Volume Exclusion and H-Bonding Dominate the Thermodynamics and Solvation of Trimethylamine-N-oxide in Aqueous Urea

Abstract

Trimethylamine-N-oxide (TMAO) and urea represent the extremes among the naturally occurring organic osmolytes in terms of their ability to stabilize/destabilize proteins. Their mixtures are found in nature and have generated interest in terms of both their physiological role and their potential use as additives in various applications (crystallography, drug formulation, etc.). Here we report experimental density and activity coefficient data for aqueous mixtures of TMAO with urea. From these data we derive the thermodynamics and solvation properties of the osmolytes, using Kirkwood–Buff theory. Strong hydrogen-bonding at the TMAO oxygen, combined with volume exclusion, accounts for the thermodynamics and solvation of TMAO in aqueous urea. As a result, TMAO behaves in a manner that is surprisingly similar to that of hard-spheres. There are two mandatory solvation sites. In plain water, these sites are occupied with water molecules, which are seamlessly replaced by urea, in proportion to its volume fraction. We discuss how this result gives an explanation both for the exceptionally strong exclusion of TMAO from peptide groups and for the experimentally observed synergy between urea and TMAO.

Probing volumetric properties of biomolecular systems by pressure perturbation calorimetry (PPC) – The effects of hydration, cosolvents and crowding

“…m o is highest for 1 M TMAO and lowest for 1 M urea. These results are in good agreement with previous data reported for room temperature [42,43].…”

Counteracting chemical chaperone effects on the single-molecule α-synuclein structural landscape

“…Isotopic substitution neutron-scattering measurements of ternary-component urea-TMAO mixtures in water previously suggested that TMAO interacts directly with urea through preferential hydrogen bonding (59). However, recent studies show that the interaction affinity between the two osmolytes is too weak to be relevant even at high osmolyte concentrations (60,61). Therefore, we favor a model in which the free energy of transfer of the protein in TMAO solutions is opposite of that in urea solutions, with the free energies being additive in urea-TMAO mixtures.…”

A current perspective on the compensatory effects of urea and methylamine on protein stability and function

“…In another development, it has also been shown that TMAO becomes more potent stabilizer of protein in the presence of urea, making urea less efficient in the presence of TMAO. This is because TMAO has two mandatory solvation sites that can be occupied by urea substantially enlarging the effective hard sphere diameter (Rosgen and Jackson-Atogi, 2012).…”