The small fungal protein hydrophobin from Pisolithus tinctorius (HYDPt-1) has been used to modify electrode surfaces by self-assembly from aqueous solutions. Voltammetric experiments performed with Fe(CN)63- as an electrochemical probe have shown that hydrophobin can be immobilized on hydrophobic surfaces of a glassy carbon electrode (GCE), or on a thin mercury film electrode (TMFE), or on hydrophilic surfaces such as a gold electrode (GE). Higher efficiency of coverage is obtained for GCE or TMFE. The hydrophobin-modified electrodes were in turn functionalized with the electroactive ubiquinone (Q10), quinone (Q0), and azobenzene. While Q10 was adsorbed on the surface of the hydrophobin layer, smaller molecules, azobenzene and Q0, penetrated into the pores of the layer. In all cases, a stable attachment of the electroactive reagents to the electrode via the hydrophobin was achieved.