Measurement of the contributions of 1D and 3D pathways to the translocation of a protein along DNA

Abstract

Proteins that act at specific DNA sequences bind DNA randomly and then translocate to the target site. The translocation is often ascribed to the protein sliding along the DNA while maintaining continuous contact with it. Proteins also can move on DNA by multiple cycles of dissociation͞reassociation within the same chain. To distinguish these pathways, a strategy was developed to analyze protein motion between DNA sites. The strategy reveals whether the protein maintains contact with the DNA as it transfers from one site to another by sliding or whether it loses contact by a dissociation͞reassociation step. In reactions at low salt, the test protein stayed on the DNA as it traveled between sites, but only when the sites were <50 bp apart. Transfers of >30 bp at in vivo salt, and over distances of >50 bp at any salt, always included at least one dissociation step. Hence, for this enzyme, 1D sliding operates only over short distances at low salt, and 3D dissociation͞ reassociation is its main mode of translocation.DNA-protein interaction ͉ recognition sequence ͉ restriction enzyme

Dynamical model of DNA-protein interaction: Effect of protein charge distribution and mechanical properties

“…More precisely, the improved model predicts, like our original one, that DNA sampling proceeds via a succession of 3D motion in the cell, 1D sliding along the DNA sequence, short or long hops between neighboring or more widely separated sites, and intersegmental transfers. This behavior is confirmed by recent experiments [10][11][12][13][14][15][16][17][18][19] and is one the key assumptions of kinetic models. Quite interestingly, this behavior is however not an assumption for molecular mechanical models but rather a consequence of the form of DNA-protein interactions and evolution equations.…”

“…Still, it should be mentioned that the average number of beads visited during each sliding event (5 to 10 beads, that is from 75 to 150 base pairs) is in fairly good agreement with experimental results, which lie in the range 30 to 170 base pairs [18,19]. (3.3)) applies only to diffusive motion.…”

“…The development of new techniques for in vivo microscopy has recently permitted the direct visualization of the motion of proteins inside the cell and the precise determination of their diffusion coefficients [10][11][12][13][14][15][16][17][18][19]. Nonetheless, the exact mechanism of non-specific DNA-protein interactions still remains unclear, because proteins come into a…”

Stochastic models of intracellular transport

“…The BHW model and its extensions provide a plausible mechanism for facilitated diffusion that has some support from experimental studies, which demonstrate that proteins do indeed slide along DNA (Gorman and Greene, 2008;Gowers et al, 2005;Li et al, 2009;Tafvizi et al, 2008;Winter et al, 1981). In particular, recent advances in single-molecule spectroscopy means that the motion of flourescently labeled proteins along DNA chains can be quantified with high precision, although it should be noted that most of these studies have been performed in vitro.…”

Protein–DNA Electrostatics

“…In Ref. [49], the same S. E. Halford and co-workers showed for the restriction enzyme ecoRV that at low salt, the protein only slides continuously on DNA for distances shorter than 50 base pairs. Transfers of more than 30 base pairs at in vivo salt, and over distances of more than 50 base pairs at any salt, always included at least one dissociation step.…”