Mass spectrometry-based peptide sequencing and an evaluation of the enzymatic properties of the I. walleriana extrafloral nectar proteome




Richardson, Luke

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For many years, ecologists have observed how plants mediate mutualist interactions with aggressive defenders against herbivores, such as ants, through production of metabolite-rich extrafloral nectar (EFN) as a benefit to the ant defender. However, this nectar can also support unwanted microbial contamination detrimental to the plant. Recent studies have elucidated enzymatic mechanisms in nectar by which plants protect vulnerable tissues from microbial infestation. As a novel subject of interest, relatively few nectar proteomes have been fully evaluated and characterized. The EFN proteome of Impatiens walleriana, a common flowering plant native to East Africa, falls into this group. Given its abundant secretion of EFN, attractiveness to insects, including mosquitoes, and robust expression of nectar proteins, I. walleriana was determined by the Kearney lab to be an excellent candidate to serve as a transgenic model system for study of nectar-mediated delivery of insecticidal proteins. In pursuance of this objective, the purposes of this study are twofold: firstly, to generate and identify nectar protein, and then genomic, sequence data, and, secondly, evaluate the enzymatic properties of the I. walleriana EFN proteome in its defense of nectary tissues against microbial infestation. We collected extensive peptide sequence data via bottom-up proteomic analysis of I. walleriana EFN proteins with tandem mass spectrometry in expectation of identifying peptides with a transcriptome-derived protein sequence database. Additionally, I found that the EFN had mild antimicrobial activity on E. coli but a much more pronounced effect on yeast growth. Furthermore, activity indicating the presence of peroxidase class enzyme was detected and quantified in the I. walleriana EFN and implicated in microbial defense.



Bottom-up proteomics, Tandem mass spectrometry, peroxidase, Impatiens walleriana