Contribution of the active site aspartic acid to catalysis in the bacterial neuraminidase from Micromonospora viridifaciens
A recombinant D92G mutant sialidase from Micromonospora viridifaciens has been cloned, expressed and purified. Kinetic studies reveal that the replacement of the conserved aspartic acid with glycine results in a catalytically competent retaining sialidase that possesses significant activity against...
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| Veröffentlicht in: | FEBS letters 2004-11, Vol.577 (1), p.265-269 |
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| creator | Watson, Jacqueline N. Newstead, Simon Dookhun, Veedeeta Taylor, Garry Bennet, Andrew J. |
| description | A recombinant D92G mutant sialidase from
Micromonospora viridifaciens has been cloned, expressed and purified. Kinetic studies reveal that the replacement of the conserved aspartic acid with glycine results in a catalytically competent retaining sialidase that possesses significant activity against activated substrates. The contribution of this aspartate residue to the free energy of hydrolysis for natural substrates is greater than 19 kJ/mol. The three dimensional structure of the D92G mutant shows that the removal of aspartic acid 92 causes no significant re-arrangement of the active site, and that an ordered water molecule substitutes for the carboxylate group of D92. |
| doi_str_mv | 10.1016/j.febslet.2004.10.016 |
| format | Article |
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Micromonospora viridifaciens has been cloned, expressed and purified. Kinetic studies reveal that the replacement of the conserved aspartic acid with glycine results in a catalytically competent retaining sialidase that possesses significant activity against activated substrates. The contribution of this aspartate residue to the free energy of hydrolysis for natural substrates is greater than 19 kJ/mol. The three dimensional structure of the D92G mutant shows that the removal of aspartic acid 92 causes no significant re-arrangement of the active site, and that an ordered water molecule substitutes for the carboxylate group of D92.</description><identifier>ISSN: 0014-5793</identifier><identifier>EISSN: 1873-3468</identifier><identifier>DOI: 10.1016/j.febslet.2004.10.016</identifier><identifier>PMID: 15527797</identifier><language>eng</language><publisher>England: Elsevier B.V</publisher><subject>Aspartic Acid - metabolism ; Bacteria ; Base Sequence ; Binding Sites ; Carbohydrate Sequence ; Catalysis ; Catalytic mechanism ; DNA Primers ; Magnetic Resonance Spectroscopy ; Micromonospora ; Micromonospora - enzymology ; Micromonospora viridifaciens ; MU-αNeu5Ac, 4-methylumbelliferyl α-d-N-acetylneuraminide ; Mutagenesis ; Neuraminidase ; Neuraminidase - chemistry ; Neuraminidase - metabolism ; Sialidase</subject><ispartof>FEBS letters, 2004-11, Vol.577 (1), p.265-269</ispartof><rights>2004 Federation of European Biochemical Societies</rights><rights>FEBS Letters 577 (2004) 1873-3468 © 2015 Federation of European Biochemical Societies</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c4573-85218df26e2ae5afb248e29e200d65b9b8c2c00ce0b2ccfedf3c032fc868df23</citedby><cites>FETCH-LOGICAL-c4573-85218df26e2ae5afb248e29e200d65b9b8c2c00ce0b2ccfedf3c032fc868df23</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://onlinelibrary.wiley.com/doi/pdf/10.1016%2Fj.febslet.2004.10.016$$EPDF$$P50$$Gwiley$$H</linktopdf><linktohtml>$$Uhttps://www.sciencedirect.com/science/article/pii/S0014579304012463$$EHTML$$P50$$Gelsevier$$Hfree_for_read</linktohtml><link.rule.ids>314,776,780,1411,1427,3537,27901,27902,45550,45551,46384,46808,65534</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/15527797$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Watson, Jacqueline N.</creatorcontrib><creatorcontrib>Newstead, Simon</creatorcontrib><creatorcontrib>Dookhun, Veedeeta</creatorcontrib><creatorcontrib>Taylor, Garry</creatorcontrib><creatorcontrib>Bennet, Andrew J.</creatorcontrib><title>Contribution of the active site aspartic acid to catalysis in the bacterial neuraminidase from Micromonospora viridifaciens</title><title>FEBS letters</title><addtitle>FEBS Lett</addtitle><description>A recombinant D92G mutant sialidase from
Micromonospora viridifaciens has been cloned, expressed and purified. Kinetic studies reveal that the replacement of the conserved aspartic acid with glycine results in a catalytically competent retaining sialidase that possesses significant activity against activated substrates. The contribution of this aspartate residue to the free energy of hydrolysis for natural substrates is greater than 19 kJ/mol. The three dimensional structure of the D92G mutant shows that the removal of aspartic acid 92 causes no significant re-arrangement of the active site, and that an ordered water molecule substitutes for the carboxylate group of D92.</description><subject>Aspartic Acid - metabolism</subject><subject>Bacteria</subject><subject>Base Sequence</subject><subject>Binding Sites</subject><subject>Carbohydrate Sequence</subject><subject>Catalysis</subject><subject>Catalytic mechanism</subject><subject>DNA Primers</subject><subject>Magnetic Resonance Spectroscopy</subject><subject>Micromonospora</subject><subject>Micromonospora - enzymology</subject><subject>Micromonospora viridifaciens</subject><subject>MU-αNeu5Ac, 4-methylumbelliferyl α-d-N-acetylneuraminide</subject><subject>Mutagenesis</subject><subject>Neuraminidase</subject><subject>Neuraminidase - chemistry</subject><subject>Neuraminidase - metabolism</subject><subject>Sialidase</subject><issn>0014-5793</issn><issn>1873-3468</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2004</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqNkUFv1DAQha2Kql0KPwHkE7ddbCdOnBOCVUuRWnGgd8uxx2JWSbzYzlYr_jwOuxLHcrL99M0bzzxC3nG24Yw3H3cbD30aIG8EY3XRNkW9ICuu2mpd1Y16RVaM8Xot2666Jq9T2rHyVry7ItdcStG2Xbsiv7dhyhH7OWOYaPA0_wRqbMYD0IS53NPexIy2iOhoDtSabIZjwkRx-kv3BYeIZqATzNGMOKEzCaiPYaSPaMsRppD2IRp6wIgOffGCKb0hl94MCd6ezxvydHf7tL1fP3z_-m37-WFta1mGUVJw5bxoQBiQxveiViA6KHO7RvZdr6ywjFlgvbDWg_OVZZXwVjVLWXVDPpxs9zH8miFlPWKyMAxmgjAn3bSslrx0egnkXcs6JbsCyhNYZkspgtf7iKOJR82ZXtLRO31ORy_pLHJRS937c4O5H8H9qzrHUYD7E_CMAxz_z1Xf3X4RP5aol6RZzbiom6pYfTpZQdnsASHqtCzdgsMINmsX8IXf_gG5dbwx</recordid><startdate>20041105</startdate><enddate>20041105</enddate><creator>Watson, Jacqueline N.</creator><creator>Newstead, Simon</creator><creator>Dookhun, Veedeeta</creator><creator>Taylor, Garry</creator><creator>Bennet, Andrew J.</creator><general>Elsevier B.V</general><scope>6I.</scope><scope>AAFTH</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>7QL</scope><scope>7T7</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>P64</scope><scope>7X8</scope></search><sort><creationdate>20041105</creationdate><title>Contribution of the active site aspartic acid to catalysis in the bacterial neuraminidase from Micromonospora viridifaciens</title><author>Watson, Jacqueline N. ; 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| ispartof | FEBS letters, 2004-11, Vol.577 (1), p.265-269 |
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| source | MEDLINE; Wiley Online Library Journals; Elsevier ScienceDirect Journals Complete; Wiley Free Archive; Alma/SFX Local Collection; EZB Electronic Journals Library |
| subjects | Aspartic Acid - metabolism Bacteria Base Sequence Binding Sites Carbohydrate Sequence Catalysis Catalytic mechanism DNA Primers Magnetic Resonance Spectroscopy Micromonospora Micromonospora - enzymology Micromonospora viridifaciens MU-αNeu5Ac, 4-methylumbelliferyl α-d-N-acetylneuraminide Mutagenesis Neuraminidase Neuraminidase - chemistry Neuraminidase - metabolism Sialidase |
| title | Contribution of the active site aspartic acid to catalysis in the bacterial neuraminidase from Micromonospora viridifaciens |
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