MALDI MS Imaging of Chickpea Seeds ( Cicer arietinum ) and Crab's Eye Vine ( Abrus precatorius ) after Tryptic Digestion Allows Spatially Resolved Identification of Plant Proteins.

Abstract

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) imaging following in situ enzymatic digestion is a versatile analytical method for the untargeted investigation of protein distributions, which has rarely been used for plants so far. The present study describes a workflow for in situ tryptic digestion of plant seed tissue for MALDI MS imaging. Substantial modifications to the sample preparation procedure for mammalian tissues were necessary to cater to the specific properties of plant materials. For the first time, distributions of tryptic peptides were successfully visualized in plant tissue using MS imaging with accurate mass detection. Sixteen proteins were visualized and identified in chickpea seeds showing different distribution patterns, e.g., in the cotyledons, radicle, or testa. All tryptic peptides were detected with a mass resolution higher than 60,000 as well as a mass accuracy better than 1.5 ppm root-mean-square error and were matched to results from complementary liquid chromatography-MS/MS (LC-MS/MS) data. The developed method was also applied to crab's eye vine seeds for targeted MS imaging of the toxic protein abrin, showing the presence of abrin-a in all compartments. Abrin (59 kDa), as well as the majority of proteins visualized in chickpeas, was larger than 50 kDa and would thus not be readily accessible by top-down MS imaging. Since antibodies for plant proteins are often not readily available, in situ digestion MS imaging provides unique information, as it makes the distribution and identification of larger proteins in plant tissues accessible in an untargeted manner. This opens up new possibilities in the field of plant science as well as to assess the nutritional quality and/or safety of crops.

Plant phenotyping relevance

植物種子組織中のタンパク質分布という植物状態を可視化するMALDI MSイメージング手法を開発し、複数種子で適用・検証しており、測定法が研究の中心である。

Code and data availability

The article describes MALDI MS imaging of plant proteins but contains no public deposit of imaging data, phenotype datasets, or author analysis code. The imzML Converter, imzML Analyzer, Mirion, and Pep-Calc are generic third-party tools; the Supporting Information is a PDF of supplementary figures/tables, not a data或码

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