CO metabolism of carboxydothermus hydrogenoformans and archaeoglobus fulgidus

Microbial CO metabolism was studied in detail with the ultimate aim to assess the feasibility of a biotechnological process that could replace the existing water gas shift technology in the production of a fuel cell grade hydrogen gas from synthesis gas. It is expected that a biotechnological process is less sensitive to impurities present in synthesis gas and can reach lower CO thresholds, and thus might be more cost effective than conventional catalysts. Low CO thresholds are especially required for polymer electrolyte membrane fuel cells. These fuel cells have a broad application potential in a future hydrogen economy. The bulk of hydrogen produced today is derived from natural gas in a steam reforming process that forms CO besides H 2 . CO is removed by its conversion to H 2 in the water gas shift reaction. However, due to thermodynamic limitations of the process operation temperature, the required CO thresholds are not obtained (<10 ppm). CO conversion at more ambient temperatures is in that respect advantageous for biological CO conversion to obtain a fuel cell grade H 2 .It was demonstrated in Chapter 2 that batch cultures of Carboxydothermus hydrogenoformans converted CO to levels below 2 ppm while accumulating H 2 . In these cultures it was necessary to remove CO 2 from the gas phase. Without removal of CO 2 , CO thresholds approached 100 ppm. CO limits that are generally communicated for PEM-FC, indicate 10 ppm as allowable threshold for CO in H 2 gas. The potential of a biotechnological process is thus supported, however, the time needed to reach these thresholds was considerable and needs attention. Biomass activity and gas/liquid mass transfer are possible rate limiting factors in the biological water gas shift reaction. As demonstrated in Chapter 3, higher conversion rates are possible in batch culture than achieved with the cultivation technique used in Chapter 2. In fact, it appears that only in the early stage of batch cultivation the biomass is limiting CO conversion rates. After some time biomass has grown to sufficient density and gas/mass transfer becomes limiting. Even in the most turbulent shaking regime allowed by the incubator, gas/liquid mass transfer remained rate limiting. Success of a tentative biotechnological process therefore likely depends on gas/liquid mass transfer rates of specific reactor types and associated restrictions dictated by process economics.Physiological PerspectiveIn recent years it has become clear that CO is used as a substrate by a diverse group of strict anaerobic micro-organisms. While it previously was believed that CO predominantly inhibited the growth of many anaerobes, albeit at higher partial pressures than aerobes. Currently research has indicated that CO is a versatile substrate which is effectively used in many microbial metabolisms. The advances that were made especially show that many electron acceptors can be reduced by a wide variety of micro organisms with CO as electron donor. The advances could be made due to the fact that CO is generally neglected as a substrate in physiological studies of novel anaerobic isolates. Genome sequencing projects also demonstrate that the enzymes, CO dehydrogenases, involved in the CO metabolism are present in already well known organisms that have never been tested with CO. An example of this is Archaeoglobus fulgidus , which was tested for its ability to grow with CO in Chapter 5. While it was speculated that A. fulgidus could oxidise CO coupled to the reduction of protons to H 2 , it grew acetogenically with CO instead. In fact, A. fulgidus is the first true homo-acetogenic archaeon known. Remarkable of its metabolism was the intermediate accumulation of formate. It was proposed that A. fulgidus forms acetate via the acetyl-CoA pathway. In this pathway, CO 2 is reduced to form the methyl group of acetate. However, formate is not an intermediate in the expected pathway in which CO 2 is reduced to form formyl-methanofuran and subsequently formyl-tertahydromethanopterin. Besides the formation of acetate, A. fulgidus can reduce sulfate to sulfide with CO as electron donor. It distinguishes itself from bacterial sulfate reducers by its tolerance to CO. While most known sulfate-reducing bacteria are inhibited by elevated levels of CO, A. fulgidus was not noticeably inhibited in the presence of 136 kPa CO. Since A. fulgidus is not capable of growth with H 2 and CO 2 or sulfate as electron acceptors, this organism could be employed to selectively oxidise CO in gas mixtures containing H 2 and remove trace amounts of CO to levels below 10 ppm.Selective oxidation of CO is not possible with C. hydrogenoformans . Although C. hydrogenoformans can reduce various electron acceptors with CO, it also does so with H 2 (Chapter 4). C. hydrogenoformans is a true CO specialist. It is able to grow hydrogenogenically with CO, to reduce various other electron acceptors with CO, and to use CO as sole source of energy and carbon. C. hydrogenoformans contains five CO dehydrogenase genes. Besides the identified activities in H 2 formation, in NADPH generation and in autotrophic carbon fixation for three of these CO dehydrogenase, two CO dehydrogenases are still without function (Chapter 5). Conditions that may lead to expression of these CO dehydrogenases are proposed. In nitrate amended cultures a CO dehydrogenase was present, that was not present in cells grown with other substrates. Wu et al. (2005) already indicated that a CO dehydrogenase of C. hydrogenoformans might be involved in oxidative stress response and be expressed under micro aerophilic conditions. Clearly the microbial physiology regarding CO is still incomplete and is more diverse than thought up to now. Further studies are needed in this respect.