Extremophiles

, Volume 22, Issue 3, pp 395–405 | Cite as

Crystal structure of the NADP+ and tartrate-bound complex of l-serine 3-dehydrogenase from the hyperthermophilic archaeon Pyrobaculum calidifontis

  • Kazunari Yoneda
  • Haruhiko Sakuraba
  • Tomohiro Araki
  • Toshihisa Ohshima
Original Paper

Abstract

A gene encoding l-serine dehydrogenase (l-SerDH) that exhibits extremely low sequence identity to the Agrobacterium tumefaciens l-SerDH was identified in the hyperthermophilic archaeon Pyrobaculum calidifontis. The predicted amino acid sequence showed 36% identity with that of Pseudomonas aeruginosa l-SerDH, suggesting that P. calidifontis l-SerDH is a novel type of l-SerDH, like Ps. aeruginosa l-SerDH. The overexpressed enzyme appears to be the most thermostable l-SerDH described to date, and no loss of activity was observed by incubation for 30 min at temperatures up to 100 °C. The enzyme showed substantial reactivity towards d-serine, in addition to l-serine. Two different crystal structures of P. calidifontis l-SerDH were determined using the Se-MAD and MR method: the structure in complex with NADP+/sulfate ion at 1.18 Å and the structure in complex with NADP+/l-tartrate (substrate analog) at 1.57 Å. The fold of the catalytic domain showed similarity with that of Ps. aeruginosa l-SerDH. However, the active site structure significantly differed between the two enzymes. Based on the structure of the tartrate, l- and d-serine and 3-hydroxypropionate molecules were modeled into the active site and the substrate binding modes were estimated. A structural comparison suggests that the wide cavity at the substrate binding site is likely responsible for the high reactivity of the enzyme toward both l- and d-serine enantiomers. This is the first description of the structure of the novel type of l-SerDH with bound NADP+ and substrate analog, and it provides new insight into the substrate binding mechanism of l-SerDH. The results obtained here may be very informative for the creation of l- or d-serine-specific SerDH by protein engineering.

Keywords

Archaea Crystal structure l-Serine 3-dehydrogenase Hyperthermophile Pyrobaculum calidifontis strain JCM 11548/VA1 

Abbreviations

l-SerDH

l-Serine 3-dehydrogenase

MDHP

Methyl-2,2-dimethyl-3-hydroxypropionate

Se-MAD

Selenium multiple-wavelength anomalous dispersion

MR

Molecular replacement

RMSD

Root-mean-square deviation

PDB

Protein data bank

Notes

Acknowledgements

The synchrotron-radiation experiment was performed at the Photon Factory BL-1A and 5A, Tsukuba, Japan. We are grateful to the staff of the Photon Factory for assistance with data collection, which was approved by the Photon Factory Program Advisory Committee (Proposal no. 2016G502). We also thank Ms. Y. Chijiiwa at the Department of Bioscience, School of Agriculture, Tokai University, for helping us with the N-terminal amino acid sequence analysis. This work was supported by a grant for JSPS KAKENHI, Grant Number 16K18689, a grant from the Tokai University Educational System, and a grant from the Institute for Fermentation, Osaka (IFO).

Author contributions

KY performed the experiments and wrote the manuscript. KY, HS, and TO designed the study, analyzed the data, and wrote the manuscript. KY, HS, TA, and TO participated in discussions during the preparation of the manuscript. All authors had final approval of the submitted and published versions.

Compliance with ethical standards

Conflict of interest

The authors declare no conflicts of interest.

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Copyright information

© Springer Japan KK, part of Springer Nature 2018

Authors and Affiliations

  • Kazunari Yoneda
    • 1
  • Haruhiko Sakuraba
    • 2
  • Tomohiro Araki
    • 1
  • Toshihisa Ohshima
    • 3
  1. 1.Department of Bioscience, School of AgricultureTokai UniversityKumamotoJapan
  2. 2.Department of Applied Biological Science, Faculty of AgricultureKagawa UniversityKagawaJapan
  3. 3.Department of Biomedical Engineering, Faculty of EngineeringOsaka Institute of TechnologyOsakaJapan

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