Important role of fungal intracellular laccase for melanin synthesis: purification and characterization of an intracellular laccase from Lentinula edodes fruit bodies
Iwate Biotechnology Research Center, 22-174-4 Narita, Kitakami, Iwate 024-0003, Japan Correspondence Masaru Nagai nagai{at}ibrc.or.jp A laccase (EC 1.10.3.2) was isolated from the fully browned gills of Lentinula edodes fruit bodies. The enzyme was purified to a homogeneous preparation using hydroph...
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| Veröffentlicht in: | Microbiology (Society for General Microbiology) 2003-09, Vol.149 (9), p.2455-2462 |
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| creator | Nagai, Masaru Kawata, Maki Watanabe, Hisayuki Ogawa, Machiko Saito, Kumiko Takesawa, Toshikazu Kanda, Katsuhiro Sato, Toshitsugu |
| description | Iwate Biotechnology Research Center, 22-174-4 Narita, Kitakami, Iwate 024-0003, Japan
Correspondence Masaru Nagai nagai{at}ibrc.or.jp
A laccase (EC 1.10.3.2) was isolated from the fully browned gills of Lentinula edodes fruit bodies. The enzyme was purified to a homogeneous preparation using hydrophobic, cation-exchange and size-exclusion chromatography. SDS-PAGE analysis showed the purified laccase, Lcc 2, to be a monomeric protein of 58·0 kDa. The enzyme had an isoelectric point of around pH 6·9. The optimum pH for enzyme activity was around 3·0 against 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt (ABTS), and it was most active at 40 °C and stable up to 50 °C. The enzyme contained 8·6 % carbohydrate and some copper atoms. The enzyme oxidized ABTS, p -phenylenediamine, pyrogallol, guaiacol, 2,6-dimethoxyphenol, catechol and ferulic acid, but not veratryl alcohol and tyrosine. -(3,4-Dihydroxyphenyl)alanine ( L -DOPA), which was not oxidized by a laccase previously reported from the culture filtrate of L. edodes , was also oxidized by Lcc 2, and the oxidative product of L -dopa was identified as L -DOPA quinone by HPLC analysis. Lcc 2 was able to oxidize phenolic compounds extracted from fresh gills to brown-coloured products, suggesting a role for laccase in melanin synthesis in this strain.
Abbreviations: ABTS, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt; DHN, dihydroxynaphthalene; L -DOPA, -(3,4-dihydroxyphenyl)alanine; GDHB, -glutaminyl-3,4-dihydroxybenzene; GHB, -glutaminyl-4-hydroxybenzene; Lcc, laccase; Tyr, tyrosinase; PB, 10mM sodium phosphate buffer |
| doi_str_mv | 10.1099/mic.0.26414-0 |
| format | Article |
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Correspondence Masaru Nagai nagai{at}ibrc.or.jp
A laccase (EC 1.10.3.2) was isolated from the fully browned gills of Lentinula edodes fruit bodies. The enzyme was purified to a homogeneous preparation using hydrophobic, cation-exchange and size-exclusion chromatography. SDS-PAGE analysis showed the purified laccase, Lcc 2, to be a monomeric protein of 58·0 kDa. The enzyme had an isoelectric point of around pH 6·9. The optimum pH for enzyme activity was around 3·0 against 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt (ABTS), and it was most active at 40 °C and stable up to 50 °C. The enzyme contained 8·6 % carbohydrate and some copper atoms. The enzyme oxidized ABTS, p -phenylenediamine, pyrogallol, guaiacol, 2,6-dimethoxyphenol, catechol and ferulic acid, but not veratryl alcohol and tyrosine. -(3,4-Dihydroxyphenyl)alanine ( L -DOPA), which was not oxidized by a laccase previously reported from the culture filtrate of L. edodes , was also oxidized by Lcc 2, and the oxidative product of L -dopa was identified as L -DOPA quinone by HPLC analysis. Lcc 2 was able to oxidize phenolic compounds extracted from fresh gills to brown-coloured products, suggesting a role for laccase in melanin synthesis in this strain.
Abbreviations: ABTS, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt; DHN, dihydroxynaphthalene; L -DOPA, -(3,4-dihydroxyphenyl)alanine; GDHB, -glutaminyl-3,4-dihydroxybenzene; GHB, -glutaminyl-4-hydroxybenzene; Lcc, laccase; Tyr, tyrosinase; PB, 10mM sodium phosphate buffer</description><identifier>ISSN: 1350-0872</identifier><identifier>EISSN: 1465-2080</identifier><identifier>DOI: 10.1099/mic.0.26414-0</identifier><identifier>PMID: 12949171</identifier><language>eng</language><publisher>Reading: Soc General Microbiol</publisher><subject>Biological and medical sciences ; Fundamental and applied biological sciences. Psychology ; Growth, nutrition, metabolism, transports, enzymes. Molecular biology ; Hydrogen-Ion Concentration ; Laccase - isolation & purification ; Laccase - metabolism ; Lentinula edodes ; Melanins - biosynthesis ; Microbiology ; Mycology ; Oxidoreductases - antagonists & inhibitors ; Oxidoreductases - isolation & purification ; Oxidoreductases - metabolism ; Shiitake Mushrooms - enzymology ; Substrate Specificity</subject><ispartof>Microbiology (Society for General Microbiology), 2003-09, Vol.149 (9), p.2455-2462</ispartof><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c450t-4cb9f4e81eb8ce345633612bfb3a22b9421f1113170d746d205069e7db7f82493</citedby><cites>FETCH-LOGICAL-c450t-4cb9f4e81eb8ce345633612bfb3a22b9421f1113170d746d205069e7db7f82493</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,27901,27902</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=15364629$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/12949171$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Nagai, Masaru</creatorcontrib><creatorcontrib>Kawata, Maki</creatorcontrib><creatorcontrib>Watanabe, Hisayuki</creatorcontrib><creatorcontrib>Ogawa, Machiko</creatorcontrib><creatorcontrib>Saito, Kumiko</creatorcontrib><creatorcontrib>Takesawa, Toshikazu</creatorcontrib><creatorcontrib>Kanda, Katsuhiro</creatorcontrib><creatorcontrib>Sato, Toshitsugu</creatorcontrib><title>Important role of fungal intracellular laccase for melanin synthesis: purification and characterization of an intracellular laccase from Lentinula edodes fruit bodies</title><title>Microbiology (Society for General Microbiology)</title><addtitle>Microbiology</addtitle><description>Iwate Biotechnology Research Center, 22-174-4 Narita, Kitakami, Iwate 024-0003, Japan
Correspondence Masaru Nagai nagai{at}ibrc.or.jp
A laccase (EC 1.10.3.2) was isolated from the fully browned gills of Lentinula edodes fruit bodies. The enzyme was purified to a homogeneous preparation using hydrophobic, cation-exchange and size-exclusion chromatography. SDS-PAGE analysis showed the purified laccase, Lcc 2, to be a monomeric protein of 58·0 kDa. The enzyme had an isoelectric point of around pH 6·9. The optimum pH for enzyme activity was around 3·0 against 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt (ABTS), and it was most active at 40 °C and stable up to 50 °C. The enzyme contained 8·6 % carbohydrate and some copper atoms. The enzyme oxidized ABTS, p -phenylenediamine, pyrogallol, guaiacol, 2,6-dimethoxyphenol, catechol and ferulic acid, but not veratryl alcohol and tyrosine. -(3,4-Dihydroxyphenyl)alanine ( L -DOPA), which was not oxidized by a laccase previously reported from the culture filtrate of L. edodes , was also oxidized by Lcc 2, and the oxidative product of L -dopa was identified as L -DOPA quinone by HPLC analysis. Lcc 2 was able to oxidize phenolic compounds extracted from fresh gills to brown-coloured products, suggesting a role for laccase in melanin synthesis in this strain.
Abbreviations: ABTS, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt; DHN, dihydroxynaphthalene; L -DOPA, -(3,4-dihydroxyphenyl)alanine; GDHB, -glutaminyl-3,4-dihydroxybenzene; GHB, -glutaminyl-4-hydroxybenzene; Lcc, laccase; Tyr, tyrosinase; PB, 10mM sodium phosphate buffer</description><subject>Biological and medical sciences</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Growth, nutrition, metabolism, transports, enzymes. Molecular biology</subject><subject>Hydrogen-Ion Concentration</subject><subject>Laccase - isolation & purification</subject><subject>Laccase - metabolism</subject><subject>Lentinula edodes</subject><subject>Melanins - biosynthesis</subject><subject>Microbiology</subject><subject>Mycology</subject><subject>Oxidoreductases - antagonists & inhibitors</subject><subject>Oxidoreductases - isolation & purification</subject><subject>Oxidoreductases - metabolism</subject><subject>Shiitake Mushrooms - enzymology</subject><subject>Substrate Specificity</subject><issn>1350-0872</issn><issn>1465-2080</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2003</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkU2LFDEQhoMo7rp69Cq5KOyhx1SS_oi3ZfFjYcCLnkM6XZmJdCdjkmZZf5C_04wzsCfxlPDWU29V8RLyGtgGmFLvF283bMM7CbJhT8glyK5tOBvY0_oXLWvY0PML8iLnH4zVIoPn5AK4kgp6uCS_75ZDTMWEQlOckUZH3Rp2ZqY-lGQszvM6m0RnY63JSF1MdMHZBB9ofghlj9nnD_SwJu-8NcXHQE2YqN2b2l0w-V8nsRqb8C_TFBe6xVB8qDrFKU6Yq7r6Qsc4ecwvyTNn5oyvzu8V-f7p47fbL8326-e725ttY2XLSiPtqJzEAXAcLArZdkJ0wEc3CsP5qCQHBwACejb1sps4a1mnsJ_G3g1cKnFF3p18Dyn-XDEXvfh83NcEjGvWfbWDvh3-C8IwKAGir2BzAm2KOSd0-pD8YtKDBqaPCdZGq5n-m6BmlX9zNl7HBadH-hxZBd6eAZOtmV0ywfr8yLWikx0_nnJ94vZ-t7_3CfUOQ52V4ujjcShIpZXmsm3FH_zgtdQ</recordid><startdate>20030901</startdate><enddate>20030901</enddate><creator>Nagai, Masaru</creator><creator>Kawata, Maki</creator><creator>Watanabe, Hisayuki</creator><creator>Ogawa, Machiko</creator><creator>Saito, Kumiko</creator><creator>Takesawa, Toshikazu</creator><creator>Kanda, Katsuhiro</creator><creator>Sato, Toshitsugu</creator><general>Soc General Microbiol</general><general>Society for General Microbiology</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>M7N</scope><scope>7X8</scope></search><sort><creationdate>20030901</creationdate><title>Important role of fungal intracellular laccase for melanin synthesis: purification and characterization of an intracellular laccase from Lentinula edodes fruit bodies</title><author>Nagai, Masaru ; Kawata, Maki ; Watanabe, Hisayuki ; Ogawa, Machiko ; Saito, Kumiko ; Takesawa, Toshikazu ; Kanda, Katsuhiro ; Sato, Toshitsugu</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c450t-4cb9f4e81eb8ce345633612bfb3a22b9421f1113170d746d205069e7db7f82493</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2003</creationdate><topic>Biological and medical sciences</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Growth, nutrition, metabolism, transports, enzymes. Molecular biology</topic><topic>Hydrogen-Ion Concentration</topic><topic>Laccase - isolation & purification</topic><topic>Laccase - metabolism</topic><topic>Lentinula edodes</topic><topic>Melanins - biosynthesis</topic><topic>Microbiology</topic><topic>Mycology</topic><topic>Oxidoreductases - antagonists & inhibitors</topic><topic>Oxidoreductases - isolation & purification</topic><topic>Oxidoreductases - metabolism</topic><topic>Shiitake Mushrooms - enzymology</topic><topic>Substrate Specificity</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Nagai, Masaru</creatorcontrib><creatorcontrib>Kawata, Maki</creatorcontrib><creatorcontrib>Watanabe, Hisayuki</creatorcontrib><creatorcontrib>Ogawa, Machiko</creatorcontrib><creatorcontrib>Saito, Kumiko</creatorcontrib><creatorcontrib>Takesawa, Toshikazu</creatorcontrib><creatorcontrib>Kanda, Katsuhiro</creatorcontrib><creatorcontrib>Sato, Toshitsugu</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>MEDLINE - Academic</collection><jtitle>Microbiology (Society for General Microbiology)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Nagai, Masaru</au><au>Kawata, Maki</au><au>Watanabe, Hisayuki</au><au>Ogawa, Machiko</au><au>Saito, Kumiko</au><au>Takesawa, Toshikazu</au><au>Kanda, Katsuhiro</au><au>Sato, Toshitsugu</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Important role of fungal intracellular laccase for melanin synthesis: purification and characterization of an intracellular laccase from Lentinula edodes fruit bodies</atitle><jtitle>Microbiology (Society for General Microbiology)</jtitle><addtitle>Microbiology</addtitle><date>2003-09-01</date><risdate>2003</risdate><volume>149</volume><issue>9</issue><spage>2455</spage><epage>2462</epage><pages>2455-2462</pages><issn>1350-0872</issn><eissn>1465-2080</eissn><abstract>Iwate Biotechnology Research Center, 22-174-4 Narita, Kitakami, Iwate 024-0003, Japan
Correspondence Masaru Nagai nagai{at}ibrc.or.jp
A laccase (EC 1.10.3.2) was isolated from the fully browned gills of Lentinula edodes fruit bodies. The enzyme was purified to a homogeneous preparation using hydrophobic, cation-exchange and size-exclusion chromatography. SDS-PAGE analysis showed the purified laccase, Lcc 2, to be a monomeric protein of 58·0 kDa. The enzyme had an isoelectric point of around pH 6·9. The optimum pH for enzyme activity was around 3·0 against 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt (ABTS), and it was most active at 40 °C and stable up to 50 °C. The enzyme contained 8·6 % carbohydrate and some copper atoms. The enzyme oxidized ABTS, p -phenylenediamine, pyrogallol, guaiacol, 2,6-dimethoxyphenol, catechol and ferulic acid, but not veratryl alcohol and tyrosine. -(3,4-Dihydroxyphenyl)alanine ( L -DOPA), which was not oxidized by a laccase previously reported from the culture filtrate of L. edodes , was also oxidized by Lcc 2, and the oxidative product of L -dopa was identified as L -DOPA quinone by HPLC analysis. Lcc 2 was able to oxidize phenolic compounds extracted from fresh gills to brown-coloured products, suggesting a role for laccase in melanin synthesis in this strain.
Abbreviations: ABTS, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt; DHN, dihydroxynaphthalene; L -DOPA, -(3,4-dihydroxyphenyl)alanine; GDHB, -glutaminyl-3,4-dihydroxybenzene; GHB, -glutaminyl-4-hydroxybenzene; Lcc, laccase; Tyr, tyrosinase; PB, 10mM sodium phosphate buffer</abstract><cop>Reading</cop><pub>Soc General Microbiol</pub><pmid>12949171</pmid><doi>10.1099/mic.0.26414-0</doi><tpages>8</tpages></addata></record> |
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| subjects | Biological and medical sciences Fundamental and applied biological sciences. Psychology Growth, nutrition, metabolism, transports, enzymes. Molecular biology Hydrogen-Ion Concentration Laccase - isolation & purification Laccase - metabolism Lentinula edodes Melanins - biosynthesis Microbiology Mycology Oxidoreductases - antagonists & inhibitors Oxidoreductases - isolation & purification Oxidoreductases - metabolism Shiitake Mushrooms - enzymology Substrate Specificity |
| title | Important role of fungal intracellular laccase for melanin synthesis: purification and characterization of an intracellular laccase from Lentinula edodes fruit bodies |
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