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Characterization of a thermostable dihydrodipicolinate synthase from Thermoanaerobacter tengcongensis

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Abstract

Dihydrodipicolinate synthase (DHDPS) catalyses the first reaction of the (S)-lysine biosynthesis pathway in bacteria and plants. The hypothetical gene for dihydrodipicolinate synthase (dapA) of Thermoanaerobacter tengcongensis was found in a cluster containing several genes of the diaminopimelate lysine–synthesis pathway. The dapA gene was cloned in Escherichia coli, DHDPS was subsequently produced and purified to homogeneity. The T. tengcongensis DHDPS was found to be thermostable (T 0.5 = 3 h at 90°C). The specific condensation of pyruvate and (S)-aspartate-β -semialdehyde was catalyzed optimally at 80°C at pH 8.0. Enzyme kinetics were determined at 60°C, as close as possible to in vivo conditions. The established kinetic parameters were in the same range as for example E. coli dihydrodipicolinate synthase. The specific activity of the T. tengcongensis DHDPS was relatively high even at 30°C. Like most dihydrodipicolinate synthases known at present, the DHDPS of T. tengcongensis seems to be a tetramer. A structural model reveals that the active site is well conserved. The binding site of the allosteric inhibitor lysine appears not to be conserved, which agrees with the fact that the DHDPS of T. tengcongensis is not inhibited by lysine under physiological conditions.

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Abbreviations

DAP:

Diaminopimelate

DIBAL:

Diisobutylaluminium hydride

DHDP:

Dihydrodipicolinate

DHDPR:

Dihydrodipicolinate reductase

DHDPS:

Dihydrodipicolinate synthase

HTPA:

(4S)-4–hydroxy-2,3,4,5-tetrahydro-(2S)-dipicolinic acid

NAL:

N-acetylneuraminate lyase

(S)-ASA:

(S)-aspartate-β-semialdehyde

TCA:

Trichloroacetic acid

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Acknowledgments

We want to thank Dr. Gabriella M. Preda (West University of Timisoara) for the synthesis of a batch of (S)-ASA. This research is performed as part of the IBOS Programme (Integration of Biosynthesis & Organic Synthesis) of Advanced Chemical Technologies for Sustainability (ACTS).

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Authors and Affiliations

  1. Laboratory of Microbiology, Wageningen University, Dreijenplein 10, 6703 HB, Wageningen, The Netherlands

    Suzanne Wolterink-van Loo, Mark Levisson & John van der Oost

  2. Laboratory of Organic Chemistry, Wageningen University, Dreijenplein 8, 6703 HB, Wageningen, The Netherlands

    Maud C. Cabrières & Maurice C. R. Franssen

Authors
  1. Suzanne Wolterink-van Loo
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  2. Mark Levisson
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  3. Maud C. Cabrières
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  4. Maurice C. R. Franssen
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  5. John van der Oost
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Corresponding author

Correspondence to John van der Oost.

Additional information

Communicated by G. Antranikian.

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Wolterink-van Loo, S., Levisson, M., Cabrières, M.C. et al. Characterization of a thermostable dihydrodipicolinate synthase from Thermoanaerobacter tengcongensis . Extremophiles 12, 461–469 (2008). https://doi.org/10.1007/s00792-008-0152-z

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