Conformational Change of Transcription Factors from Search to Specific Binding: A lac Repressor Case Study

Abstract

In a process known as facilitated diffusion, DNA-binding proteins find their target sites by combining three-dimensional diffusion and one-dimensional scanning of the DNA. Following the trade-off between speed and stability, agile exploration of DNA requires loose binding, whereas, at the DNA target site, the searching protein needs to establish tight interactions with the DNA. To enable both efficient search and stable binding, DNA-binding proteins and DNA often switch conformations upon recognition. Here, we study the one-dimensional diffusion and DNA binding of the dimeric lac repressor (LacI), which was reported to adopt two different conformations when binding different conformations of DNA. Using coarse-grained molecular dynamic simulations, we studied the diffusion and the sequence-specific binding of these conformations of LacI, as well as their truncated or monomeric variants, with two DNA conformations: straight and bent. The simulations were compared to experimental observables. This study supports that linear diffusion along DNA combines tight rotation-coupled groove tracking and rotation-decoupled hopping, where the protein briefly dissociates and reassociates just a few base pairs away. Tight groove tracking is crucial for target-site recognition, while hopping speeds up the overall search process. We investigated the diffusion of different LacI conformations on DNA and show how the flexibility of LacI’s hinge regions ensures agility on DNA as well as faithful groove tracking. If the hinge regions instead form α-helices at the protein–DNA interface, tight groove tracking is not possible. On the contrary, the helical hinge region is essential for tight binding to bent, specific DNA, for the formation of the specific complex. Based on our study of different encounter complexes, we argue that the conformational change in LacI and DNA bending are somewhat coupled. Our findings underline the importance of two distinct protein conformations for facilitated diffusion and specific binding, respectively.

Can molecular dynamics be used to simulate biomolecular recognition?

“…64 Despite this, coarse-grained simulations have shown that the protonation of His29 is essential for the sliding motion during the search process. 37 The systems were simulated in the NPT ensemble at 310 K, the optimal growth temperature for E. coli. 65 The protein and DNA were solvated in a solution with a 150 mM concentration of KCl and a 5 mM concentration of magnesium ions.…”

“…All these processes are crucial for the repressor to find the target site quickly and efficiently. [34][35][36][37] In fact, it would seem that the processes the repressor performs are balanced in a way that maximizes the likelihood of encountering the operator site once the repressor is non-specifically bound to the DNA in the proximity of the operator.…”

“…The most important of these mechanisms for our purposes, however, is sliding because the repressor recognizes the specific site and consequently transitions into the specific complex while undergoing this process. 35,37 That recognition only takes place while the repressor is sliding is evidenced by the so-called antenna effect, which effectively increases the size of the target to the sliding length. 29,31,39 Figure 3 illustrates the free energy landscape for the reaction mechanism that was described in the last paragraph.…”

Diffusion-Controlled Reactions: An Overview

“…Varying p from 0 to 1 covers the whole range of reactivities from 0 to +∞. In the biochemical context, conformational changes of a macromolecule between nearly isoenergetic folded states can alter its function; this mechanism is primarily important for protein-ligand and protein-protein recognition [48][49][50]. When such a protein arrives onto the catalytic surface (its reaction partner), it has to be in an appropriate conformational state (with probability p) to be able to initiate the reaction (4); otherwise, the protein leaves the catalytic surface and restarts its bulk diffusion [51].…”