Differential S-nitrosylation and characterization of purified S-nitrosoglutathione reductase (GSNOR) from Brassica juncea shows multiple forms of the enzyme
Copyright © 2024 Elsevier Masson SAS. All rights reserved.
| Veröffentlicht in: | Plant physiology and biochemistry : PPB. - 1991. - 207(2024) vom: 15. Feb., Seite 108404 |
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| Format: | Online-Aufsatz |
| Sprache: | English |
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2024
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| Zugriff auf das übergeordnete Werk: | Plant physiology and biochemistry : PPB |
| Schlagworte: | Journal Article Biotin switch technique Denitrosylation GSNOR S-nitrosylation Oxidoreductases EC 1.- formaldehyde dehydrogenase, glutathione-independent EC 1.2.1.46 Aldehyde Oxidoreductases mehr... |
| Zusammenfassung: | Copyright © 2024 Elsevier Masson SAS. All rights reserved. S-nitrosoglutathione reductase (GSNOR). a master regulator of NO homeostasis, is a single-copy gene in most plants. In Lotus japonicus, two GSNOR isoforms were identified exhibiting similar kinetic properties but differential tissue-specific expressions. Previously, a genome-wide identification in Brassica juncea revealed four copies of GSNOR, each encoding proteins that vary in subunit molecular weights and pI. Here, we report multiple forms of GSNOR using 2D immunoblot which showed 4 immunopositive spots of 41.5 kDa (pl 5.79 and 6.78) and 43 kDa (pl 6.16 and 6.23). To confirm, purification of GSNOR using anion-exchange chromatography yielded 2 distinct pools (GSNOR-A & GSNOR-B) with GSNOR activities. Subsequently, affinity-based purification resulted in 1 polypeptide from GSNOR-A and 2 polypeptides from GSNOR-B. Size exclusion-HPLC confirmed 3 GSNORs with molecular weight of 87.48 ± 2.74 KDa (GSNOR-A); 87.36 ± 3.25 and 82.74 ± 2.75 kDa (GSNOR-B). Kinetic analysis showed Km of 118 ± 11 μM and Vmax of 287 ± 22 nkat/mg for GSNOR-A, whereas Km of 96.4 ± 8 μM and Vmax of 349 ± 15 nkat/mg for GSNOR-B. S-nitrosylation and inhibition by NO showed redox regulation of all BjGSNORs. Both purified GSNORs exhibited variable denitrosylation efficiency as depicted by Biotin Switch assay. To the best of our knowledge, this is the first report confirming multiple isoforms of GSNOR in B. juncea |
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| Beschreibung: | Date Completed 18.03.2024 Date Revised 18.03.2024 published: Print-Electronic Citation Status MEDLINE |
| ISSN: | 1873-2690 |
| DOI: | 10.1016/j.plaphy.2024.108404 |