A new chemical methodology to detect enzyme activity at nanomolar concentrations is described. The sensing mechanism relies on the use of a catalytic Au MPC, which highly efficiently cleaves HPNPP causing the release of p-nitrophenolate, a yellow reporter molecule. Catalysis is completely inhibited in the presence of low micromolar concentrations of biologically important oligoanions, such as ATP, ADP or an Asp-tripeptide, which bind with high affinities to the Au MPC surface. The change in free energy of binding, ΔG, increases in a perfect linear manner as a function of the number of charges present in the analyte. The enzymatic hydrolysis of an Asp-tripeptide turns on the catalytic production of p-nitrophenolate by the catalytic Au MPC, since the products of hydrolysis have a lower affinity for the Au MPC surface. The cascade of the two catalytic processes causes a strong signal amplification allowing the detection of nanomolar enzyme concentrations.

Detection of Enzyme Activity through Catalytic Signal Amplification with Functionalized Gold Nanoparticles

BONOMI, RENATO;CAZZOLARO, ALESSANDRO;SANSONE, ANNA;SCRIMIN, PAOLO MARIA;PRINS, LEONARD JAN
2011

Abstract

A new chemical methodology to detect enzyme activity at nanomolar concentrations is described. The sensing mechanism relies on the use of a catalytic Au MPC, which highly efficiently cleaves HPNPP causing the release of p-nitrophenolate, a yellow reporter molecule. Catalysis is completely inhibited in the presence of low micromolar concentrations of biologically important oligoanions, such as ATP, ADP or an Asp-tripeptide, which bind with high affinities to the Au MPC surface. The change in free energy of binding, ΔG, increases in a perfect linear manner as a function of the number of charges present in the analyte. The enzymatic hydrolysis of an Asp-tripeptide turns on the catalytic production of p-nitrophenolate by the catalytic Au MPC, since the products of hydrolysis have a lower affinity for the Au MPC surface. The cascade of the two catalytic processes causes a strong signal amplification allowing the detection of nanomolar enzyme concentrations.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11577/2480689
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