A new radioactive cross-linking reagent for studying the interactions of proteins.
We have developed a photoactivable, heterobifunctional, reversible, radioactively labeled, chemical cross-linking reagent, 3-[(2-nitro-4-azidophenyl)-2-aminoethyldithio]-N-succinimidyl propionate, for studying the interactions of proteins in situ. When reacted in the dark with a purified protein, it forms a covalent derivative which can be purified and reconstituted into biological systems. This derivative will form cross-links to neighboring macromolecules only upon photolysis; reduction cleaves the cross-link and transfers the radiolabel to the second molecule, which can then be identified by standard techniques. We have tested the cross-linker using the binding of gelatin to fibronectin. The cross-linker gives the proper chemical behavior under biological conditions, reacts with high yield and with a very low level of nonspecific cross-linking, and can be used to identify protein-protein and other interactions at the cell surface and elsewhere. The advantages, limitations and possible uses for this reagent are discussed.
D001386:Azides; D003432:Cross-Linking Reagents / Q000138:chemical synthesis*; D005353:Fibronectins; D005780:Gelatin; D007202:Indicators and Reagents; D008722:Methods; D010782:Photolysis; D011506:Proteins; D012685:Sepharose; D013329:Structure-Activity Relationship; D013388:Succinimides / Q000138:chemical synthesis*
D001386:Azides; D003432:Cross-Linking Reagents; D005353:Fibronectins; D007202:Indicators and Reagents; D011506:Proteins; D013388:Succinimides; C033692:3-((2-nitro-4-azidophenyl)-2-aminoethyldithio)-N-succinimidyl propionate; D005780:Gelatin; D012685:Sepharose