Wednesday, January 17, 2007

What urea does to water

Urea is a model small hydrophilic solute, but also a denaturant for proteins. Both of these things make it of much interest to know how urea is solvated. It’s commonly said that urea acts as a ‘structure breaker’ of water, promoting the formation of a dense, disordered liquid structure. That suggestion is reiterated, but then challenged, in a paper in JPC B by Yoshihito Hayashi and colleagues from the labs of Sony in Japan. They’ve used dielectric spectroscopy to study the solvation of urea, and conclude that urea ‘fits’ rather easily into the tetrahedrally H-bonded structure of water and thus doesn’t greatly disrupt the structure. It seems that dielectric spectroscopy is a somewhat blunt tool for probing such questions, but as Hayashi et al. point out, their conclusions are consistent with those of Alan Soper and colleagues (Biophys. Chem. 105, 649; 2003), who studied the problem using neutron scattering. Strangely, perhaps, Hayashi et al. examine the question using the framework of the two-state model of Frank and Franks (J. Chem. Phys. 48, 4746; 1968), which posits an ‘ice-like ordered’ phase of water coexisting with a ‘dense disordered’ phase. I don’t know that anyone really thinks such a model is realistic any longer for water at room temperature… Neither am I convinced that the whole concept of structure-making and -breaking is very helpful for understanding hydration of solutes.

Anyway, here’s what I say about Soper’s study, and the more general issue of urea hydration, in my review article:

A neutron-scattering study shows that the urea molecule can ‘substitute’ quite readily for water in the hydrogen-bonded network: the radial distribution function of urea around water in a 1:4 solution looks remarkably like that of water around water. Although urea has nearly three times the molecular volume of water, the structure of liquid water is sufficiently ‘open’ that a urea molecule appears to displace just two waters, offering up to eight hydrogen bonds in place of the displaced pair. Despite this apparently ‘easy’ substitution, however, incorporating urea into the network appears to disrupt it, creating a local compression of the second hydration shell around water molecules in a manner similar to the effect of high pressure on the liquid. Yet the orientational dynamics of the water molecules seem largely unaffected even at urea concentrations high enough for all the water to be part of hydration shells. Only one water molecule per urea, on average, has a significantly slower reorientational time constant (about six times that of bulk water) – which can be rationalized according to a hydration structure in which one water is complexed to the urea molecule via two hydrogen bonds.

Postscript: Dave Thirumalai at the University of Maryland has drawn my attention to some previous papers that he and others have published on the solvation of urea and its influence in denaturation. In JACS 120, 427 (1998), he finds from MD simulations that urea, far from destabilizing the hydrophobic interaction, actually stabilizes the attraction between two solvated methane molecules, which would lead to the expectation of it acting as a renaturant. But the situation is changed for hydrophilic/charged species, to which urea becomes absorbed. In proteins, this would lead to a repulsion between hydrophilic groups, causing swelling, which would expose and destabilize the ‘buried’ hydrophobic residues. This electrostatic origin of urea denaturation was supported by further calculations in 2003 (JACS 125, 1950; 2003), where Dave and Ray Mountain simulated a hydrocarbon chain in water with slight charges at each end. Urea solvates these charged chain ends, which destablizes the collapsed chain.
For the hydration of urea itself, Dave and Ray found in 2004 (JPC B 108, 6826; 2004) that hydrogen-bonding is not the whole story – there is an important excluded-volume contribution too: the larger size of urea limits its ability to form as many hydrogen bonds as is theoretically possible. The simulations also showed that the water H-bond network around urea is perturbed significantly – it is too simplistic to say merely that urea slots nicely into the network.

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