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A novel Bicine running buffer system for doubled sodium dodecyl sulfate – polyacrylamide gel electrophoresis of membrane proteins
Author(s) -
Williams Taufika Islam,
Combs Jennifer C.,
Thakur Anup P.,
Strobel Herbert J.,
Lynn Bert C.
Publication year - 2006
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.200500730
Subject(s) - sodium dodecyl sulfate , buffer (optical fiber) , polyacrylamide gel electrophoresis , gel electrophoresis , chromatography , chemistry , gel electrophoresis of proteins , sodium , electrophoresis , membrane protein , polyacrylamide , two dimensional gel electrophoresis , membrane , biochemistry , proteomics , enzyme , computer science , gene , telecommunications , organic chemistry , polymer chemistry
A novel, Bicine‐based SDS‐PAGE buffer system was developed for the analysis of membrane proteins. The method involves molecular weight‐based separations of fully denatured and solubilized proteins in two dimensions. This doubled SDS‐PAGE (dSDS‐PAGE) approach produced a diagonal arrangement of protein spots and successfully circumvented problems associated with membrane proteome analysis involving traditional gel‐based methods. Membrane proteins from the anaerobic bacterium Clostridium thermocellum were used for these investigations. Tricine‐dSDS‐PAGE and the newly developed Bicine‐dSDS‐PAGE were compared with the standard glycine‐dSDS‐PAGE (Laemmli protocol) in their suitability to separate C. thermocellum membrane proteins. Large‐format gel experiments using optimized gel preparation and running buffer conditions revealed a 112% increase in protein spot count for Tricine‐dSDS‐PAGE and a 151% increase for Bicine‐dSDS‐PAGE, compared to glycine‐dSDS‐PAGE. The data clearly indicated that Bicine‐dSDS‐PAGE is a superior method for the analysis of membrane proteins, providing enhanced resolution and protein representation.