Solvent effects on allosteric equilibria: stabilization of T and R conformations of Escherichia coli aspartate transcarbamylase by organic solvents

Abstract

The activity of E. coli aspartate transcarbamylase (ATCase) is markedly influenced by the addition or organic solvents to the assay medium. The cosolvents tested, which include simple aliphatic alcohols, amides and ureas, as well as acetone and dioxane, fall into 2 different classes: the most polar ones (formamide, acetamide, N-methylformamide and urea) stimulate the enzyme activity for all concentrations tested. Solvents that are less polar than water inhibit the enzyme at low concentrations but stimulate it at higher concentrations. No comparable effects are observed in the case of the isolated catalytic subunits, a nonregulated form of ATCase. Extensive kinetic studies on ATCase and on 2 of its Michaelian derivatives, 2-thioU-ATCase and carbamylated ATCase, indicate that solvents modulate the same allosteric transition that is responsible for homotropic interactions between the catalytic sites. The stabilization of the R state of ATCase by comparatively high concentrations of cosolvents is reminiscent of similar findings made on Hb and glycogen phosphorylase, suggesting a common underlying mechanism. Addition of organic cosolvents to water is known to reduce hydrophobic interactions, and this effect may preferentially stabilize the more relaxed conformations of allosteric proteins, because they have a larger surface exposed to solvent. The stabilization of the T state by low concentrations of all but the most polar cosolvents simply reflects stronger electrostatic interactions in this conformation.