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Open AccessFeature PaperArticle

Protein Extraction, Enrichment and MALDI MS and MS/MS Analysis from Bitter Orange Leaves (Citrus aurantium)

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Department of Chemistry and Chemical Technologies, University of Calabria, 87036 Arcavacata di Rende, Italy
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Department of Pharmacy, Health and Nutritional Sciences, University of Calabria, 87036 Arcavacata di Rende, Italy
3
Department of Chemistry, Università degli Studi di Roma La Sapienza, 00185 Rome, Italy
*
Author to whom correspondence should be addressed.
Molecules 2020, 25(7), 1485; https://doi.org/10.3390/molecules25071485
Received: 26 February 2020 / Revised: 21 March 2020 / Accepted: 23 March 2020 / Published: 25 March 2020
Citrus aurantium is a widespread tree in the Mediterranean area, and it is mainly used as rootstock for other citrus. In the present study, a vacuum infiltration centrifugation procedure, followed by solid phase extraction matrix-assisted laser desorption ionization tandem mass spectrometry (SPE MALDI MS/MS) analysis, was adopted to isolate proteins from leaves. The results of mass spectrometry (MS) profiling, combined with the top-down proteomics approach, allowed the identification of 78 proteins. The bioinformatic databases TargetP, SignalP, ChloroP, WallProtDB, and mGOASVM-Loc were used to predict the subcellular localization of the identified proteins. Among 78 identified proteins, 20 were targeted as secretory pathway proteins and 36 were predicted to be in cellular compartments including cytoplasm, nucleus, and cell membrane. The largest subcellular fraction was the secretory pathway, accounting for 25% of total proteins. Gene Ontology (GO) of Citrus sinensis was used to simplify the functional annotation of the proteins that were identified in the leaves. The Kyoto Encyclopedia of Genes and Genomes (KEGG) showed the enrichment of metabolic pathways including glutathione metabolism and biosynthesis of secondary metabolites, suggesting that the response to a range of environmental factors is the key processes in citrus leaves. Finally, the Lipase GDSL domain-containing protein GDSL esterase/lipase, which is involved in plant development and defense response, was for the first time identified and characterized in Citrus aurantium. View Full-Text
Keywords: Citrus aurantium; vacuum infiltration centrifugation; MALDI MS/MS; GDSLs; SPE enrichments Citrus aurantium; vacuum infiltration centrifugation; MALDI MS/MS; GDSLs; SPE enrichments
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Aiello, D.; Siciliano, C.; Mazzotti, F.; Di Donna, L.; Risoluti, R.; Napoli, A. Protein Extraction, Enrichment and MALDI MS and MS/MS Analysis from Bitter Orange Leaves (Citrus aurantium). Molecules 2020, 25, 1485.

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