Proximity Labeling

1. Front Matter

   Pages i-x

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2. The EMARS Reaction for Proximity Labeling

   • Koichi Honke, Arisa Miyagawa-Yamaguchi, Norihiro Kotani

   Pages 1-12

3. Proteomic Proximity Labeling to Reveal Interactions Between Biomolecules

   • Johanna Susan Rees

   Pages 13-28

4. Establishing Cell Culture-Based Experimental Setups for Proximity Labeling Using Ascorbate Peroxidase (APEX)

   • David U. Mick

   Pages 29-39

5. APEX Peroxidase-Catalyzed Proximity Labeling and Multiplexed Quantitative Proteomics

   • Marian Kalocsay

   Pages 41-55

6. Identification of Lipid Droplet Proteomes by Proximity Labeling Proteomics Using APEX2

   • Kirill Bersuker, James A. Olzmann

   Pages 57-72

7. Antibody-Driven Proximity Labeling in Fixed Tissues

   • Daniel Z. Bar, Francis S. Collins

   Pages 73-81

8. BioID: A Method to Generate a History of Protein Associations

   • Danielle G. May, Kyle J. Roux

   Pages 83-95

9. Direct Identification of Biotinylated Proteins from Proximity Labeling (Spot-BioID)

   • Song-Yi Lee, Jeong Kon Seo, Hyun-Woo Rhee

   Pages 97-105

10. Identifying Synaptic Proteins by In Vivo BioID from Mouse Brain

    • Akiyoshi Uezu, Scott Soderling

    Pages 107-119

11. A Proximity Ligation-Based Method to Detect RNA-DNA Association

    • Tyler K. Fenstermaker, Guizhi Sun, Alexander Mazo, Svetlana Petruk

    Pages 121-129

12. Enzymatic Transfer of Photo-Cross-Linkers for RNA-Protein Photo-Cross-Linking at the mRNA 5′-Cap

    • Nils Muthmann, Fabian Muttach, Andrea Rentmeister

    Pages 131-146

13. Proximity-Induced Bioorthogonal Chemistry Using Inverse Electron Demand Diels-Alder Reaction

    • Jasper S. Möhler, Philipp Werther, Richard Wombacher

    Pages 147-163

14. Overcoming the Limitations of Sortase with Proximity-Based Sortase-Mediated Ligation (PBSL)

    • Hejia Henry Wang, Andrew Tsourkas

    Pages 165-177

15. Proximity-Driven Site-Specific and Covalent Labeling of Proteins with a TexasRed Fluorophore Reacting (ReacTR) Peptide Tag

    • Murat Sunbul, Andres Jäschke

    Pages 179-190

16. Affinity Conjugation for Rapid and Covalent Labeling of Proteins in Live Cells

    • Xi Chen, Fu Li, Yao-Wen Wu

    Pages 191-202

17. Ligand-Directed N-Sulfonyl Pyridone Chemistry for Selective Native Protein Labeling and Imaging in Live Cell

    • Marie Masuda, Kazuya Matsuo, Itaru Hamachi

    Pages 203-224

18. Back Matter

    Pages 225-231

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This book provides detailed protocols and untold tips and tricks regarding the most well-known examples of proximity labeling methods, in which the protein of interest is genetically fused to or labeled with an enzyme that can generate short-lived reactive species to non-specifically label molecules within a certain radius of up to twenty nanometers. Beginning with peroxidase-based proximity labeling methods, the volume continues with BioID, proximity labeling methods that describe the proximity ligation assay to detect RNA-DNA interactions, UV cross-linking to demonstrate RNA-protein interactions, and how chemical and enzymatic reactivities can be improved upon DNA-DNA and protein-protein interactions, as well as “proximity-induced self-labeling,” where the radius of labeling is zero. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. 

Authoritative and practical, Proximity Labeling: Methods and Protocols serves as an ideal guide for researchers exploring the crucial roles that proximity-driven reactions play in biological systems.

• Peroxidase-based labeling
• BioID
• Proximity ligation assay
• Biophysical properties
• Subcellular architecture
• Biomolecules
• Protein-protein interactions
• RNA-protein interactions

• Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, Heidelberg, Germany

  Murat Sunbul, Andres Jäschke

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