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Modifications of the activity of nitrate reductase from cucumber roots

  • Published:
Biologia Plantarum

Abstract

The regulatory properties of NADH-dependent nitrate reductase (NR) in desalted root extracts from hydroponically grown cucumber (Cucumis sativus L.) seedlings were examined. The lowest activity of NR was detected in extracts incubated with Mg2+ and ATP. An inhibitory effect of Mg-ATP was cancelled in the presence of staurosporine (the protein kinase inhibitor) and completely reversed after addition of ethylenediaminetetraacetate (EDTA) as well as AMP into reaction mixture. Reactivation of enzyme due to AMP presence, contrary to the chelator-dependent NR activation, was sensitive to microcystin LR (the protein phosphatase inhibitor). Above results indicated that the nitrate reductase in cucumber roots was regulated through reversible phosphorylation of enzyme protein. A drop in the activity of NR was also observed after incubation of enzyme at low pH. At low pH, the presence of ATP alone in the incubation medium was sufficient to inactivate NR, indicating that H+ can substitute the Mg2+ in formation of an inactive complex of enzyme. ATP-dependent inactivation of NR at low pH was prevented by staurosporine and reversed by AMP. However, AMP action was not altered by microcystin LR suggesting that in low pH the nucleotide induced reactivation of NR is not limited to the protein phosphorylation.

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Abbreviations

NR:

nitrate reductase

BSA:

bovine serum albumin

DTT:

dithiotreitol

DMSO:

dimethylsulfoxide

EDTA:

ethylenediaminetetraacetate

Hepes:

hydroxyethylpiperazine ethanesulfonic acid

PMSF:

phenylmethylsulphonyl fluoride

PVPP:

polyvinylpolypyrrolidone

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Authors and Affiliations

  1. Plant Physiology Department, Institute of Plant Biology, Wroclaw University, Kanonia 6/8, PL-50328, Wroclaw, Poland

    M. Reda & G. Klobus

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  1. M. Reda
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  2. G. Klobus
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Correspondence to M. Reda.

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Reda, M., Klobus, G. Modifications of the activity of nitrate reductase from cucumber roots. Biol Plant 50, 42–47 (2006). https://doi.org/10.1007/s10535-005-0072-z

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  • DOI: https://doi.org/10.1007/s10535-005-0072-z

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