Journal Highlight: A robust protocol for the isolation of cellular proteins from Xanthomonas campestris to analyze the methionine effect in 2D-gel experiments

Skip to Navigation

Ezine

  • Published: Oct 30, 2017
  • Author: separationsNOW
  • Channels: Proteomics & Genomics
thumbnail image: Journal Highlight: A robust protocol for the isolation of cellular proteins from <em>Xanthomonas campestris</em> to analyze the methionine effect in 2D-gel experiments

A phenol extraction protocol developed for the purification of bacterial proteins that provides excellent 2D gel separation has been illustrated by examining the effects of methionine on the cellular proteome of Xanthomonas campestris.

A robust protocol for the isolation of cellular proteins from Xanthomonas campestris to analyze the methionine effect in 2D-gel experiments

Electrophoresis, 2017, 38, 2603-2609
Fabian Schulte, Markus Hardt and Karsten Niehaus

Abstract: Two-dimensional PAGE (2D-PAGE) is a key technique for the separation of complex protein samples to survey the protein inventory of prokaryotic microorganisms. Although the preparation of proteins is a critical step in 2D gel experiments, its effect on the outcome of proteome data is often underestimated. In this work, we show that the choice of protein isolation method can have a profound impact on the quality of 2D gels of protein extracts from prokaryotes. Based on Xanthomonas campestris, a commercially relevant producer of the thickening agent xanthan, we validated a phenol extraction protocol for the purification of bacterial proteins that provides excellent 2D gel separation. As a proof of concept, this method was used to study the effect of methionine—a medium compound that reduces the xanthan output of industrial fermentations—on the cellular proteome of Xanthomonas. The detection of nine regulated proteins associated with sulfur metabolism (Cgl, CysI, CysJ, CysK, MetH1, MetY) and sugar nucleotide biosynthesis (Pgi, Ugd, XanA) proved the efficiency of phenol extraction for the screening of statistically significant abundance changes in 2D gel spots and MALDI-TOF-MS based identification in bacteria. Since this method is very robust, it may be useful for the study of other prokaryotes that are relevant in industrial biotechnology.

  • This paper is free to view for all users registered on separationsNOW.com until the end of January 2018.
    After this time, you can purchase it using Pay-Per-View on Wiley Online Library.

Follow us on Twitter!

Social Links

Share This Links

Bookmark and Share

Microsites

Suppliers Selection
Societies Selection

Banner Ad

Click here to see
all job opportunities

Most Viewed

Copyright Information

Interested in spectroscopy? Visit our sister site spectroscopyNOW.com

Copyright © 2017 John Wiley & Sons, Inc. All Rights Reserved