Electrokinetic detection of single-molecule phosphorylation

Current single-molecule fluorescence experiments lack direct sensitivity to molecular phosphorylation state. We show that electrokinetic properties measured with an anti-Brownian trap resolve the number of phosphorylated sites on individual biomolecules and can monitor phosphorylation cycles with single kinase turn overs. As an application, we directly visualize the rate limiting step in the kinase catalytic cycle.