Sequence-Dependent Mechanical and Electrochemical Properties of Gold Binding Peptide Fused to FAD-dependent Glucose Dehydrogenase in Enzyme-Electrode System

Abstract

Enzyme-based bioelectronics have been increasingly developed in the research fields such as biofuel cells, biosensors, and bioelectrosynthesis. To enhance the performance of the enzyme-electrode that is basis of enzyme-based bioelectronics, the immobilization technique of enzyme to the electrode surface is key factor. From the perspective of single enzyme on the electrode, enzyme immobilization is a factor that determines the interfacial electron transfer efficiency between enzyme and electrode. Furthermore, in the multi-enzyme system where chain reaction occurs between enzymes, the immobilization technology is a factor that determines the efficiency of mass transfer of intermediates between enzymes. To satisfy all these requirements, the immobilization method that could control the enzymatic positioning toward the desired site must be developed. In this study, the selective immobilization strategy is proposed via genetic fusion of gold binding peptides (GBP) to FAD-dependent glucose dehydrogenase (FAD-GDH) as a model enzyme. Further, binding affinity and selectivity of GBP-fused FAD-GDH was analysed on multi-materials, varying the sequence of GBPs. To compare the binding selectivity of GBPs fused to FAD-GDH, five types of peptide sequence known as GBP from other literature was selected. Then, for the preparation of synthetic FADGDH tagged with GBP candidates, plasmid construction for expression of synthetic FADGDHs was conducted and then, expression and purification of synthetic FAD-GDHs was performed. Next, the binding selectivity test was studied with target material (Au) as well as other materials (Pt, Ti, and SiO2). Then, the electrochemical performance of the synthetic enzyme is tested. This research suggests a new generation of enzyme immobilization platform technology usable to various enzyme based bioelectrochemical system.