Bio-inspired conducting peptide nanowires

Abstract

Peptides are nature’s molecular building blocks for protein self-assembly, consisting of a sequence of amino acids covalently bound through a peptide bond. By varying the quantity, position, and type of amino acids in the peptide sequence, peptides can be tailored to deliver various chemical and physical properties. Through such tailoring, peptides become useful candidates for material self-assembly in a variety of applications, such as biotechnology, drug delivery, and bioelectronics. Typically, peptides are considered as insulators (i.e., not electrically conductive); however, microbial nanowires, composed entirely of protein, in bacteria such as G. sulfurreducens have impressive electrical properties. We have designed peptides inspired by characteristic sequences of these microbial nanowires to make conductive peptide structures. This will help us to better understand the underlying mechanisms of electron transfer in complex biological structures and understand the parameters that contribute to and promote electrical conductivity in peptides.Based on the literature, the presence of aromatic residues and the physical structure and configuration of the peptides within that structure play an important role in conferring conductivity to microbial nanowires. Therefore, we designed a 21-mer negative control peptide design (without any aromatic residues), labelled L6, with a predominantly α-helical structure that self-assembles into a fibrillar structure. To introduce electrical conductivity, the aliphatic residues in the hydrophobic core of the L6 coiled coil structure were substituted with different types and combinations of selected aromatic residues (phenylalanine (Phe) and/or tryptophan (Trp)). The introduction of aromatic residues disturbed a portion of the peptide’s α-helical structure: the design containing six Trp residues, labelled W6, showed the lowest α-helical structure and resulted in non-fibrillar assemblies, while peptides containing Phe residues resulted in fibrillar assemblies. F4 and F6 (with four and six Phe residues, respectively) retained more of the α-helical structure compared to the Trp containing peptides, F4W2 and W6 (with four and six Trp residues, respectively). Current-voltage-time (I/V-t) measurements were used to measure the intrinsic conductivity of the peptide by applying a DC voltage for 100 s over a dried peptide film deposited on interdigitated microelectrode arrays with 5 μm gaps. All aromatic containing designs showed improved conductivity over the L6 control. W6 showed the highest improvement in conductivity, followed by F4W2, indicating that including Trp in the peptide design improves the peptide electrical conductivity more than Phe. This result is in agreement with the literature, reporting that Trp amino acids have more efficient electron transfer properties thanPhe amino acids. A longer peptide was designed to stabilise the secondary structure, called W4-29mer, increasing the peptide length to 29-mer. Adding another heptad repeat (based on the L6 design) and placing four Trp residues in the second and third heptad repeats resulted in a fibril-forming peptide with an α-helical structure similar to L6, overcoming the destabilising effect of Trp. The conductivity of the W4-29mer was between that of F4W2 and W6, improving conductivity while retaining the α-helical structure. However, conductivity values were insignificant (nS.cm-1 scale) compared to the highest conductive peptide reported in the literature, ACC-Hex (mS.cm-1 range). To compare the results with the literature, the 29-mer ACC-Hex peptide was tested under the same conditions used for the peptides designed here. In addition, to improve the ACC-Hex conductivity further, a new peptide was designed (ACC-HexW) with all four Phe residues in ACC-Hex substituted with Trp residues. The I/V-t results for both peptides were found to be similar to F6 and F4. Applying the same experimental procedure for assemblies of ACC-Hex in the literature still resulted in similar conductivity values, highlighting the technical challenges of replicating data in this field. Hence, a contactless method was trialled as a proof-of-concept to measure assemblies’ conductivity by terahertz (THz) spectroscopy. W6 and L6 showed a similar trend using THz spectroscopy compared to I/V-t measurement, where W6 found to be more conductive than L6.In addition to the design parameters of the peptide, the effect of external conditions such as environmental conditions and the addition of other chemical compounds are crucial to designing functional bioelectronic materials. Increasing humidity levels increased peptide conductivity while measuring the I/V-t of all 21-mer designs. This was attributed to the ability of charged polar residues to contribute to proton conduction. Trp-containing peptides showed a higher increase in conductivity relative to humidity levels compared to other peptides, likely due to the hydrogen bonding ability of the amide group in the Trp indole ring contributing to proton conduction. Addition of chemical compounds expected to alter the electron-carrying capacity of the assemblies, such as pyrene and carbon nanotubes (CNTs), were investigated on F4W2 and W6 using the established I/V-t method. While the addition of pyrene, an aromatic compound, did not improve the conductivity of F4W2 and W6, the addition of CNT improved peptide conductivity. Additionally, the peptides dispersed and debundled the CNTs which may be promising for using these peptides in composite bioelectronic and biosensor applications.