Showing papers by "James W. Jorgenson published in 2016"
TL;DR: The most popular techniques for extraction, enrichment, solubilization, and digestion are compared, resulting in an overall improved workflow for the insoluble portion of Saccharomyces cerevisiae cell lysate.
Abstract: The importance of membrane proteins in biological systems is indisputable; however, their amphipathic nature makes them difficult to analyze. In this study, the most popular techniques for extraction, enrichment, solubilization, and digestion are compared, resulting in an overall improved workflow for the insoluble portion of Saccharomyces cerevisiae cell lysate. Yeast cells were successfully lysed using a French press pressure cell at 20 000 psi, and resulting proteins were fractionated prior to digestion to reduce sample complexity. The proteins were best solubilized with the addition of ionic detergent sodium deoxycholate (1%) and through the application of high-frequency sonication prior to a tryptic digestion at 37 °C. Overall, the improved membrane proteomic workflow resulted in a 26% increase in membrane protein identifications for baker’s yeast. In addition, more membrane protein identifications were unique to the improved protocol. When comparing membrane proteins that were identified in the impr...
36 citations
TL;DR: The effect of sonication while packing very high slurry concentrations is presented and three of the columns underwent sonication during packing and yielded highly efficient separations with reduced plate heights as low as 1.05.
34 citations
TL;DR: This work introduces a system that operates at a constant pressure and is capable of separations at pressures up to 45kpsi, and is assessed with a complex peptide mixture and a range of microcapillary columns packed with sub-2μm C18 particles.
32 citations
TL;DR: The results suggest adjustments of this concentration are required with regard to particle size, surface roughness, column dimensions, slurry liquid, and external effects utilized during the packing process (pressure protocol, ultrasound, electric fields).
28 citations
TL;DR: The development of a silica based IMER for on-line protein digestion, which produced rapid digestions in the presence of organic mobile phase for both model proteins and a complex sample consisting of the insoluble portion of a yeast cell lysate is presented.
27 citations