EVIDENCE FOR A POTENTIAL-DEPENDENT REVERSIBLE INACTIVATION OF UREASE ADSORBED ON A GOLD ELECTRODE

The potentiometric collection mode of the scanning electrochemical mic roscope has been applied to a kinetic study of changes in immobilised enzyme activity. Urease adsorbs spontaneously onto gold surfaces from aqueous solution and the immobilised enzyme was shown to retain activi ty as a catalyst for the hydrolysis of urea to ammonium and bicarbonat e ions. Urease activity was monitored using an ion-selective potentiom etric tip electrode to detect a product (NH4+) of the enzymatic reacti on and the use of a 50 mu m radius gold electrode to immobilise the en zyme results in a steady state diffusion field for the reaction produc ts and enables quantitative interpretation of the data. The ammonium i on flux determined from the tip potential decreased by approximately 4 0% when the potential of the gold was scanned or stepped across the ra nge from 0.0 to + 0.4 V vs. Ag/AgCl. The decrease in flux is reversibl e and the original value almost completely recovered after the potenti al was returned to 0 V. The flux decrease is attributed to inactivatio n of the enzyme. The timescale of the inactivation is limited by the e stablishment of a steady state concentration profile of ammonium ions at the 50 pm radius source. The kinetics of the reactivation are signi ficantly slower with an apparent first order rate constant of 0.022 s( -1). No faradaic processes were observed at the gold electrode over th e potential range from 0.0 to +0.4 V. We therefore suggest that change s in the enzyme quaternary structure due to the changing electric fiel d and ionic composition at the interface are responsible.