Drug Target Identification Using an iTRAQ-Based Quantitative Chemical Proteomics Approach-Based on a Target Profiling Study of Andrographolide
Identifying the cellular binding targets of drugs and other bioactive small molecules is a crucial step for understanding their molecular mechanisms of action as well as potential off-target effects. The field of chemical proteomics is an emerging discipline in chemical biology using synthetic chemi...
Gespeichert in:
Veröffentlicht in: | Methods in enzymology 2017, Vol.586, p.291-309 |
---|---|
Hauptverfasser: | , , , , , , |
Format: | Artikel |
Sprache: | eng |
Schlagworte: | |
Online-Zugang: | Volltext |
Tags: |
Tag hinzufügen
Keine Tags, Fügen Sie den ersten Tag hinzu!
|
container_end_page | 309 |
---|---|
container_issue | |
container_start_page | 291 |
container_title | Methods in enzymology |
container_volume | 586 |
creator | Wang, J Wong, Y K Zhang, J Lee, Y-M Hua, Z-C Shen, H-M Lin, Q |
description | Identifying the cellular binding targets of drugs and other bioactive small molecules is a crucial step for understanding their molecular mechanisms of action as well as potential off-target effects. The field of chemical proteomics is an emerging discipline in chemical biology using synthetic chemistry and high-throughput detection techniques to study small molecule-protein interactions. In this chapter, we describe a quantitative chemical proteomics protocol combining bioorthogonal click chemistry and quantitation by isobaric tags for relative and absolute quantification (iTRAQ) to identify the specific binding targets of drugs and bioactive small molecules such as natural products. A modified drug probe with a click chemistry-enabling addition is synthesized and used in live cell treatments where it undergoes covalent interactions with its cognate cellular targets. The probes are then ligated to biotin through click chemistry and enriched with avidin beads, followed by iTRAQ labeling and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis for protein identification and relative quantitation discriminating specific targets from nonspecific binding proteins. The presented protocol has been used to successfully profile prominent drugs and natural products including andrographolide, aspirin, curcumin, etc., and can be a powerful tool to study the molecular mechanisms of bioactive small molecules. |
doi_str_mv | 10.1016/bs.mie.2016.09.049 |
format | Article |
fullrecord | <record><control><sourceid>proquest_pubme</sourceid><recordid>TN_cdi_proquest_miscellaneous_1863218547</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><sourcerecordid>1863218547</sourcerecordid><originalsourceid>FETCH-LOGICAL-c260t-a1b27d33401d0234bba86c79a0625b5cbbfdf5d1a99a3aff227d14c6d8f1db693</originalsourceid><addsrcrecordid>eNo10MlOwzAQgGELCdGyvAAH5COXBNvZnGMoW6VKUGjP0Ti2U6NsxA5SX4Jnxoj2NHP49Gs0CF1TElJC0zthw9aokPk9JHlI4vwEzWmSZEGWcz5D59Z-EsIyntMzNGOcRlmS8jn6eRinGm9grJXDS6k6Z7SpwJm-w1truhpDh83mvVgH92CVxOsJvHFefCu82KnW6wa_jb1Tvd8tLoZh7KHaHbzvwLHvlTbNX_TDTXKPe42LTo59PcKw6xsj1SU61dBYdXWYF2j79LhZvASr1-flolgFFUuJC4AKlskoigmVhEWxEMDTKsuBpCwRSSWEljqRFPIcItCaeU3jKpVcUynSPLpAt_9df-vXpKwrW2Mr1TTQqX6yJeVpxChP4szTmwOdRKtkOYymhXFfHn8Y_QL6KnUU</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>1863218547</pqid></control><display><type>article</type><title>Drug Target Identification Using an iTRAQ-Based Quantitative Chemical Proteomics Approach-Based on a Target Profiling Study of Andrographolide</title><source>MEDLINE</source><source>Elsevier ScienceDirect Journals Complete</source><creator>Wang, J ; Wong, Y K ; Zhang, J ; Lee, Y-M ; Hua, Z-C ; Shen, H-M ; Lin, Q</creator><creatorcontrib>Wang, J ; Wong, Y K ; Zhang, J ; Lee, Y-M ; Hua, Z-C ; Shen, H-M ; Lin, Q</creatorcontrib><description>Identifying the cellular binding targets of drugs and other bioactive small molecules is a crucial step for understanding their molecular mechanisms of action as well as potential off-target effects. The field of chemical proteomics is an emerging discipline in chemical biology using synthetic chemistry and high-throughput detection techniques to study small molecule-protein interactions. In this chapter, we describe a quantitative chemical proteomics protocol combining bioorthogonal click chemistry and quantitation by isobaric tags for relative and absolute quantification (iTRAQ) to identify the specific binding targets of drugs and bioactive small molecules such as natural products. A modified drug probe with a click chemistry-enabling addition is synthesized and used in live cell treatments where it undergoes covalent interactions with its cognate cellular targets. The probes are then ligated to biotin through click chemistry and enriched with avidin beads, followed by iTRAQ labeling and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis for protein identification and relative quantitation discriminating specific targets from nonspecific binding proteins. The presented protocol has been used to successfully profile prominent drugs and natural products including andrographolide, aspirin, curcumin, etc., and can be a powerful tool to study the molecular mechanisms of bioactive small molecules.</description><identifier>EISSN: 1557-7988</identifier><identifier>DOI: 10.1016/bs.mie.2016.09.049</identifier><identifier>PMID: 28137568</identifier><language>eng</language><publisher>United States</publisher><subject>Alkynes - chemistry ; Chromatography, Liquid ; Click Chemistry ; Diterpenes - chemistry ; HCT116 Cells ; Humans ; Protein Binding ; Proteome - chemistry ; Proteome - isolation & purification ; Proteomics ; Staining and Labeling ; Tandem Mass Spectrometry</subject><ispartof>Methods in enzymology, 2017, Vol.586, p.291-309</ispartof><rights>2017 Elsevier Inc. All rights reserved.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c260t-a1b27d33401d0234bba86c79a0625b5cbbfdf5d1a99a3aff227d14c6d8f1db693</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>315,781,785,4025,27928,27929,27930</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/28137568$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Wang, J</creatorcontrib><creatorcontrib>Wong, Y K</creatorcontrib><creatorcontrib>Zhang, J</creatorcontrib><creatorcontrib>Lee, Y-M</creatorcontrib><creatorcontrib>Hua, Z-C</creatorcontrib><creatorcontrib>Shen, H-M</creatorcontrib><creatorcontrib>Lin, Q</creatorcontrib><title>Drug Target Identification Using an iTRAQ-Based Quantitative Chemical Proteomics Approach-Based on a Target Profiling Study of Andrographolide</title><title>Methods in enzymology</title><addtitle>Methods Enzymol</addtitle><description>Identifying the cellular binding targets of drugs and other bioactive small molecules is a crucial step for understanding their molecular mechanisms of action as well as potential off-target effects. The field of chemical proteomics is an emerging discipline in chemical biology using synthetic chemistry and high-throughput detection techniques to study small molecule-protein interactions. In this chapter, we describe a quantitative chemical proteomics protocol combining bioorthogonal click chemistry and quantitation by isobaric tags for relative and absolute quantification (iTRAQ) to identify the specific binding targets of drugs and bioactive small molecules such as natural products. A modified drug probe with a click chemistry-enabling addition is synthesized and used in live cell treatments where it undergoes covalent interactions with its cognate cellular targets. The probes are then ligated to biotin through click chemistry and enriched with avidin beads, followed by iTRAQ labeling and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis for protein identification and relative quantitation discriminating specific targets from nonspecific binding proteins. The presented protocol has been used to successfully profile prominent drugs and natural products including andrographolide, aspirin, curcumin, etc., and can be a powerful tool to study the molecular mechanisms of bioactive small molecules.</description><subject>Alkynes - chemistry</subject><subject>Chromatography, Liquid</subject><subject>Click Chemistry</subject><subject>Diterpenes - chemistry</subject><subject>HCT116 Cells</subject><subject>Humans</subject><subject>Protein Binding</subject><subject>Proteome - chemistry</subject><subject>Proteome - isolation & purification</subject><subject>Proteomics</subject><subject>Staining and Labeling</subject><subject>Tandem Mass Spectrometry</subject><issn>1557-7988</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2017</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNo10MlOwzAQgGELCdGyvAAH5COXBNvZnGMoW6VKUGjP0Ti2U6NsxA5SX4Jnxoj2NHP49Gs0CF1TElJC0zthw9aokPk9JHlI4vwEzWmSZEGWcz5D59Z-EsIyntMzNGOcRlmS8jn6eRinGm9grJXDS6k6Z7SpwJm-w1truhpDh83mvVgH92CVxOsJvHFefCu82KnW6wa_jb1Tvd8tLoZh7KHaHbzvwLHvlTbNX_TDTXKPe42LTo59PcKw6xsj1SU61dBYdXWYF2j79LhZvASr1-flolgFFUuJC4AKlskoigmVhEWxEMDTKsuBpCwRSSWEljqRFPIcItCaeU3jKpVcUynSPLpAt_9df-vXpKwrW2Mr1TTQqX6yJeVpxChP4szTmwOdRKtkOYymhXFfHn8Y_QL6KnUU</recordid><startdate>2017</startdate><enddate>2017</enddate><creator>Wang, J</creator><creator>Wong, Y K</creator><creator>Zhang, J</creator><creator>Lee, Y-M</creator><creator>Hua, Z-C</creator><creator>Shen, H-M</creator><creator>Lin, Q</creator><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>7X8</scope></search><sort><creationdate>2017</creationdate><title>Drug Target Identification Using an iTRAQ-Based Quantitative Chemical Proteomics Approach-Based on a Target Profiling Study of Andrographolide</title><author>Wang, J ; Wong, Y K ; Zhang, J ; Lee, Y-M ; Hua, Z-C ; Shen, H-M ; Lin, Q</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c260t-a1b27d33401d0234bba86c79a0625b5cbbfdf5d1a99a3aff227d14c6d8f1db693</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2017</creationdate><topic>Alkynes - chemistry</topic><topic>Chromatography, Liquid</topic><topic>Click Chemistry</topic><topic>Diterpenes - chemistry</topic><topic>HCT116 Cells</topic><topic>Humans</topic><topic>Protein Binding</topic><topic>Proteome - chemistry</topic><topic>Proteome - isolation & purification</topic><topic>Proteomics</topic><topic>Staining and Labeling</topic><topic>Tandem Mass Spectrometry</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Wang, J</creatorcontrib><creatorcontrib>Wong, Y K</creatorcontrib><creatorcontrib>Zhang, J</creatorcontrib><creatorcontrib>Lee, Y-M</creatorcontrib><creatorcontrib>Hua, Z-C</creatorcontrib><creatorcontrib>Shen, H-M</creatorcontrib><creatorcontrib>Lin, Q</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>MEDLINE - Academic</collection><jtitle>Methods in enzymology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Wang, J</au><au>Wong, Y K</au><au>Zhang, J</au><au>Lee, Y-M</au><au>Hua, Z-C</au><au>Shen, H-M</au><au>Lin, Q</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Drug Target Identification Using an iTRAQ-Based Quantitative Chemical Proteomics Approach-Based on a Target Profiling Study of Andrographolide</atitle><jtitle>Methods in enzymology</jtitle><addtitle>Methods Enzymol</addtitle><date>2017</date><risdate>2017</risdate><volume>586</volume><spage>291</spage><epage>309</epage><pages>291-309</pages><eissn>1557-7988</eissn><abstract>Identifying the cellular binding targets of drugs and other bioactive small molecules is a crucial step for understanding their molecular mechanisms of action as well as potential off-target effects. The field of chemical proteomics is an emerging discipline in chemical biology using synthetic chemistry and high-throughput detection techniques to study small molecule-protein interactions. In this chapter, we describe a quantitative chemical proteomics protocol combining bioorthogonal click chemistry and quantitation by isobaric tags for relative and absolute quantification (iTRAQ) to identify the specific binding targets of drugs and bioactive small molecules such as natural products. A modified drug probe with a click chemistry-enabling addition is synthesized and used in live cell treatments where it undergoes covalent interactions with its cognate cellular targets. The probes are then ligated to biotin through click chemistry and enriched with avidin beads, followed by iTRAQ labeling and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis for protein identification and relative quantitation discriminating specific targets from nonspecific binding proteins. The presented protocol has been used to successfully profile prominent drugs and natural products including andrographolide, aspirin, curcumin, etc., and can be a powerful tool to study the molecular mechanisms of bioactive small molecules.</abstract><cop>United States</cop><pmid>28137568</pmid><doi>10.1016/bs.mie.2016.09.049</doi><tpages>19</tpages></addata></record> |
fulltext | fulltext |
identifier | EISSN: 1557-7988 |
ispartof | Methods in enzymology, 2017, Vol.586, p.291-309 |
issn | 1557-7988 |
language | eng |
recordid | cdi_proquest_miscellaneous_1863218547 |
source | MEDLINE; Elsevier ScienceDirect Journals Complete |
subjects | Alkynes - chemistry Chromatography, Liquid Click Chemistry Diterpenes - chemistry HCT116 Cells Humans Protein Binding Proteome - chemistry Proteome - isolation & purification Proteomics Staining and Labeling Tandem Mass Spectrometry |
title | Drug Target Identification Using an iTRAQ-Based Quantitative Chemical Proteomics Approach-Based on a Target Profiling Study of Andrographolide |
url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2024-12-10T18%3A28%3A57IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-proquest_pubme&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=Drug%20Target%20Identification%20Using%20an%20iTRAQ-Based%20Quantitative%20Chemical%20Proteomics%20Approach-Based%20on%20a%20Target%20Profiling%20Study%20of%20Andrographolide&rft.jtitle=Methods%20in%20enzymology&rft.au=Wang,%20J&rft.date=2017&rft.volume=586&rft.spage=291&rft.epage=309&rft.pages=291-309&rft.eissn=1557-7988&rft_id=info:doi/10.1016/bs.mie.2016.09.049&rft_dat=%3Cproquest_pubme%3E1863218547%3C/proquest_pubme%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=1863218547&rft_id=info:pmid/28137568&rfr_iscdi=true |