Universität zu Köln

Mohn, Marie Agatha, Thaqi, Besarta and Fischer-Schrader, Katrin ORCID: 0000-0003-4532-8918 (2019). Isoform-Specific NO Synthesis by Arabidopsis thaliana Nitrate Reductase. Plants-Basel, 8 (3). BASEL: MDPI. ISSN 2223-7747

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Abstract

Nitrate reductase (NR) is important for higher land plants, as it catalyzes the rate-limiting step in the nitrate assimilation pathway, the two-electron reduction of nitrate to nitrite. Furthermore, it is considered to be a major enzymatic source of the important signaling molecule nitric oxide (NO), that is produced in a one-electron reduction of nitrite. Like many other plants, the model plant Arabidopsis thaliana expresses two isoforms of NR (NIA1 and NIA2). Up to now, only NIA2 has been the focus of detailed biochemical studies, while NIA1 awaits biochemical characterization. In this study, we have expressed and purified functional fragments of NIA1 and subjected them to various biochemical assays for comparison with the corresponding NIA2-fragments. We analyzed the kinetic parameters in multiple steady-state assays using nitrate or nitrite as substrate and measured either substrate consumption (nitrate or nitrite) or product formation (NO). Our results show that NIA1 is the more efficient nitrite reductase while NIA2 exhibits higher nitrate reductase activity, which supports the hypothesis that the isoforms have special functions in the plant. Furthermore, we successfully restored the physiological electron transfer pathway of NR using reduced nicotinamide adenine dinucleotide (NADH) and nitrate or nitrite as substrates by mixing the N-and C-terminal fragments of NR, thus, opening up new possibilities to study NR activity, regulation and structure.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Mohn, Marie Agatha UNSPECIFIED UNSPECIFIED UNSPECIFIED Thaqi, Besarta UNSPECIFIED UNSPECIFIED UNSPECIFIED Fischer-Schrader, Katrin UNSPECIFIED orcid.org/0000-0003-4532-8918 UNSPECIFIED
URN:	urn:nbn:de:hbz:38-153636
DOI:	10.3390/plants8030067
Journal or Publication Title:	Plants-Basel
Volume:	8
Number:	3
Date:	2019
Publisher:	MDPI
Place of Publication:	BASEL
ISSN:	2223-7747
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language CHLORATE-RESISTANT MUTANT; NITRIC-OXIDE SYNTHASE; L MERR LEAVES; WILD-TYPE; DIFFERENTIAL EXPRESSION; NITROGEN OXIDE(S); STOMATAL CLOSURE; KINETIC-ANALYSIS; TOBACCO-LEAVES; IN-VITRO Multiple languages Plant Sciences Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/15363