Dynamics of Pseudomonas aeruginosa azurin and its Cys3Ser mutant at single crystal gold surfaces investigated by cyclic voltammetry and atomic force microscopy

Cyclic voltammetry of Pseudomonas aeruginosa azurin on polycrystalline gold is reversible (E0=360mV vs she;50mM ammonium acetate) but the voltammetric signals decay with time constants of about 3x10-3 s-1. No signal is observed for monocrystalline Au(111). Cys3Ser azurin is electrochemically inactive on either type of gold electrode but shows a reversible although decaying peak (362mV, 50mM ammonium acetate; decay time constant ~ 2x10-3 s-1) on edge-plane pyrolytic graphite.Ex situ and in situ atomic force microscopy (AFM) of the azurins on Au(111) show initially arrays of protein structures of lateral 100-200Å and vertical ~ 50Å extension. These could be individual molecular images convoluted with the tip curvature. As scanning proceeds the structures in the ex situ mode collect into large two-dimensional assemblies while the adsorbed protein in the in situ mode is largely swept into the solution, recovering the free Au(111) surface. The cyclic voltammetry and AFM data are consistent with time dependent adsorption of the azurins on gold via the disulphide bridge (wild-type) or free thiol group (Cys3Ser mutant).