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Volume 169, Issue 10

Asuc_0142 of 130Z is the -aspartate/C4-dicarboxylate exchanger DcuA Open Access

Abstract

Anaerobic bacteria often use antiporters DcuB (malate/succinate antiport) or DcuA (-aspartate/succinate antiport) for the excretion of succinate during fumarate respiration. The rumen bacterium is able to produce large amounts of succinate by fumarate respiration, using the DcuB-type transporter DcuE for -malate/succinate antiport. Asuc_0142 was annotated as a second DcuB-type transporter. Deletion of Asuc_0142 decreased the uptake rate for -[C]aspartate into cells. Properties of transport by heterologously expressed Asuc_0142 were investigated in an mutant deficient of anaerobic C4DC transporters. Expression of Asuc_0142 resulted in high uptake activity for -[C]fumarate or -[C]aspartate, but the former showed a strong competitive inhibition by -aspartate. In loaded with -[C]aspartate, [C]succinate or [C]fumarate, extracellular C4DCs initiated excretion of the intracellular substrates, with a preference for -aspartate/succinate or -aspartate/fumarate antiport. These findings indicate that Asuc_0142 represents a DcuA-type transporter for -aspartate uptake and -aspartate/C4DC antiport, differentiating it from the DcuB-type transporter DcuE for -malate/succinate antiport. Sequence analysis and predicted structural characteristics confirm structural similarity of Asuc_0142 to DcuA, and Asuc_0142 was thus re-named as DcuA. The bovine rumen fluid contains -aspartate (99.6 µM), whereas fumarate and -malate are absent. Therefore, bovine rumen colonisers depend on -aspartate as an exogenous substrate for fumarate respiration. encodes HemG (protoporphyrinogen oxidase) and PyrD (dihydroorotate dehydrogenase) for haem and pyrimidine biosynthesis. The enzymes require fumarate as an electron acceptor, suggesting an essential role for -aspartate, DcuA, and fumarate respiration for growing in the bovine rumen.

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Keyword(s): Actinobacillus succinogenes , bovine rumen , C4DC , DcuA and L-aspartate
Funding
This study was supported by the:
  • National Research Foundation of Korea (Award NRF-2019R1A2C1008066)
    • Principle Award Recipient: OkBin Kim
  • National Research Foundation of Korea (Award NRF-2021R1A6A1A10039823)
    • Principle Award Recipient: OkBin Kim
© 2023 The Authors
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License.
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2023-10-31
2024-12-02
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References

  1. Guest JR. Anaerobic growth of Escherichia coli K12 with fumarate as terminal electron acceptor. Genetic studies with menaquinone and fluoroacetate-resistant mutants. Microbiology 1979; 115:259–271 [View Article] [PubMed]
     [Google Scholar]
  2. Unden G, Strecker A, Kleefeld A, Kim OB. C4-dicarboxylate utilization in aerobic and anaerobic growth. EcoSal Plus 2016; 7: [View Article] [PubMed]
     [Google Scholar]
  3. Engel P, Krämer R, Unden G. Transport of C4-dicarboxylates by anaerobically grown Escherichia coli. Energetics and mechanism of exchange, uptake and efflux. Eur J Biochem 1994; 222:605–614 [View Article] [PubMed]
     [Google Scholar]
  4. Schubert C, Unden G. C4-dicarboxylates as growth substrates and signaling molecules for commensal and pathogenic enteric bacteria in mammalian intestine. J Bacteriol 2022; 204:e0054521 [View Article] [PubMed]
     [Google Scholar]
  5. Nguyen BD, Cuenca VM, Hartl J, Gül E, Bauer R et al. Import of aspartate and malate by DcuABC drives H2/fumarate respiration to promote initial Salmonella gut-lumen colonization in mice. Cell Host Microbe 2020; 27:922–936 [View Article] [PubMed]
     [Google Scholar]
  6. Schubert C, Winter M, Ebert-Jung A, Kierszniowska S, Nagel-Wolfrum K et al. C4-dicarboxylates and l-aspartate utilization by Escherichia coli K-12 in the mouse intestine: l-aspartate as a major substrate for fumarate respiration and as a nitrogen source. Environ Microbiol 2021; 23:2564–2577 [View Article] [PubMed]
     [Google Scholar]
  7. Janausch IG, Zientz E, Tran QH, Kröger A, Unden G. C4-dicarboxylate carriers and sensors in bacteria. Biochim Biophys Acta 2002; 1553:39–56 [View Article] [PubMed]
     [Google Scholar]
  8. Six S, Andrews SC, Unden G, Guest JR. Escherichia coli possesses two homologous anaerobic C4-dicarboxylate membrane transporters (DcuA and DcuB) distinct from the aerobic dicarboxylate transport system (Dct). J Bacteriol 1994; 176:6470–6478 [View Article] [PubMed]
     [Google Scholar]
  9. Rhie MN, Cho YB, Lee YJ, Kim OB. High-affinity l-malate transporter DcuE of Actinobacillus succinogenes catalyses reversible exchange of C4-dicarboxylates. Environ Microbiol Rep 2019; 11:129–139 [View Article] [PubMed]
     [Google Scholar]
  10. Rhie MN, Park B, Ko H-J, Choi I-G, Kim OB. Transcriptome analysis and anaerobic C4 -dicarboxylate transport in Actinobacillus succinogenes. MicrobiologyOpen 2018; 7:e00565 [View Article] [PubMed]
     [Google Scholar]
  11. Strecker A, Schubert C, Zedler S, Steinmetz P, Unden G. DcuA of aerobically grown Escherichia coli serves as a nitrogen shuttle (L-aspartate/fumarate) for nitrogen uptake. Mol Microbiol 2018; 109:801–811 [View Article] [PubMed]
     [Google Scholar]
  12. Koolman J, Röhm K, Sadikin M. Atlas Berwarna & Teks Biokimia Jakarta: Penerbit Hipokrates; 1994
     [Google Scholar]
  13. Schubert C, Unden G. Fumarate, a central electron acceptor for Enterobacteriaceae beyond fumarate respiration and energy conservation. In Advances in Microbial Physiology vol 82 Academic Press; 2023 pp 267–299 https://doi.org/10.1016/bs.ampbs.2022.10.002
     [Google Scholar]
  14. Schubert C, Zedler S, Strecker A, Unden G. L-aspartate as a high-quality nitrogen source in Escherichia coli: regulation of L-aspartase by the nitrogen regulatory system and interaction of L-aspartase with GlnB. Mol Microbiol 2021; 115:526–538 [View Article] [PubMed]
     [Google Scholar]
  15. Zientz E, Janausch IG, Six S, Unden G. Functioning of DcuC as the C4-dicarboxylate carrier during glucose fermentation by Escherichia coli. J Bacteriol 1999; 181:3716–3720 [View Article] [PubMed]
     [Google Scholar]
  16. Jones SA, Chowdhury FZ, Fabich AJ, Anderson A, Schreiner DM et al. Respiration of Escherichia coli in the mouse intestine. Infect Immun 2007; 75:4891–4899 [View Article] [PubMed]
     [Google Scholar]
  17. Litsanov B, Brocker M, Oldiges M, Bott M. Succinic acid. In Bioprocessing of Renewable Resources to Commodity Bioproducts 2014 pp 435–472 [View Article]
     [Google Scholar]
  18. McKinlay JB, Laivenieks M, Schindler BD, McKinlay AA, Siddaramappa S et al. A genomic perspective on the potential of Actinobacillus succinogenes for industrial succinate production. BMC Genomics 2010; 11:680 [View Article] [PubMed]
     [Google Scholar]
  19. Rhie MN, Yoon HE, Oh HY, Zedler S, Unden G et al. A Na+-coupled C4-dicarboxylate transporter (Asuc_0304) and aerobic growth of Actinobacillus succinogenes on C4-dicarboxylates. Microbiology 2014; 160:1533–1544 [View Article] [PubMed]
     [Google Scholar]
  20. Bertin Y, Segura A, Jubelin G, Dunière L, Durand A et al. Aspartate metabolism is involved in the maintenance of enterohaemorrhagic Escherichia coli O157:H7 in bovine intestinal content. Environ Microbiol 2018; 20:4473–4485 [View Article] [PubMed]
     [Google Scholar]
  21. McKinlay JB, Zeikus JG, Vieille C. Insights into Actinobacillus succinogenes fermentative metabolism in a chemically defined growth medium. Appl Environ Microbiol 2005; 71:6651–6656 [View Article] [PubMed]
     [Google Scholar]
  22. Kim OB, Unden G. The L-tartrate/succinate antiporter TtdT (YgjE) of L-tartrate fermentation in Escherichia coli. J Bacteriol 2007; 189:1597–1603 [View Article] [PubMed]
     [Google Scholar]
  23. Miller J. A short course in bacterial genetics: A laboratory manual and handbood for Escherichia coli and related bacteria. In A Short Course in Bacterial Genetics: A Laboratory Manual and Handbood for Escherichia Coli and Related Bacteria 1992 pp 143–149
     [Google Scholar]
  24. Zientz E, Six S, Unden G. Identification of a third secondary carrier (DcuC) for anaerobic C4-dicarboxylate transport in Escherichia coli: roles of the three Dcu carriers in uptake and exchange. J Bacteriol 1996; 178:7241–7247 [View Article] [PubMed]
     [Google Scholar]
  25. Bauer J, Fritsch MJ, Palmer T, Unden G. Topology and accessibility of the transmembrane helices and the sensory site in the bifunctional transporter DcuB of Escherichia coli. Biochemistry 2011; 50:5925–5938 [View Article] [PubMed]
     [Google Scholar]
  26. Golby P, Kelly DJ, Guest JR, Andrews SC. Transcriptional regulation and organization of the dcuA and dcuB genes, encoding homologous anaerobic C4-dicarboxylate transporters in Escherichia coli. J Bacteriol 1998; 180:6586–6596 [View Article] [PubMed]
     [Google Scholar]
  27. Xu J, Zhang Y. How significant is a protein structure similarity with TM-score = 0.5?. Bioinformatics 2010; 26:889–895 [View Article] [PubMed]
     [Google Scholar]
  28. Reva BA, Finkelstein AV, Skolnick J. What is the probability of a chance prediction of a protein structure with an rmsd of 6å?. Fold Des 1998; 3:141–147 [View Article] [PubMed]
     [Google Scholar]
  29. Jones SA, Gibson T, Maltby RC, Chowdhury FZ, Stewart V et al. Anaerobic respiration of Escherichia coli in the mouse intestine. Infect Immun 2011; 79:4218–4226 [View Article] [PubMed]
     [Google Scholar]
  30. Taylor RG, Walker DC, McInnes RR. E. coli host strains significantly affect the quality of small scale plasmid DNA preparations used for sequencing. Nucleic Acids Res 1993; 21:1677–1678 [View Article]
     [Google Scholar]
  31. Datsenko KA, Wanner BL. One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A 2000; 97:6640–6645 [View Article]
     [Google Scholar]
  32. Guzman LM, Belin D, Carson MJ, Beckwith JON. Tight regulation, modulation, and high-level expression by vectors containing the arabinose PBAD promoter. J Bacteriol 1995; 177:4121–4130 [View Article]
     [Google Scholar]
  33. Guex N, Peitsch MC. SWISS-MODEL and the SWISS-PdbViewer: an environment for comparative protein modeling. Electrophoresis 1997; 18:2714–2723 [View Article] [PubMed]
     [Google Scholar]
/content/journal/micro/10.1099/mic.0.001411
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Asuc_0142 of Actinobacillus succinogenes 130Z is the l-aspartate/C4-dicarboxylate exchanger DcuA
Microbiology 169, 001411 (2023); https://doi.org/10.1099/mic.0.001411
/content/journal/micro/10.1099/mic.0.001411
/content/journal/micro/10.1099/mic.0.001411
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