In Situ Monitoring of Membrane Protein Electron Transfer via Surface-Enhanced Resonance Raman Spectroscopy

In situ analysis of membrane protein-ligand interactions under physiological conditions is of significance for both fundamental and applied science, but it is still a big challenge due to the limits in sensitivity and selectivity. Here, we demonstrate the potential of surface-enhanced resonance Raman spectroscopy (SERRS) for the investigation of membrane protein-protein interactions. Lipid biolayers are successfully coated on silver nanoparticles through electrostatic interactions, and a highly sensitive and biomimetic membrane platform is obtained in vitro. Self-assembly and immobilization of the reduced cytochrome b(5) on the coated membrane are achieved and protein native biological functions are preserved. Owing to resonance effect, the Raman fingerprint of the immobilized cytochrome b(5) redox center is selectively enhanced, allowing for in situ and real-time monitoring of the electron transfer process between cytochrome b(5) and their partners, cytochrome c and myoglobin. This study provides a sensitive analytical approach for membrane proteins and paves the way for in situ exploration of their structural basis and functions.