Biochemical Characterization of a Psychrophilic Phytase from an Artificially Cultivable Morel Morchella importuna
2
조회
0
다운로드
0
인용
0
참고문헌
키워드
초록
Psychrophilic phytases suitable for aquaculture are rare. In this study, a phytase of the histidine acid phosphatase (HAP) family was identified in Morchella importuna, a psychrophilic mushroom. The phytase showed 38% identity with Aspergillus niger PhyB, which was the closest hit. The M. importuna phytase was overexpressed in Pichia pastoris, purified, and characterized. The phytase had an optimum temperature at 25°C, which is the lowest among all the known phytases to our best knowledge. The optimum pH (6.5) is higher than most of the known HAP phytases, which is fit for the weak acidic condition in fish gut. At the optimum pH and temperature, MiPhyA showed the maximum activity level (2,384.6 ± 90.4 μmol·min-1·mg-1, suggesting that the enzyme possesses a higher activity level over many known phytases at low temperatures. The phytate-degrading efficacy was tested on three common feed materials (soybean meal/rapeseed meal/corn meal) and was compared with the well-known phytases of Escherichia coli and A. niger. When using the same amount of activity units, MiPhyA could yield at least 3× more inorganic phosphate than the two reference phytases. When using the same weight of protein, MiPhyA could yield at least 5× more inorganic phosphate than the other two. Since it could degrade phytate in feed materials efficiently under low temperature and weak acidic conditions, which are common for aquacultural application, MiPhyA might be a promising candidate as a feed additive enzyme.
Abstract
Psychrophilic phytases suitable for aquaculture are rare. In this study, a phytase of the histidine acid phosphatase (HAP) family was identified in Morchella importuna, a psychrophilic mushroom. The phytase showed 38% identity with Aspergillus niger PhyB, which was the closest hit. The M. importuna phytase was overexpressed in Pichia pastoris, purified, and characterized. The phytase had an optimum temperature at 25°C, which is the lowest among all the known phytases to our best knowledge. The optimum pH (6.5) is higher than most of the known HAP phytases, which is fit for the weak acidic condition in fish gut. At the optimum pH and temperature, MiPhyA showed the maximum activity level (2,384.6 ± 90.4 μmol·min-1·mg-1, suggesting that the enzyme possesses a higher activity level over many known phytases at low temperatures. The phytate-degrading efficacy was tested on three common feed materials (soybean meal/rapeseed meal/corn meal) and was compared with the well-known phytases of Escherichia coli and A. niger. When using the same amount of activity units, MiPhyA could yield at least 3× more inorganic phosphate than the two reference phytases. When using the same weight of protein, MiPhyA could yield at least 5× more inorganic phosphate than the other two. Since it could degrade phytate in feed materials efficiently under low temperature and weak acidic conditions, which are common for aquacultural application, MiPhyA might be a promising candidate as a feed additive enzyme.
참고문헌 (49)
- Gulati HK, "Production, purification and characterization of thermostable phytase from thermophilic fungus Thermomyces lanuginosus TL-7", Acta Microbiol. Immunol. Hung, 54, pp.121~138 (2007)
- Borgi M, "The attractive recombinant phytase from Bacillus licheniformis: biochemical and molecular characterization", Appl. Microbiol. Biotechnol, 98, pp.5937~5947 (2014)
- Hao Tan, "Cloning, Overexpression, and Characterization of a Metagenome-Derived Phytase with Optimal Activity at Low pH", Journal of Microbiology and Biotechnology, 25(6), pp.930~935 (2015), 한국미생물·생명공학회[DOI]
- Cao L, "Application of microbial phytase in fish feed", Enzyme Microb. Technol, 40, pp.497~507 (2007)
- Tan H, "A novel phytase derived from an acidic peat-soil microbiome showing high stability under acidic plus pepsin conditions", J. Mol. Microbiol. Biotechnol, 26, pp.291~301 (2016)
- Ushasree MV, "Microbial phytase: impact of advances in genetic engineering in revolutionizing its properties and applications", Bioresour. Technol, 245, pp.1790~1799 (2017)
- Tran TT, "A thermostable phytase from Bacillus sp. MD2: cloning, expression and high-level production in Escherichia coli", J. Ind. Microbiol. Biotechnol, 37, pp.279~287 (2010)
- Tan H, "Identification and characterization of a mesophilic phytase highly resilient to high-temperatures from a fungus-garden associated metagenome", Appl. Microbiol. Biotechnol, 100, pp.2225~2241 (2016)
- Lei X-G, "Phytase, a new life for an 'old' enzyme", Annu. Rev. Anim. Biosci, 1, pp.283~309 (2013)
- Rebello S, "Molecular advancements in the development of thermostable phytases", Appl. Microbiol. Biotechnol, 101, pp.2677~2689 (2017)
- Huang Huoqing, "Novel Low-Temperature-Active Phytase from Erwinia carotovora var. carotovota ACCC 10276", Journal of Microbiology and Biotechnology, 19(10), pp.1085~1091 (2009), 한국미생물·생명공학회
- Huang H, "A novel beta-propeller phytase from Pedobacter nyackensis MJ11 CGMCC 2503 with potential as an aquatic feed additive", Appl. Microbiol. Biotechnol, 83, pp.249~259 (2009)
- Yu P, "Purification and characterization of a novel cold-adapted phytase from Rhodotorula mucilaginosa strain JMUY14 isolated from Antarctic", J. Basic Microbiol, 55, pp.1029~1039 (2015)
- Peng W, "Status analysis of morel artificial cultivation in Sichuan", Edible Med. Mushrooms, 24, pp.145~150 (2016)
- Schmidt EL, "Spore germination of and carbohydrate colonization by Morchella esculenta at different soil temperatures", Mycologia, 75, pp.870~875 (1983)
- Xu H, "Optimization of cultivation conditions for extracellular polysaccharide and mycelium biomass by Morchella esculenta As51620", Biochem. Eng. J, 39, pp.66~73 (2008)
- Goldway M, "Morchella conica exhibiting a long fruiting season", Mycol. Res, 104, pp.1000~1004 (2000)
- Hong DK, "Gene cloning and characterization of a psychrophilic phthalate esterase with organic solvent tolerance from an Arctic bacterium Sphingomonas glacialis PAMC 26605", J. Mol. Catal. B Enzyme, 133, pp.S337~S345 (2017)
- Feller G, "Psychrophilic enzymes: hot topics in cold adaptation", Nat. Rev. Microbiol, 1, pp.200~208 (2003)
- Feller G, "Psychrophilic enzymes: molecular basis of cold adaptation", Cell. Mol. Life Sci, 53, pp.830~841 (1997)
- Parvizpour S, "Structural and functional analysis of a novel psychrophilic β-mannanase from Glaciozyma antarctica PI12", J. Comput. Aided Mol. Des, 28, pp.685~698 (2014)
- Petersen TN, "SignalP 4.0: discriminating signal peptides from transmembrane regions", Nat. Methods, 8, pp.785~786 (2011)
- Tan H, "Long-term phosphorus fertilisation increased the diversity of the total bacterial community and the phoD phosphorus mineraliser group in pasture soils", Biol. Fertil. Soils, 49, pp.661~672 (2013)
- Letunic I, "Interactive Tree Of Life (iTOL): an online tool for phylogenetic tree display and annotation", Bioinformatics, 23, pp.127~128 (2006)
- Fu D, "Catalytic efficiency of HAP phytases is determined by a key residue in close proximity to the active site", Appl. Microbiol. Biotechnol, 90, pp.1295~1302 (2011)
- Huang H, "A novel phytase from Yersinia rohdei with high phytate hydrolysis activity under low pH and strong pepsin conditions", Appl. Microbiol. Biotechnol, 80, pp.417~426 (2008)
- Hao Tan, "Enhancing the Thermal Resistance of a Novel Acidobacteria-Derived Phytase by Engineering of Disulfide Bridges", Journal of Microbiology and Biotechnology, 26(10), pp.1717~1722 (2016), 한국미생물·생명공학회[DOI]
- Bradford MM., "A rapid and sensitive method for the quantization of microgram quantities of protein utilizing the principle of protein-dye binding", Anal. Biochem, 72, pp.248~254 (1976)
- Lim D, "Crystal structures of Escherichia coli phytase and its complex with phytate", Nat. Struct. Biol, 7, pp.108~113 (2000)
- Uchida H, "Expression of Aspergillus oryzae phytase gene in Aspergillus oryzae RIB40 niaD−", J. Biosci. Bioeng, 102, pp.564~567 (2006)
- Heinonen JK, "A new and convenient colorimetric determination of inorganic orthophosphate and its application to the assay of inorganic pyrophosphatase", Anal. Biochem, 113, pp.313~317 (1981)
- Yao MZ, "Phytases: crystal structures, protein engineering and potential biotechnological applications", J. Appl. Microbiol, 112, pp.1~14 (2012)
- Lassen SF, "Expression, gene cloning, and characterization of five novel phytases from four basidiomycete fungi: Peniophora lycii, Agrocybe pediades, a Ceriporia sp., and Trametes pubescens", Appl. Environ. Microbiol, 67, pp.4701~4707 (2001)
- Ushasree M, "Gene cloning and soluble expression of Aspergillus niger phytase in E. coli cytosol via chaperone co-expression", Biotechnol. Lett, 36, pp.85~91 (2014)
- Pasamontes L, "Gene cloning, purification, and characterization of a heat-stable phytase from the fungus Aspergillus fumigatus", Appl. Environ. Microbiol, 63, pp.1696~1700 (1997)
- Berka RM, "Molecular characterization and expression of a phytase gene from the thermophilic fungus Thermomyces lanuginosus", Appl. Environ. Microbiol, 64, pp.4423~4427 (1998)
- Hao Tan, "Identification of Novel Phytase Genes from an Agricultural Soil-Derived Metagenome", Journal of Microbiology and Biotechnology, 24(1), pp.113~118 (2014), 한국미생물·생명공학회[DOI]
- Niu C, "Engineering the residual side chains of HAP phytases to improve their pepsin resistance and catalytic efficiency", Sci. Rep, 7, pp.e42133~ (2017)
- Shao, Na, "Cloning, Expression, and Characterization of a New Phytase from the Phytopathogenic Bacterium Pectobacterium wasabiae DSMZ 18074", Journal of Microbiology and Biotechnology, 18(7), pp.1221~1226 (2008), 한국미생물·생명공학회
- Fu D, "A highly pH-stable phytase from Yersinia kristeensenii:cloning, expression, and characterization", Enzyme Microb. Technol, 42, pp.499~505 (2008)
- Shi P, "A novel phytase gene appA from Buttiauxella sp. GC21isolated from grass carp intestine", Aquaculture, 275, pp.70~75 (2008)
- Hu ang H, "A novel phytase with preferable characteristics from Yersinia intermedia", Biochem. Biophys. Res. Commun, 350, pp.884~889 (2006)
- Vats P, "Biochemical characterisation of extracellular phytase (myo-inositol hexakisphosphate phospho hydrolase) from a hyper-producing strain of Aspergillus niger van Teighem", J. Ind. Microbiol. Biotechnol, 32, pp.141~147 (2005)
- Singh B, "Characterization of a HAPphytase from a thermophilic mould Sporotrichum thermophile", Bioresour. Technol, 100, pp.2046~2051 (2009)
- Kim H-W, "Isolation and characterization of a phytase with improved properties from Citrobacter braakii", Biotechnol. Lett, 25, pp.1231~1234 (2003)
- Weaver JD, "Impact of assay conditions on activity estimate and kinetics comparison of Aspergillus niger PhyA and Escherichia coli AppA2 phytases", J. Agric. Food Chem, 57, pp.5315~5320 (2009)
- Kumar V, "Phytate and phytase in fish nutrition", J. Anim. Physiol. Anim. Nutr, 96, pp.335~364 (2012)
- Nwanna LC, "Effect of wetincubation of dietary plant feedstuffs with phytases on growth and mineral digestibility by common carp (Cyprinus carpio L)", Aquaculture, 271, pp.461~468 (2007)
- Evaluation report on the analytical methods submitted in connection with the application for authorisation as a feed additive according to Regulation (EC) No 1831, "2003: Optiphos®", González de la Huebra MJ, 2005