Abstract
Two yeast strains, which have the ability to degrade biodegradable plastic films, were isolated from the larval midgut of a stag beetle, Aegus laevicollis. Both of them are most closely related to Cryptococcus magnus and could degrade biodegradable plastic (BP) films made of poly(butylene succinate) (PBS) and poly(butylene succinate-co-adipate) (PBSA) effectively. A BP-degrading enzyme was purified from the culture broth of one of the isolated strains employing a newly developed affinity purification method based on the binding action of the enzyme to the substrate (emulsified PBSA) and its subsequent degradative action toward the substrate. Partial amino acid sequences of this enzyme suggested that it belongs to the cutinase family, and thus, the enzyme was named CmCut1. It has a molecular mass of 21 kDa and a degradative activity for emulsified PBSA which was significantly enhanced by the simultaneous presence of Ca2+ or Mg2+ at a concentration of about 2.5 mM. Its optimal pH was 7.5, and the optimal temperature was 40 °C. It showed a broad substrate specificity for p-nitrophenyl (pNP)-fatty acid esters ranging from pNP-acetate (C2) to pNP-stearate (C18) and films of PBSA, PBS, poly(ε-caprolactone), and poly(lactic acid).
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Acero EH, Ribitsch D, Steinkellner G, Gruber K, Greimel K, Eiteljoerg I, Trotscha E, Wei R, Zimmermann W, Zinn M, Cavaco-Paulo A, Freddi G, Schwab H, Guebitz G (2011) Enzymatic surface hydrolysis of PET: Effect of structural diversity on kinetic properties of cutinases from Thermobifida. Macromolecules 44(12):4632–4640. doi:10.1021/Ma200949p
Araya K (1993) Relationship between the decay types of dead wood and occurrence of lucanid beetles (Coleoptera: Lucanidae). Appl Entomol Zool 28(1):27–33
Breznak JA, Brune A (1994) Role of microorganisms in the digestion of lignocellulose by termites. Annu Rev Entomol 39:453–487. doi:10.1146/annurev.en.39.010194.002321
Chatterjee S, Roy B, Roy D, Banerjee R (2010) Enzyme-mediated biodegradation of heat treated commercial polyethylene by Staphylococcal species. Polym Degrad Stab 95(2):195–200. doi:10.1016/j.polymdegradstab.2009.11.025
Douglas AE (1998) Nutritional interactions in insect-microbial symbioses: Aphid and their symbiotic bacteria Buchnera. Annu Rev Entomol 43:17–37. doi:10.1146/annurev.ento.43.1.17
Elbanna K, Lütke-Eversloh T, Jendrossek D, Luftmann H, Steinbüchel A (2004) Studies on the biodegradability of polythioester copolymers and homopolymers by polyhydroxyalkanoate (PHP)-degrading bacteria and PHA depolymerases. Arch Microbiol 182:212–225. doi:10.1007/s00203-004-0715-z
Fell JW, Boekhout T, Fonseca A, Scorzetti G, Statzell-Tallman A (2000) Biodiversity and systematics of basidiomycetous yeasts as determined by large-subunit rDNA D1/D2 domain sequence analysis. Int J Syst Evol Microbiol 50:1351–1371. doi:10.1099/00207713-50-3-1351
Fonseca Á, Boekhout T, Fell JW (2011) Cryptococcus Vuillemin (1901). In: Kurtzman CP, Fell JW, Boekhout T (eds) The yeasts: a taxonomic study, 5th edn. Elsevier, New York, pp 1661–1737
Gross RA, Kalra B (2002) Biodegradable polymers for the environment. Science 297(5582):803–807. doi:10.1126/science.297.5582.803
Iefuji H, Iimura Y, Obata T (1994) Isolation and characterization of a yeast Cryptococcus sp. S-2 that produces raw starchdigesting alpha-amylase, xylanase, and polygalacturonase. Biosci Biotechnol Biochem 58(12):2261–2262. doi:10.1271/bbb.58.2261
Iyer S, Shah R, Sharma A, Jendrossek D, Desai A (2000) Purification of Aspergillus fumigatus (Pdf1) poly(β-hydroxybutyrate) (PHB) depolymerase using a new, single-step substrate affinity chromatography method: characterization of the PHB depolymerase exhibiting novel self-aggregation behavior. J Polym Environ 8(4):197–203. doi:10.1023/A:1015249811314
Kamini NR, Fujii T, Kurosu T, Iefuji H (2000) Production, purification and characterization of an extracellular lipase from the yeast, Cryptococcus sp S-2. Process Biochem 36(4):317–324. doi:10.1016/S0032-9592%2800%2900228-4
Kasuya K, Ishii N, Inoue Y, Yazawa K, Tagaya T, Yotsumoto T, Kazahaya J, Nagai D (2009) Characterization of a mesophilic aliphatic-aromatic copolyester-degrading fungus. Polym Degrad Stab 94(8):1190–1196. doi:10.1016/j.polymdegradstab.2009.04.013
Kitamoto HK, Shinozaki Y, Cao XH, Morita T, Konishi M, Tago K, Kajiwara H, Koitabashi M, Yoshida S, Watanabe T, Sameshima-Yamashita Y, Nakajima-Kambe T, Tsushima S (2011) Phyllosphere yeasts rapidly break down biodegradable plastics. AMB Express 1(1):44. doi:10.1186/2191-0855-1-44
Kodama Y, Masaki K, Kondo H, Suzuki M, Tsuda S, Nagura T, Shimba N, Suzuki E, Iefuji H (2009) Crystal structure and enhanced activity of a cutinase-like enzyme from Cryptococcus sp. strain S-2. Proteins 77(3):710–717. doi:10.1002/prot.22484
Koitabashi M, Noguchi MT, Sameshima-Yamashita Y, Hiradate S, Suzuki K, Yoshida S, Watanabe T, Shinozaki Y, Tsushima S, Kitamoto HK (2012) Degradation of biodegradable plastic mulch films in soil environment by phylloplane fungi isolated from gramineous plants. AMB Express 2:40. doi:10.1186/2191-0855-2-40
Kwon-Chung KJ (2011) Filobasidium Olive (1968). In: Kurtzman CP, Fell JW, Boekhout T (eds) The yeasts: a taxonomic study, 5th edn. Elsevier, New York, pp 1457–1465
Kyrikou I, Briassoulis D (2007) Biodegradation of agricultural plastic films: a critical review. J Polym Environ 15(3):227–227. doi:10.1007/s10924-007-0063-6
Lange K-B (2011) Bioplastics to pass the one million tonne mark in 2011. Press Release, European Bioplastics, Berlin
Maeda H, Yamagata Y, Abe K, Hasegawa F, Machida M, Ishioka R, Gomi K, Nakajima T (2005) Purification and characterization of a biodegradable plastic-degrading enzyme from Aspergillus oryzae. Appl Microbiol Biotechnol 67(6):778–788. doi:10.1007/s00253-004-1853-6
Masaki K, Kamini NR, Ikeda H, Iefuji H (2005) Cutinase-like enzyme from the yeast Cryptococcus sp. strain S-2 hydrolyzes polylactic acid and other biodegradable plastics. Appl Environ Microbiol 71(11):7548–7550. doi:10.1128/AEM.71.11.7548-7550.2005
Masaki K, Tsuchioka H, Hirano T, Kato M, Ikeda H, Iefuji H (2012) Construction of a new recombinant protein expression system in the basidiomycetous yeast Cryptococcus sp. strain S-2 and enhancement of the production of a cutinase-like enzyme. Appl Microbiol Biotechnol 93(4):1627–1636. doi:10.1007/s00253-011-3680-x
Papaneophytou CP, Pantazaki AA, Kyriakidis DA (2009) An extracellular polyhydroxybutyrate depolymerase in Thermus thermophiles HB8. Appl Microbiol Biotechnol 83:659–668. doi:10.1007/s00253-008-1842-2
Papaneophytou CP, Pantazaki AA (2011) A novel affinity chromatographic material for the purification of extracellular polyhydroxybutyrate depolymerases. J Polym Environ 19:876–886. doi:10.1007/s10924-011-0345-x
Park DS, Oh HW, Jeong WJ, Kim H, Park HY, Bae KS (2007) A culture-based study of the bacterial communities within the guts of nine longicorn beetle species and their exo-enzyme producing properties for degrading xylan and pectin. J Microbiol 45(5):394–401
Ronkvist AM, Lu WH, Feder D, Gross RA (2009) Cutinase-catalyzed deacetylation of poly(vinyl acetate). Macromolecules 42(16):6086–6097. doi:10.1021/Ma900530j
Shevchenko A, Wilm M, Vorm O, Mann M (1996) Mass spectrometric sequencing of proteins silver-stained polyacrylamide gels. Anal Chem 68(5):850–858. doi:10.1021/ac950914h
Shinozaki Y, Morita T, Cao XH, Yoshida S, Koitabashi M, Watanabe T, Suzuki K, Sameshima-Yamashita Y, Nakajima-Kambe T, Fujii T, Kitamoto HK (2012a) Biodegradable plastic-degrading enzyme from Pseudozyma antarctica: cloning, sequencing, and characterization. Appl Microbiol Biotechnol. doi:10.1007/s00253-012-4188-8
Shinozaki Y, Watanabe T, Nakajima-Kambe T, Kitamoto HK (2012b) Rapid and simple colorimetric assay for detecting the enzymatic degradation of biodegradable plastic films. J Biosci Bioeng. doi:10.1016/j.jbiosc.2012.08.010
Suh SO, McHugh JV, Pollock DD, Blackwell M (2005) The beetle gut: a hyperdiverse source of novel yeasts. Mycol Res 109(Pt 3):261–265. doi:10.1017/S0953756205002388
Tanahashi M, Kubota K, Matsushita N, Togashi K (2010) Discovery of mycangia and the associated xylose-fermenting yeasts in stag beetles (Coleoptera: Lucanidae). Naturwissenschaften 97:311–317. doi:10.1007/s00114-009-0643-5
Uchida H, Nakajima-Kambe T, Shigeno-Akutsu Y, Nomura N, Tokiwa Y, Nakahara T (2000) Properties of a bacterium which degrades solid poly(tetramethylene succinate)-co-adipate, a biodegradable plastic. FEMS Microbiol Lett 189(1):25–29. doi:10.1111/j.1574-6968.2000.tb09201.x
Vega FE, Dowd PF (2005) The role of yeasts as insect endosymbionts. In: Vega FE, Blackwell M (eds) Insect–fungal associations: ecology and evolution. Oxford University Press, New York, pp 211–243
Acknowledgments
We thank Showa Denko K. K. for generously supplying the Bionolle materials (PBS and PBSA) and Ms. C. Xiao-Hong for the technical assistance. We also acknowledge Dr. H. Iefuji and Dr. T. Nakajima-Kambe as well as Dr. E. Suto for the valuable comments in this research. This research was financially supported by the Ministry of the Environment, KAKENHI (235658083), and the National Institute for Agro-Environmental Sciences, Japan.
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Suzuki, K., Sakamoto, H., Shinozaki, Y. et al. Affinity purification and characterization of a biodegradable plastic-degrading enzyme from a yeast isolated from the larval midgut of a stag beetle, Aegus laevicollis . Appl Microbiol Biotechnol 97, 7679–7688 (2013). https://doi.org/10.1007/s00253-012-4595-x
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DOI: https://doi.org/10.1007/s00253-012-4595-x