▎ 摘　　要

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 preQ(0). Density functional theory analysis of QueF's catalytic cycle with BnCN shows similar transition-state barriers to preQ(0), but differences in the formation of the initial thioimidate covalent bonding ( increment G(double dagger) = 19.0 kcal mol(-1) for preQ(0) vs 27.7 kcal mol(-1) for BnCN) and final disassociation step ( increment G = -24.3 kcal mol(-1) for preQ(0) vs increment 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.