Enzymatic properties of the ferredoxin-dependent nitrite reductase from chlamydomonas reinhardtii. Evidence for hydroxylamine as a late intermediate in ammonia production

Masakazu Hirasawa, Jatindra N. Tripathy, Frederik Sommer, Ramasamy Somasundaram, Jung Sung Chung, Matthew Nestander, Mahima Kruthiventi, Masoud Zabet-Moghaddam, Michael K. Johnson, Sabeeha S. Merchant, James P. Allen, David B. Knaff

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24 Citations (Scopus)

Abstract

The ferredoxin-dependent nitrite reductase from the green alga Chlamydomonas reinhardtii has been cloned, expressed in Escherichia coli as a His-tagged recombinant protein, and purified to homogeneity. The spectra, kinetic properties and substrate-binding parameters of the C. reinhardtii enzyme are quite similar to those of the ferredoxin-dependent spinach chloroplast nitrite reductase. Computer modeling, based on the published structure of spinach nitrite reductase, predicts that the structure of C. reinhardtii nitrite reductase will be similar to that of the spinach enzyme. Chemical modification studies and the ionic-strength dependence of the enzyme's ability to interact with ferredoxin are consistent with the involvement of arginine and lysine residues on C. reinhardtii nitrite reductase in electrostatically-stabilized binding to ferredoxin. The C. reinhardtii enzyme has been used to demonstrate that hydroxylamine can serve as an electron-accepting substrate for the enzyme and that the product of hydroxylamine reduction is ammonia, providing the first experimental evidence for the hypothesis that hydroxylamine, bound to the enzyme, can serve as a late intermediate during the reduction of nitrite to ammonia catalyzed by the enzyme.

Original languageEnglish
Pages (from-to)67-77
Number of pages11
JournalPhotosynthesis Research
Volume103
Issue number2
DOIs
Publication statusPublished - Jan 1 2010

Keywords

  • Ferredoxin
  • Hydroxylamine reduction
  • NII1
  • Nitrite reductase
  • PETF
  • Tertiary structure

ASJC Scopus subject areas

  • Biochemistry
  • Plant Science
  • Cell Biology

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  • Cite this

    Hirasawa, M., Tripathy, J. N., Sommer, F., Somasundaram, R., Chung, J. S., Nestander, M., Kruthiventi, M., Zabet-Moghaddam, M., Johnson, M. K., Merchant, S. S., Allen, J. P., & Knaff, D. B. (2010). Enzymatic properties of the ferredoxin-dependent nitrite reductase from chlamydomonas reinhardtii. Evidence for hydroxylamine as a late intermediate in ammonia production. Photosynthesis Research, 103(2), 67-77. https://doi.org/10.1007/s11120-009-9512-5