Infrared laser spectroscopy of proteins at the nanoscale

The intricate mechanisms governing protein activity, crucial in life processes and diverse applications like bioelectronics and biomedicine, necessitate experimental advancements for a comprehensive understanding. Nanosciences present innovative pathways to investigate certain unexplored aspects influencing protein functionality. One example of this is the effect of externally applied electric fields, especially on proteins naturally exposed to such fields, like membrane proteins within cell membranes. Among these, proton/ion transporters such as photosensitive microbial rhodopsins appear as relevant examples. However, experimental techniques capable of correlating the investigations of protein conformational changes with the controlled voltage potentials to which the proteins are subjected are currently lacking. To bridge this gap, here it is proposed a novel technique based on the state-of-theart AFM-IR nanospectroscopy platform. It integrates the sensitivity of infrared (IR) spectroscopy to protein conformation with electric field control, exploiting a metallic atomic force microscope (AFM) tip as both a mechanical IR detector and a nanoelectrode. Initial experiments on the prototype photosensitive protein Bacteriorhodopsin demonstrate its potential, with future perspectives aimed at exploring the effects of transmembrane electric potential on protein dynamics. In a broader sense, this innovative work stands as a significant advancement toward unraveling the intricate interplay between molecular structure and electric fields at the nanoscale.