Ivanka M Karadzic | PubMed Health

PubMed2005

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

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.

Acetone↗

PubMed2004

Proteasomes: perspectives from the Archaea.

The development of whole systems approaches to microbiology (e.g. genomics and proteomics) has facilitated a global view of archaeal physiology. Surprisingly, as archaea respond to environmental signals, the majority of protein concentration changes that occur are not reflected at the mRNA level. This incongruity highlights the importance of post-transcription control mechanisms in these organisms. One of the central players in proteolysis is the proteasome, a multicatalytic energy-dependent protease. Proteasomes serve both proteolytic and non-proteolytic roles in protein quality control and in the regulation of cell function. The proteolytic active sites of these enzymes are housed within a central chamber of an elaborate nanocompartment termed the 20S proteasome or core particle. Axial gates, positioned at each end of this particle, restrict the type of substrate that can access the proteolytic active sites. Assortments of regulatory AAA complexes are predicted to recognize/bind and unfold substrate proteins, open the axial gates, and translocate substrate into the 20S core particle.

Archaea↗