Control Of Protein Conformation And Orientation On Graphene

Graphene-based biosensors have attracted considerable attention due to their advantages of label-free detection and high sensitivity. Many such biosensors utilize noncovalent van der Waals force to attach proteins onto graphene surface while preserving graphene's high conductivity. Maintaining the protein structure without denaturation/substantial conformational change and controlling proper protein orientation on the graphene surface are critical for biosensing applications of these biosensors fabricated with proteins on graphene. Based on the knowledge we obtained from our previous experimental study and computer modeling of amino acid residual level interactions between graphene and peptides, here we systemically redesigned an important protein for better conformational stability and desirable orientation on graphene. In this paper, immunoglobulin G (IgG) antibody-binding domain of protein G (protein GB1) was studied to demonstrate how we can preserve the protein native structure and control the protein orientation on graphene surface by redesigning protein mutants. Various experimental tools including sum frequency generation vibrational spectroscopy, attenuated total refection-Fourier transform infrared spectroscopy, fluorescence spectroscopy, and circular dichroism spectroscopy were used to study the protein GB1 structure on graphene, supplemented by molecular dynamics simulations. By carefully designing the protein GB1 mutant, we can avoid strong unfavorable interactions between protein and graphene to preserve protein conformation and to enable the protein to adopt a preferred orientation. The methodology developed in this study is general and can be applied to study different proteins on graphene and beyond. With the knowledge obtained from this research, one could apply this method to optimize protein function on surfaces (e.g., to enhance biosensor sensitivity).