Role of the NiFe Hydrogenase Hya in Oxidative Stress Defense in Geobacter sulfurreducens

Geobacter sulfurreducens, an Fe(III)-reducing deltaproteobacterium found in anoxic subsurface environments, contains 4 NiFe hydrogenases. Hyb, a periplasmically oriented membrane-bound NiFe hydrogenase, is essential for hydrogen-dependent growth. The functions of the three other hydrogenases are unknown.We show here that the other periplasmically oriented membrane-bound NiFe hydrogenase, Hya, is necessary for growth after exposure to oxidative stress when hydrogen or a highly limiting concentration of acetate is the electron source. The beneficial impact of Hya on growth was dependent on the presence of H2 in the atmosphere. Moreover, the Hya-deficient strain was more sensitive to the presence of superoxide or hydrogen perox-ide. Hya was also required to safeguard Hyb hydrogen oxidation activity after exposure to O2. Overexpression studies demon-strated that Hya was more resistant to oxidative stress than Hyb. Overexpression of Hya also resulted in the creation of a recom-binant strain better fitted for exposure to oxidative stress than wild-type G. sulfurreducens. These results demonstrate that one of the physiological roles of the O2-resistant Hya is to participate in the oxidative stress defense of G. sulfurreducens. Hydrogenases are ubiquitous enzymes catalyzing the reversibleinterconversion of H2 with 2H and 2e (40, 42). Hydroge-nases are separated into three classes depending on themetal com-position of their active site: NiFe hydrogenases, FeFe hydroge-nases, and Fe hydrogenases (40, 42). NiFe and FeFe hydrogenases are involved in hydrogen respiration, hydrogen production dur