Sliding of a single lac repressor protein along DNA is tuned by DNA sequence and molecular switching

In any living cell, genome maintenance is carried out by DNA-binding proteins that recognize specific sequences among a vast amount of DNA. This in- cludes fundamental processes such as DNA replica- tion, DNA repair, and gene expression and regulation. Here, we study the mechanism of DNA target search by a single lac repressor protein (LacI) with ultrafast force-clamp spectroscopy, a sub-millisecond and few base-pair resolution technique based on laser tweez- ers. We measure 1D-diffusion of proteins on DNA at physiological salt concentrations with 20 bp res- olution and find that sliding of LacI along DNA is sequence dependent. We show that only allosteri- cally activated LacI slides along non-specific DNA se- quences during target search, whereas the inhibited conformation does not support sliding and weakly in- teracts with DNA. Moreover, we find that LacI under- goes a load-dependent conformational change when it switches between sliding and strong binding to the target sequence. Our data reveal how DNA sequence and molecular switching regulate LacI target search process and provide a comprehensive model of fa- cilitated diffusion for LacI.