Binding Interaction of Organofluorine–Serum Albumin: A Comparative Ligand-Detected ¹⁹F NMR Analysis

Abstract

In the present study, we attempt to characterize fluorinated ligand-serum albumin interaction in solution by a set of one-dimensional F ligand-based experiments. In this regard, a model system diflunisal (DFL)-human serum albumin (HSA) has been chosen to benchmark the utility ofF relaxation and diffusion-based experiments in deciphering ligand-protein interactions. Further, we extend the application of a similar set of F experiments to unravel the molecular interaction in an unexplored system of 2,6-difluorobenzoic acid (DFBA)-bovine serum albumin (BSA). Interaction analysis of DFBA-SA is of particular interest because DFBA is not only a stable metabolite of a number of pesticides but also used as the starting reagent of many fluorinated drugs. Observation ofF-H & H-H saturation transfer difference effects confirmed binding of the ligands to SA. Further, these ligand-protein complexes were probed in terms of the dissociation constant ( K), number of binding sites ( n), bound fraction of the ligand ( P), the complex lifetime (τ), and exchange rate ( K). Although Carr-Purcell-Meiboom-Gill (CPMG)-based transverse relaxation and diffusion analysis quantified the former three quantities, the latter two were determined by the constant time fast pulsing CPMG method. Additionally, F competition binding experiments performed with well-characterized BSA site markers and DFBA indicated nonspecific binding of DFBA to BSA, whereas similar measurements in the case of HSA with DFL and DFBA revealed superior binding interaction of DFL with SA.

Self-Assembly of Precisely Fluorinated Albumin for Dual Imaging-Guided Synergistic Chemo–Photothermal–Photodynamic Cancer Therapy

“…It is worth mentioning that EG 12 -BSA displayed much shorter 19 F T 1 (441.9 vs 630.0 ms) and much longer 19 F T 2 (6.2 ms vs too short to be measured) than the fluorinated albumin reported by Yu and coworkers, 44 which is beneficial for 19 F MRI. Because T 2 of fluorinate compounds is highly sensitive to BSA binding, 48 the neglectable T 2 changes showed no apparent interaction between the bulky fluorocarbon and BSA. Third, the aggregation of the partially fluorinated BSAs in water was studied with dynamic light scattering (DLS).…”

“…Then, Yu et al developed perfluoro- tert -butoxylated tags and selectively conjugated them to the 34-cysteine of albumin (Scheme ), which gave multiple broad 19 F NMR peaks with ultra-short transverse relaxation times, hindering their 19 F MRI application. The severe 19 F signal splitting and line broadening were probably caused by the short linkers and albumin binding of the fluorocarbon. − Therefore, precise modification of albumin with non-binding fluorocarbons through a long and flexible linker may deliver fluorinated albumin with high 19 F MRI sensitivity and preferred physicochemical properties.…”