Conventional stainless steel fermentors cannot be used to grow bacteria at high temperatures, with high pressures of (flammable or toxic) gases, in corrosive media (with high sulfide concentrations or at extremes of pH), or in media with a defined metal content (because of metal leaching from the fermentor walls). In this proposal we request two enamel-lined, metal- free, fermentors of 63 liter and 400 liter capacity, a centrifuge for cell harvesting, and a metal-free water system. The fermentors are rated to 150oC and 6 atmospheres and are specifically designed to grow bacteria under extreme conditions. They will be used to grow recently discovered extremely thermophilic bacteria at 90-110oC for the purpose of characterizing their metalloenzymes involved in the metabolism of various gases (H2, N2, H2S, CH4 and CO). In addition, these and various fermentative, acetogenic, methanogenic and sulfide-producing bacteria will be grown in defined metal media. This allows the expression of certain enzymes, e.g. V-nitrogenase, and the use of expensive isotopes (Fe, Mo, W, Ni) for spectroscopic analyses of various metalloenzymes. The overall objectives are to elucidate the structures of a variety of metal clusters found in proteins and the roles that these clusters play in electron transfer and catalytic activity. If funded, the University of Georgia would become the only facility in the US with the capability of growing large scale bacterial cultures under extreme and metal-defined conditions.
