethanol. Lactic acid bacteria also occur regularly in must or wine. They are mostly undesirable due to their capacity to produce wine-spoiling compounds. Especially strains of Lactobacillus brevis are able to produce biogenic amines as well as precursors of ethyl carbamate and different off-flavours (N-heterocycles, volatile phenols). By excessive formation of acetic acid some lactobacilli may even induce slow/stuck grape juice fermentations. In conventional winemaking, additions of sulphite or lysozyme are used to inhibit the growth of spoilage microorganisms. There is a strong interest in finding alternatives, because of the reduced activity of lysozyme in the wine milieu, a limited growth-inhibiting activity against lactic acid bacteria, and some health risks described regarding the application of sulphite. We found that a culture supernatant of Streptomyces albidoflavus B 578 lysed all bacteria previously isolated from must and wine samples (Acetobacter sp., Lactobacillus sp., Leuconostoc sp., Oenococcus oeni, Pediococcus sp., Staphylococcus sp.) including 35 strains of L. brevis. Two bacteriolytic exoenzymes were isolated and characterized from the streptomycete: a muramidase (24 kDa) and a protease (17 kDa). Both hydrolyzed cell wall components of L. brevis (peptidoglycan, S-layer proteins) and were active under wine-relevant conditions.

Keywords

Streptomyces albidoflavus, Lactobacillus brevis, Wine Spoilage, Stuck Fermentations, Cell Wall, Muramidase, Protease, S-Layer

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1]	Konig, H., Unden, G. and Frohlich, J. (2009) Biology of Microorganisms on Grapes, in Must and in Wine. Springer-Verlag, Heidelberg.
[2]	Bartowsky, E.J. (2009) Bacterial Spoilage of Wine and Approaches to Minimize It. Letters in Applied Microbiology, 48, 149-156. http://dx.doi.org/10.1111/j.1472-765X.2008.02505.x
[3]	Rodas, A.M., Ferrer, S. and Pardo, I. (2005) Polyphasic Study of Wine Lactobacillus Strains: Taxonomic Implications. International Journal of Systematic and Evolutionary Microbiology, 55, 197-207. http://dx.doi.org/10.1099/ijs.0.63249-0
[4]	Dicks, L.M.T. and Endo, A. (2009) Taxonomic Status of Lactic Acid Bacteria in Wine and Key Characteristics to Differentiate Species. South African Journal of Enology and Viticulture, 30, 72-90.
[5]	Sebastian, P., Herr, P., Fischer, U. and Konig, H. (2011) Molecular Identification of Lactic Bacteria Occurring in Must and Wine. South African Journal of Enology and Viticulture, 32, 300-309.
[6]	Araque, I., Bordons, A. and Reguant, C. (2013) Effect of Ethanol and Low pH on Citrulline and Ornithine Excretion and arc Gene Expression by Strains of Lactobacillus brevis and Pediococcus pentosaceus. Food Microbiology, 33, 107-113. http://dx.doi.org/10.1016/j.fm.2012.09.005
[7]	Petri, A., Pfannebecker, J., Frohlich, J. and Konig, H. (2013) Fast Identification of Wine Related Lactic Acid Bacteria by Multiplex PCR. Food Microbiology, 33, 48-54. http://dx.doi.org/10.1016/j.fm.2012.08.011
[8]	Couto, J.A., Campos, F.M., Figueiredo, A.R. and Hogg, T.A. (2006) Ability of Lactic Acid Bacteria to Produce Volatile Phenols. American Journal of Enology and Viticulture, 57, 166-171.
[9]	Costello, P.J., Lee, T.H. and Henschke, P.A. (2001) Ability of Lactic Acid Bacteria to Produce N-Heterocycles Causing Mausy Off-Flavour in Wine. Australian Journal of Grape and Wine Research, 7, 160-167. http://dx.doi.org/10.1111/j.1755-0238.2001.tb00205.x
[10]	Ribéreau-Gayon, P., Glories, Y., Maujean, A. and Dubourdieu, D. (2006) Handbook of Enology Vol. 2—The Chemistry of Wine Stabilization and Treatments. 2nd Edition, John Wiley & Sons, Ltd., Hoboken. http://dx.doi.org/10.1002/0470010398
[11]	Lin, S.C. and Georgiou, G. (2005) A Biocatalyst for the Removal of Sulfite from Alcoholic Beverages. Biotechnology and Bioengineering, 89, 123-127. http://dx.doi.org/10.1002/bit.20307
[12]	Logardt, I.M. and Neujahr, H.Y. (1975) Lysis of Modified Walls of Lactobacillus fermentum. Journal of Bacteriology, 124, 73-77.
[13]	Pérez-Calderón, R., Gonzalo-Garijo, M.A., Lamilla-Yerga, A., Mangas-Santos, R. and Morena-Gastón, I. (2007) Recurrent Angiodema Due to Lysozyme Allergy. Journal of Investigational Allergology and Clinical Immunology, 17, 264-266.
[14]	Blattel, V., Wirth, K., Claus, H., Schlott, B., Pfeiffer, P. and Konig, H. (2009) A Lytic Cocktail from Streptomyces sp. B578 for the Control of Lactic and Acetic Acid Bacterial in Wine. Applied Microbiology and Biotechnology, 83, 839-848. http://dx.doi.org/10.1007/s00253-009-1926-7
[15]	De Man, J.C., Rogosa, M. and Shape, M.R. (1960) A Medium for the Cultivation of Lactobacilli. Journal of Applied Bacteriology, 23, 130-135. http://dx.doi.org/10.1111/j.1365-2672.1960.tb00188.x
[16]	Hopwood, D.A., Bibb, M.J., Chater, K.F., Kieder, T., Bruton, C.J., Kieser, H.M., Lydiate, D.J., Smith, C.P., Ward, J.M. and Schrempf, H. (1985) Genetic Manipulation of Streptomycetes—A Laboratory Manual. The John Innes Foundation, Norwich.
[17]	Ohbuchi, K., Hasegawa, K., Hamachi, M., Ozeki, K. and Kumagai, C. (2001) Isolation of a New Lytic Enzyme for Hiochi Bacteria and Other Lactic Acid Bacteria. Journal of Bioscience and Bioengineering, 91, 487-492.
[18]	Laemmli, U.K. (1970) Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4. Nature, 227, 680-685. http://dx.doi.org/10.1038/227680a0
[19]	Claus, H., Tenzer, S., Sobe, M., Schlander, M., Konig, H. and Frohlich, J. (2014) Effect of Carboxymethyl Cellulose on Tartrate Salt, Protein and Colour Stability of Red Wine. Australian Journal of Grape and Wine Research, 20, 186-193. http://dx.doi.org/10.1111/ajgw.12070
[20]	Schleifer, K.H. and Kandler, O. (1972) Peptidoglycan Types of Bacterial Cell Walls and Their Taxonomic Implications. Bacteriological Reviews, 36, 407-477.
[21]	Miller, G.L. (1959) Use of Dinitrosalicylic Acid Reagent for Determination of Reducing Sugar. Analytical Chemistry, 31, 426-428. http://dx.doi.org/10.1021/ac60147a030
[22]	Dohm, N., Petri, A., Schlander, M., Schlott, B., Konig, H. and Claus, H. (2011) Molecular and Biochemical Properties of the S-Layer Protein from the Wine Bacterium Lactobacillus hilgardii B706. Archives of Microbiology, 193, 251-261. http://dx.doi.org/10.1007/s00203-010-0670-9
[23]	Gifford, A., Klippenstein, J. and Moore, M. (2002) Serum Stimulates Growth of and Proteinase Secretion by Aspergillus fuminatus. Infection and Immunity, 70, 19-26. http://dx.doi.org/10.1128/IAI.70.1.19-26.2002
[24]	Vollmer, W., Joris, B., Charlier, P. and Foster, S. (2008) Bacterial Peptidoglycan (Murein) Hydrolases. FEMS Micro-biological Reviews, 32, 259-286. http://dx.doi.org/10.1111/j.1574-6976.2007.00099.x
[25]	Delfini, C., Cersosimo, M., Del Prete, V., Strano, M., Gaetano, G., Pagliara, A. and Ambró, S. (2004) Resistance Screening Assay of Wine Lactic Acid Bacteria on Lysozyme: Efficacy of Lysozyme in Unclarified Musts. Journal of Agricultural and Food Chemistry, 52, 1861-1866. http://dx.doi.org/10.1021/jf034824m
[26]	Wirth, K., Sebastian, P., Schneider, I. and Konig, H. (2011) Vorkommen von Essigsaurebakterien in Jungweinen: Isolierung neuer Arten. Der Deutsche Weinbau, 6, 36-37.
[27]	Yokogawa, K., Kawata, S. and Yoshimura, Y. (1973) Studies on Lytic Enzyme against Cariogenic Streptococci. II. Lytic Enzyme from Streptomyces globisporus 1829 Strain. Agricultural and Biological Chemistry, 37, 799-808. http://dx.doi.org/10.1271/bbb1961.37.799
[28]	Salazar, O. and Asenjo, J.A. (2007) Enzymatic Lysis of Microbial Cells. Biotechnological Letters, 29, 984-994. http://dx.doi.org/10.1007/s10529-007-9345-2
[29]	Callewaert, L., Walmagh, M., Michiels, C.W. and Livigne, R. (2011) Food Applications of Bacterial Cell-Wall Hydrolases. Current Opinions in Biotechnology, 22, 164-171. http://dx.doi.org/10.1016/j.copbio.2010.10.012
[30]	Szweda, P., Schielmann, M., Kotlowski, R., Gorczyca, G., Zalewska, M. and Milewski, S. (2012) Peptidoglycan Hydrolases—Potential Weapons against Staphylococcus aureus. Applied Microbiology and Biotechnology, 96, 1157-1174. http://dx.doi.org/10.1007/s00253-012-4484-3
[31]	Rau, A., Hogg, T., Marquardt, R. and Hilgenfeld, R. (2001) A New Lysozyme Fold. Crystal Structure of the Muramidase from Streptomyces coelicolor at 1.65 A. Journal of Biological Chemistry, 276, 31994-31999. http://dx.doi.org/10.1074/jbc.M102591200
[32]	Seo, H.J., Shimonishi, T., Ohmiya, K. and Hayashi, K. (2001) Characterization of N-acetylmuramidase M-1 of Streptomyces globisporus Produced by Escherichia coli BL21 (DE3) pLysS. Journal of Bioscience and Bioengineering, 92, 472-474.
[33]	Yokogawa, K., Kawata, S. and Yoshimura, Y. (1976) Purification and Properties of a Lytic Enzyme from Streptomyces griseus H-402. Agricultural and Biological Chemistry, 40, 661-667. http://dx.doi.org/10.1271/bbb1961.40.661
[34]	Hayashi, K., Kasumi, T., Kubo, N. and Tsumura, N. (1981) Purification and Characterization of the Lytic Enzyme Produced by Streptomyces rutgersensis H-46. Agricultural and Biological Chemistry, 45, 2289-2300. http://dx.doi.org/10.1271/bbb1961.45.2289
[35]	Clarke, A. and Dupont, C. (1992) O-Acetylated Peptidoglycan: Its Occurrence, Pathobiological Significance and Biosynthesis. Canadian Journal of Microbiology, 38, 85-91. http://dx.doi.org/10.1139/m92-014
[36]	Delcour, J., Ferain, T., Deghorain, M., Palumbo, E. and Hols, P. (1999) The Biosynthesis and Functionality of the Cell-Wall of Lactic Bacteria. Antonie van Leeuwenhoek, 76, 159-184. http://dx.doi.org/10.1023/A:1002089722581
[37]	Seo, H.J., Kitaoka, M., Ohmiya, K. and Hayashi, K. (2003) Substrate Specificity of the N,6-O-Diacetylmuramidase from Streptomyces globisporus. Journal of Bioscience and Bioengineering, 95, 313-316. http://dx.doi.org/10.1016/S1389-1723(03)80037-3
[38]	Sidhu, S.S., Kalmar, G.B., Willis, L.G. and Borgford, T.J. (1994) Streptomyces griseus Protease C. Journal of Biological Chemistry, 269, 20167-20171.
[39]	Wahlby, S. and Engstrom, L. (1968) Studies on Streptomyces griseus Protease: II. The Amino Acid Sequence around the Reactive Serine Residue of DFP-Sensitive Components with Esterase Activity. Biochimica et Biophysica Acta, 151, 402-408. http://dx.doi.org/10.1016/0005-2744(68)90107-1
[40]	Lee, T.W. and James, M.N.G. (2008) 1.2Angstrom-Resolution Crystal Structures Reveal the Second Tetrahedral Intermediates of Streptogrisin B (SGPB). Biochimica et Biophysica Acta (BBA)-Proteins and Proteomics, 1784, 319-334. http://dx.doi.org/10.1016/j.bbapap.2007.11.012
[41]	Thorne, K.J., Oliver, R.C. and Barrett, A.J. (1976) Lysis and Killing of Bacteria by Lysosomal Proteinases. Infection and Immunity, 14, 555-563.
[42]	Claus, H., Akca, E., Debaerdemaeker, T., Evrard, C., Declecq, J.P., Harris, J.R., Schlott, B. and Konig, H. (2005) Molecular Organization of Selected Prokaryotic S-Layer Proteins. Canadian Journal of Microbiology, 51, 731-743. http://dx.doi.org/10.1139/w05-093
[43]	Hynonen, U. and Palva, A. (2013) Lactobacillus Surface Layer Proteins: Structure, Function and Applications. Applied Microbiology and Biotechnology, 97, 5225-5243. http://dx.doi.org/10.1007/s00253-013-4962-2
[44]	Waters, E.J. and Colby, C.B. (2009) Proteins. In: Morena-Arribas, M.V. and Polo, M.C., Eds., Wine Chemistry and Biochemistry, Springer Science & Business Media, New York, 213-230. http://dx.doi.org/10.1007/978-0-387-74118-5_11
[45]	Edwards, C.G., Reynolds, A.G., Rodriguez, A.V., Semon, M.J. and Mills, J.M. (1999) Implication of Acetic Acid in the Induction of Slow/Stuck Grape Juice Fermentations and Inhibition of Yeast by Lactobacillus sp. American Journal of Enology and Viticulture, 50, 204-210.