Fluorimetric-Based Method to Detect and Quantify Total S-Nitrosothiols (SNOs) in Plant Samples
Accumulating experimental evidence indicates that S-nitrosylation (technically S-nitrosation) events have a central role in plant biology, presumably accounting for much of the widespread influence of nitric oxide (NO) on developmental, metabolic, and stress-related plant responses. Therefore, the accurate detection and quantification of S-nitrosylated proteins and peptides can be particularly useful to determine the relevance of this class of compounds in the ever-increasing number of NO-dependent signaling events described in plant systems. Up to now, the quantification of S-nitrosothiols (SNOs) in plant samples has mostly relied on the Saville reaction and the ozone-based chemiluminescence method, which lacks sensitivity and are very time-consuming, respectively. Taking advantage of the photolytic properties of S-nitrosylated proteins and peptides, the method described in this chapter allows simple, fast, and high-throughput detection of SNOs in plant samples.
Key wordsOzone chemiluminescence Saville S-nitrosothiols S-nitrosoglutathione Nitric oxide Nitrosylated proteins
Funding: This work was supported by Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) [grant numbers 2011/50637-0, 2013/18056-2 and 2013/15108-1], Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) [grant numbers 442045/2014-0, and 309504/2014-7], Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) [grant number 99999.006262/2015-05] and CNCSIS/UEFISCDI [grant PN-II-RU-TE-2014-4-2555]. Research in FJC laboratory is supported by the ERDF-cofinanced grants from the Ministry of Economy and Competitiveness (AGL2015-65104-P) and Junta de Andalucía (group BIO192), Spain.
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