DETECTION OF THE GENE ENCODING THE SMALL SUBUNIT OF THE CO DEHYDROGENASE ENZYME IN THE H_2-EVOLVING BACTERIUM RUBRIVIVAX GELATINOSUS CBS

摘要

Rubrivivax (formerly Rhodocyclus) gelatinosus CBS is a purple non-sulfur bacterium that offers great potential for biological H_2 production on a commercial scale. When cultured in the presence of CO in the dark, Rubrivivax gelatinosus CBS catalyzes a water-gas shift reaction whereby CO (and H_2O) is shifted into H_2 (and CO_2) in near stoichiometric quantities. Using CO generated from gasified waste biomass, the bacterial water-gas shift reaction could be employed to produce H_2 as a clean alternative fuel. Three key enzymes involved in the CO-water shift reaction, as well as the genes encoding them, have been characterized in a closely related purple non-sulfur bacterium, Rhodospirillum rubrum: a CO dehydrogenase (CODH), the ferredoxin-like electron-carrier small subunit of the CODH enzyme, and an H_2-evolving hydrogenase. The genes for the hydrogenase are encoded on a transcriptional unit that is separate from the genes encoding the CODH and its ferredoxin-like small subunit. Using the polymerase chain reaction (PCR), we amplified a fragment of the Rhodospirillum rubrum ferredoxin-like subunit gene (cooF). This was then used as a probe in Southern blots of restriction endonuclease digested genomic DNA extracted from Rubrivivax gelatinosus CBS. Fragments from both Sall and Xhol digested Rubrivivax gelatinosus DNA hybridized with the probe. The sizes of the Sall fragments detected were 1.78 Kb and 511 bp. The size of the Xhol fragment detected was 2.21 Kb. These bands represent a potential identification of the gene for a ferredoxin-like subunit in the CO oxidative pathway in Rubrivivax gelatinosus CBS that was not formerly known. This identification is currently being confirmed, and the gene sequenced.