- Original article
- Open Access
Characterization of β-N-acetylhexosaminidase (LeHex20A), a member of glycoside hydrolase family 20, from Lentinula edodes (shiitake mushroom)
© Konno et al.; licensee Springer. 2012
- Received: 1 May 2012
- Accepted: 13 May 2012
- Published: 1 June 2012
We purified and cloned a β-N-acetylhexosaminidase, LeHex20A, with a molecular mass of 79 kDa from the fruiting body of Lentinula edodes (shiitake mushroom). The gene lehex20a gene had 1,659 nucleotides, encoding 553 amino acid residues. Sequence analysis indicated that LeHex20A belongs to glycoside hydrolase (GH) family 20, and homologues of lehex20a are broadly represented in the genomes of basidiomycetes. Purified LeHex20A hydrolyzed the terminal monosaccharide residues of β-N-acetylgalactosaminides and β-N-acetylglucosaminides, indicating that LeHex20A is a β-N-acetylhexosaminidase classified into EC 220.127.116.11. The maximum LeHex20A activity was observed at pH 4.0 and 50°C. The kinetic constants were estimated using chitooligosaccharides with degree of polymerization 2-6. GH20 β-N-acetylhexosaminidases generally prefer chitobiose among natural substrates. However, LeHex20A had the highest catalytic efficiency (kcat/Km) for chitotetraose, and the Km values for GlcNAc6 were 3.9-fold lower than for chitobiose. Furthermore, the enzyme partially hydrolyzed amorphous chitin polymers. These results indicate that LeHex20A can produce N-acetylglucosamine from long-chain chitomaterials.
- Fungal cell wall
- Glycoside hydrolase family 20
Chitin, composed of β-1,4 linked N- acetylglucosamine (GlcNAc) units, is mainly found in crustaceans, insects and fungi. Enzymatic degradation of chitin is catalyzed by a two-component chitinolytic enzyme system. One component is chitinases (EC 18.104.22.168), which hydrolyze β-1,4 linkages in chitin polymers, endolytically producing chitooligosaccharides, especially chitobiose (Brurberg et al. 1996; Tanaka et al. 2001). The other is β-N-acetylhexosaminidases (EC 22.214.171.124), which typically have no activity against chitin polymers, and instead degrade chitooligosaccharides formed by chitinases into monomers. Because the enzymes prefer short β-N-acetylglucosaminide substrates, chitobiose and p-nitrophenyl-N-acetyl-β-D-glucosaminide (pNP-GlcNAc), they are also called chitobiases (Drouillard et al. 1997; Tews et al. 1996). β-N-acetylhexosaminidases are widely distributed in animal tissues (Korneluk et al. 1986), insects (Hogenkamp et al. 2008; Yang et al. 2008), plants (Meli et al. 2010), bacteria (Clarke et al. 1995; Mark et al. 2001) and fungi (Cannon et al. 1994; López-Mondéjar et al. 2009; Jones and Kosman 1980), and belong to glycoside hydrolase (GH) families 3, 20 and 84 as categorized in the CAZy database (http://www.cazy.org/index.html). The GH20 enzymes hydrolyze nonreducing terminal monosaccharide residues of β-N-acetylgalactosaminides and β-N-acetylglucosaminides.
In fungi, chitin is a main cell-wall component, together with β-glucans (Iten and Matile 1970; Vetter 2007), and most filamentous fungi such as ascomycetes and basidiomycetes produce chitinolytic enzymes. Some mycoparasitic fungi such as Trichoderma species produce extracellular chitinolytic enzymes for degradation of host cell walls during their mycoparasitic attack (Carsolio et al. 1994; Peterbauer et al. 1996; Seidl et al. 2006). On the other hand, some fungal chitinolytic enzymes act on their own cell walls during changes in morphology, which are an essential process in the fungal cell cycle (Mitchell and Sabar 1966; Seiler and Plamann 2003). For example, some chitinases (Rast et al. 1991; Shin et al. 2009) and β-N-acetylhexosaminidases (Cannon et al. 1994; Kim et al. 2002) from filamentous fungi such as Aspergillus and Mucor species are suggested to have roles in processes such as hyphal autolysis, growth and branching. However, little information is known about the physiological function and role of fungal chitinolytic enzymes. Moreover, there have been no reports of cloned and characterized chitinolytic enzymes from basidiomycetes.
Most basidiomycetes form a fruiting body (mushroom) as part of their usual life cycle. The cell walls of the fruiting body are constructed mainly from chitin and β-glucans, and these polysaccharides are self-degraded by enzymes associated with cell walls during morphological changes (Shida et al. 1981; Minato et al. 2004). Recently, we reported identification of four β-1,3-glucanases (EXG1, EXG2, TLG1 and GLU1) and one β-1,6-glucanase (LePus30A) from the Lentinula edodes fruiting body, the shiitake mushroom (Sakamoto et al. 2005a; 2005b; 2006; 2011; Konno and Sakamoto 2011). While enzymes involved in cell wall metabolism of L. edodes have been reported only for those acting on β-glucan, the presence of chitinolytic enzymes were suggested in our previous study (Sakamoto et al. 2009). In the present study, we purified and characterized a β-N-acetylhexosaminidase, LeHex20A, from the fruiting body of L. edodes.
L. edodes cultivated dikaryotic strain H600 (Hokken. Co., Ltd) was used in all experiments (Sakamoto et al., 2005a). Fruiting bodies for RNA and protein extraction were prepared using the method of Nagai et al. (2003). Mature fruiting bodies were separated into pileus, gill and stipe. Harvested mature fruiting bodies were immediately transferred to a desiccator at 25°C and 80% humidity for post-harvest preservation. All samples were stored at -80°C.
Colloidal chitin was prepared according to Hsu and Lockwood (1975). Mechanochemically ground chitin was kindly provided by the Department of Chemical Engineering, Ichinoseki National College of Technology (Nakagawa et al. 2011).
Purification of β-N-acetylhexosaminidase
Proteins were extracted from 320 g of fresh fruiting bodies. Samples were crushed in liquid nitrogen, suspended in 320 ml of 10 mM sodium phosphate buffer (pH 7.0), and incubated with rotation for 30 min at room temperature. Ammonium sulfate was added until the concentration reached 70% saturation. The resulting precipitates were collected by centrifugation (30 min, 4,500 × g) and dissolved in 10 mM sodium phosphate buffer (pH 7.0) containing ammonium sulfate at 30% saturation. The supernatant was applied to a Phenyl-Toyopearl column (1.6 × 10 cm, Tosoh Co., Ltd., Tokyo, Japan) equilibrated with 10 mM sodium phosphate buffer (pH 7.0) containing ammonium sulfate at 30% saturation. The column was washed with 45 ml of the same buffer, and proteins were eluted in 45 ml of a linear concentration gradient (30-0% saturation) of ammonium sulfate at a flow rate of 1.5 ml/min. Fractions containing β-N-acetylhexosaminidase activity were collected and concentrated using an Amicon Ultra 5,000 NMWL filter (Millipore, Billerica, MA, USA), and then applied to a MonoQ 5/50 GL anion exchange column (0.5 × 5 cm, GE Healthcare, Little Chalfont, UK). Adsorbed proteins were eluted using a linear concentration gradient of NaCl (0–0.5 M) at a flow rate of 0.5 ml/min. The eluted enzyme was then applied to a DEAE-Toyopearl Pak 650S anion exchange column (0.8 × 7.5 cm, Tosoh Co., Ltd.) equilibrated with 10 mM sodium phosphate buffer. The enzyme was eluted with a linear concentration gradient of NaCl (60 ml, 0–0.5 M) at a flow rate of 0.5 ml/min. Fractions containing activity were collected and concentrated. Concentrated proteins were then applied to a Superdex 75 10/30 gel filtration column (GE Healthcare) equilibrated in 10 mM sodium phosphate buffer (pH 7.0) with 0.1 M NaCl, and proteins were eluted with the same buffer at a flow rate of 0.4 ml/min. Purified LeHex20A was analyzed by SDS-PAGE, and the N-terminal amino acid sequence of LeHex20A was analyzed as described in Sakamoto et al. (2005a).
Cloning and sequencing of the lehex20a gene
cDNA was synthesized from total RNA extracted from fresh fruiting bodies using the SMART PCR RACE kit (BD Bioscience, CA, USA), according to the manufacturer’s protocol. 3′-RACE was performed using degenerate primers (chi4-3U: 5′-ACN GYN GYN ATG GTN TGG AT-3′ and chi4-4U: 5′-TGG TGY GAY CCN TTY AAR AC-3′) designed against conserved amino acid sequences of GH family 20 in filamentous fungi. cDNA for the 5′-RACE PCR template was synthesized from the RNA using a GeneRacer kit (Invitrogen, CA, USA), and PCR was performed as described previously (Sakamoto et al., 2005b) using specific primers (chi4-56-RACEL: 5′-AGT TTA GCT TGA GCA TCA GTC AAA T-3′ and chi4-93-RACEL: 5′-CTC GGT CCA AAG TAG GTG TTC T-3′) and GeneRacer primers (Invitrogen). The presence of a signal peptide in the deduced amino sequence was predicted using the SignalP server (http://www.cbs.dtu.dk/services/SignalP/). Comparative analysis of homology with enzymes registered in the GenBank databases was carried out using an NCBI BLAST search (http://www.ncbi.nlm.nih.gov/BLAST) with the default parameters.
β-N-Acetylhexosaminidase activity was assayed in 20 mM sodium acetate buffer (pH 4.2) at 37°C for 15 min. For purification, activity was determined using 0.32 mM of 4-methylumbelliferyl β-D-N,N',N"-triacetylchitotrioside (4MU-GlcNAc3, an analogue of the natural substrate, GlcNAc4; Sigma-Aldrich Inc., St. Louis, MO, USA) as substrate (Hood 1991). The reaction was quenched with 0.4 M Na2CO3, and the released 4MU was measured by spectrophotofluorimetry with excitation at 365 nm and emission at 445 nm. The effects of pH (pH 3-9) and temperature (10-80°C) on enzyme activity were analyzed as described previously (Konno and Sakamoto 2011). To elucidate the substrate specificity of the enzyme, assays were performed using the following substrates: p-nitrophenyl-N-acetyl-β-D-glucosaminide (pNP-GlcNAc), p-nitrophenyl-N-acetyl-beta-D-galactosaminide (pNP-GalNAc), p-nitrophenyl-D-glucoside (pNP-Glc) (Sigma-Aldrich), chitooligosaccharides, (GlcNAc2-6, Seikagaku Biobusiness Co., Tokyo, Japan), the complex N-glycan (GlcNAcβ-1,2Manα-1,6) (GlcNAcβ-1,2Manα-1,3) Manβ-1,4GlcNAcβ-1,4GlcNAc-PA (TaKaRa Bio Inc., Shiga, Japan), chitin (Wako Pure Chemicals Co., Osaka, Japan), ethylene glycol chitin (Seikagaku kogyo, Co., Tokyo, Japan), colloidal chitin and the mechanochemically ground chitin. To assay pNP, the amount of pNP was determined spectrophotometrically at 405 nm. The extinction coefficient of pNP was assumed to be 17,100 M-1 cm-1. The amount of GlcNAc released from chitin oligomers and polymers was determined by the Morgan-Elson assay according to the method of Keyhani and Roseman (1996). One unit (U) of enzyme activity was defined as the amount of enzyme that produces 1 μmol GlcNAc per minute under the above conditions. To determine the kinetic properties of LeHex20A, the reactions were performed with 0.05-0.5 mM of substrate and 3.1 nM of purified LeHex20A in 20 mM sodium acetate buffer (pH 4.2) at 37°C for 15 min. In these reactions, the products formed from the chitooligosaccharides (GlcNAc2-6) were monosaccharides (GlcNAc) and oligosaccharides shortened by one GlcNAc unit (HPLC analyses, data not shown). The values of kcat and Km were estimated using Lineweaver-Burk plots.
To study the degradation of natural substrates, 2% (w/v) amorphous chitins (colloidal chitin or mechanochemically grinded chitin) or 0.5 mM chitooligosaccharides were incubated at 30 °C in 20 mM sodium acetate buffer (pH 4.2), using 0.92 nM purified LeHex20A for the chitins and 0.40 nM LeHex20A for the chitooligosaccharides. The hydrolysis products were analyzed using an HPLC system equipped with a TSKgel Amide-80 column (4.6 × 250 mm, Tosoh). The mobile phase was 65% (v/v) acetonitrile at a flow rate of 1.0 ml/min, and the column temperature was 80°C. Eluted carbohydrates were detected by monitoring UV absorption at 205 nm.
Nucleotide sequence accession number
The nucleotide sequence encoding LeHex20A has been deposited in the DDBJ/EMBL/GenBank databases under the accession number [DDBJ: AB703443].
Purification of LeHex20A and cloning of its gene, lehex20a
According to the results of SignalP analysis, the first 17 amino acid residues in the N-terminal region are expected to be a signal peptide, indicating that the mature protein, consisting of 536 amino acids, is an extracellular or cell wall protein. LeHex20A has a calculated molecular mass of 58 kDa, suggesting that the protein is glycosylated. Indeed, the amino acid sequence had 13 possible N-glycosylation sites (Asn-Xxx-Thr/Ser http://www.cbs.dtu.dk/services/NetNGlyc/). In addition, there were many possible sites for O-glycosylation (http://www.cbs.dtu.dk/services/NetOGlyc/). The sequence was analyzed using searches on the Pfam database (http://pfam.sanger.ac.uk/search/sequence). Search results for the sequence showed that the amino acid sequence contains GH20 domains. The deduced amino acid sequence was analyzed using the blastp algorithm of the NCBI protein database. The BLAST search showed that the sequence had up to 61% sequence identity to putative GH20 proteins (containing putative β-N-acetylhexosaminidase sequences) from basidiomycetous species such as Serpula lacrymans (EGN97893), Coprinopsis cinerea (XP_001835638) and Postia placenta (XP_002472465). The sequence showed homology to putative GH20 proteins from ascomycetes such as Leptosphaeria maculans (CBX95932) and Trichoderma reesei (EGR50812), although in these cases, sequence identity levels were only about 30%. These results suggest that LeHex20A belongs to GH20. Further sequence analyses were carried out using the blastp algorithm in genome sequence databases of basidiomycetes (http://genome.jgi-psf.org/programs/fungi/index.jsf). These searches revealed that LeHex20A has high levels of similarity to proteins of basidiomycetes including Fomitopsis pinicola (ID number from DOE Joint Genome Institute, 129075; similarity, 61%), Heterobasidion annosum (151266; 58%), Agaricus bisporus (120598; 58%) and Pleurotus ostreatus (57387/87509; 58%). Thus, homologues of lehex20a seem to be conserved in basidiomycetes.
Enzymatic properties of LeHex20A
Effects of pH and temperature on enzyme activity were examined using 4MU-GlcNAc3 as a substrate. The maximum LeHex20A activity was observed at pH 4.0 in 20 mM sodium acetate buffer and at 50°C. The enzyme was stable across a pH range from 5 to 8 when incubated at 4°C for 20 h. The enzyme was inactivated after incubation at 60°C for 30 min.
Kinetic parameters of LeHex20A
0.34 ± 0.01
335 ± 10
0.43 ± 0.03
177 ± 8
0.42 ± 0.04
242 ± 14
0.14 ± 0.001
236 ± 1
0.07 ± 0.008
119 ± 5
0.08 ± 0.001
97 ± 1
0.1 ± 0.002
97 ± 1
Knowledge of chitinolytic enzymes from basidiomycetes is important because these enzymes are likely to play roles on the autolysis of cell-wall chitin, which may result in morphological changes and affect the product quality of fruiting bodies (Kamada et al. 1982; Kües 2000; Mitchell and Sabar 1966; Sakamoto et al. 2009; Sone and Misaki 1978). Nevertheless, little is known about chitinolytic enzymes from basidiomycetes. In this study, we purified a protein with β-N-acetylhexosaminidase activity from fresh L. edodes fruiting bodies, and the analysis of its primary structure showed that the enzyme belongs to GH20. Therefore, we named the protein LeHex20A. To the best of our knowledge, this is the first report of a gene encoding a GH20 β-N-acetylhexosaminidase from basidiomycetes.
Most of the GH20 β-N-acetylhexosaminidases have the highest catalytic efficiency for GlcNAc2 among natural substrates, and do not hydrolyze long-chain chitomaterials (Keyhani and Roseman 1996; Koga et al. 1996; Ueda and Arai 1992; Yang et al. 2008). However, the catalytic efficiency of LeHex20A for GlcNAc6 was greater than for GlcNAc2, due to its high affinity for GlcNAc6. Moreover, the enzyme partially hydrolyzed two kinds of amorphous chitin polymers, resulting in production of GlcNAc. There are few reports about degradation of chitin polymers by β-N-acetylhexosaminidases (Suginta et al. 2010). Indeed, only six β-N-acetylhexosaminidases (EC 126.96.36.199) that are able to degrade chitin polymer are listed in the BRENDA database (http://www.brenda-enzymes.info/index.php4). Thus, LeHex20A showed unique enzymatic properties with respect to typical GH20 β-N-acetylhexosaminidases. Because β-N-acetylhexosaminidases hydrolyze nonreducing terminal monosaccharide residues of substrates, binding between the subsite (-1) of the enzymes and the nonreducing terminal β-N-acetylhexosaminides of substrates seems to be essential for the catalysis. Suginta et al. (2010) reported that GH20 β-N-acetylhexosaminidases from the bacterium Vibrio harveyi show hydrolytic activity toward colloidal chitin, and suggested that the enzyme has a binding pocket containing four GlcNAc binding subsites. Therefore, LeHex20A might have other GlcNAc binding subsites, as suggested for this protein by Suginta et al.
As described in this report, LeHex20A can produce GlcNAc from longer-chain chitomaterials compared with typical β-N-acetylhexosaminidases. Moreover, the L. edodes fruiting body, the shiitake mushroom, is a very popular edible cultivated mushroom. Therefore, LeHex20A may be valuable for efficient and safe enzymatic production of GlcNAc from native chitin materials.
We thank to Miss Ayumi Obara for her help with experiments. We are grateful to Vincent G H Eijsink of Norwegian University of Life Sciences for his suggestion and comments. We also thank to Kazuhide Totani, Mitsuru Nikaido, Yuko S. Nakagawa of the Department of Chemical Engineering, Ichinoseki National College of Technology, who provide us mechanochemically ground chitin.
This research was supported by a Grant-in-Aid for Scientific Research to N.K. (no. 23780169) from the Japan Society for the Promotion of Science (JSPS), and Grants for project research (Development of fundamental technology for analysis and evaluation of functional agricultural products and functional foods).
- Brurberg MB, Nes IF, Eijsink VG: Comparative studies of chitinases A and B from Serratia marcescens. Microbiology 1996, 142: 1581–1589. 10.1099/13500872-142-7-1581View ArticlePubMedGoogle Scholar
- Cannon RD, Niimi K, Jenkinson HF, Shepherd MG: Molecular cloning and expression of the Candida albicans β-N-acetylglucosaminidase (HEX1) gene. J Bacteriol 1994, 176: 2640–2647.PubMed CentralPubMedGoogle Scholar
- Carsolio C, Gutiérrez A, Jiménez B, Van Montagu M, Herrera-Estrella A: Characterization of ech-42, a Trichoderma harzianum endochitinase gene expressed during mycoparasitism. Proc Natl Acad Sci U S A 1994, 91: 10903–10907. 10.1073/pnas.91.23.10903PubMed CentralView ArticlePubMedGoogle Scholar
- Clarke VA, Platt N, Butters TD: Cloning and expression of the β-N-acetylglucosaminidase gene from Streptococcus pneumoniae. Generation of truncated enzymes with modified aglycon specificity. J Biol Chem 1995, 270: 8805–8814. 10.1074/jbc.270.15.8805View ArticlePubMedGoogle Scholar
- Drouillard S, Armand S, Davies GJ, Vorgias CE, Henrissat B: Serratia marcescens chitobiase is a retaining glycosidase utilizing substrate acetamido group participation. Biochem J 1997, 328: 945–949.PubMed CentralView ArticlePubMedGoogle Scholar
- Hogenkamp DG, Arakane Y, Kramer KJ, Muthukrishnan S, Beeman RW: Characterization and expression of the β-N-acetylhexosaminidase gene family of Tribolium castaneum. Insect Biochem Mol Biol 2008, 38: 478–489. 10.1016/j.ibmb.2007.08.002View ArticlePubMedGoogle Scholar
- Hood MA: Comparison of four methods for measuring chitinase activity and the application of the 4-MUF assay in aquatic environments. J Microbiol Methods 1991, 13: 151–160. 10.1016/0167-7012(91)90015-IView ArticleGoogle Scholar
- Hsu SC, Lockwood JL: Powdered chitin agar as a selective medium for enumeration of actinomycetes in water and soil. Appl Microbiol 1975, 29: 422–426.PubMed CentralPubMedGoogle Scholar
- Intra J, Pavesi G, Horner DS: Phylogenetic analyses suggest multiple changes of substrate specificity within the glycosyl hydrolase 20 family. BMC Evol Biol 2008, 8: 214. 10.1186/1471-2148-8-214PubMed CentralView ArticlePubMedGoogle Scholar
- Iten W, Matile P: Role of chitinase and other lysosomal enzymes of Coprinus Zagopus in the autolysis of fruiting bodies. J Gen Microbiol 1970, 61: 301–309.View ArticleGoogle Scholar
- Jones CS, Kosman DJ: Purification, properties, kinetics, and mechanism of β-N-acetylglucosamidase from Aspergillus niger. J Biol Chem 1980, 255: 11861–11869.PubMedGoogle Scholar
- Kamada T, Hamada Y, Takemaru T: Autolysis in vitro of the stipe cell wall in Coprinus macrorhizus. Microbiology 1982, 128: 1041–1046. 10.1099/00221287-128-5-1041View ArticleGoogle Scholar
- Keyhani NO, Roseman S: The chitin catabolic cascade in the marine bacterium Vibrio furnissii. Molecular cloning, isolation, and characterization of a periplasmic β-N-acetylglucosaminidase. J Biol Chem 1996, 271: 33425–33434. 10.1074/jbc.271.52.33425View ArticlePubMedGoogle Scholar
- Kim S, Matsuo I, Ajisaka K, Nakajima H, Kitamoto K: Cloning and characterization of the nagA gene that encodes β-N-acetylglucosaminidase from Aspergillus nidulans and its expression in Aspergillus oryzae. Biosci Biotechnol Biochem 2002, 66: 2168–2175. 10.1271/bbb.66.2168View ArticlePubMedGoogle Scholar
- Koga D, Hoshika H, Matsushita M, Tanaka A, Ide A, Kono M: Purification and characterization of β-N-acetylhexosaminidase from the liver of a prawn, Penaeus japonicus. Biosci Biotechnol Biochem 1996, 60: 194–199. 10.1271/bbb.60.194View ArticlePubMedGoogle Scholar
- Konno N, Sakamoto Y: An endo-β-1,6-glucanase involved in Lentinula edodes fruiting body autolysis. Appl Microbiol Biotechnol 2011, 91: 1365–1373. 10.1007/s00253-011-3295-2View ArticlePubMedGoogle Scholar
- Korneluk RG, Mahuran DJ, Neote K, Klavins MH, O’Dowd BF, Tropak M, Willard HF, Anderson MJ, Lowden JA, Gravel RA: Isolation of cDNA clones coding for the α-subunit of human β-hexosaminidase. J Biol Chem 1986, 261: 8407–8413.PubMedGoogle Scholar
- Kües U: Life history and developmental processes in the basidiomycete Coprinus cinereus. Microbiol Mol Biol Rev 2000, 64: 316–353. 10.1128/MMBR.64.2.316-353.2000PubMed CentralView ArticlePubMedGoogle Scholar
- López-Mondéjar R, Catalano V, Kubicek CP, Seidl V: The β-N-acetylglucosaminidases NAG1 and NAG2 are essential for growth of Trichoderma atroviride on chitin. FEBS J 2009, 276: 5137–5148. 10.1111/j.1742-4658.2009.07211.xView ArticlePubMedGoogle Scholar
- Mark BL, Vocadlo DJ, Knapp S, Triggs-Raine BL, Withers SG, James MN: Crystallographic evidence for substrate-assisted catalysis in a bacterial β-hexosaminidase. J Biol Chem 2001, 276: 10330–10337. 10.1074/jbc.M011067200View ArticlePubMedGoogle Scholar
- Mayer C, Vocadlo DJ, Mah M, Rupitz K, Stoll D, Warren RA, Withers SG: Characterization of a β-N-acetylhexosaminidase and a β-N-acetylglucosaminidase/β-glucosidase from Cellulomonas fimi. FEBS J 2006, 273: 2929–2941. 10.1111/j.1742-4658.2006.05308.xView ArticlePubMedGoogle Scholar
- Meli VS, Ghosh S, Prabha TN, Chakraborty N, Chakraborty S, Datta A: Enhancement of fruit shelf life by suppressing N-glycan processing enzymes. Proc Natl Acad Sci U S A 2010, 107: 2413–2418. 10.1073/pnas.0909329107PubMed CentralView ArticlePubMedGoogle Scholar
- Minato K, Kawakami S, Nomura K, Tsuchida H, Mizuno M: An exo β-1,3 glucanase synthesized de novo dgrades lentinan during storage of Lentinula edodes and diminishes immunomodulationg activity of the mushroom. Carbohydr Polym 2004, 56: 279–286. 10.1016/j.carbpol.2003.11.016View ArticleGoogle Scholar
- Mitchell R, Sabar N: Autolytic enzymes in fungal cell walls. J Gen Microbiol 1966, 42: 39–42.View ArticlePubMedGoogle Scholar
- Nagai M, Kawata M, Watanabe H, Ogawa M, Saito K, Takesawa T, Kanda K, Sato T: Important role of fungal intracellular laccase for melanin synthesis: purification and characterization of an intracellular laccase from Lentinula edodes fruit bodies. Microbiology 2003, 149: 2455–2462. 10.1099/mic.0.26414-0View ArticlePubMedGoogle Scholar
- Nakagawa YS, Oyama Y, Kon N, Nikaido M, Tanno K, Kogawa J, Inomata S, Masui A, Yamamura A, Kawaguchi M, Matahira Y, Totani K: Development of innovative technologies to decrease the environmental burdens associated with using chitin as a biomass resource: mechanochemical grinding and enzymatic degradation. Carbohydr Polym 2011, 83: 1843–1849. 10.1016/j.carbpol.2010.10.050View ArticleGoogle Scholar
- Peterbauer CK, Lorito M, Hayes CK, Harman GE, Kubicek CP: Molecular cloning and expression of the nag1 gene (N -acetyl-β-D-glucosaminidase-encoding gene) from Trichoderma harzianum P1. Curr Genet 1996, 30: 325–331. 10.1007/s002940050140View ArticlePubMedGoogle Scholar
- Rast DM, Horsch M, Furter R, Gooday GW: A complex chitinolytic system in exponentially growing mycelium of Mucor rouxii: properties and function. J Gen Microbiol 1991, 137: 2797–2810.View ArticlePubMedGoogle Scholar
- Sakamoto Y, Irie T, Sato T: Isolation and characterization of a fruiting body-specific exo-β-1,3-glucanase-encoding gene, exg1, from Lentinula edodes. Curr Genet 2005, 47: 244–252. 10.1007/s00294-005-0563-7View ArticlePubMedGoogle Scholar
- Sakamoto Y, Minato K, Nagai M, Kawakami S, Mizuno M, Sato T: Characterization of the Lentinula edodes exg2 gene encoding a lentinan-degrading exo-β-1,3-glucanase. Curr Genet 2005, 48: 195–203. 10.1007/s00294-005-0002-9View ArticlePubMedGoogle Scholar
- Sakamoto Y, Watanabe H, Nagai M, Nakade K, Takahashi M, Sato T: Lentinula edodes tlg1 encodes a thaumatin-like protein that is involved in lentinan degradation and fruiting body senescence. Plant Physiol 2006, 141: 793–801. 10.1104/pp.106.076679PubMed CentralView ArticlePubMedGoogle Scholar
- Sakamoto Y, Nakade K, Sato T: Characterization of the post-harvest changes in gene transcription in the gill of the Lentinula edodes fruiting body. Curr Genet 2009, 55: 409–423. 10.1007/s00294-009-0255-9View ArticlePubMedGoogle Scholar
- Sakamoto Y, Nakade K, Konno N: Endo-β-1,3-glucanase GLU1, from the fruiting body of Lentinula edodes, belongs to a new glycoside hydrolase family. Appl Environ Microbiol 2011, 77: 8350–8354. 10.1128/AEM.05581-11PubMed CentralView ArticlePubMedGoogle Scholar
- Seidl V, Druzhinina IS, Kubicek CP: A screening system for carbon sources enhancing β-N-acetylglucosaminidase formation in Hypocrea atroviridis (Trichoderma atroviride). Microbiology 2006, 152: 2003–2012. 10.1099/mic.0.28897-0View ArticlePubMedGoogle Scholar
- Seiler S, Plamann M: The genetic basis of cellular morphogenesis in the filamentous fungus Neurospora crassa. Mol Biol Cell 2003, 14: 4352–4364. 10.1091/mbc.E02-07-0433PubMed CentralView ArticlePubMedGoogle Scholar
- Shida M, Ushioda Y, Nakajima T, Matsuda K: Structure of the alkali-insoluble skeletal glucan of Lentinus edodes. J Biochem 1981, 90: 1093–1100.PubMedGoogle Scholar
- Shin KS, Kwon NJ, Kim YH, Park HS, Kwon GS, Yu JH: Differential roles of the ChiB chitinase in autolysis and cell death of Aspergillus nidulans. Eukaryot Cell 2009, 8: 738–746. 10.1128/EC.00368-08PubMed CentralView ArticlePubMedGoogle Scholar
- Sone Y, Misaki A: Purification and characterization of β-D-mannosidase and β-N-acetyl-D-hexosaminidase of Tremella fuciformis. J Biochem 1978, 83: 1135–1144.PubMedGoogle Scholar
- Suginta W, Chuenark D, Mizuhara M, Fukamizo T: Novel β-N-acetylglucosaminidases from Vibrio harveyi 650: cloning, expression, enzymatic properties, and subsite identification. BMC Biochem 2010, 11: 40. 10.1186/1471-2091-11-40PubMed CentralView ArticlePubMedGoogle Scholar
- Tanaka T, Fukui T, Imanaka T: Different cleavage specificities of the dual catalytic domains in chitinase from the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1. J Biol Chem 2001, 276: 35629–35635. 10.1074/jbc.M105919200View ArticlePubMedGoogle Scholar
- Tews I, Perrakis A, Oppenheim A, Dauter Z, Wilson KS, Vorgias CE: Bacterial chitobiase structure provides insight into catalytic mechanism and the basis of Tay-Sachs disease. Nat Struct Biol 1996, 3: 638–648. 10.1038/nsb0796-638View ArticlePubMedGoogle Scholar
- Ueda M, Arai M: Purification and some properties of β-N-Acetylglucosaminidase from Aeromonas sp. 10S-24. Biosci Biotechnol Biochem 1992, 56: 1204–1207. 10.1271/bbb.56.1204View ArticleGoogle Scholar
- Vetter J: Chitin content of cultivated mushrooms Agaricus bisporus, Pleurotus ostreatus and Lentinula edodes. Food Chem Volume 2007, 102: 6–9. 10.1016/j.foodchem.2006.01.037View ArticleGoogle Scholar
- Williams SJ, Mark BL, Vocadlo DJ, James MN, Withers SG: Aspartate 313 in the Streptomyces plicatus hexosaminidase plays a critical role in substrate-assisted catalysis by orienting the 2-acetamido group and stabilizing the transition state. J Biol Chem 2002, 277: 40055–40065. 10.1074/jbc.M206481200View ArticlePubMedGoogle Scholar
- Yang Q, Liu T, Liu F, Qu M, Qian X: A novel β-N-acetyl-D-hexosaminidase from the insect Ostrinia furnacalis (Guenée). FEBS J 2008, 275: 5690–5702. 10.1111/j.1742-4658.2008.06695.xView ArticlePubMedGoogle Scholar
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