Abstract 1050 Protein Identification and Characterization via Binding Based Proteomic Profiling (BBPP) of Fatty acid amides in Drosophila melanogaster

Fatty acid amides (FAAs) have become of particular interest since the discovery and work done on anandamide. Though this is a vast category of compounds, there has not been much work done on them. The role of these compounds endogenously has been of particular interest and as a result, we have employed the use of binding-based proteomic profiling (BBPP) to elucidate the same. Fatty acid analog probes are synthesized with a diazirine photoreactive functional group and an alkyne handle to facilitate click chemistry. These probes are used in the labeling, identification, and characterization of FAA binding proteins in Drosophila melanogaster. The diazirine group, when exposed to UV irradiation will prompt the formation of a carbene that will cause the formation of a covalent bond between the probe and the protein to which it is bound, allowing selective labeling of the proteome. The alkyne handle provided the ability to use click chemistry to attach rhodamine azide for fluorescent imaging or biotin azide for western blot and subsequent protein enrichment via the use of an avidin column. The enriched proteins can then be identified by means of high-resolution LC-MS/MS for eventual characterization. BBPP offers the opportunity to create FAA analogs of various fatty acid chains that can be used to understand the proteins they preferentially bind to and their roles biologically. Thank you to the National Institutes of Health for Funding this research project. NIH R15-GM107864.