Chemistry for the analysis of protein-protein interactions: Rapid and efficient cross-linking triggered by long wavelength light

摘要

Chemical cross-linking is a potentially useful technique for probing the architecture of multiprotein com- plexes. However, analyses using typical bifunctional cross-linkers often suffer from poor yields, and large-scale modification of nucleophilic side chains can result in artifactual results attrib- utable to structural destabilization. We report here the de novo design and development of a type of protein cross-linking reac- tion that uses a photogenerated oxidant to mediate rapid and efficient cross-linking of associated proteins. The process in- volves brief photolysis of tris-bipyridylruthenium(II) dication with visible light in the presence of the electron acceptor am- monium persulfate and the proteins of interest. Very high yields of cross-linked products can be obtained with irradiation times of <1 second. This chemistry obviates many of the problems associated with standard cross-liuking reagents.