Aspects of the biochemistry of the thermophilic microorganism Bacillus stearothermophilus.

Thermophilic microorganisms present two fundamental proolems. (I) How are they able to grow at temperatures greatly above those normally considered suitable for life. (2) Why do thermophiles not grow at more moderate temperatures. This thesis is an account of experiments directed towards resolution of these problems using a prototrophic strain of the thermophile Bacillus stearothermophilus. Although virtually all the thermophile enzymes isolated to date have proved to be thermostable compared with their mesophile counterparts the possibility has remained open that part of the mechanism of thermophily is a rapid resynthesis of denatured proteins. It is shown here that protein turnover does occur during growth out the rate is too low to be compatible with this ''rapid repair" hypothesis. Studies of the stabilities of individual enzymes in intact cells confirm the results of the turnover experiments. This organisms' ability to grow on minimal media facilitated an examination of the energetics of growth. The energy required for growth is shown to be comparable with that of related mesophiles with no evidence for a massive diversion of energy for resynthesis of denatured macromolecules. These growth yield studies also show that the efficiency of substrate utilization decreases as the temperature rises. This is largely the result of changes in the extent of oxidation of the substrate. The effects of temperature on the regulation of a key enzyme, isolated from the thermophile, pyruvate kinase, have been studied to test the hypothesis that the temperature limits for growth of a particular microorganism are the result of an accumulation of derangements of metabolic regulation at temperature extremes. In most respects except thermostability the thermophile enzyme resembles those from mesophiles. Temperature effects on kinetic and regulatory parameters are too small to be implicated in the lower limit for growth but are probably involved in the temperature effects on growth yield.