Elucidation of Ionic and Electronic Charge Transport for Sensitive Electronic Characterization in Phage Films

Long range electron transport is a rare trait in nature. Using rational design on naturally non-conductive proteins, we can apply our current mechanistic understanding of this not fully understood process and learn more about it in a bottom-up approach. However, several difficulties occur in measuring very low conductive proteins, often orders of magnitude lower than natively conductive protein nanowires. More complex analyses are needed to sensitively track performance changes, and to better understand the diversity of charge transfer mechanisms involved in these heterogeneous environments.<br/>We selected M13 bacteriophages as models for detailed electronic characterization in natively non-conductive proteins, for their dual potential of tolerating mutagenesis and structural organization into nanotubes for potential charge transport. Phages insert their genetic code into bacteria, messaging the host to create multiple copies of its parasite. In the case of M13 bacteriophage, the <i>E. coli</i> host creates virus particles self-assembled into protein-based nanotubes 800 nm long and 5 nm in diameter. Along its coat, up to 2700 copies of pVIII alpha helical proteins form the majority of the rod.<br/>The multiple repeats of pVIII protein, their self-assembly into a nanorod, and the phage’s liquid crystal properties allowing it to reach high ordered supramolecular structures make the phage an intriguing candidate for inferring conductivity. This follows multiple efforts to genetically engineer more conductive protein-based self-assembling materials.<br/>Using M13 bacteriophage films as a model, we cast them on interdigitated micro electrodes to study their electrical properties. The micro features and multiple digits of the electrodes lead to enhanced geometry to amplify the current read from low conductive material. In addition, to better distinguish between and understand different electric circuit parameters, we developed a simplified Randles circuit model based on IV sweeps and transient current we observed. To then deconvolute ionic from electronic charge transport, we adapted transient current measurements with humidity control.<br/>These techniques allow us to study ionic and electronic charge transfer separately. At low relative humidity (10%) and after the current reaches steady state, few ions are mobile enough to migrate, and they reach equilibrium with the electric field. At this point, only electrons are moving through the material. Additionally, with our simplified Randles circuit model, the transient current at such a low humidity could be principally attributed to electronic parallel plate capacitance. We were able to mathematically derive different parameters from the model, such as film and contact resistance, and capacitance. As humidity increases, the transient current decay becomes dominated by ionic migration (double-layer capacitance-like). The dependence of the amplitude of the decay was used to identify the extent of ionic charge transport present in the material.<br/>This method could be potentially applied to diverse proteins. We can compare between different proteins with different conductivity enhancement strategies, or between mutants obtained from the same origin through different design approaches. This allows us to get a better mechanistic understanding of complex bioprocessed protein environments, containing salts from growth and purification, and diverse amino acid chemistry. In addition, this technique, which uses microfabricated interdigitated electrodes, has high enough sensitivity to discriminate between materials with less than two-fold differences in conductivity. Therefore, it would allow the identification of small improvements in conductivity and guide towards the evolution of more conductive proteins with potentially unexplored charge transport mechanisms.