Proteomic quantification of carrier proteins for different free fatty acids in mouse plasma

Free fatty acids (FFAs) are hydrophobic, thus requiring binding to carrier proteins to dissolve in plasma. Our recent study indicated that plasma FFAs may be transported by a diverse array of proteins, based on results from a pulldown assay using palmitic acid-conjugated agarose. However, there was a technical limitation that plasma proteins were isolated based on their interaction with a single FFA species, while a broad range of FFAs circulate in vivo. Although palmitic acid is a major FFA, a study using a variety of FFA classes is needed to understand if FFA carrier proteins have specificity for beads reacted with certain fatty acid classes. In the present study, we used beads coated by covalent linkage with palmitic acid, oleic acid, linoleic acid, or α-linolenic acid to isolate plasma proteins that can bind to each FFA. Proteomics identified 231 mouse plasma proteins that were pulled down by FFA-coated beads. The overall proteome characteristics were generally similar between the FFA species, with strong correlations between protein abundances isolated by each FFA. Only 35 proteins showed different relative abundances between bead types, mostly (33 proteins) found to be lower in α-linolenic acid compared to other FFAs; the total abundance of these proteins only accounted for 0.22-0.87 % of the isolated proteins, indicating that >99 % of the FFA carrier proteome was captured similarly by the different bead types. In conclusion, plasma proteins isolated by one FFA species in a pulldown assay may reasonably depict the collection of FFA carrier proteins in plasma.