A GFET Nitrile Sensor Using a Graphene‐Binding Fusion Protein.

A new route to single‐step and non‐covalent immobilization of proteins on graphene is exemplified with the first biosensor for nitriles based on a graphene field‐effect transistor (GFET). The biological recognition element is a fusion protein consisting of nitrile reductase QueF from Escherichia coli with an N‐terminal self‐assembling and graphene‐binding dodecapeptide. Atomic force microscopy and analysis using a quartz crystal microbalance show that both the oligopeptide and the fusion protein incorporating it form a single adlayer of monomeric enzyme on graphene. The fusion protein has a 6.3‐fold increase in binding affinity for benzyl cyanide (BnCN) versus wild‐type QueF and a 1.4‐fold increase for affinity for the enzyme's natural substrate preQ0. Density functional theory analysis of QueF's catalytic cycle with BnCN shows similar transition‐state barriers to preQ0, but differences in the formation of the initial thioimidate covalent bonding (∆G‡ = 19.0 kcal mol−1 for preQ0 vs 27.7 kcal mol−1 for BnCN) and final disassociation step (∆G = −24.3 kcal mol−1 for preQ0 vs ∆G = +4.6 kcal mol−1 for BnCN). Not only do these results offer a single‐step route to GFET modification, but they also present new opportunities in the biocatalytic synthesis of primary amines from nitriles. [ABSTRACT FROM AUTHOR]