Allosteric Sensing of Fatty Acid Binding by NMR: Application to Human Serum Albumin

Human serum albumin (HSA) serves not only as a physiological oncotic pressure regulator and a ligand carrier but also as a biomarker for pathologies ranging from ischemia to diabetes. Moreover, HSA is a biopharmaceutical with a growing repertoire of putative clinical applications from hypovolemia to Alzheimer’s disease. A key determinant of the physiological, diagnostic, and therapeutic functions of HSA is the amount of long chain fatty acids (LCFAs) bound to HSA. Here, we propose to utilize ¹³C-oleic acid for the NMR-based assessment of albumin-bound LCFA concentration (CONFA). ¹³C-Oleic acid primes HSA for a LCFA-dependent allosteric transition that modulates the frequency separation between the two main ¹³C NMR peaks of HSA-bound oleic acid (Δν_AB). On the basis of Δν_AB, the overall [¹²C-LCFA]_Tot/[HSA]_Tot ratio is reproducibly estimated in a manner that is only minimally sensitive to glycation, albumin concentration, or redox potential, unlike other methods to quantify HSA-bound LCFAs such as the albumin–cobalt binding assay.