Purification of thermostable α-galactosidase from Irpex lacteus and its use for hydrolysis of oligosaccharides
Yajie Guo
College of Biological Sciences, China Agricultural University, Beijing, P.R. China
Search for more papers by this authorYi Song
School of Public Health, Peking University, Beijing, P.R. China
Search for more papers by this authorYi Qiu
College of Biological Sciences, China Agricultural University, Beijing, P.R. China
Search for more papers by this authorXiaoming Shao
Beijing key Laboratory of Biodiversity and Organic Farming, College of Resources and Environmental Sciences, China Agricultural University, Beijing, P.R. China
Search for more papers by this authorHexiang Wang
College of Biological Sciences, China Agricultural University, Beijing, P.R. China
Search for more papers by this authorCorresponding Author
Yuan Song
College of Biological Sciences, China Agricultural University, Beijing, P.R. China
Correspondence: Yuan Song, College of Biological Sciences, China Agricultural University, Beijing 100193, P.R. China
E-mail:[email protected]
Search for more papers by this authorYajie Guo
College of Biological Sciences, China Agricultural University, Beijing, P.R. China
Search for more papers by this authorYi Song
School of Public Health, Peking University, Beijing, P.R. China
Search for more papers by this authorYi Qiu
College of Biological Sciences, China Agricultural University, Beijing, P.R. China
Search for more papers by this authorXiaoming Shao
Beijing key Laboratory of Biodiversity and Organic Farming, College of Resources and Environmental Sciences, China Agricultural University, Beijing, P.R. China
Search for more papers by this authorHexiang Wang
College of Biological Sciences, China Agricultural University, Beijing, P.R. China
Search for more papers by this authorCorresponding Author
Yuan Song
College of Biological Sciences, China Agricultural University, Beijing, P.R. China
Correspondence: Yuan Song, College of Biological Sciences, China Agricultural University, Beijing 100193, P.R. China
E-mail:[email protected]
Search for more papers by this authorAbstract
A monomeric α-galactosidase (ILGI) from the mushroom Irpex lacteus was purified 94.19-fold to electrophoretic homogeneity. ILGI exhibited a specific activity of 18.36 U mg−1 and demonstrated a molecular mass of 60 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). ILGI was optimally active at 80 °C and pH 5.0, and it was stable over a temperature range of 4–70 °C and a wide pH range of 2.0–12.0. ILGI was completely inactivated by Ag+ and Hg2+ ions and N-bromosuccinimide (NBS). Moreover, ILGI exhibited good resistance to proteases. Galactose acted as a noncompetitive inhibitor with Ki and Kis of 3.34 and 0.29 mM, respectively. The α-galactosidase presented a broad substrate specificity, which included p-nitrophenyl α-D-galactopyranoside (pNPGal), melibiose, stachyose, and raffinose with Km values of 1.27, 3.24, 7.1, and 22.12 mM, correspondingly. ILGI exhibited efficient and complete hydrolysis to raffinose and stachyose. The aforementioned features of this enzyme suggest its potential value in food and feed industries.
References
- 1 Kotiguda, G., Kapnoor, S., Kulkarni, D., Mulimani, V., 2007. Degradation of raffinose oligosaccharides in soymilk by immobilized alpha-galactosidase of Aspergillus oryzae. J. Microbiol. Biotechnol., 17, 1430–1436.
- 2 Thippeswamy, S., Mulimani, V.H., 2002. Enzymic degradation of raffinose family oligosaccharides in soymilk by immobilized α-galactosidase from Gibberella fujikuroi. Process Biochem., 38, 635–640.
- 3 Anisha, G.S., Prema, P., 2006. Production of α-galactosidase by a novel actinomycete Streptomyces griseoloalbus and its application in soymilk hydrolysis. World J. Microbiol. Biotechnol., 23, 859–864.
- 4 Roopashri, A.N., Varadaraj, M.C., 2014. Hydrolysis of flatulence causing oligosaccharides by α-d-galactosidase of a probiotic Lactobacillus plantarum MTCC 5422 in selected legume flours and elaboration of probiotic attributes in soy-based fermented product. Eur. Food Res. Technol., 239, 99–115.
- 5 Chen, Z., Yan, Q., Jiang, Z., Liu, Y., et al., 2015. High-level expression of a novel alpha-galactosidase gene from Rhizomucor miehei in Pichia pastoris and characterization of the recombinant enyzme. Protein Expr. Purif., 110, 107–114.
- 6 Wang, H., Shi, P., Luo, H., Huang, H., et al., 2014. A thermophilic alpha-galactosidase from Neosartorya fischeri P1 with high specific activity, broad substrate specificity and significant hydrolysis ability of soymilk. Bioresour. Technol., 153, 361–364.
- 7 Singh, N., Kayastha, A.M., 2012. Purification and characterization of alpha-galactosidase from white chickpea (Cicer arietinum). J. Agric. Food Chem., 60, 3253–3259.
- 8 Schiffmann, R., 2009. Fabry disease. Pharmacol. Ther., 122, 65––77.
- 9 Linden, J.C., 1982. Immobilized alpha-d-galactosidase in the sugar-beet industry. Enzyme Microb. Technol., 4, 130–136.
- 10 Clarke, J.H., Davidson, K., Rixon, J.E., Halstead, J.R., et al., 2000. A comparison of enzyme-aided bleaching of softwood paper pulp using combinations of xylanase, mannanase and alpha-galactosidase. Appl. Microbiol. Biotechnol., 53, 661–667.
- 11 Mi, S., Meng, K., Wang, Y., Bai, Y., et al., 2007. Molecular cloning and characterization of a novel α-galactosidase gene from Penicillium sp. F63 CGMCC 1669 and expression in Pichia pastoris. Enzyme Microb. Technol., 40, 1373–1380.
- 12 Bayraktar, H., Önal, S., 2013. Concentration and purification of α-galactosidase from watermelon (Citrullus vulgaris) by three phase partitioning. Sep. Purif. Technol., 118, 835–841.
- 13 Unzueta, U., Vázquez, F., Accardi, G., Mendoza, R., Toledo-Rubio, V., et al., 2015. Strategies for the production of difficult-to-express full-length eukaryotic proteins using microbial cell factories: production of human alpha-galactosidase A. Appl. Microbiol. Biotechnol., 99, 5863–5874.
- 14 Cantarel, B.L., Coutinho, P.M., Rancurel, C., Bernard, T., et al., 2009. The carbohydrate-active enzymes database (CAZy): an expert resource for glycogenomics. Nucleic. Acids Res., 37, D233–D238.
- 15 Chauhan, A., Siddiqi, N.J., Sharma, B., 2014. A novel promising strain of Trichoderma evansii (WF-3) for extracellular alpha-galactosidase production by utilizing different carbon sources under optimized culture conditions. Biomed. Res. Int., 2014, 1–12.
- 16 Novotný, Č., Erbanová, P., Cajthaml, T., Rothschild, N., et al., 2000. Irpex lacteus, a white rot fungus applicable to water and soil bioremediation. Appl. Microbiol. Biotechnol., 54, 850–853.
- 17 Na, Z., Yan, L., Jia-hui, L., Juan, W., et al., 2012. Isolation, purification and bioactivities of polysaccharides from Irpex lacteus. Chem. Res. Chinese U., 28, 249–254.
- 18 Kikuchi, E., Kobayashi, H., Kusakabe, I., Murakami, K., 1988. Fiber-structured cheese making with Irpex lacteus milk-clotting enzyme. Agric. Biol. Chem., 52, 1277–1278.
- 19 Kobayashi, H., Kusakabe, I., Murakami, K., 1983. Purification and characterization of two milk-clotting enzymes from Irpex lacteus. Agric. Biol. Chem., 47, 551–558.
- 20 Malhotra, O.P., Dey, P.M., 1967. Purification and physical properties of sweet-almond α-galactosidase. Biochem. J., 103, 508–513.
- 21
Miller, G.L.,
1957.
Use of dinitrosaIicyIic acid reagent for determination of reducing sugar.
Anal. Chem.,
31, 426–428.
10.1021/ac60147a030 Google Scholar
- 22 Anisha, G.S., John, R.P., Prema, P., 2009. Biochemical and hydrolytic properties of multiple thermostable α-galactosidases from Streptomyces griseoloalbus: obvious existence of a novel galactose-tolerant enzyme. Process Biochem., 44, 327–333.
- 23 Kurakake, M., Okumura, T., Morimoto, Y., 2015. Synthesis of galactosyl glycerol from guar gum by transglycosylation of alpha-galactosidase from Aspergillus sp, MK14. Food Chem., 172, 150–154.
- 24 Simila, J., Gernig, A., Murray, P., Fernandes, S., Tuohy, M.G., 2010. Cloning and expression of a thermostable alpha-galactosidase from the thermophilic fungus Talaromyces emersonii in the methylotrophic yeast Pichia pastoris. J. Microbiol. Biotechnol., 20, 1653–1663.
- 25 Kandari, S., Choi, Y.J., Lee, B.H., 2014. Purification and characterization of hydrolytic and transgalactosyl α-galactosidase from Lactobacillus helveticus ATCC 10797. Eur. Food Res. Technol., 239, 877–884.
- 26 Kotake, T., Kitazawa, K., Takata, R., Okabe, K., et al., 2009. Molecular cloning and expression in Pichia pastoris of a Irpex lacteus exo-beta-(1->3)-galactanase gene. Biosci. Biotechnol. Biochem., 73, 2303–2309.
- 27 Turner, P., Mamo, G., Karlsson, E.N., 2007. Potential and utilization of thermophiles and thermostable enzymes in biorefining. Microb. Cell Fact., 6, 9.
- 28 Shivam, K., Mishra, S.K., 2010. Purification and characterization of a thermostable α-galactosidase with transglycosylation activity from Aspergillus parasiticus MTCC-2796. Process Biochem., 45, 1088–1093.
- 29 Wang, H., Ma, R., Shi, P., Huang, H., et al., 2015. Insights into the substrate specificity and synergy with mannanase of family 27 alpha-galactosidases from Neosartorya fischeri P1. Appl. Microbiol. Biotechnol., 99, 1261–1272.
- 30 Katrolia, P., Jia, H., Yan, Q., Song, S., et al., 2012. Characterization of a protease-resistant alpha-galactosidase from the thermophilic fungus Rhizomucor miehei and its application in removal of raffinose family oligosaccharides. Bioresour. Technol., 110, 578–586.
- 31 Fujimoto, Z., Kaneko, S., Momma, M., Kobayashi, H., et al., 2003. Crystal structure of rice alpha-galactosidase complexed with d-galactose. J. Biol. Chem., 278, 20313–20318.
- 32 Gote, M.M., Khan, M.I., Khire, J.M., 2007. Active site directed chemical modification of α-galactosidase from Bacillus stearothermophilus (NCIM 5146): involvement of lysine, tryptophan and carboxylate residues in catalytic site. Enzyme Microb. Technol., 40, 1312–1320.
- 33 Viana, P.A., de Rezende, S.T., Marques, V.M., Trevizano, L.M., et al., 2006. Extracellular alpha-galactosidase from Debaryomyces hansenii UFV-1 and its use in the hydrolysis of raffinose oligosaccharides. J. Agric. Food. Chem., 54, 2385–2391.
- 34 Ferreira, J.G., Reis, A.P., Guimaraes, V.M., Falkoski, D.L., et al., 2011. Purification and characterization of Aspergillus terreus alpha-galactosidases and their use for hydrolysis of soymilk oligosaccharides. Appl. Biochem. Biotechnol., 164, 1111–1125.
- 35 Cao, Y., Wang, Y., Luo, H., Shi, P., et al., 2009. Molecular cloning and expression of a novel protease-resistant GH-36 alpha-galactosidase from Rhizopus sp. F78 ACCC 30795. J. Microbiol. Biotechnol., 19, 129–300.
- 36 Zhou, J., Dong, Y., Li, J., Zhang, R., et al., 2012. Cloning, heterologous expression, and characterization of novel protease-resistant alpha-galactosidase from new Sphingomonas strain. J. Microbiol. Biotechnol., 22, 1532–1539.
- 37 Dey, P.M., Patel, S., Brownleader, M.D., 1993. Induction of alpha-galactosidase in Penicillium ochrochloron by guar (Cyamopsis tetragonobola) gum. Biotechnol. Appl. Biochem., 17, 361–371.
- 38 Zhou, J., Shi, P., Huang, H., Cao, Y., et al., 2010. A new alpha-galactosidase from symbiotic Flavobacterium sp. TN17 reveals four residues essential for alpha-galactosidase activity of gastrointestinal bacteria. Appl. Microbiol. Biotechnol., 88, 1297–1309.
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