Quantitative evaluation of the biosynthetic pathways leading to δ-aminolevulinic acid from the Shemin precursor glycine via the C5 pathway in Arthrobacter hyalinus by analysis of ~(13)C-labeled coproporphyrinogen III biosynthesized from [2-~(13)C]glycine, [1- ~(13)C]acetate, and [2-~(13)C]acetate using ~(13)C NMR spectroscopy

摘要

The biosynthetic pathways leading to δ-aminolevulinic acid (ALA) from the Shemin precursor glycine via the C5 pathway in Arthrobacter hyalinus were quantitatively evaluated by means of feeding experiments with [2- ~(13)C]glycine, sodium [1-~(13)C]acetate, and sodium [2- ~(13)C]acetate, followed by analysis of the labeling patterns of coproporphyrinogen III (Copro'gen III) (biosynthesized from ALA) using ~(13)C NMR spectroscopy. Two biosynthetic pathways leading to ALA from glycine via the C5 pathway were identified: i.e., transformation of glycine to l-serine catalyzed by glycine hydroxymethyltransferase, and glycine synthase-catalyzed catabolism of glycine to N ~5,N ~(10)-methylene-tetrahydrofolic acid (THF), which reacts with another molecule of glycine to afford l-serine. l-Serine is transformed to acetyl-CoA via pyruvic acid. Acetyl-CoA enters the tricarboxylic acid cycle, affording 2-oxoglutaric acid, which in turn is transformed to l-glutamic acid. The l-glutamic acid enters the C5 pathway, affording ALA in A. hyalinus. A ~(13)C NMR spectroscopic comparison of the labeling patterns of Copro'gen III obtained after feeding of [2-~(13)C]glycine, sodium [1-~(13)C]acetate, and sodium [2-~(13)C]acetate showed that [2-~(13)C]glycine transformation and [2-~(13)C]glycine catabolism in A. hyalinus proceed in the ratio of 52 and 48 %. The reaction of [2-~(13)C]glycine and N ~5,N ~(10)-methylene-THF, that of glycine and N ~5,N ~(10)-[methylene-~(13)C] methylene-THF generated from the [2-~(13)C]glycine catabolism, and that of [2-~(13)C]glycine and N ~5,N ~(10)-[methylene- ~(13)C]methylene-THF transformed the fed [2-~(13)C]glycine to [1-~(13)C]acetyl-CoA, [2-~(13)C]acetyl-CoA, and [1,2- ~(13)C_2]acetyl-CoA in the ratios of 42, 37, and 21 %, respectively. These labeled acetyl-CoAs were then incorporated into ALA. Our results provide a quantitative picture of the pathways of biosynthetic transformation to ALA from glycine in A. hyalinus.