Curvature-tuning membrane mimetics as novel sensing interface to probe affinity fluctuation of bridging integrator 1...

Curvature-sensitive proteins play important roles in cellular physiology. To study their interaction properties, reliable platforms of curved membrane mimics are key but current systems are limited by the requirement for fluorescent labels, which can modify membrane conformation and activity, leading to obscured description of interaction behavior as compared to native conditions. We report here the construction and application of a robust and tunable curved membrane interface for label-free surface plasmon resonance (SPR) based probing of the interactions with bridging integrator 1 (BIN1), a BAR-domain containing protein that has been linked to cancer and autoimmune diseases. The curvature was tuned with varying vesicle size, and surface charge was controlled by doping different content of ganglioside GM1. A twentyfold enhancement of BIN1 response was observed on 50-nm vesicles as compared to the flat SLB. Replacement of GM₁ with neutral GA₁ or reducing GM₁ content from 1 % to 0.1 % in vesicles led to much lower response signal, suggesting the preference of negatively charged membranes in BIN1 binding. To demonstrate the application in complex media and for biomarker monitoring, we measured BIN1 in spiked urine samples, which showed excellent results. Small loss in signal was observed as compared to that in PBS buffer, which have been attributed to ionic compounds present in the urine sample that interfere with BIN1-membrane interactions. This work shows that the vesicle-based curved membrane proves to be an effective platform for investigating membrane-curvature sensing proteins and their biophysical interactions, and for quantifying protein markers in biological matrices by SPR.