An electrochemical method of hydrogenase activity measurement is developed. It permits a new approach to the activation/inactivation process of the Desulfovibrio gigas hydrogenase. A monolayer of hydrogenase is grafted onto a glassy carbon electrode which is both the support of the enzyme and the detector of the activity. The physicochemical composition of the enzyme microenvironment is thus well defined and easily controlled by the electrode potential. Successive periods of inactivation and activation are applied to the same hydrogenase molecules, thus the activity can be correlated to the chronology of the experiments. We distinguish two kinds of activation/inactivation processes. The first one, already described for the enzyme stored for some months in aerobic conditions, is a slow activation by molecular hydrogen or a reducing medium (half-reaction time = 2 h). The second one is an anaerobic inactivation by an oxidizing potential. This first order inactivation (half-reaction time = 10 min) is fully reversible. This modulation of the activity level is controlled by an Ni(III)/Ni(II) redox couple (Eh = -455 mV/calomel-saturated KCl electrode at pH 8.3) involving one electron and one proton. This work proposes an explanation for the activation of the hydrogenase taking into account the participation of an [Fe-S] cluster and of the nickel atom.