Capillary sieving electrophoresis-micellar electrokinetic chromatography fully automated two-dimensional capillary electrophoresis analysis of Deinococcus radiodurans protein homogenate.
We report the one- and two-dimensional (1-D and 2-D) capillary electrophoresis separation of Deinococcus radiodurans protein homogenate. Proteins are labeled with the fluorogenic reagent 3-(2-furoyl)quinoline-2-carboxaldehyde (FQ), which reacts with lysine residues and creates a highly fluorescent product. Detection is by laser-induced fluorescence. 1-D capillary sieving electrophoresis (CSE) produces over 150,000 plates and micellar electrokinetic capillary chromatography (MEKC) produces over 900,000 plates for components in a D. radiodurans protein homogenate. In a 2-D separation, proteins are first separated by CSE. Fractions are repetitively transferred to a second capillary for further separation based on MEKC. The 2-D separation has a approximately 550 spot capacity. Over 150 components are partially resolved from the homogenate. Resolution is limited in the first dimension by diffusion of proteins during the long separation period and in the second dimension by the combination of a long fraction-transfer time and short separation period.
Department of Chemistry, University of Washington, Seattle, WA 98915-1700, USA.
David A
DA
Michels
Shen
S
Hu
K Amy
KA
Dambrowitz
Michael J
MJ
Eggertson
Kevin
K
Lauterbach
Norman J
NJ
Dovichi
D001331:Automation; D001426:Bacterial Proteins / Q000032:analysis*; D020374:Chromatography, Micellar Electrokinetic Capillary / Q000379:methods*; D034301:Deinococcus / Q000737:chemistry*; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*
D001426:Bacterial Proteins