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Tomotake Morita - One of the best experts on this subject based on the ideXlab platform.
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Disruption of protease A and B orthologous genes in the basidiomycetous yeast Pseudozyma antarctica GB-4(0) yields a stable extracellular Biodegradable Plastic-degrading enzyme
PloS one, 2021Co-Authors: Natsuki Omae, Yuka Sameshima-yamashita, Hiroko Kitamoto, Hideaki Koike, Kazunori Ushimaru, Tomotake MoritaAbstract:The yeast Pseudozyma antarctica (currently designated Moesziomyces antarcticus) secretes a xylose-induced Biodegradable Plastic-degrading enzyme (PaE). To suppress degradation of PaE during production and storage, we targeted the inhibition of proteolytic enzyme activity in P. antarctica. Proteases A and B act as upper regulators in the proteolytic network of the model yeast, Saccharomyces cerevisiae. We searched for orthologous genes encoding proteases A and B in the genome of P. antarctica GB-4(0) based on the predicted amino acid sequences. We found two gene candidates, PaPRO1 and PaPRO2, with conserved catalytically important domains and signal peptides indicative of vacuolar protease function. We then prepared gene-deletion mutants of strain GB-4(0), ΔPaPRO1 and ΔPaPRO2, and evaluated PaE stability in culture by immunoblotting analysis. Both mutants exhibited sufficient production of PaE without degradation fragments, while the parent strain exhibited the degradation fragments. Therefore, we concluded that the protease A and B orthologous genes are related to the degradation of PaE. To produce a large quantity of PaE, we made a PaPRO2 deletion mutant of a PaE-overexpression strain named XG8 by introducing a PaE high-production cassette into the strain GB-4(0). The ΔPaPRO2 mutant of XG8 was able to produce PaE without the degradation fragments during large-scale cultivation in a 3-L jar fermenter for 3 days at 30°C. After terminating the agitation, the PaE activity in the XG8 ΔPaPRO2 mutant culture was maintained for the subsequent 48 h incubation at 25°C regardless of remaining cells, while activity in the XG8 control was reduced to 55.1%. The gene-deleted mutants will be useful for the development of industrial processes of PaE production and storage.
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deficiency of Biodegradable Plastic degrading enzyme production in a gene deletion mutant of phyllosphere yeast pseudozyma antarctica defective in mannosylerythritol lipid biosynthesis
AMB Express, 2019Co-Authors: Azusa Saika, Hideaki Koike, Hiroko Kitamoto, Tohru Yarimizu, Tomotake MoritaAbstract:The basidiomycetous yeast Pseudozyma antarctica (currently designated Moesziomyces antarcticus) produces extracellular enzymes and glycolipids, including mannosylerythritol lipids (MELs), which are biosurfactants. Strain GB-4(0) of this species was previously isolated from rice husks and produces Biodegradable Plastic-degrading enzyme (Pseudozyma antarctica esterase; PaE). In this study, we generated a MEL biosynthesis-deficient strain (∆PaEMT1) by deleting the gene PaEMT1, which is essential to MEL biosynthesis in strain GB-4(0). The resulting ∆PaEMT1 strain showed deficient PaE activity, and the corresponding signal was hardly detected in its culture supernatant through western blotting analysis using rabbit anti-PaE serum. On the other hand, the relative expression of the gene PaCLE1, encoding PaE, was identical between GB-4(0) and ∆PaEMT1 based on quantitative real-time PCR. When strain ∆PaEMT1 was grown in culture media supplemented with various surfactants, i.e., Tween20, BRIJ35 and TritonX-100, and MELs, PaE activity and secretion recovered. We also attempted to detect intracellular PaE using cell-free extract, but observed no signal in the soluble or insoluble fractions of ∆PaEMT1. This result suggested that the PaCLE1 gene was not translated to PaE, or that expressed PaE was degraded immediately in ∆PaEMT1. Based on these results, MEL biosynthesis is an important contributor to PaE production.
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Degradation profiles of Biodegradable Plastic films by Biodegradable Plastic-degrading enzymes from the yeast Pseudozyma antarctica and the fungus Paraphoma sp. B47-9
Polymer Degradation and Stability, 2017Co-Authors: Shun Sato, Yukiko Shinozaki, Tomotake Morita, Yuka Sameshima-yamashita, Azusa Saika, Ken Suzuki, Tokuma Fukuoka, Hiroshi Habe, Hiroko KitamotoAbstract:Esterases from the yeast Pseudozyma antarctica (PaE) and the fungus Paraphoma sp. B47-9 (PCLE) can degrade Biodegradable Plastics (Shinozaki et al., 2013; Suzuki et al., 2014). The degradation profiles of Plastic films composed of poly(butylene succinate), poly(butylene succinate-co-adipate), or poly(butylene adipate) by these enzymes were characterized by liquid chromatography-mass spectroscopy in terms of the molecular structures and molecular weights of the degradation products. Monomers and oligomers with molecular weights corresponding to dimers to octamers were identified as products of degradation by PaE in an aqueous reaction solution, irrespective of the type of Biodegradable Plastic film. Size-exclusion chromatography indicated that the number-average molecular weight of degraded films decreased with reaction time, suggesting that PaE degraded polyester films randomly into monomer units (endo-type degradation). PCLE also degraded polyester films randomly into monomer units, albeit more slowly than did PaE.
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Biodegradable Plastic-degrading enzyme from Pseudozyma antarctica: Cloning, sequencing, and characterization
Applied Microbiology and Biotechnology, 2013Co-Authors: Yukiko Shinozaki, Tomotake Morita, Yuka Sameshima-yamashita, Toshiaki Nakajima-kambe, Ken Suzuki, Motoo Koitabashi, Shigenobu Yoshida, Xiao-hong Cao, Takeshi FujiiAbstract:Pseudozyma antarctica JCM 10317 exhibits a strong degradation activity for Biodegradable Plastics (BPs) such as agricultural mulch films composed of poly(butylene succinate) (PBS) and poly(butylene succinate-co-adipate) (PBSA). An enzyme named PaE was isolated and the gene encoding PaE was cloned from the strain by functional complementation in Saccharomyces cerevisiae. The deduced amino acid sequence of PaE contains 198 amino acids with a predicted molecular weight of 20,362.41. High identity was observed between this sequence and that of cutinase-like enzymes (CLEs) (61-68 %); therefore, the gene encoding PaE was named PaCLE1. The specific activity of PaE against emulsified PBSA was 54.8 +/- 6.3 U/mg. In addition to emulsified BPs, PaE degraded solid films of PBS, PBSA, poly(epsilon-caprolactone), and poly(lactic acid).
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Phyllosphere yeasts rapidly break down Biodegradable Plastics
AMB Express, 2011Co-Authors: Hiroko K. Kitamoto, Yukiko Shinozaki, Tomotake Morita, Hideyuki Kajiwara, Masaaki Konishi, Motoo Koitabashi, Kazutami Tago, Xiao-hong Cao, Shigenobu YoshidaAbstract:The use of Biodegradable Plastics can reduce the accumulation of environmentally persistent Plastic wastes. The rate of degradation of Biodegradable Plastics depends on environmental conditions and is highly variable. Techniques for achieving more consistent degradation are needed. However, only a few microorganisms involved in the degradation process have been isolated so far from the environment. Here, we show that Pseudozyma spp. yeasts, which are common in the phyllosphere and are easily isolated from plant surfaces, displayed strong degradation activity on films made from poly-butylene succinate or poly-butylene succinate-co-adipate. Strains of P. antarctica isolated from leaves and husks of paddy rice displayed strong degradation activity on these films at 30°C. The type strain, P. antarctica JCM 10317, and Pseudozyma spp. strains from phyllosphere secreted a Biodegradable Plastic-degrading enzyme with a molecular mass of about 22 kDa. Reliable source of Biodegradable Plastic-degrading microorganisms are now in our hands.
Tina Treude - One of the best experts on this subject based on the ideXlab platform.
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microbial colonization and degradation of polyethylene and Biodegradable Plastic bags in temperate fine grained organic rich marine sediments
Marine Pollution Bulletin, 2016Co-Authors: Alice Nauendorf, Nikolaus K. Bigalke, Elena V Gorb, Martin Wahl, Matthias Haeckel, Stanislav N Gorb, Stefan Krause, Tina TreudeAbstract:Highlights • Polypropylene and Biodegradable Plastic bags were incubated in marine sediments. • Bacterial colonization was highest on Biodegradable Plastic bags. • None of the two bag types showed signs of degradation after 98 days. • Marine sediments probably represent a long-term sink for both types of litter. Abstract To date, the longevity of Plastic litter at the sea floor is poorly constrained. The present study compares colonization and biodegradation of Plastic bags by aerobic and anaerobic benthic microbes in temperate fine-grained organic-rich marine sediments. Samples of polyethylene and Biodegradable Plastic carrier bags were incubated in natural oxic and anoxic sediments from Eckernforde Bay (Western Baltic Sea) for 98 days. Analyses included (1) microbial colonization rates on the bags, (2) examination of the surface structure, wettability, and chemistry, and (3) mass loss of the samples during incubation. On average, Biodegradable Plastic bags were colonized five times higher by aerobic and eight times higher by anaerobic microbes than polyethylene bags. Both types of bags showed no sign of biodegradation during this study. Therefore, marine sediment in temperate coastal zones may represent a long-term sink for Plastic litter and also supposedly compostable material.
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Microbial colonization and degradation of polyethylene and Biodegradable Plastic bags in temperate fine-grained organic-rich marine sediments
Marine Pollution Bulletin, 2016Co-Authors: Alice Nauendorf, Nikolaus K. Bigalke, Elena V Gorb, Martin Wahl, Matthias Haeckel, Stanislav N Gorb, Stefan Krause, Tina TreudeAbstract:To date, the longevity of Plastic litter at the sea floor is poorly constrained. The present study compares colonization and biodegradation of Plastic bags by aerobic and anaerobic benthic microbes in temperate fine-grained organic-rich marine sediments. Samples of polyethylene and Biodegradable Plastic carrier bags were incubated in natural oxic and anoxic sediments from Eckernförde Bay (Western Baltic Sea) for 98 days. Analyses included (1) microbial colonization rates on the bags, (2) examination of the surface structure, wettability, and chemistry, and (3) mass loss of the samples during incubation. On average, Biodegradable Plastic bags were colonized five times higher by aerobic and eight times higher by anaerobic microbes than polyethylene bags. Both types of bags showed no sign of biodegradation during this study. Therefore, marine sediment in temperate coastal zones may represent a long-term sink for Plastic litter and also supposedly compostable material.
Yukiko Shinozaki - One of the best experts on this subject based on the ideXlab platform.
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Degradation profiles of Biodegradable Plastic films by Biodegradable Plastic-degrading enzymes from the yeast Pseudozyma antarctica and the fungus Paraphoma sp. B47-9
Polymer Degradation and Stability, 2017Co-Authors: Shun Sato, Yukiko Shinozaki, Tomotake Morita, Yuka Sameshima-yamashita, Azusa Saika, Ken Suzuki, Tokuma Fukuoka, Hiroshi Habe, Hiroko KitamotoAbstract:Esterases from the yeast Pseudozyma antarctica (PaE) and the fungus Paraphoma sp. B47-9 (PCLE) can degrade Biodegradable Plastics (Shinozaki et al., 2013; Suzuki et al., 2014). The degradation profiles of Plastic films composed of poly(butylene succinate), poly(butylene succinate-co-adipate), or poly(butylene adipate) by these enzymes were characterized by liquid chromatography-mass spectroscopy in terms of the molecular structures and molecular weights of the degradation products. Monomers and oligomers with molecular weights corresponding to dimers to octamers were identified as products of degradation by PaE in an aqueous reaction solution, irrespective of the type of Biodegradable Plastic film. Size-exclusion chromatography indicated that the number-average molecular weight of degraded films decreased with reaction time, suggesting that PaE degraded polyester films randomly into monomer units (endo-type degradation). PCLE also degraded polyester films randomly into monomer units, albeit more slowly than did PaE.
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production of a Biodegradable Plastic degrading enzyme from cheese whey by the phyllosphere yeast pseudozyma antarctica gb 4 1 w
Journal of Bioscience and Bioengineering, 2014Co-Authors: Takashi Watanabe, Yukiko Shinozaki, Ken Suzuki, Motoo Koitabashi, Shigenobu Yoshida, Yuka Sameshimayamashita, Hiroko K. KitamotoAbstract:Cheese whey is a by-product of cheese production and has high concentrations of lactose (about 5%) and other nutrients. Pseudozyma antarctica produces a unique cutinase-like enzyme, named PaE, that efficiently degrades Biodegradable Plastics. A previous study showed that a combination of 1% oil and 0.5% lactose increased cutinase-like enzyme production by another species of yeast. In this study, to produce PaE from cheese whey, we investigated the effects of soybean oil on PaE production (expressed as Biodegradable Plastic-degrading activity) by P. antarctica growing on lactose or cheese whey. In flask cultures, the final PaE activity was only 0.03 U/ml when soybean oil was used as the sole carbon source, but increased to 1.79 U/ml when a limited amount of soybean oil (under 0.5%) was combined with a relatively high concentration of lactose (6%). Using a 5-L jar fermentor with lactose fed-batch cultivation and periodic soybean oil addition, about 14.6 U/ml of PaE was obtained after 5 days of cultivation. When the lactose was replaced with cheese whey, PaE production was 10.8 U/ml after 3 days of cultivation.
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purification characterization and cloning of the gene for a Biodegradable Plastic degrading enzyme from paraphoma related fungal strain b47 9
Applied Microbiology and Biotechnology, 2014Co-Authors: Ken Suzuki, Yukiko Shinozaki, Motoo Koitabashi, Takeshi Fujii, Shigenobu Yoshida, Yuka Sameshimayamashita, Masako Tsujimoto Noguchi, Hiroko KitamotoAbstract:Paraphoma-related fungal strain B47-9 secreted a Biodegradable Plastic (BP)-degrading enzyme which amounted to 68 % (w/w) of the total secreted proteins in a culture medium containing emulsified poly(butylene succinate-co-adipate) (PBSA) as sole carbon source. The gene for this enzyme was found to be composed of an open reading frame consisting of 681 nucleotides encoding 227 amino acids and two introns. Southern blot analysis showed that this gene exists as a single copy. The deduced amino acid sequence suggested that this enzyme belongs to the cutinase (E.C.3.1.1.74) family; thus, it was named P araphoma-related fungus cutinase-like enzyme (PCLE). It degraded various types of BP films, such as poly(butylene succinate), PBSA, poly(butylene adipate-co-terephthalate), poly(e-caprolactone), and poly(dl-lactic acid). It has a molecular mass of 19.7 kDa, and an optimum pH and temperature for degradation of emulsified PBSA of 7.2 and 45 °C, respectively. Ca2+ ion at a concentration of about 1.0 mM markedly enhanced the degradation of emulsified PBSA.
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Biodegradable Plastic-degrading enzyme from Pseudozyma antarctica: Cloning, sequencing, and characterization
Applied Microbiology and Biotechnology, 2013Co-Authors: Yukiko Shinozaki, Tomotake Morita, Yuka Sameshima-yamashita, Toshiaki Nakajima-kambe, Ken Suzuki, Motoo Koitabashi, Shigenobu Yoshida, Xiao-hong Cao, Takeshi FujiiAbstract:Pseudozyma antarctica JCM 10317 exhibits a strong degradation activity for Biodegradable Plastics (BPs) such as agricultural mulch films composed of poly(butylene succinate) (PBS) and poly(butylene succinate-co-adipate) (PBSA). An enzyme named PaE was isolated and the gene encoding PaE was cloned from the strain by functional complementation in Saccharomyces cerevisiae. The deduced amino acid sequence of PaE contains 198 amino acids with a predicted molecular weight of 20,362.41. High identity was observed between this sequence and that of cutinase-like enzymes (CLEs) (61-68 %); therefore, the gene encoding PaE was named PaCLE1. The specific activity of PaE against emulsified PBSA was 54.8 +/- 6.3 U/mg. In addition to emulsified BPs, PaE degraded solid films of PBS, PBSA, poly(epsilon-caprolactone), and poly(lactic acid).
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Affinity purification and characterization of a Biodegradable Plastic-degrading enzyme from a yeast isolated from the larval midgut of a stag beetle, Aegus laevicollis
Applied microbiology and biotechnology, 2012Co-Authors: Ken Suzuki, Yukiko Shinozaki, Takeshi Fujii, Hironori Sakamoto, Jun Tabata, Atsushi Mochizuki, Seiya Tsushima, Hiroko KitamotoAbstract: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(e-caprolactone), and poly(lactic acid).
Shigenobu Yoshida - One of the best experts on this subject based on the ideXlab platform.
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production of a Biodegradable Plastic degrading enzyme from cheese whey by the phyllosphere yeast pseudozyma antarctica gb 4 1 w
Journal of Bioscience and Bioengineering, 2014Co-Authors: Takashi Watanabe, Yukiko Shinozaki, Ken Suzuki, Motoo Koitabashi, Shigenobu Yoshida, Yuka Sameshimayamashita, Hiroko K. KitamotoAbstract:Cheese whey is a by-product of cheese production and has high concentrations of lactose (about 5%) and other nutrients. Pseudozyma antarctica produces a unique cutinase-like enzyme, named PaE, that efficiently degrades Biodegradable Plastics. A previous study showed that a combination of 1% oil and 0.5% lactose increased cutinase-like enzyme production by another species of yeast. In this study, to produce PaE from cheese whey, we investigated the effects of soybean oil on PaE production (expressed as Biodegradable Plastic-degrading activity) by P. antarctica growing on lactose or cheese whey. In flask cultures, the final PaE activity was only 0.03 U/ml when soybean oil was used as the sole carbon source, but increased to 1.79 U/ml when a limited amount of soybean oil (under 0.5%) was combined with a relatively high concentration of lactose (6%). Using a 5-L jar fermentor with lactose fed-batch cultivation and periodic soybean oil addition, about 14.6 U/ml of PaE was obtained after 5 days of cultivation. When the lactose was replaced with cheese whey, PaE production was 10.8 U/ml after 3 days of cultivation.
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purification characterization and cloning of the gene for a Biodegradable Plastic degrading enzyme from paraphoma related fungal strain b47 9
Applied Microbiology and Biotechnology, 2014Co-Authors: Ken Suzuki, Yukiko Shinozaki, Motoo Koitabashi, Takeshi Fujii, Shigenobu Yoshida, Yuka Sameshimayamashita, Masako Tsujimoto Noguchi, Hiroko KitamotoAbstract:Paraphoma-related fungal strain B47-9 secreted a Biodegradable Plastic (BP)-degrading enzyme which amounted to 68 % (w/w) of the total secreted proteins in a culture medium containing emulsified poly(butylene succinate-co-adipate) (PBSA) as sole carbon source. The gene for this enzyme was found to be composed of an open reading frame consisting of 681 nucleotides encoding 227 amino acids and two introns. Southern blot analysis showed that this gene exists as a single copy. The deduced amino acid sequence suggested that this enzyme belongs to the cutinase (E.C.3.1.1.74) family; thus, it was named P araphoma-related fungus cutinase-like enzyme (PCLE). It degraded various types of BP films, such as poly(butylene succinate), PBSA, poly(butylene adipate-co-terephthalate), poly(e-caprolactone), and poly(dl-lactic acid). It has a molecular mass of 19.7 kDa, and an optimum pH and temperature for degradation of emulsified PBSA of 7.2 and 45 °C, respectively. Ca2+ ion at a concentration of about 1.0 mM markedly enhanced the degradation of emulsified PBSA.
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Biodegradable Plastic-degrading enzyme from Pseudozyma antarctica: Cloning, sequencing, and characterization
Applied Microbiology and Biotechnology, 2013Co-Authors: Yukiko Shinozaki, Tomotake Morita, Yuka Sameshima-yamashita, Toshiaki Nakajima-kambe, Ken Suzuki, Motoo Koitabashi, Shigenobu Yoshida, Xiao-hong Cao, Takeshi FujiiAbstract:Pseudozyma antarctica JCM 10317 exhibits a strong degradation activity for Biodegradable Plastics (BPs) such as agricultural mulch films composed of poly(butylene succinate) (PBS) and poly(butylene succinate-co-adipate) (PBSA). An enzyme named PaE was isolated and the gene encoding PaE was cloned from the strain by functional complementation in Saccharomyces cerevisiae. The deduced amino acid sequence of PaE contains 198 amino acids with a predicted molecular weight of 20,362.41. High identity was observed between this sequence and that of cutinase-like enzymes (CLEs) (61-68 %); therefore, the gene encoding PaE was named PaCLE1. The specific activity of PaE against emulsified PBSA was 54.8 +/- 6.3 U/mg. In addition to emulsified BPs, PaE degraded solid films of PBS, PBSA, poly(epsilon-caprolactone), and poly(lactic acid).
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degradation of Biodegradable Plastic mulch films in soil environment by phylloplane fungi isolated from gramineous plants
AMB Express, 2012Co-Authors: Motoo Koitabashi, Yukiko Shinozaki, Ken Suzuki, Shigenobu Yoshida, Seiya Tsushima, Masako T Noguchi, Yuka Sameshimayamashita, Syuntaro Hiradate, Hiroko K. KitamotoAbstract:To improve the biodegradation of Biodegradable Plastic (BP) mulch films, 1227 fungal strains were isolated from plant surface (phylloplane) and evaluated for BP-degrading ability. Among them, B47-9 a strain isolated from the leaf surface of barley showed the strongest ability to degrade poly-(butylene succinate-co-butylene adipate) (PBSA) and poly-(butylene succinate) (PBS) films. The strain grew on the surface of soil-mounted BP films, produced breaks along the direction of hyphal growth indicated that it secreted a BP-degrading enzyme, and has directly contributing to accelerating the degradation of film. Treatment with the culture filtrate decomposed 91.2 wt%, 23.7 wt%, and 14.6 wt% of PBSA, PBS, and commercially available BP polymer blended mulch film, respectively, on unsterlized soil within 6 days. The PCR-DGGE analysis of the transition of soil microbial community during film degradation revealed that the process was accompanied with drastic changes in the population of soil fungi and Acantamoeba spp., as well as the growth of inoculated strain B47-9. It has a potential for application in the development of an effective method for accelerating degradation of used Plastics under actual field conditions.
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Phyllosphere yeasts rapidly break down Biodegradable Plastics
AMB Express, 2011Co-Authors: Hiroko K. Kitamoto, Yukiko Shinozaki, Tomotake Morita, Hideyuki Kajiwara, Masaaki Konishi, Motoo Koitabashi, Kazutami Tago, Xiao-hong Cao, Shigenobu YoshidaAbstract:The use of Biodegradable Plastics can reduce the accumulation of environmentally persistent Plastic wastes. The rate of degradation of Biodegradable Plastics depends on environmental conditions and is highly variable. Techniques for achieving more consistent degradation are needed. However, only a few microorganisms involved in the degradation process have been isolated so far from the environment. Here, we show that Pseudozyma spp. yeasts, which are common in the phyllosphere and are easily isolated from plant surfaces, displayed strong degradation activity on films made from poly-butylene succinate or poly-butylene succinate-co-adipate. Strains of P. antarctica isolated from leaves and husks of paddy rice displayed strong degradation activity on these films at 30°C. The type strain, P. antarctica JCM 10317, and Pseudozyma spp. strains from phyllosphere secreted a Biodegradable Plastic-degrading enzyme with a molecular mass of about 22 kDa. Reliable source of Biodegradable Plastic-degrading microorganisms are now in our hands.
Hiroko K. Kitamoto - One of the best experts on this subject based on the ideXlab platform.
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production of a Biodegradable Plastic degrading enzyme from cheese whey by the phyllosphere yeast pseudozyma antarctica gb 4 1 w
Journal of Bioscience and Bioengineering, 2014Co-Authors: Takashi Watanabe, Yukiko Shinozaki, Ken Suzuki, Motoo Koitabashi, Shigenobu Yoshida, Yuka Sameshimayamashita, Hiroko K. KitamotoAbstract:Cheese whey is a by-product of cheese production and has high concentrations of lactose (about 5%) and other nutrients. Pseudozyma antarctica produces a unique cutinase-like enzyme, named PaE, that efficiently degrades Biodegradable Plastics. A previous study showed that a combination of 1% oil and 0.5% lactose increased cutinase-like enzyme production by another species of yeast. In this study, to produce PaE from cheese whey, we investigated the effects of soybean oil on PaE production (expressed as Biodegradable Plastic-degrading activity) by P. antarctica growing on lactose or cheese whey. In flask cultures, the final PaE activity was only 0.03 U/ml when soybean oil was used as the sole carbon source, but increased to 1.79 U/ml when a limited amount of soybean oil (under 0.5%) was combined with a relatively high concentration of lactose (6%). Using a 5-L jar fermentor with lactose fed-batch cultivation and periodic soybean oil addition, about 14.6 U/ml of PaE was obtained after 5 days of cultivation. When the lactose was replaced with cheese whey, PaE production was 10.8 U/ml after 3 days of cultivation.
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degradation of Biodegradable Plastic mulch films in soil environment by phylloplane fungi isolated from gramineous plants
AMB Express, 2012Co-Authors: Motoo Koitabashi, Yukiko Shinozaki, Ken Suzuki, Shigenobu Yoshida, Seiya Tsushima, Masako T Noguchi, Yuka Sameshimayamashita, Syuntaro Hiradate, Hiroko K. KitamotoAbstract:To improve the biodegradation of Biodegradable Plastic (BP) mulch films, 1227 fungal strains were isolated from plant surface (phylloplane) and evaluated for BP-degrading ability. Among them, B47-9 a strain isolated from the leaf surface of barley showed the strongest ability to degrade poly-(butylene succinate-co-butylene adipate) (PBSA) and poly-(butylene succinate) (PBS) films. The strain grew on the surface of soil-mounted BP films, produced breaks along the direction of hyphal growth indicated that it secreted a BP-degrading enzyme, and has directly contributing to accelerating the degradation of film. Treatment with the culture filtrate decomposed 91.2 wt%, 23.7 wt%, and 14.6 wt% of PBSA, PBS, and commercially available BP polymer blended mulch film, respectively, on unsterlized soil within 6 days. The PCR-DGGE analysis of the transition of soil microbial community during film degradation revealed that the process was accompanied with drastic changes in the population of soil fungi and Acantamoeba spp., as well as the growth of inoculated strain B47-9. It has a potential for application in the development of an effective method for accelerating degradation of used Plastics under actual field conditions.
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Phyllosphere yeasts rapidly break down Biodegradable Plastics
AMB Express, 2011Co-Authors: Hiroko K. Kitamoto, Yukiko Shinozaki, Tomotake Morita, Hideyuki Kajiwara, Masaaki Konishi, Motoo Koitabashi, Kazutami Tago, Xiao-hong Cao, Shigenobu YoshidaAbstract:The use of Biodegradable Plastics can reduce the accumulation of environmentally persistent Plastic wastes. The rate of degradation of Biodegradable Plastics depends on environmental conditions and is highly variable. Techniques for achieving more consistent degradation are needed. However, only a few microorganisms involved in the degradation process have been isolated so far from the environment. Here, we show that Pseudozyma spp. yeasts, which are common in the phyllosphere and are easily isolated from plant surfaces, displayed strong degradation activity on films made from poly-butylene succinate or poly-butylene succinate-co-adipate. Strains of P. antarctica isolated from leaves and husks of paddy rice displayed strong degradation activity on these films at 30°C. The type strain, P. antarctica JCM 10317, and Pseudozyma spp. strains from phyllosphere secreted a Biodegradable Plastic-degrading enzyme with a molecular mass of about 22 kDa. Reliable source of Biodegradable Plastic-degrading microorganisms are now in our hands.