Patch Fluorometry

Abstract

Patch fluorometry is a biophysical technique that combines the power of patch clamping and optical recording, with the aim of correlating local conformational rearrangements in ion channel protein with the channel gating process. This is achieved through simultaneous recordings of fluorescence and current signals from the same population of ion channels in a membrane patch. The method can be applied to studies of ion channel structure–function relationship as well as membrane protein dynamics. Unlike nuclear magnetic resonance or other biophysical methods for structural study, patch fluorometry provides real‐time dynamic structural information from ion channels in their native membrane environment whereas they carry out their functions. Recent advances greatly expand the scope of this powerful method, allowing researchers to follow structural changes in channel protein with unprecedented spatial and temporal resolutions and accuracy. Key Concepts: Ion channels achieve their cell‐sensor functions through changes in conformation in response to physical or chemical stimuli. Conformational changes in an ion channel protein do not always alter the current, and are thus invisible to patch clamp recording. Fluorescence emission is highly sensitive to the local environment of fluorophores. Site‐directed fluorophores serve as sensors for local structural changes. Patch fluorometry allows direct correlation between channel protein conformational changes and the functional states of the channel.

Sensor Devices Inspired by the Five Senses: A Review

“…Structure of TRPs.T he inner C-a nd N-termini composition change between different TRP subfamilies.A dapted from [229]. either temperature,c hemicals or pH change [230].A sf or the voltage-gated ion channels,s ome TRPs are voltage sensitive.I nt he voltage-gatedi on channels,t he change in the electric field across the membrane generated by the ion concentration is sensed by positively charged residues in the S4 transmembraned omain.T he movement of charged residues is translated into the movement of the subunits and the openingo ft he pore channel. On the other hand, most of the TRPs lack the positively charged amino acids,t hus the voltages ensitivity is lower compared to that of voltage-gated ion channels.T he voltage range sensed by TRPso ften falls beyond the physiological range ofa normal cell and the voltage response can be affectedb yothers timuli, like temperature and chemicals.…”

“…Thec oupling of aw eak voltage sensitivity and the response to other stimuli shift the voltage rangetoaphysiological range relevant to the channel activity. Them olecular basisf or the coupling of voltage-dependent and temperature-dependent sensing is still under debate [230].…”