Dipeptidyl peptidase-IV activity assay and inhibitor screening using a gold nanoparticle-modified gold electrode with an immobilized enzyme substrate

We report on an electrochemical biosensor for the determination of the activity of dipeptidyl peptidase-IV (DPP-IV), and on a method for screening the effect of its inhibitors. An enzyme substrate (Fc-peptide) was immobilized on the surface of a gold electrode, and double signal amplification was accomplished via an additional layer consisting of phenyl rings and gold nanoparticles. The activity of DPP-IV was determined at levels as low as 39 nU·mL⁻¹ and over a linear detection range as wide as from 0.5 μU·mL⁻¹ to 2.5 mU·mL⁻¹. The inhibitory effects of diprotin A and the His-Leu dipeptide on the activity of DPP-IV also were tested and gave IC₅₀ values of 93.5 and 95.5 μM, respectively. The assay is rapid, precise and selective. It may be extended to other peptidases and, possibly, proteases and their inhibitors. (author)