Solution 13C nuclear magnetic resonance spectroscopic analysis of the amino acids of Methanosphaera stadtmanae: Biosynthesis and origin of one- carbon units from acetate and carbon dioxide

Abstract

We found that general pathways for amino acid synthesis of Methanosphaera stadtmanae, a methanogen that forms CH4 from H2 and methanol, resembled those of methanogens that form CH4 from CO2 or from the methyl group of acetate. We determined the incorporation of 14C-labeled CO2, formate, methanol, methionine, serine, and acetate into cell macromolecules. Labeling of amino acid carbons was determined by solution nuclear magnetic resonance spectroscopy after growth with 13C-labeled acetate, CO2, serine, and methanol. The α and β carbons of serine and alanine were formed from carboxyl and methyl carbons of acetate, respectively, and the amino acid carboxyl groups were formed from CO2. This indicates that pyruvate was formed by reductive carboxylation of acetate. Labeling of the methyl carbon of methionine indicated that the major route of synthesis was from the hydroxymethyl carbon of serine that arises from the methyl carbon of acetate. Methanol was a minor source of the methyl of methionine. Unambiguous assignment was made of the sources of all carbons of histidine. Labeling of the histidine 7 position (ε carbon) was consistent with formation from the C-2 of the purine ring of ATP and the origin of the C-2 from a formyl unit derived from the hydroxymethyl carbon of serine.