1887

Abstract

sp. ATCC 51142 is an aerobic N-fixing and hydrogen-producing cyanobacterium. Isotopomer analysis of its amino acids revealed an identical labelling profile for leucine and isoleucine when 51142 was grown mixotrophically using 2-C-labelled glycerol as the main carbon source. This indicated that 51142 employs the atypical alternative citramalate pathway for isoleucine synthesis, with pyruvate and acetyl-CoA as precursors. Utilization of the citramalate pathway was confirmed by an enzyme assay and LC-MS/MS analysis. Furthermore, the genome sequence of 51142 shows that the gene encoding the key enzyme (threonine ammonia-lyase) in the normal isoleucine pathway is missing. Instead, the cce_0248 gene in 51142 exhibits 53 % identity to the gene encoding citramalate synthase (CimA, GSU1798) from . Reverse-transcription PCR indicated that the cce_0248 gene is expressed and its transcriptional level is lower in medium with isoleucine than in isoleucine-free medium. Additionally, a search for citramalate synthase and threonine ammonia-lyase implies that this alternative isoleucine synthesis pathway may be present in other cyanobacteria, such as and . This suggests that the pathway is more widespread than originally thought, as previous identifications of the citramalate pathway are limited to mostly anaerobic bacteria or archaea. Furthermore, this discovery opens the possibility that such autrotrophic micro-organisms may be engineered for robust butanol and propanol production from 2-ketobutyrate, which is an intermediate in the isoleucine biosynthesis pathway.

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2010-02-01
2024-03-29
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References

  1. Atsumi S., Liao J. C. 2008; Directed evolution of Methanococcus jannaschii citramalate synthase for biosynthesis of 1-propanol and 1-butanol by Escherichia coli. Appl Environ Microbiol 74:7802–7808
    [Google Scholar]
  2. Charon N. W., Johnson R. C., Peterson D. 1974; Amino acid biosynthesis in the spirochete Leptospira: evidence for a novel pathway of isoleucine biosynthesis. J Bacteriol 117:203–211
    [Google Scholar]
  3. Colón-López M. S., Sherman L. A. 1998; Transcriptional and translational regulation of photosystem I and II genes in light-dark- and continuous-light-grown cultures of the unicellular cyanobacterium Cyanothece sp. strain ATCC 51142. J Bacteriol 180:519–526
    [Google Scholar]
  4. Eikmanns B., Linder D., Thauer R. K. 1983; Unusual pathway of isoleucine biosynthesis in Methanobacterium thermoautotrophicum. Arch Microbiol 136:111–113
    [Google Scholar]
  5. Feng X., Mouttaki H., Lin L., Huang R., Wu B., Hemme C. L., He Z., Zhang B., Hicks L. M. other authors 2009; Characterization of the central metabolic pathways in Thermoanaerobacter sp. X514 via isotopomer-assisted metabolite analysis. Appl Environ Microbiol 75:5001–5008
    [Google Scholar]
  6. Heinig K., Henion J. 1999; Fast liquid chromatographic-mass spectrometric determination of pharmaceutical compounds. J Chromatogr B Biomed Sci Appl 732:445–458
    [Google Scholar]
  7. Howell D. M., Xu H. M., White R. H. 1999; ( R)-Citramalate synthase in methanogenic archaea. J Bacteriol 181:331–333
    [Google Scholar]
  8. Jahn U., Huber H., Eisenreich W., Hugler M., Fuchs G. 2007; Insights into the autotrophic CO2 fixation pathway of the archaeon Ignicoccus hospitalis: comprehensive analysis of the central carbon metabolism. J Bacteriol 189:4108–4119
    [Google Scholar]
  9. Johnson D. R., Lee P. K. H., Holmes V. F., Fortin A. C., Alvarez-Cohen L. 2005; Transcriptional expression of the tceA gene in a Dehalococcoides-containing microbial enrichment. Appl Environ Microbiol 71:7145–7151
    [Google Scholar]
  10. Kisumi M., Komatsubara S., Chibata I. 1977; Pathway for isoleucine formation form pyruvate by leucine biosynthetic enzymes in leucine-accumulating isoleucine revertants of Serratia marcescens. J Biochem 82:95–103
    [Google Scholar]
  11. Risso C., Van Dien S. J., Orloff A., Lovley D. R., Coppi M. V. 2008; Elucidation of an alternate isoleucine biosynthesis pathway in Geobacter sulfurreducens. J Bacteriol 190:2266–2274
    [Google Scholar]
  12. Schäfer S., Paalme T., Vilu R., Fuchs G. 1989; 13C-NMR study of acetate assimilation in Thermoproteus neutrophilus. Eur J Biochem 186:695–700
    [Google Scholar]
  13. Shastri A. A., Morgan J. A. 2007; A transient isotopic labeling methodology for 13C metabolic flux analysis of photoautotrophic microorganisms. Phytochemistry 68:2302–2312
    [Google Scholar]
  14. Stöckel J., Welsh E. A., Liberton M., Kunnvakkam R., Aurora R., Pakrasi H. B. 2008; Global transcriptomic analysis of Cyanothece 51142 reveals robust diurnal oscillation of central metabolic processes. Proc Natl Acad Sci U S A 105:6156–6161
    [Google Scholar]
  15. Tang Y. J., Chakraborty R., Martin H. G., Chu J., Hazen T. C., Keasling J. D. 2007; Flux analysis of central metabolic pathways in Geobacter metallireducens during reduction of soluble Fe(III)-NTA. Appl Environ Microbiol 73:3859–3864
    [Google Scholar]
  16. Tang Y. J., Martin H. G., Myers S., Rodriguez S., Baidoo E. E., Keasling J. D. 2009a; Advances in analysis of microbial metabolic fluxes via 13C isotopic labeling. Mass Spectrom Rev 28:362–375
    [Google Scholar]
  17. Tang Y. J., Sapra R., Joyner D., Hazen T. C., Myers S., Reichmuth D., Blanch H., Keasling J. D. 2009b; Analysis of metabolic pathways and fluxes in a newly discovered thermophilic and ethanol-tolerant Geobacillus strain. Biotechnol Bioeng 102:1377–1386
    [Google Scholar]
  18. Tang Y. J., Yi S., Zhuang W.-Q., Zinder S. H., Keasling J. D., Alvarez-Cohen L. 2009c; Investigation of carbon metabolism in “ Dehalococcoides ethenogenes” strain 195 via isotopic and transcriptomic analyses. J Bacteriol 191:5224–5231
    [Google Scholar]
  19. Toepel J., Welsh E., Summerfield T. C., Pakrasi H. B., Sherman L. A. 2008; Differential transcriptional analysis of the cyanobacterium Cyanothece sp strain ATCC 51142 during light-dark and continuous-light growth. J Bacteriol 190:3904–3913
    [Google Scholar]
  20. Wahl S. A., Dauner M., Wiechert W. 2004; New tools for mass isotopomer data evaluation in 13C flux analysis: mass isotope correction, data consistency checking, and precursor relationships. Biotechnol Bioeng 85:259–268
    [Google Scholar]
  21. Welsh E. A., Liberton M., Stöckel J., Loh T., Elvitigala T., Wang C., Wollam A., Fulton R. S., Clifton S. W. other authors 2008; The genome of Cyanothece 51142, a unicellular diazotrophic cyanobacterium important in the marine nitrogen cycle. Proc Natl Acad Sci U S A 105:15094–15099
    [Google Scholar]
  22. Westfall H. N., Charon N. W., Peterson D. E. 1983; Multiple pathways for isoleucine biosynthesis in the spirochete Leptospira. J Bacteriol 154:846–853
    [Google Scholar]
  23. Xu H., Zhang Y. Z., Guo X. K., Ren S., Staempfli A. A., Chiao J., Jiang W., Zhao G. 2004; Isoleucine biosynthesis in Leptospira interrogans serotype lai strain 56601 proceeds via a threonine-independent pathway. J Bacteriol 186:5400–5409
    [Google Scholar]
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