(R)-Cysteate-nitrogen assimilation by Cupriavidus necator H16 with excretion of 3-sulfolactate: a patchwork pathway.
Cupriavidus necator H16 grew exponentially with (R)-cysteate, a structural analogue of aspartate, as sole source of nitrogen in succinate-salts medium. Utilization of cysteate was quantitative and concomitant with growth and with the excretion of the deaminated product (R)-sulfolactate, which was identified thoroughly. The deaminative pathway started with transport of (R)-cysteate into the cell, which we attributed to an aspartate transporter. Transamination to sulfopyruvate involved an aspartate/(R)-cysteate:2-oxoglutarate aminotransferase (Aoa/Coa) and regeneration of the amino group acceptor by NADP⁺-coupled glutamate dehydrogenase. Reduction of sulfopyruvate to (R)-sulfolactate was catalyzed by a (S)-malate/(R)-sulfolactate dehydrogenase (Mdh/Sdh). Excretion of the sulfolactate could be attributed to the sulfite/organosulfonate exporter TauE, which was co-encoded and co-expressed, with sulfoacetaldehyde acetyltransferase (Xsc), though Xsc was irrelevant to the current pathway. The metabolic enzymes could be assayed biochemically. Aoa/Coa and Mdh/Sdh were highly enriched by protein separation, partly characterized, and the relevant locus-tags identified by peptide-mass fingerprinting. Finally, RT-PCR was used to confirm the transcription of all appropriate genes. We thus demonstrated that Cupriavidus necator H16 uses a patchwork pathway by recruitment of 'housekeeping' genes and sulfoacetaldehyde-degradative genes to scavenge for (R)-cysteate-nitrogen.
Department of Biology, University of Konstanz, 78457 Konstanz, Germany.
Jutta
J
Mayer
Karin
K
Denger
Klaus
K
Hollemeyer
David
D
Schleheck
Alasdair M
AM
Cook
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D020118:Cupriavidus necator / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D003544:Cysteic Acid / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D007773:Lactates / Q000378:metabolism*; D009584:Nitrogen / Q000378:metabolism*
D001426:Bacterial Proteins; D007773:Lactates; D003544:Cysteic Acid; D009584:Nitrogen