An electroosmotic flow-free two-direction migration strategy enables fast affinity capillary electrophoresis to study...

Affinity Capillary Electrophoresis (ACE) is a useful analytical tool to study noncovalent interactions. However, it remains challenging for ACE to measure weak and unstable interactions due to the fast dissociation of the binding complex and the possible destruction of the complex by a high electric field. In this study, we proposed a two-direction migration strategy that enables ACE to detect weak and unstable but important interactions by decreasing the migration distance of the binding complex and controlling the opposite migration direction of the free probe. By synthesizing a polyacrylamide-coated neutral capillary, free of electroosmotic flow, two-direction CE migration of basic peptides (positively charged) and peptide-RNA complexes (negatively charged) was achieved. Furthermore, the weak interactions between small nuclear U2 RNA and histone peptides were detected by this two-direction migration CE approach. The effects of the methylation states of histone peptides on the weak peptide-RNA interactions were also explored by this new approach. Collectively, the suggested modification of the ACE method is able to qualitatively characterize weak interactions.