Abstract
Various secreted peptides, including peptide hormones, are present in the apoplast, but their biochemical characterization remains a challenge due to their low abundance, difficulty in extraction, and interference from numerous secondary metabolites. Here, we describe a simple and straightforward protocol for the extraction of apoplastic peptides with a high purity. This protocol takes advantage of the fact that apoplastic peptides diffuse and accumulate in the culture medium when Arabidopsis seedlings are subjected to whole-plant submerged culture. The peptides in the culture medium are efficiently recovered by o-chlorophenol extraction followed by acetone precipitation. The recovered peptides can be subjected to nano-liquid chromatography coupled to tandem mass spectrometry (nano-LC-MS/MS) without any additional clean-up. This procedure enables the structural elucidation of mature peptide hormones in the apoplast with the use of Arabidopsis plants that overexpress peptide hormone genes.
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© 2024 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature
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Ogawa-Ohnishi, M., Matsubayashi, Y. (2024). Extraction of Apoplastic Peptides for the Structural Elucidation of Mature Peptide Hormones in Arabidopsis. In: Schaller, A. (eds) Plant Peptide Hormones and Growth Factors. Methods in Molecular Biology, vol 2731. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3511-7_6
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DOI: https://doi.org/10.1007/978-1-0716-3511-7_6
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Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-3510-0
Online ISBN: 978-1-0716-3511-7
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