Chemoproteomic Profiling of Isoprenoid Pyrophosphate Interacting Proteins with Photoaffinity Probes

Isoprenoid pyrophosphates are key intermediates in the biosynthesis of many biologically important molecules and in the prenylation of proteins involved in various biological processes. Except for these well-studied functions, new roles of isoprenoid pyrophosphates have been reported continuously. To study their functions on the proteome scale, we developed a series of isoprenoid pyrophosphate photoaffinity probes for the covalent labeling of their interacting proteins. Through “click” reaction with the alkyne handle on the probes, fluorophores could be attached to the labeled proteins for the gel-based fluorescence analysis or cell imaging. A biotin moiety could be used instead to enrich labeled protein. Combining with SILAC-based quantitative proteomic analysis, several known and putative interacting proteins for farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP) were identified, including many nucleocytoplasmic transport proteins. We have validated the interaction between transportin-1 with FPP and GGPP in vitro, suggesting potential new function of FPP and GGPP in protein transportation. This probe-labeling method could be further applied for evaluating other isoprenoid interacting proteins and exploring their cellular functions and localization, which will greatly facilitate biochemical research of isoprenoids.