Highly Efficient Single-Step Enrichment of Low Abundance Phosphopeptides from Plant Membrane Preparations

Overview

Journal Front Plant Sci

Date 2017 Oct 19

PMID 29042862

Citations 10

Authors

Xu Na Wu

Lin Xi

Heidi Pertl-Obermeyer

Zhi Li

Liang-Cui Chu

Waltraud X Schulze

Affiliations

Soon will be listed here.

Abstract

Mass spectrometry (MS)-based large scale phosphoproteomics has facilitated the investigation of plant phosphorylation dynamics on a system-wide scale. However, generating large scale data sets for membrane phosphoproteins usually requires fractionation of samples and extended hands-on laboratory time. To overcome these limitations, we developed "ShortPhos," an efficient and simple phosphoproteomics protocol optimized for research on plant membrane proteins. The optimized workflow allows fast and efficient identification and quantification of phosphopeptides, even from small amounts of starting plant materials. "ShortPhos" can produce label-free datasets with a high quantitative reproducibility. In addition, the "ShortPhos" protocol recovered more phosphorylation sites from membrane proteins, especially plasma membrane and vacuolar proteins, when compared to our previous workflow and other membrane-based data in the PhosPhAt 4.0 database. We applied "ShortPhos" to study kinase-substrate relationships within a nitrate-induction experiment on roots. The "ShortPhos" identified significantly more known kinase-substrate relationships compared to previous phosphoproteomics workflows, producing new insights into nitrate-induced signaling pathways.

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