Shotgun Proteome Analysis for Insight into Secondary Metabolite Production in Muscadine Grape

Ripening of non-climacteric fruit involves complex biochemical changes that have profound importance on accumulation of phenolic compounds beneficial to human health. Muscadine grape (Vitis rotundifolia Michx.) has gained popularity because of its health benefits. Although muscadine grape berry components have been characterized, enzymes and proteins coding for enologically- and nutraceutically-important components and their expression pattern during berry ripening is poorly understood. In this study, proteome of a white muscadine (cv. Carlos) grape cultivar commonly used for wine production was characterized during berry ripening.  Berries were harvested at pre-veraison and three ripening stages, protein extracted and resolved by electrophoresis.  Trypsin digested peptides from individual gel replicates and samples were subjected to LC/MS/MS using a Thermo LTQ HPLC-nESI-LIT. The raw files were analyzed using Proteome Discoverer (version 1.4) with SequestHT and Mascot search nodes using Vitis species FASTA database, and the data was further validated by ‘Scaffold version 4.0’ software. A total of 1411 proteins showing quantitative differences between maturation stages were identified. Differential analysis of these proteins revealed that 563 proteins were involved in various biological processes and metabolic pathways. Of these, 88 proteins were involved in the production of phenolics and are expressed after veraison. Some of these proteins were also found to be responsible for flavonoid, anthocyanin, stilbene, limonoid and terpenoid biosynthesis. In addition to providing data on biochemical changes that characterize muscadine berry ripening, this study also delivers information for use in marker-assisted selection aimed to ameliorate phenolics content of muscadine grape. Supported by USDA/NIFA/CBG.