Proteomics

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Sample preparation procedure for mass spectrometric analysis of plant histones


ABSTRACT: The protocol for preparing samples of plant histones prior to LC-MS/MS analysis was established, using a filter-aided sample preparation (FASP) technique to remove natural contaminants and chemical additives incompatible with MS-based procedures. The protocol consists of nuclei isolation, histone extraction into acidic solvent, protein propionylation and on-membrane digestion, then peptide propionylation in solution. Two variants of the derivatization protocol, in-solution propionylation in the vial (PROP-in-SOL) and propionylation on the filter unit (PROP-on-FILTER), were compared by efficiency of the labeling and by abundance of identified histone peptide forms.

INSTRUMENT(S): LTQ Orbitrap Elite

ORGANISM(S): Arabidopsis Thaliana (mouse-ear Cress)

TISSUE(S): Plant Cell

SUBMITTER: Gabriela Lochmanova  

LAB HEAD: Zbynek Zdrahal

PROVIDER: PXD010102 | Pride | 2018-09-20

REPOSITORIES: Pride

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Publications

Filter-Aided Sample Preparation Procedure for Mass Spectrometric Analysis of Plant Histones.

Ledvinová Dominika D   Mikulášek Kamil K   Kuchaříková Hana H   Brabencová Sylva S   Fojtová Miloslava M   Zdráhal Zbyněk Z   Lochmanová Gabriela G  

Frontiers in plant science 20180919


Characterization of histone post-translational modifications (PTMs) is still challenging, and robust histone sample preparation is essential for convincing evaluation of PTMs by mass spectrometry. An effective protocol for extracting plant histone proteins must also avoid excessive co-extraction of the numerous potential interfering compounds, including those related to secondary metabolism. Currently, the co-existence of histone marks is addressed mostly by shotgun proteomic analysis following  ...[more]

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