Static and Dynamic Investigations of RecA-DNA Complex with a Solid-State Nanopore

Abstract

In recent years, artificial nanometer-sized pores (nanopores) have been successfully employed as a new tool to detect the passage of DNA molecules. When a molecule is driven through the nanopore by an externally applied electric field, it blocks part of the ionic current, resulting in a temporal current blockade. So far, only translocations of bare DNA have been reported. In this work, we extend this to protein-coated DNA molecules by including the well-studied RecA protein, which plays an essential role in the central steps of homologous recombination. To accomplish its DNA strand exchange activities, RecA polymerizes onto DNA to form a stiff helical nucleoprotein filament. These filaments translocate through solid-state pores, as we demonstrate unambiguously. Additionally, we use the nanopore system in conjunction with an integrated optical tweezer, which allows us to insert the RecA-DNA complex into the nanopore. We will discuss results as they pertain to both the charge structure and conformation of the complex. This work also lays the groundwork for future experiments on sequential screening of proteins, say transcription factors, which locally coat DNA.