Biochemical Characterization of the First Fungal Glycoside Hydrolyase Family 3 β‑N‑Acetylglucosaminidase from Rhizomucor miehei
A novel β-N-acetylglucosaminidase gene (RmNag) from Rhizomucor miehei was cloned and expressed in Escherichia coli. RmNag shares the highest identity of 37% with a putative β-N-acetylglucosaminidase from Aspergillus clavatus. The recombinant enzyme was purified to homogeneity. The optimal pH and tem...
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| Veröffentlicht in: | Journal of agricultural and food chemistry 2014-06, Vol.62 (22), p.5181-5190 |
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| creator | Yang, Shaoqing Song, Shuang Yan, Qiaojuan Fu, Xing Jiang, Zhengqiang Yang, Xinbin |
| description | A novel β-N-acetylglucosaminidase gene (RmNag) from Rhizomucor miehei was cloned and expressed in Escherichia coli. RmNag shares the highest identity of 37% with a putative β-N-acetylglucosaminidase from Aspergillus clavatus. The recombinant enzyme was purified to homogeneity. The optimal pH and temperature of RmNag were pH 6.5 and 50 °C, respectively. It was stable in the pH range 6.0–8.0 and at temperatures below 45 °C. RmNag exhibited strict substrate specificity for p-nitrophenyl β-N-acetylglucosaminide (pNP-GlcNAc) and N-acetyl chitooligosaccharides. The apparent K m of RmNag toward pNP-GlcNAc was 0.13 mM. The purified enzyme displayed an exo-type manner as it released the only end product of GlcNAc from all the tested N-acetyl chitooligosaccharides. Besides, RmNag exhibited relatively high N-acetyl-β-d-glucosaminide tolerance with an inhibition constant K i value of 9.68 mM. The excellent properties may give the enzyme great potential in industries. This is the first report on a glycoside hydrolyase family 3 β-N-acetylglucosaminidase from a fungus. |
| doi_str_mv | 10.1021/jf500912b |
| format | Article |
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RmNag shares the highest identity of 37% with a putative β-N-acetylglucosaminidase from Aspergillus clavatus. The recombinant enzyme was purified to homogeneity. The optimal pH and temperature of RmNag were pH 6.5 and 50 °C, respectively. It was stable in the pH range 6.0–8.0 and at temperatures below 45 °C. RmNag exhibited strict substrate specificity for p-nitrophenyl β-N-acetylglucosaminide (pNP-GlcNAc) and N-acetyl chitooligosaccharides. The apparent K m of RmNag toward pNP-GlcNAc was 0.13 mM. The purified enzyme displayed an exo-type manner as it released the only end product of GlcNAc from all the tested N-acetyl chitooligosaccharides. Besides, RmNag exhibited relatively high N-acetyl-β-d-glucosaminide tolerance with an inhibition constant K i value of 9.68 mM. The excellent properties may give the enzyme great potential in industries. This is the first report on a glycoside hydrolyase family 3 β-N-acetylglucosaminidase from a fungus.</description><identifier>ISSN: 0021-8561</identifier><identifier>EISSN: 1520-5118</identifier><identifier>DOI: 10.1021/jf500912b</identifier><identifier>PMID: 24811866</identifier><language>eng</language><publisher>United States: American Chemical Society</publisher><subject>Acetylglucosaminidase - chemistry ; Acetylglucosaminidase - genetics ; Acetylglucosaminidase - metabolism ; Amino Acid Sequence ; Aspergillus clavatus ; Base Sequence ; chitooligosaccharides ; Cloning, Molecular ; Enzyme Stability ; Escherichia coli ; Fungal Proteins - chemistry ; Fungal Proteins - genetics ; Fungal Proteins - metabolism ; fungi ; genes ; Glycoside Hydrolases - chemistry ; Glycoside Hydrolases - genetics ; Glycoside Hydrolases - metabolism ; glycosides ; industry ; Kinetics ; Molecular Sequence Data ; Rhizomucor - chemistry ; Rhizomucor - enzymology ; Rhizomucor - genetics ; Rhizomucor miehei ; Sequence Alignment ; Substrate Specificity ; temperature</subject><ispartof>Journal of agricultural and food chemistry, 2014-06, Vol.62 (22), p.5181-5190</ispartof><rights>Copyright © 2014 American Chemical Society</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-a339t-6dc195e4d44102550cdecf603f59aa9f70f4ad3cdbd6c8fb12bc6d9b66bdc32b3</citedby><cites>FETCH-LOGICAL-a339t-6dc195e4d44102550cdecf603f59aa9f70f4ad3cdbd6c8fb12bc6d9b66bdc32b3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://pubs.acs.org/doi/pdf/10.1021/jf500912b$$EPDF$$P50$$Gacs$$H</linktopdf><linktohtml>$$Uhttps://pubs.acs.org/doi/10.1021/jf500912b$$EHTML$$P50$$Gacs$$H</linktohtml><link.rule.ids>314,780,784,2765,27076,27924,27925,56738,56788</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/24811866$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Yang, Shaoqing</creatorcontrib><creatorcontrib>Song, Shuang</creatorcontrib><creatorcontrib>Yan, Qiaojuan</creatorcontrib><creatorcontrib>Fu, Xing</creatorcontrib><creatorcontrib>Jiang, Zhengqiang</creatorcontrib><creatorcontrib>Yang, Xinbin</creatorcontrib><title>Biochemical Characterization of the First Fungal Glycoside Hydrolyase Family 3 β‑N‑Acetylglucosaminidase from Rhizomucor miehei</title><title>Journal of agricultural and food chemistry</title><addtitle>J. Agric. Food Chem</addtitle><description>A novel β-N-acetylglucosaminidase gene (RmNag) from Rhizomucor miehei was cloned and expressed in Escherichia coli. RmNag shares the highest identity of 37% with a putative β-N-acetylglucosaminidase from Aspergillus clavatus. The recombinant enzyme was purified to homogeneity. The optimal pH and temperature of RmNag were pH 6.5 and 50 °C, respectively. It was stable in the pH range 6.0–8.0 and at temperatures below 45 °C. RmNag exhibited strict substrate specificity for p-nitrophenyl β-N-acetylglucosaminide (pNP-GlcNAc) and N-acetyl chitooligosaccharides. The apparent K m of RmNag toward pNP-GlcNAc was 0.13 mM. The purified enzyme displayed an exo-type manner as it released the only end product of GlcNAc from all the tested N-acetyl chitooligosaccharides. Besides, RmNag exhibited relatively high N-acetyl-β-d-glucosaminide tolerance with an inhibition constant K i value of 9.68 mM. The excellent properties may give the enzyme great potential in industries. This is the first report on a glycoside hydrolyase family 3 β-N-acetylglucosaminidase from a fungus.</description><subject>Acetylglucosaminidase - chemistry</subject><subject>Acetylglucosaminidase - genetics</subject><subject>Acetylglucosaminidase - metabolism</subject><subject>Amino Acid Sequence</subject><subject>Aspergillus clavatus</subject><subject>Base Sequence</subject><subject>chitooligosaccharides</subject><subject>Cloning, Molecular</subject><subject>Enzyme Stability</subject><subject>Escherichia coli</subject><subject>Fungal Proteins - chemistry</subject><subject>Fungal Proteins - genetics</subject><subject>Fungal Proteins - metabolism</subject><subject>fungi</subject><subject>genes</subject><subject>Glycoside Hydrolases - chemistry</subject><subject>Glycoside Hydrolases - genetics</subject><subject>Glycoside Hydrolases - metabolism</subject><subject>glycosides</subject><subject>industry</subject><subject>Kinetics</subject><subject>Molecular Sequence Data</subject><subject>Rhizomucor - chemistry</subject><subject>Rhizomucor - enzymology</subject><subject>Rhizomucor - genetics</subject><subject>Rhizomucor miehei</subject><subject>Sequence Alignment</subject><subject>Substrate Specificity</subject><subject>temperature</subject><issn>0021-8561</issn><issn>1520-5118</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2014</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpt0E9u1DAUBnCrArVDYdELFG8qwSLlOYk98bIdMS1SRSWga8vxn4lHTlzsZJGuuuACXIWDcAhOgkdTumJhvcX303vyh9AJgXMCJfmwtRSAk7I9QAtCSygoIc0LtIAcFg1l5Ai9SmkLAA1dwiE6KusmC8YW6MelC6ozvVPS41Uno1Sjie5Bji4MOFg8dgavXUwjXk_DJqMrP6uQnDb4etYx-FmmLGTv_Iwr_PvXn8efn_O7UGac_cZPGedwcHrnbAw9_tK5h9DnIOLemc641-illT6ZN0_zGN2tP35bXRc3t1efVhc3hawqPhZMK8KpqXVd529TCkobZRlUlnIpuV2CraWulG41U41tcyGKad4y1mpVlW11jN7t997H8H0yaRS9S8p4LwcTpiQIrUpOYVnzTN_vqYohpWisuI-ul3EWBMSudPFceranT2untjf6Wf5rOYO3e2BlEHITXRJ3X0sgDIAQ3sDu3NleSJXENkxxyD3859Rf30qXOQ</recordid><startdate>20140604</startdate><enddate>20140604</enddate><creator>Yang, Shaoqing</creator><creator>Song, Shuang</creator><creator>Yan, Qiaojuan</creator><creator>Fu, Xing</creator><creator>Jiang, Zhengqiang</creator><creator>Yang, Xinbin</creator><general>American Chemical Society</general><general>American Chemical Society, Books and Journals Division</general><scope>FBQ</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>7X8</scope></search><sort><creationdate>20140604</creationdate><title>Biochemical Characterization of the First Fungal Glycoside Hydrolyase Family 3 β‑N‑Acetylglucosaminidase from Rhizomucor miehei</title><author>Yang, Shaoqing ; Song, Shuang ; Yan, Qiaojuan ; Fu, Xing ; Jiang, Zhengqiang ; Yang, Xinbin</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-a339t-6dc195e4d44102550cdecf603f59aa9f70f4ad3cdbd6c8fb12bc6d9b66bdc32b3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2014</creationdate><topic>Acetylglucosaminidase - chemistry</topic><topic>Acetylglucosaminidase - genetics</topic><topic>Acetylglucosaminidase - metabolism</topic><topic>Amino Acid Sequence</topic><topic>Aspergillus clavatus</topic><topic>Base Sequence</topic><topic>chitooligosaccharides</topic><topic>Cloning, Molecular</topic><topic>Enzyme Stability</topic><topic>Escherichia coli</topic><topic>Fungal Proteins - chemistry</topic><topic>Fungal Proteins - genetics</topic><topic>Fungal Proteins - metabolism</topic><topic>fungi</topic><topic>genes</topic><topic>Glycoside Hydrolases - chemistry</topic><topic>Glycoside Hydrolases - genetics</topic><topic>Glycoside Hydrolases - metabolism</topic><topic>glycosides</topic><topic>industry</topic><topic>Kinetics</topic><topic>Molecular Sequence Data</topic><topic>Rhizomucor - chemistry</topic><topic>Rhizomucor - enzymology</topic><topic>Rhizomucor - genetics</topic><topic>Rhizomucor miehei</topic><topic>Sequence Alignment</topic><topic>Substrate Specificity</topic><topic>temperature</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Yang, Shaoqing</creatorcontrib><creatorcontrib>Song, Shuang</creatorcontrib><creatorcontrib>Yan, Qiaojuan</creatorcontrib><creatorcontrib>Fu, Xing</creatorcontrib><creatorcontrib>Jiang, Zhengqiang</creatorcontrib><creatorcontrib>Yang, Xinbin</creatorcontrib><collection>AGRIS</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Journal of agricultural and food chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Yang, Shaoqing</au><au>Song, Shuang</au><au>Yan, Qiaojuan</au><au>Fu, Xing</au><au>Jiang, Zhengqiang</au><au>Yang, Xinbin</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Biochemical Characterization of the First Fungal Glycoside Hydrolyase Family 3 β‑N‑Acetylglucosaminidase from Rhizomucor miehei</atitle><jtitle>Journal of agricultural and food chemistry</jtitle><addtitle>J. Agric. Food Chem</addtitle><date>2014-06-04</date><risdate>2014</risdate><volume>62</volume><issue>22</issue><spage>5181</spage><epage>5190</epage><pages>5181-5190</pages><issn>0021-8561</issn><eissn>1520-5118</eissn><abstract>A novel β-N-acetylglucosaminidase gene (RmNag) from Rhizomucor miehei was cloned and expressed in Escherichia coli. RmNag shares the highest identity of 37% with a putative β-N-acetylglucosaminidase from Aspergillus clavatus. The recombinant enzyme was purified to homogeneity. The optimal pH and temperature of RmNag were pH 6.5 and 50 °C, respectively. It was stable in the pH range 6.0–8.0 and at temperatures below 45 °C. RmNag exhibited strict substrate specificity for p-nitrophenyl β-N-acetylglucosaminide (pNP-GlcNAc) and N-acetyl chitooligosaccharides. The apparent K m of RmNag toward pNP-GlcNAc was 0.13 mM. The purified enzyme displayed an exo-type manner as it released the only end product of GlcNAc from all the tested N-acetyl chitooligosaccharides. Besides, RmNag exhibited relatively high N-acetyl-β-d-glucosaminide tolerance with an inhibition constant K i value of 9.68 mM. The excellent properties may give the enzyme great potential in industries. This is the first report on a glycoside hydrolyase family 3 β-N-acetylglucosaminidase from a fungus.</abstract><cop>United States</cop><pub>American Chemical Society</pub><pmid>24811866</pmid><doi>10.1021/jf500912b</doi><tpages>10</tpages></addata></record> |
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| subjects | Acetylglucosaminidase - chemistry Acetylglucosaminidase - genetics Acetylglucosaminidase - metabolism Amino Acid Sequence Aspergillus clavatus Base Sequence chitooligosaccharides Cloning, Molecular Enzyme Stability Escherichia coli Fungal Proteins - chemistry Fungal Proteins - genetics Fungal Proteins - metabolism fungi genes Glycoside Hydrolases - chemistry Glycoside Hydrolases - genetics Glycoside Hydrolases - metabolism glycosides industry Kinetics Molecular Sequence Data Rhizomucor - chemistry Rhizomucor - enzymology Rhizomucor - genetics Rhizomucor miehei Sequence Alignment Substrate Specificity temperature |
| title | Biochemical Characterization of the First Fungal Glycoside Hydrolyase Family 3 β‑N‑Acetylglucosaminidase from Rhizomucor miehei |
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