# Solution Ionic Strength Can Modulate Functional Loop Conformations in E. coli Dihydrofolate Reductase

## TL;DR

This study shows that the ionic strength of the solution can affect the shape of a functional loop in the E. coli DHFR enzyme, influencing its structure and function.

## Contribution

The study reveals that solution ionic strength modulates the conformational behavior of the M20 loop in ecDHFR, independent of ligand binding.

## Key findings

- High ionic strength creates a free energy barrier between occluded and closed M20 loop states.
- Structures of ecDHFR at high ionic strength may not reflect physiological conditions.
- Lower ionic strength allows the M20 loop to adopt open or partially closed conformations.

## Abstract

The observation of multiple conformations of a functional
loop
(termed M20) in the Escherichia coli dihydrofolate reductase (ecDHFR) enzyme triggered
the proposition that large-scale motions of protein structural elements
contribute to enzyme catalysis. The transition of the M20 loop from
a closed conformation to an occluded conformation was thought to aid the rate-limiting release of the
products. However, the influence of charged species in the solution
environment on the observed M20 loop conformations, independent of
charged ligands bound to the enzyme, had not been considered. Molecular
dynamics simulations of ecDHFR in model CaCl2 solutions of varying molar ionic strengths IM reveal a substantial free energy barrier between occluded and closed M20 loop states at IM exceeding the E. coli threshold (∼0.24 M). This barrier may facilitate crystallization
of ecDHFR in the occluded state,
consistent with ecDHFR structures obtained at IM exceeding 0.3 M. At lower IM (≤0.15 M), the M20 loop can explore the occluded state, but prefers an open/partially closed conformation, again consistent with ecDHFR structures. Our findings caution against using ecDHFR structures obtained at nonphysiological ionic strengths
in interpreting catalytic events or in structure-based drug design.