Abstract
Methanothermobacter thermoautotrophicus is an archaeon that reduces CO2 into methane with hydrogen as the electron donor. Under natural and laboratory conditions, hydrogen concentrations may vary over orders of magnitude. The organism has to adapt to these changes. In this thesis, the adaptation of M. thermoautotrophicus to varying hydrogen concentrations is investigated at the bioenergetic and physiological levels. The study includes the development of new methods for the determination of intracellular pH and membrane potential in the organism and for the quantitative detection of the cofactors methanofuran, HS-CoM, HS-HTP and their derivatives. The thermodynamic and bio-energetic properties of several key reactions, including F420 reduction, formyl-methanofuran synthesis and CoM-S-S-CoB reduction reactions, were studied in vivo in relation to the applied hydrogen partial pressure. Cell suspensions of M. thermoautotrophicus maintain the concentration ratio between reduced and oxidized F420 in thermodynamic equilibrium with the applied hydrogen partial pressure. Changes in hydrogen concentrations affect the ion-translocation stoichiometry of the heterodisulfide reductase and formyl-methanofuran dehydrogenase reactions in different ways. The bio-energetic changes have been investigated in direct connection with the growth physiology of the organism. By use of a set of theoretically derived mathematical and experimentally verified expressions, growth physiology is described in terms of growth rates, methane-forming activities, growth yields and 'specific maintenance coefficients'.Fed-batch and chemostat culture experiments showed that growth proceeded in three different growth phases, characterized by specific relations between growth rates, growth yields and maintenance coefficients. The transitions between the growth phases reflect changes in ion-translocation stoichiometries resulting from changes in the hydrogen concentration. Furthermore, the transition of one growth phase to another is accompanied by stepwise changes in intracellular pH and proton motive force that remain invariant during each phase
