Hydrogen metabolism in aerobic hydrogen-oxidizing bacteria

Abstract

A survey on organisms able to use molecular hydrogen as electron donor in the energy-yielding process is presented. In the group of the aerobic hydrogen-oxidizing bacteria so far two types of hydrogenases have been encountered, a NAD-reducing, soluble enzyme (H2 : NAD oxidoreductase) and a membrane-bound enzyme unable to reduce pyridine nucleotides. With respect to the distribution of both types of hydrogenases three groups of hydrogen-oxidizing bacteria can be diffentiated containing (i) both types (Alcaligenes eutrophus), (ii) a soluble enzyme only (Nocardia opaca lb), and (iii) a membrane-bound hydrogenase only (majority of genera and species). The results of studies on the NAD-specific hydrogenase of A. eutrophus are summarized. Results on the solubilization and purification of the membrane-bound hydrogenase of A. eutrophus are presented in detail. The enzyme was solubilized from purified membranes by Triton X-100 and sodium desoxycholate or phospholipase D. The crude membrane extract was fractionated by ammonium sulfate precipitation and chromatography on carboxymethylcellulose at pH 5.5. The enzyme was stable in potassium phosphate buffer; it resembles the soluble enzyme with respect to stability under oxidizing conditions. Further biochemical and immunological data indicate, however, that both enzymes are different with respect to their native structure.

Reduction of Unusual Iron-Sulfur Clusters in the H2-sensing Regulatory Ni-Fe Hydrogenase from Ralstonia eutropha H16

“…Because the RH operates under aerobic conditions, similar [4Fe-3S-3O] cluster formation may readily occur. A [4Fe-3S-3O] cluster has also been found in the so-called "prismane" protein (43 2 and is also unable to bind the PAS domain of the HoxJ protein (14). The HoxB C terminus may, thus, mediate these protein interactions and stabilize the Fe-S species appearing to be lost in RH stop between RH WT and RH stop .…”

“…1B) was purified from R. eutropha HF574(pGE567) as a Strep-tag II fusion protein. 2 Starting with 18 g of cells yielded 1.5 mg of RH stop with a specific activity of 1.6 units/mg of protein.…”

“…2B). Flushing of the RH with H 2 generates the Ni-C state 2 T. Buhrke, O. Lenz, and B. Friedrich, manuscript in preparation. Signal quantification revealed that greater than 90% of the nickel was converted to Ni(III) in both preparations after incubation with H 2 for 10 min, implying similarly effective heterolytic hydrogen cleavage.…”

“…The chemolithoautotrophic ␤-proteobacterium bacterium Ralstonia eutropha H16 houses three different Ni-Fe hydrogenases that are physiologically active in the presence of O 2 (2,3). The membrane-bound and soluble NAD-reducing enzymes are involved in energy conversion (4,5).…”

“…Several unusual properties of the RH (3,10,11) are remarkably different from those of the so-called standard Ni-Fe hydrogenases from inter alia, Desulfovibrio gigas and Allochromatium vinosum (1,12,13). In contrast to the dimeric standard Ni-Fe hydrogenases, the RH forms a double dimer [HoxBC] 2 (Fig. 1A) that is connected to a tetramer of the HoxJ protein (14).…”