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  • Improvement of two-dimensional gel electrophoresis proteome maps of the haloarchaeon Haloferax volcanii.

Improvement of two-dimensional gel electrophoresis proteome maps of the haloarchaeon Haloferax volcanii.

Proteomics (2005-01-04)
Ivanka M Karadzic, Julie A Maupin-Furlow
ABSTRACT

Proteins of haloarchaea are remarkably unstable in low-ionic-strength solvents and tend to aggregate under standard two-dimensional (2-D) gel electrophoresis conditions, causing strong horizontal streaking. We have developed a new approach to generate 2-D maps of halophilic proteins which included washing cells with 1.5 M Tris-HCl buffer. In addition, proteins were precipitated with acetone, solubilized with urea and thiourea in the presence of the sulfobetaine detergent 3-[(3-cholamidopropyl)dimethylamino]-1-propanesulfonate (CHAPS), reduced with tributylphosphine (TBP), and separated with microrange strips of immobilized pH gradients (pH 3.9-5.1). This combination enabled the construction of highly reproducible 2-D maps of Haloferax volcanii proteins.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Tri-n-butylphosphine, 99%
Sigma-Aldrich
Tributylphosphine solution, 200 mM (in N-methyl-2-pyrrolidinone), liquid
Sigma-Aldrich
NDSB 211, ≥98% (TLC)