Electrochemical Activity of Molecular Hybrids: Self-Assembly of (XH)4 Peptides and Hemin on Graphite Electrodes

The development of artificial enzymes has gained interest in electrochemical biosensors because natural enzymes have a relatively short lifetime and high cost for practical applications. Peptides with a small number of amino acids compared with proteins can be synthesized and chemically stable, thus they can be a good candidate as an artificial enzyme. Short-chain peptides that self-assemble into fiber-like structures in liquid are known to be catalytically active by forming complex structures with cofactors [1]. These catalytically-active peptides often contain histidine, which plays a role in stabilizing the structure and coordinating with cofactors at the imidazole group on the side chain, enhancing the catalytic activity. In our previous work, we found that catalytic peptides containing histidine exhibited their self-assembly on graphite surfaces and revealed electrocatalytic activity combined with the coenzyme hemin [2]. More recent work showed that newly-designed peptides (XH peptides) consisting of a repeating structure of two amino acids with histidine and another amino acid, “X”, also exhibited self-assembly on graphite surfaces. These peptides are a useful tool for understanding the role of the counter amino acid X on their surface self-assembly and in tuning the hydrophobicity of the graphite surface [3].<br/>However, the capability of this XH peptide to immobilize cofactor hemins has not yet been investigated. In this study, we studied the interaction between hemin and the (XH)4 peptide on the surface of a graphite electrode by atomic force microscopy (AFM). Three counter amino acids, tyrosine (Y), valine (V), and leucine (L), were employed as counter amino acids X for this study, e.g. YHYHYHYH. AFM showed that each peptide formed uniform linear structures with a height of a few nanometers. They also showed a hexagonal symmetry on the graphite surface, which is commensurate with the lattice structures of graphite. When the cofactor hemin was added, dot-like structures with a height of 0.5 nm~1 nm appeared on the peptide nanowires. The peptide with the highest number of particles on the peptide and the smallest particle size was LH4, followed by VH4 and YH4. Assuming that the particles on the wire were hemins, the XH peptide can be a suitable scaffold for immobilizing hemin. We also performed the electrochemical measurement to evaluate the hydrogen peroxide reduction capacity of the prepared XH peptide-hemin electrodes, and it was found that these peptides with a simple sequence of XH have enzymatic activity. The observations implied that the activity is closely related to the interactions with the hemin and peptides regarding the surface density of the immobilized hemins and their structural stability.<br/><br/>[1] Zozulia. et al. Chem Soc Rev, 2018, 47(10), 3621-3639.<br/>[2] Wei Luo. et al. Nanoscale, 2022, 14, 8326-8331.<br/>[3] Wei Luo. et al. Langmuir 2023, 39, 20, 7057–7062