Products of CO2 fixation and 14C labelling pattern of alanine in Methanobacterium thermoautotrophicum pulse-labelled with 14CO2

The incorporation of ¹⁴CO₂ by an exponentially growing culture of the autotrophic bacterium Methanobacterium thermoautotrophicum has been studied. The distribution of radioactivity during 2s–120s incubation periods has been analyzed by chromatography and radioautography. After a 2 s incubation most of the radioactivity of the ethanolsoluble fraction was present in the amino acids alanine, glutamate, glutamine and aspartate, whereas phosphorylated compounds were only weakly labelled. The percentage of the total radioactivity fixed, which was contained in the principal early labelled amino acid alanine, increased in the first 20 s and only then decreased, indicating that alanine is derived from primary products of CO₂ fixation.The labelling patterns of alanine produced during various incubation times have been determined by degradation. After a 2 s ¹⁴CO₂ pulse, 61% of the radioactivity was located in C-1, 23% in C-2, and 16% in C-3. The results are consistent with the operation of a previously proposed autotrophic CO₂ assimilation pathway which involves the formation of acetyl CoA from 2 CO₂ via one-carbon unit intermediates, followed by the reductive carboxylation of acetyl CoA to pyruvate.