Electrochemical Immunosensing Platform for DNA Methyltransferase Activity Analysis and Inhibitor Screening
In this paper, we developed a novel electrochemical method to quantify DNA methyltransferase (MTase) and analyze DNA MTase activity. After the double DNA helix structure was assembled on the surface of gold nanoparticle modified glassy carbon electrode, it was first methylated by M. SssI MTase and t...
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| Veröffentlicht in: | Analytical chemistry (Washington) 2012-11, Vol.84 (21), p.9072-9078 |
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| creator | Wang, Mo Xu, Zhenning Chen, Lijian Yin, Huanshun Ai, Shiyun |
| description | In this paper, we developed a novel electrochemical method to quantify DNA methyltransferase (MTase) and analyze DNA MTase activity. After the double DNA helix structure was assembled on the surface of gold nanoparticle modified glassy carbon electrode, it was first methylated by M. SssI MTase and then digested by restriction endonuclease HpaII, which could not recognize the methylated CpG site. Successively, anti-5-methylcytosine antibody was specifically conjugated on the CpG methylation site and horseradish peroxidase labeled goat antimouse IgG (HRP-IgG) was conjugated on anti-5-methylcytosine antibody. In the detection buffer solution containing H2O2 and hydroquinone, HRP-IgG can catalyze hydroquinone oxidation by H2O2 to generate benzoquinone, resulting in a highly electrochemical reduction signal. Consequently, the activity of M. SssI MTase was assayed, and DNA methylation was detected using the signal change with and without methylation. Furthermore, the inhibition investigation demonstrated that, in the presence of 160 μM S-adenosyl-l-methionine as methyl donor, 5-aza-2′-deoxycytidine, procaine, epicatechin, and caffeic acid could inhibit the M. SssI MTase activity with the IC50 values of 45.77, 410.3, 129.03, and 124.2 μM, respectively. Therefore, this study may provide a sensitive platform for screening DNA MTase inhibitors. |
| doi_str_mv | 10.1021/ac301620m |
| format | Article |
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After the double DNA helix structure was assembled on the surface of gold nanoparticle modified glassy carbon electrode, it was first methylated by M. SssI MTase and then digested by restriction endonuclease HpaII, which could not recognize the methylated CpG site. Successively, anti-5-methylcytosine antibody was specifically conjugated on the CpG methylation site and horseradish peroxidase labeled goat antimouse IgG (HRP-IgG) was conjugated on anti-5-methylcytosine antibody. In the detection buffer solution containing H2O2 and hydroquinone, HRP-IgG can catalyze hydroquinone oxidation by H2O2 to generate benzoquinone, resulting in a highly electrochemical reduction signal. Consequently, the activity of M. SssI MTase was assayed, and DNA methylation was detected using the signal change with and without methylation. Furthermore, the inhibition investigation demonstrated that, in the presence of 160 μM S-adenosyl-l-methionine as methyl donor, 5-aza-2′-deoxycytidine, procaine, epicatechin, and caffeic acid could inhibit the M. SssI MTase activity with the IC50 values of 45.77, 410.3, 129.03, and 124.2 μM, respectively. Therefore, this study may provide a sensitive platform for screening DNA MTase inhibitors.</description><identifier>ISSN: 0003-2700</identifier><identifier>EISSN: 1520-6882</identifier><identifier>DOI: 10.1021/ac301620m</identifier><identifier>PMID: 23030620</identifier><identifier>CODEN: ANCHAM</identifier><language>eng</language><publisher>Washington, DC: American Chemical Society</publisher><subject>Analytical chemistry ; Biosensing Techniques - methods ; Cell Line, Tumor ; Chemistry ; DNA Methylation ; DNA-Cytosine Methylases - antagonists & inhibitors ; DNA-Cytosine Methylases - metabolism ; Drug Evaluation, Preclinical ; Electrochemical methods ; Electrochemistry - methods ; Enzyme Assays - methods ; Enzyme Inhibitors - pharmacology ; Enzymes ; Exact sciences and technology ; General, instrumentation ; Humans ; Immunoassay - methods ; Immunoglobulins ; Molecular structure ; Nanoparticles</subject><ispartof>Analytical chemistry (Washington), 2012-11, Vol.84 (21), p.9072-9078</ispartof><rights>Copyright © 2012 American Chemical Society</rights><rights>2014 INIST-CNRS</rights><rights>Copyright American Chemical Society Nov 6, 2012</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-a373t-aac1b6c49b48e405148329a4a585efd5ad6da67839246f125cf6e37341f2ea063</citedby><cites>FETCH-LOGICAL-a373t-aac1b6c49b48e405148329a4a585efd5ad6da67839246f125cf6e37341f2ea063</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/ac301620m$$EPDF$$P50$$Gacs$$H</linktopdf><linktohtml>$$Uhttps://pubs.acs.org/doi/10.1021/ac301620m$$EHTML$$P50$$Gacs$$H</linktohtml><link.rule.ids>314,776,780,2752,27053,27901,27902,56713,56763</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=26620221$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/23030620$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Wang, Mo</creatorcontrib><creatorcontrib>Xu, Zhenning</creatorcontrib><creatorcontrib>Chen, Lijian</creatorcontrib><creatorcontrib>Yin, Huanshun</creatorcontrib><creatorcontrib>Ai, Shiyun</creatorcontrib><title>Electrochemical Immunosensing Platform for DNA Methyltransferase Activity Analysis and Inhibitor Screening</title><title>Analytical chemistry (Washington)</title><addtitle>Anal. Chem</addtitle><description>In this paper, we developed a novel electrochemical method to quantify DNA methyltransferase (MTase) and analyze DNA MTase activity. After the double DNA helix structure was assembled on the surface of gold nanoparticle modified glassy carbon electrode, it was first methylated by M. SssI MTase and then digested by restriction endonuclease HpaII, which could not recognize the methylated CpG site. Successively, anti-5-methylcytosine antibody was specifically conjugated on the CpG methylation site and horseradish peroxidase labeled goat antimouse IgG (HRP-IgG) was conjugated on anti-5-methylcytosine antibody. In the detection buffer solution containing H2O2 and hydroquinone, HRP-IgG can catalyze hydroquinone oxidation by H2O2 to generate benzoquinone, resulting in a highly electrochemical reduction signal. Consequently, the activity of M. SssI MTase was assayed, and DNA methylation was detected using the signal change with and without methylation. Furthermore, the inhibition investigation demonstrated that, in the presence of 160 μM S-adenosyl-l-methionine as methyl donor, 5-aza-2′-deoxycytidine, procaine, epicatechin, and caffeic acid could inhibit the M. SssI MTase activity with the IC50 values of 45.77, 410.3, 129.03, and 124.2 μM, respectively. Therefore, this study may provide a sensitive platform for screening DNA MTase inhibitors.</description><subject>Analytical chemistry</subject><subject>Biosensing Techniques - methods</subject><subject>Cell Line, Tumor</subject><subject>Chemistry</subject><subject>DNA Methylation</subject><subject>DNA-Cytosine Methylases - antagonists & inhibitors</subject><subject>DNA-Cytosine Methylases - metabolism</subject><subject>Drug Evaluation, Preclinical</subject><subject>Electrochemical methods</subject><subject>Electrochemistry - methods</subject><subject>Enzyme Assays - methods</subject><subject>Enzyme Inhibitors - pharmacology</subject><subject>Enzymes</subject><subject>Exact sciences and technology</subject><subject>General, instrumentation</subject><subject>Humans</subject><subject>Immunoassay - methods</subject><subject>Immunoglobulins</subject><subject>Molecular structure</subject><subject>Nanoparticles</subject><issn>0003-2700</issn><issn>1520-6882</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2012</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpl0VFPFDEQB_CGSOBAHvgCpokhwYfVabvb2328IMolqCTo82auO_V62e1C2zW5b2-VAwy-tC-_mWn_w9ipgPcCpPiARoHQEoY9NhOVhELXtXzFZgCgCjkHOGRHMW4AhMjugB1KBQpywYxtLnsyKYxmTYMz2PPlMEx-jOSj8z_5TY_JjmHg-eAfvy74F0rrbZ8C-mgpYCS-MMn9cmnLFx77bXSRo-_40q_dyqVcdWsCkc_NXrN9i32kk919zH58uvx-cVVcf_u8vFhcF6jmKhWIRqy0KZtVWVMJlShrJRsssaorsl2Fne5Qz2vVyFJbIStjNeXKUlhJCFods_OHvndhvJ8opnZw0VDfo6dxiq0QZQ6pqUFk-vYF3YxTyP_4q_JwOddNVu8elAljjIFsexfcgGHbCmj_LKB9WkC2b3Ydp9VA3ZN8TDyDsx3AmPO2OUnj4rPTGUkpnh2a-M-r_hv4GwWGmNc</recordid><startdate>20121106</startdate><enddate>20121106</enddate><creator>Wang, Mo</creator><creator>Xu, Zhenning</creator><creator>Chen, Lijian</creator><creator>Yin, Huanshun</creator><creator>Ai, Shiyun</creator><general>American Chemical Society</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>7QF</scope><scope>7QO</scope><scope>7QQ</scope><scope>7SC</scope><scope>7SE</scope><scope>7SP</scope><scope>7SR</scope><scope>7TA</scope><scope>7TB</scope><scope>7TM</scope><scope>7U5</scope><scope>7U7</scope><scope>7U9</scope><scope>8BQ</scope><scope>8FD</scope><scope>C1K</scope><scope>F28</scope><scope>FR3</scope><scope>H8D</scope><scope>H8G</scope><scope>H94</scope><scope>JG9</scope><scope>JQ2</scope><scope>KR7</scope><scope>L7M</scope><scope>L~C</scope><scope>L~D</scope><scope>P64</scope><scope>7X8</scope></search><sort><creationdate>20121106</creationdate><title>Electrochemical Immunosensing Platform for DNA Methyltransferase Activity Analysis and Inhibitor Screening</title><author>Wang, Mo ; Xu, Zhenning ; Chen, Lijian ; Yin, Huanshun ; Ai, Shiyun</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-a373t-aac1b6c49b48e405148329a4a585efd5ad6da67839246f125cf6e37341f2ea063</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2012</creationdate><topic>Analytical chemistry</topic><topic>Biosensing Techniques - methods</topic><topic>Cell Line, Tumor</topic><topic>Chemistry</topic><topic>DNA Methylation</topic><topic>DNA-Cytosine Methylases - antagonists & inhibitors</topic><topic>DNA-Cytosine Methylases - metabolism</topic><topic>Drug Evaluation, Preclinical</topic><topic>Electrochemical methods</topic><topic>Electrochemistry - methods</topic><topic>Enzyme Assays - methods</topic><topic>Enzyme Inhibitors - pharmacology</topic><topic>Enzymes</topic><topic>Exact sciences and technology</topic><topic>General, instrumentation</topic><topic>Humans</topic><topic>Immunoassay - methods</topic><topic>Immunoglobulins</topic><topic>Molecular structure</topic><topic>Nanoparticles</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Wang, Mo</creatorcontrib><creatorcontrib>Xu, Zhenning</creatorcontrib><creatorcontrib>Chen, Lijian</creatorcontrib><creatorcontrib>Yin, Huanshun</creatorcontrib><creatorcontrib>Ai, Shiyun</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>Aluminium Industry Abstracts</collection><collection>Biotechnology Research Abstracts</collection><collection>Ceramic Abstracts</collection><collection>Computer and Information Systems Abstracts</collection><collection>Corrosion Abstracts</collection><collection>Electronics & Communications Abstracts</collection><collection>Engineered Materials Abstracts</collection><collection>Materials Business File</collection><collection>Mechanical & Transportation Engineering Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Solid State and Superconductivity Abstracts</collection><collection>Toxicology Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>METADEX</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>ANTE: Abstracts in New Technology & Engineering</collection><collection>Engineering Research Database</collection><collection>Aerospace Database</collection><collection>Copper Technical Reference Library</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>Materials Research Database</collection><collection>ProQuest Computer Science Collection</collection><collection>Civil Engineering Abstracts</collection><collection>Advanced Technologies Database with Aerospace</collection><collection>Computer and Information Systems Abstracts Academic</collection><collection>Computer and Information Systems Abstracts Professional</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Analytical chemistry (Washington)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Wang, Mo</au><au>Xu, Zhenning</au><au>Chen, Lijian</au><au>Yin, Huanshun</au><au>Ai, Shiyun</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Electrochemical Immunosensing Platform for DNA Methyltransferase Activity Analysis and Inhibitor Screening</atitle><jtitle>Analytical chemistry (Washington)</jtitle><addtitle>Anal. Chem</addtitle><date>2012-11-06</date><risdate>2012</risdate><volume>84</volume><issue>21</issue><spage>9072</spage><epage>9078</epage><pages>9072-9078</pages><issn>0003-2700</issn><eissn>1520-6882</eissn><coden>ANCHAM</coden><abstract>In this paper, we developed a novel electrochemical method to quantify DNA methyltransferase (MTase) and analyze DNA MTase activity. After the double DNA helix structure was assembled on the surface of gold nanoparticle modified glassy carbon electrode, it was first methylated by M. SssI MTase and then digested by restriction endonuclease HpaII, which could not recognize the methylated CpG site. Successively, anti-5-methylcytosine antibody was specifically conjugated on the CpG methylation site and horseradish peroxidase labeled goat antimouse IgG (HRP-IgG) was conjugated on anti-5-methylcytosine antibody. In the detection buffer solution containing H2O2 and hydroquinone, HRP-IgG can catalyze hydroquinone oxidation by H2O2 to generate benzoquinone, resulting in a highly electrochemical reduction signal. Consequently, the activity of M. SssI MTase was assayed, and DNA methylation was detected using the signal change with and without methylation. Furthermore, the inhibition investigation demonstrated that, in the presence of 160 μM S-adenosyl-l-methionine as methyl donor, 5-aza-2′-deoxycytidine, procaine, epicatechin, and caffeic acid could inhibit the M. SssI MTase activity with the IC50 values of 45.77, 410.3, 129.03, and 124.2 μM, respectively. Therefore, this study may provide a sensitive platform for screening DNA MTase inhibitors.</abstract><cop>Washington, DC</cop><pub>American Chemical Society</pub><pmid>23030620</pmid><doi>10.1021/ac301620m</doi><tpages>7</tpages></addata></record> |
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| subjects | Analytical chemistry Biosensing Techniques - methods Cell Line, Tumor Chemistry DNA Methylation DNA-Cytosine Methylases - antagonists & inhibitors DNA-Cytosine Methylases - metabolism Drug Evaluation, Preclinical Electrochemical methods Electrochemistry - methods Enzyme Assays - methods Enzyme Inhibitors - pharmacology Enzymes Exact sciences and technology General, instrumentation Humans Immunoassay - methods Immunoglobulins Molecular structure Nanoparticles |
| title | Electrochemical Immunosensing Platform for DNA Methyltransferase Activity Analysis and Inhibitor Screening |
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