Amino acid biosynthesis and sodium‐dependent transport in Methanococcus voltae, as revealed by 13C NMR

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Abstract
Of several methanogenic bacteria examined only M. voltae readily incorporated exogenous amino acids into cell protein. This was easily shown, since growth in the presence of exogenous amino acids resulted in a loss of signal intensities from those Ca atoms normally labeled by [13C]acetate during biosynthesis. From 80-95% of the Ser, Lys, Pro or Val incorporated into protein could be supplied directly from the growth medium. In contrast, Asp and Glu, if supplied to the medium, accounted for only a small percentage of the total acidic amino acid used in protein synthesis. Constitutive transport systems took up a wide range of amino acids at rates of 0.1-4.1 nmol min-1 mg-1. The transport systems required Na+, with the possible exception of the basic amino acid Lys, and were inhibited by N-ethylmaleimide or 3,3'',4'',5-tetrachlorosalicylanilide. No interconversion of Ile to other amino acids was detected when cells were given [13C]Ile during growth, whereas the expected labeling of the Asp and Glu families of amino acids resulted when [13C]Asp was provided to the culture. M. voltae synthesized its aimino acids from acetate via routes fully consistent with those found in Methanospirillum hungatei. Propionate could substitute for an auxotrophic requirement for Ile, resulting in the synthesis of Ile with the .beta.-C originating from the carboxyl of acetate and the .alpha.-C from the carboxyl of propionate. No labeling of Ile from [13C]acetate could occur without the fatty acid. These results provide strong evidence for the carboxylation of propionate to form 2-oxobutyrate as intermediate in Ile biosynthesis, and show that the metabolic defect in Ile biosynthesis occurs prior to 2-oxobutyrate synthesis. The presence of constitutive amino acid trasnport systems and multiple routes for Ile biosynthesis make M. voltae an attractive methanogen for genetic studies.