One-dimensional and two-dimensional immobilized metal affinity electrophoresis

Abstract

Immobilized metal affinity electrophoresis (IMAEP) is a straightforward method in which metal ions are embedded in a polyacrylamide gel strip with a negligible electrophoretic migration. Due to the preferential binding between metal ions and the phosphate group, this method uses immobilized metal ions like iron, manganese, aluminum, or titanium to capture phosphoproteins from a mixture of phosphoprotein and nonphosphoproteins. IMAEP has also been incorporated into a traditional two-dimensional (2D) sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) system (isoelectric focusing-PAGE) to increase its resolving power. In 2D IMAEP, the metal ions in polyacrylamide gel strip are overlaid on top of the second dimensional polyacrylamide gel to stop electrophoretic migration of phosphoproteins. Data shows that there is no detrimental effect of SDS in IMAEP on the extraction of phosphoproteins from a mixture of proteins. In addition, SDS exposes phosphate groups by unfolding the phosphoproteins to facilitate metal ion-phosphate binding while supplying the protein with negative charges. © 2012 Springer Science+Business Media, LLC.