Application of capillary electrophoresis for DNA-protein binding tests

Abstract

The combination of affinity chromatography and capillary electrophoresis (CE-SDS) has been found to be a useful tool to analyse populations of proteins which specifically bind to ssor dsDNA. Proteins were extracted from tissue, cytosol or nuclei of meristems ofPisum sativum seedlings and separated on cellulose column functionalized with ss-, dsDNA (calf thymus) and ssDNA (P. sativum) at 2M concentration of sodium chloride. Electropherograms of the crude protein extracts show two fractions of proteins specific for dsDNA (calf thymus) and three fractions specific for ssDNA (calf thymus). Four and five fractions of proteins specific for ssDNA (P. sativum) were identified in the material isolated from cytosolic and nuclear extracts, respectively. Both ds-and ssDNA (calf thymus) form complexes with ca. 4.0 % of the total amount of proteins, while ssDNA (P. sativum) binds to ca. 11.0 % of cytosolic and 5.0 % of nuclear proteins.List of abbreviations: CE -capillary electrophoresis; DSB -double-stranded DNA-binding protein ds -double-stranded; ss -single-stranded; SSBsingle-stranded DNA-binding protein