HTRF-Based Assay for Microsomal Prostaglandin E2 Synthase-1 Activity
Microsomal prostaglandin E2 synthase-1 (mPGES-1) catalyzes the formation of prostaglandin E2 (PGE2) from the endoperoxide prostaglandin H 2 (PGH2). Expression of this enzyme is induced during the inflammatory response, and mouse knockout experiments suggest it may be an attractive target for antiart...
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| Veröffentlicht in: | Journal of biomolecular screening 2008-08, Vol.13 (7), p.619-625 |
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| container_title | Journal of biomolecular screening |
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| creator | Goedken, Eric R. Gagnon, Andrew I. Overmeyer, Gary T. Liu, Junjian Petrillo, Richard A. Burchat, Andrew F. Tomlinson, Medha J. |
| description | Microsomal prostaglandin E2 synthase-1 (mPGES-1) catalyzes the formation of prostaglandin E2 (PGE2) from the endoperoxide prostaglandin H 2 (PGH2). Expression of this enzyme is induced during the inflammatory response, and mouse knockout experiments suggest it may be an attractive target for antiarthritic therapies. Assaying the activity of this enzyme in vitro is challenging because of the unstable nature of the PGH 2 substrate. Here, the authors present an mPGES-1 activity assay suitable for characterization of enzyme preparations and for determining the potency of inhibitor compounds. This plate-based competition assay uses homogenous time-resolved fluorescence to measure PGE2 produced by the enzyme. The assay is insensitive to DMSO concentration up to 10% and does not require extensive washes after the initial enzyme reaction is concluded, making it a simple and convenient way to assess mPGES-1 inhibition. (Journal of Biomolecular Screening 2008:619-625) |
| doi_str_mv | 10.1177/1087057108321145 |
| format | Article |
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Expression of this enzyme is induced during the inflammatory response, and mouse knockout experiments suggest it may be an attractive target for antiarthritic therapies. Assaying the activity of this enzyme in vitro is challenging because of the unstable nature of the PGH 2 substrate. Here, the authors present an mPGES-1 activity assay suitable for characterization of enzyme preparations and for determining the potency of inhibitor compounds. This plate-based competition assay uses homogenous time-resolved fluorescence to measure PGE2 produced by the enzyme. The assay is insensitive to DMSO concentration up to 10% and does not require extensive washes after the initial enzyme reaction is concluded, making it a simple and convenient way to assess mPGES-1 inhibition. 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Expression of this enzyme is induced during the inflammatory response, and mouse knockout experiments suggest it may be an attractive target for antiarthritic therapies. Assaying the activity of this enzyme in vitro is challenging because of the unstable nature of the PGH 2 substrate. Here, the authors present an mPGES-1 activity assay suitable for characterization of enzyme preparations and for determining the potency of inhibitor compounds. This plate-based competition assay uses homogenous time-resolved fluorescence to measure PGE2 produced by the enzyme. The assay is insensitive to DMSO concentration up to 10% and does not require extensive washes after the initial enzyme reaction is concluded, making it a simple and convenient way to assess mPGES-1 inhibition. (Journal of Biomolecular Screening 2008:619-625)</description><subject>Animals</subject><subject>Arthritis - drug therapy</subject><subject>Baculoviridae - metabolism</subject><subject>Binding, Competitive</subject><subject>Gene Expression Regulation, Enzymologic</subject><subject>Humans</subject><subject>Inflammation</subject><subject>Inhibitory Concentration 50</subject><subject>Insecta</subject><subject>Intramolecular Oxidoreductases - metabolism</subject><subject>Microsomes - enzymology</subject><subject>Microsomes - metabolism</subject><subject>Peroxides - metabolism</subject><subject>Prostaglandin H2 - metabolism</subject><subject>Prostaglandin-E Synthases</subject><subject>Spectrometry, Fluorescence - methods</subject><subject>Time Factors</subject><issn>1087-0571</issn><issn>2472-5552</issn><issn>1552-454X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2008</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp1UMtKAzEUDaLYWt27kvxANHeSTNJlra0VKopWcDfkWae0M2UyFebvTWlBEFzdA-fBPQeha6C3AFLeAVWSCpkOywC4OEF9ECIjXPDP04QTTfZ8D13EuKIUWE75OeqByrMcgPXRw2zxNiX3OnqHRzHqDoe6wc-lbepYb_QavybQ6uVaV66s8CTD713VfiU9ATyybfldtt0lOgt6Hf3V8Q7Qx3SyGM_I_OXxaTyaE8uEEkRoNlRWMhmMlJniwRjDTaZy4aWzTnGvhQs2MJqncnporBHBcOtyD1ZKzwaIHnL338XGh2LblBvddAXQYj9I8XeQZLk5WLY7s_Hu13BcIAnIQRD10heretdUqcL_gT_Yr2dp</recordid><startdate>200808</startdate><enddate>200808</enddate><creator>Goedken, Eric R.</creator><creator>Gagnon, Andrew I.</creator><creator>Overmeyer, Gary T.</creator><creator>Liu, Junjian</creator><creator>Petrillo, Richard A.</creator><creator>Burchat, Andrew F.</creator><creator>Tomlinson, Medha J.</creator><general>SAGE Publications</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope></search><sort><creationdate>200808</creationdate><title>HTRF-Based Assay for Microsomal Prostaglandin E2 Synthase-1 Activity</title><author>Goedken, Eric R. ; Gagnon, Andrew I. ; Overmeyer, Gary T. ; Liu, Junjian ; Petrillo, Richard A. ; Burchat, Andrew F. ; Tomlinson, Medha J.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c3585-5a398c737fb77284fbbb4b2865e7dcd84ea5dfcf306177a9bcb5fb4cd6e1c77e3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2008</creationdate><topic>Animals</topic><topic>Arthritis - drug therapy</topic><topic>Baculoviridae - metabolism</topic><topic>Binding, Competitive</topic><topic>Gene Expression Regulation, Enzymologic</topic><topic>Humans</topic><topic>Inflammation</topic><topic>Inhibitory Concentration 50</topic><topic>Insecta</topic><topic>Intramolecular Oxidoreductases - metabolism</topic><topic>Microsomes - enzymology</topic><topic>Microsomes - metabolism</topic><topic>Peroxides - metabolism</topic><topic>Prostaglandin H2 - metabolism</topic><topic>Prostaglandin-E Synthases</topic><topic>Spectrometry, Fluorescence - methods</topic><topic>Time Factors</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Goedken, Eric R.</creatorcontrib><creatorcontrib>Gagnon, Andrew I.</creatorcontrib><creatorcontrib>Overmeyer, Gary T.</creatorcontrib><creatorcontrib>Liu, Junjian</creatorcontrib><creatorcontrib>Petrillo, Richard A.</creatorcontrib><creatorcontrib>Burchat, Andrew F.</creatorcontrib><creatorcontrib>Tomlinson, Medha J.</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><jtitle>Journal of biomolecular screening</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Goedken, Eric R.</au><au>Gagnon, Andrew I.</au><au>Overmeyer, Gary T.</au><au>Liu, Junjian</au><au>Petrillo, Richard A.</au><au>Burchat, Andrew F.</au><au>Tomlinson, Medha J.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>HTRF-Based Assay for Microsomal Prostaglandin E2 Synthase-1 Activity</atitle><jtitle>Journal of biomolecular screening</jtitle><addtitle>J Biomol Screen</addtitle><date>2008-08</date><risdate>2008</risdate><volume>13</volume><issue>7</issue><spage>619</spage><epage>625</epage><pages>619-625</pages><issn>1087-0571</issn><issn>2472-5552</issn><eissn>1552-454X</eissn><abstract>Microsomal prostaglandin E2 synthase-1 (mPGES-1) catalyzes the formation of prostaglandin E2 (PGE2) from the endoperoxide prostaglandin H 2 (PGH2). Expression of this enzyme is induced during the inflammatory response, and mouse knockout experiments suggest it may be an attractive target for antiarthritic therapies. Assaying the activity of this enzyme in vitro is challenging because of the unstable nature of the PGH 2 substrate. Here, the authors present an mPGES-1 activity assay suitable for characterization of enzyme preparations and for determining the potency of inhibitor compounds. This plate-based competition assay uses homogenous time-resolved fluorescence to measure PGE2 produced by the enzyme. The assay is insensitive to DMSO concentration up to 10% and does not require extensive washes after the initial enzyme reaction is concluded, making it a simple and convenient way to assess mPGES-1 inhibition. 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| source | MEDLINE; Alma/SFX Local Collection |
| subjects | Animals Arthritis - drug therapy Baculoviridae - metabolism Binding, Competitive Gene Expression Regulation, Enzymologic Humans Inflammation Inhibitory Concentration 50 Insecta Intramolecular Oxidoreductases - metabolism Microsomes - enzymology Microsomes - metabolism Peroxides - metabolism Prostaglandin H2 - metabolism Prostaglandin-E Synthases Spectrometry, Fluorescence - methods Time Factors |
| title | HTRF-Based Assay for Microsomal Prostaglandin E2 Synthase-1 Activity |
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