Project PXD006257

PRIDE Assigned Tags:
Biological Dataset

Summary

Title

Proteome profiling of Cumin by Q-Exactive HF mass spectrometer

Description

A shotgun proteomic approach was used to obtain comprehensive dataset of cumin proteins. For this purpose we applied a one dimensional electrophoresis coupled with high resolution mass spectrometry. The first step in our proteome analysis strategy, we evaluate different extraction methods, based on differential solubility of protein. Among those methods, Readyprep protein extraction kit provide high purity of protein extract, gave higher protein yield and typically good resolution of protein bands on one dimensional gel electrophoresis. Further nanoLC-MS/MS analysis by state-of-the-art Q Exactive HF mass spectrometer resulted in identification of very high number proteins. Bioinformatics analysis provide functional distribution of identified protein. To our knowledge, this is the first study aimed for in depth study of cumin proteome.

Sample Processing Protocol

In present study we compared three different extraction methods for protein sample preparation. First method utilize sodium acetate extraction at slightly acidic conditions (pH 5.5) and ammonium sulfate (SA/AS) precipitation. Second method uses Tris saturated Phenol solution for protein extraction followed by precipitation with methanolic ammonium acetate. We also used Readyprep total protein extraction kit from BioRad and evaluate its suitability for cumin proteins. Total extracted proteins was separated on one-dimensional gel electrophoresis (1-DE) and the entire gel lane was excised and divided into slices prior to the proteolytic digestion using trypsin. Peptides collected from tryptic digestion different slices, were subjected to further analysis on nanoLC system prior to MS/MS analysis. The use of modern state-of-the-art Q Exactive HF mass spectrometer allowed identification of several proteins.

Data Processing Protocol

All raw data files from Xcalibur software were directly analyzed by MaxQuant (version 1.3.0.5) using the Andromeda Search engine against the Viridiplantae database from UniProt. The intensity-based absolute quantification (iBAQ) in MaxQuant was performed on the identified peptides to quantify protein abundance.

Contact

Uzma Rana, Dow University of Health Sciences
Uzma Zaman, Dow research institute of biotechnology and biomedical sciences ( lab head )

Submission Date

04/04/2017

Publication Date

21/11/2017

Tissue

Not available

Instrument

Q Exactive

Software

Not available

Quantification

Not available

Experiment Type

Shotgun proteomics

Publication

    Zaman U, Urlaub H, Abbasi A. Protein Profiling of Non-model Plant Cuminum cyminum by Gel-Based Proteomic Approach. Phytochem Anal. 2017 PubMed: 29148164