Miriam Huerta1,Barituziga Banuna1,Ju-Chen Chia1,Xenofon Strakosas2,Marios Savvakis2,Daniel Simon2,Olena Vatamaniuk1,Susan Daniel1
Cornell University1,Linköping University2
Miriam Huerta1,Barituziga Banuna1,Ju-Chen Chia1,Xenofon Strakosas2,Marios Savvakis2,Daniel Simon2,Olena Vatamaniuk1,Susan Daniel1
Cornell University1,Linköping University2
Organic electrochemical transistors (OECT) based on the conductive polymer PEDOT:PSS have been used in different organisms to study and analyze different biological responses. The main advantages of the OECTs are the combination of high quality electrical signals and optical monitoring due to the transparency of PEDOT:PSS thin films. We decided to use these properties to study plant membrane proteins because, even though plant transport mechanisms have been studied for many years, the study of individual proteins is still a big challenge. One of these proteins is the copper transporter 1 (COPT1), which maintains the plants' copper homeostasis, but its function has not been totally characterized. Therefore, we formed a hybrid supported lipid bilayer (SLB) with membranes from transient GFP and GFP-COPT1-transfected <i>Arabidopsis thaliana</i> mesophyll protoplasts and fusogenic liposomes on top of PEDOT:PSS. SLB formation and fluidity were determined by fluorescence recovery after photobleaching following the diffusion coefficient calculation. Both samples were full photobleaching recovered and had similar diffusion coefficients, confirming the high quality of SLBs. Next, we combined SLBs with OECTs, and using electrochemical impedance spectroscopy (EIS) measurements, we showed that plant SLB formed a layer that blocks the passage of ions from the electrolyte into the conductive polymer, increasing the system's impedance. We used an equivalent electrical circuit modeling to extract membrane resistance (R<sub>m</sub>) from the raw impedance. Then, we tested these SLBs with different CuSO<sub>4</sub> concentrations ranging from 0-100 µM. We observed a decrease in the R<sub>m</sub> in both cases, but it was higher in those overexpressing COPT1, indicating that in the presence of copper, COPT1 transporter is open. Our platform can be used to analyze profoundly COPT1 transporter mechanism and adapted to investigate more membrane proteins involved in transport mechanisms in plants.