Preparation of Protein Lysates Using Biorthogonal Chemical Reporters for Click Reaction and in-Gel Fluorescence Analysis
Bioorthogonal chemical reporters are non-native chemical handles introduced into biomolecules of living systems, typically through metabolic or protein engineering. These functionalities can undergo bioorthogonal reactions, such as copper-catalyzed alkyne-azide cycloaddition (CuAAC), with small-mole...
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Bio-protocol LLC
2024-11-01
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| Series: | Bio-Protocol |
| Online Access: | https://bio-protocol.org/en/bpdetail?id=5114&type=0 |
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| author | Yaxin Xu Tao Peng |
| author_facet | Yaxin Xu Tao Peng |
| author_sort | Yaxin Xu |
| collection | DOAJ |
| description | Bioorthogonal chemical reporters are non-native chemical handles introduced into biomolecules of living systems, typically through metabolic or protein engineering. These functionalities can undergo bioorthogonal reactions, such as copper-catalyzed alkyne-azide cycloaddition (CuAAC), with small-molecule probes, enabling the tagging and visualization of biomolecules. This approach has greatly enhanced our understanding of cellular dynamics, enzyme targeting, and protein post-translational modifications. Herein, we report a protocol for preparing protein lysates for click reaction and in-gel fluorescence analysis, exemplified using alk-16, a terminal alkyne-functionalized stearic acid analog, to investigate proteins with fatty acylation. This protocol provides methods for the fluorescent visualization of chemical reporter–labeled proteomes or individual proteins of interest (POIs). |
| format | Article |
| id | doaj-art-1b93fe603b51448483741fb86e4e39e8 |
| institution | Kabale University |
| issn | 2331-8325 |
| language | English |
| publishDate | 2024-11-01 |
| publisher | Bio-protocol LLC |
| record_format | Article |
| series | Bio-Protocol |
| spelling | doaj-art-1b93fe603b51448483741fb86e4e39e82024-12-26T05:17:17ZengBio-protocol LLCBio-Protocol2331-83252024-11-01142210.21769/BioProtoc.5114Preparation of Protein Lysates Using Biorthogonal Chemical Reporters for Click Reaction and in-Gel Fluorescence AnalysisYaxin Xu0Tao Peng1State Key Laboratory of Chemical Oncogenomics, School of Chemical Biology and Biotechnology, Peking University Shenzhen Graduate School, Shenzhen, ChinaState Key Laboratory of Chemical Oncogenomics, School of Chemical Biology and Biotechnology, Peking University Shenzhen Graduate School, Shenzhen, ChinaBioorthogonal chemical reporters are non-native chemical handles introduced into biomolecules of living systems, typically through metabolic or protein engineering. These functionalities can undergo bioorthogonal reactions, such as copper-catalyzed alkyne-azide cycloaddition (CuAAC), with small-molecule probes, enabling the tagging and visualization of biomolecules. This approach has greatly enhanced our understanding of cellular dynamics, enzyme targeting, and protein post-translational modifications. Herein, we report a protocol for preparing protein lysates for click reaction and in-gel fluorescence analysis, exemplified using alk-16, a terminal alkyne-functionalized stearic acid analog, to investigate proteins with fatty acylation. This protocol provides methods for the fluorescent visualization of chemical reporter–labeled proteomes or individual proteins of interest (POIs).https://bio-protocol.org/en/bpdetail?id=5114&type=0 |
| spellingShingle | Yaxin Xu Tao Peng Preparation of Protein Lysates Using Biorthogonal Chemical Reporters for Click Reaction and in-Gel Fluorescence Analysis Bio-Protocol |
| title | Preparation of Protein Lysates Using Biorthogonal Chemical Reporters for Click Reaction and in-Gel Fluorescence Analysis |
| title_full | Preparation of Protein Lysates Using Biorthogonal Chemical Reporters for Click Reaction and in-Gel Fluorescence Analysis |
| title_fullStr | Preparation of Protein Lysates Using Biorthogonal Chemical Reporters for Click Reaction and in-Gel Fluorescence Analysis |
| title_full_unstemmed | Preparation of Protein Lysates Using Biorthogonal Chemical Reporters for Click Reaction and in-Gel Fluorescence Analysis |
| title_short | Preparation of Protein Lysates Using Biorthogonal Chemical Reporters for Click Reaction and in-Gel Fluorescence Analysis |
| title_sort | preparation of protein lysates using biorthogonal chemical reporters for click reaction and in gel fluorescence analysis |
| url | https://bio-protocol.org/en/bpdetail?id=5114&type=0 |
| work_keys_str_mv | AT yaxinxu preparationofproteinlysatesusingbiorthogonalchemicalreportersforclickreactionandingelfluorescenceanalysis AT taopeng preparationofproteinlysatesusingbiorthogonalchemicalreportersforclickreactionandingelfluorescenceanalysis |