Thursday, October 13, 2011

Lum-Chandler-Weeks under the microscope

Shekhar Garde and his colleagues have shown in several recent publications how hydration of solutes and its consequences, such as conformational changes in polymers, can be significantly altered near the air-water interface. The more general effect of interfaces on hydrophobic phenomena is now probed by Garde along with David Chandler, Amish Patel and others (A. Patel et al., PNAS doi 10.1073/pnas.1110703108 – paper here). They look at how interactions between hydrophobic solutes of various sizes from sub-nm to several nm are altered close to the interface of water with self-assembled monolayers of various surface chemistries, from hydrophobic to hydrophilic. They find that the driving force for the assembly of hydrophobic particles is smaller near a hydrophobic surface than it is in bulk, and decreases with increasing temperature, in contrast to the bulk (and to hydrophilic surfaces). This implies that hydrophobic surfaces should act as catalysts for the unfolding of proteins. It might also account for how chaperonins work: their initially hydrophobic surfaces help misfolded proteins to unfold, and then ATP-driven conversion of the walls to hydrophilic release the unfolded protein from the wall so that it might fold again in bulk.

Misfolding in the context of Lum-Chandler-Weeks theory is also the subject of a paper by Ruhong Zhou and colleagues (Z. Yang et al., J. Phys. Chem. B 115, 11137 (2011) – paper here). They have looked at whether dewetting transtions play a role in the assembly of two amyloidogenic beta-sheets. For the peptides considered there are two conformations that lead to stacking of beta-sheet pairs: one containing water between the sheets in a 2D slab-like geometry, the other with the water in a 1D tube-like geometry. In both cases the sheets are brought together by a drying transition, but surprisingly this is stronger for the slab-like than the tube-like case. This is attributed here to the different surface roughness of the two packing modes: the ‘staggered’ packing in the slab-like case, which is rougher, disrupts the H-bonding network of the intervening water to a greater extent.

Some further new insights into the Lum-Chandler-Weeks dewetting mechanism for hydrophobic interactions are offered by two recent papers. Li and Walker appear to see the cross-over region between the mechanisms of hydrophobic hydration at small (<1 nm) and large (>1 nm) scales that this theory predicts, by measuring the free energy of hydration of individual monomers of various size in hydrophobic polymers, using an AFM to pull the chains out from a collapsed conformation (Li, I. T. S. & Walker, G. C. PNAS 108, 16527-16532; 2011 – paper here). At a monomer size of around 1 nm in the temperature region of 50 C or so, they find a switch in the hydration entropy from negative to positive (this crossover size reduces to just 3.5 Å at 150 C). Shekhar Garde and Amish Patel have published a commentary on it (10.1073/pnas.1113256108).

And Garde and Patel have joined forces with David Chandler and others in a preprint ( – paper here) that attempts to unravel the issue of why some protein subunits (such as melittin) seem to aggregate via dewetting while others (such as BphC) do not. They find that at model (SAM) hydrophobic surfaces, simulations show that the statistics of large-amplitude fluctuations in the density of interfacial water are altered relative to the Gaussian stats of both bulk water and water at a hydrophilic surface. In other words, despite similar average interfacial water densities, the fluctuations reveal the proximity of the hydrophobic interface to a dewetting transition. This tuning of the interfacial water is, they argue, common to biological systems, where it induces a strong sensitivity to small changes in conformation, allowing the system to take advantage of the phase transition in engineering biomolecular function (in a manner analogous to the finely balanced wetting or drying of ion channels for ‘vapour-lock’ gating). Although melittin and BphC lie on opposite sides of this transition, small modifications to both can tip the balance one way or the other. This offers what seems to me to be a persuasive argument that dewetting is relevant to hydrophobic aggregation even if it does not exactly provide the mechanism for it in all (or even in most) cases: the transition is, if you like, ‘there’ even if it doesn’t manifest itself.

Alenka Luzar and her colleagues have considered this issue from the perspective of whether or not cavitation can take place at the protein-protein interface (J. Wang et al., PCCP 10.1039/c1cp22082a – paper here). They present a lattice model which offers a fast method for predicting if cavitation can happen, and find that part of the surface of melittin is sufficiently hydrophobic to permit this on a timescale that is consistent with that seen in the earlier simulations.

Alenka and her colleagues have also examined how this putative crossover length-scale for hydration behaviour is influenced by charge on the solute (J. Stat. Phys. 10.1007/s10955-011-0337-1 – paper here). They conclude that, for moderate charge, the electrostatic contribution to the solvation free energy is in fact essentially independent of solute curvature, because of a compensation between counterion shielding and the dielectric screening of water – the solvation free energy remains more or less a function only of solute surface area.

To what extent protein-ligand binding requires an atomistic description of changes in hydration is a crucial question, not least for attempts to design synthetic ligands in drug development. Ulf Ryde at Lund University and colleagues have looked at this issue by comparing the ability of continuum methods to predict binding free energies for four different protein-ligand pairs with quite different degrees of solvent exposure at the binding site (S. Genheden et al., JACS ja202972m – paper here). They find that the continuum methods often perform badly, particularly for cases with a greater degree of solvent exposure. We need to know precisely where the waters are and where they go.

And that is somewhat elucidated by Nan-jie Deng at Rutgers University and colleagues for the case of two synthetic inhibitors of HIV-1 protease, Nelfinavir and Amprenavir (N.-j. Deng et al., J. Phys. Chem. B jp204047b – paper here). The binding of these drugs is apparently entropically driven, but the question is where that entropic contribution comes from. A classical view would be inclined to attribute it to the release of water from the binding cleft, but it seems that this isn’t so: these MD simulations suggest that any entropy gain there is more than offset by the restriction of ligand rotation and vibration on binding. Instead, the favourable entropic contribution seems to come from desolvation of the ligand.

A different view of water’s influence on bimolecular recognition is provided by Stacey Wetmore and colleagues at the University of Lethbridge in Alberta (F. M. V. Leavens et al., J. Phys. Chem. B jp205424z – paper here). They have looked at how water molecules can affect the pi-pi interactions between DNA and DNA-binding proteins. Solvating water molecules seem to have essentially no influence on the strength of pi-pi stacking for histidine and adenine, but do weaken the interaction if the histidine is protonated. The latter interaction, however, remains in all cases stronger than the former.

Protons seem to be delivered to membrane proteins such as the proton pump cytochrome c oxidase via some kind of surface-enhanced, two-dimensional transport at the membrane surface. This has been previously postulated as a series of jumps between ionisable groups (phosphate and carbonyl) at the membrane surface. But that notion is challenged by Peter Pohl at the Johannes Kepler University in Linz and colleagues, whose fluorescence measurements of proton transfer at membranes show that proton transport can be equally fast in the absence of ionisable groups (A. Springer et al., PNAS 108, 14461-14466; 2011 – paper here). They conclude that it is probably the network of interfacial water molecules that is responsible instead for the rapid proton motion: as they say, “water structuring at the interface seems to be mandatory for providing the pathway”.

It has of course been long thought that protons may be delivered to the interior of an enzyme via chains of water molecules. That process is studied by heme peroxidase by Emma Lloyd Raven and coworkers at the University of Leicester (I. Efimov et al., JACS ja2007017 – paper here). Kinetic isotope effects reveal that the proton pathway utilizes a Grotthus-like shuttling of protons along a pathway towards the ferryl oxygen that involves three bound waters and two arginine residues.

A model system for studying such proton transfer in confined geometry is reported by Bradley Habenicht and Stephen Paddison at the University of Tennessee in Knoxville (J. Phys. Chem. B jp205787f – paper here). They use MD simulations to look at how protons are transported within carbon nanotubes whose inner walls are functionalized with perfluoro sulphonic acid groups. If these groups are spaced far apart (~8 Å), they tend to be individually hydrated by clusters of water molecules with little interaction between them, and correspondingly reduced acidic proton dissociation. But there is also a pronounced effect of confinement at small nanotube diameters: in the smaller tubes there are stronger interactions between the walls and the water molecules, which can lead to break-up of the hydrogen-bonded network of waters linking the sulphonic acid groups. That network may be restored by polarized charges of fluorine atoms attached to the nanotube walls.

ATP hydrolysis in the active cleft of actin plays an important role in the state of its filamentous form F-actin, affecting its rigidity and its binding of regulatory proteins. There is water in this active site, but it hasn’t previously been clear what, if anything, it does. Marissa Saunders and Greg Voth at Chicago have clarified this through MS simulations based on the crystal structure (J. Mol. Biol. 413, 279-291; 2011 – paper here). They say that the ordered waters help the protein to flatten and brings about a conformational change that promotes ATP hydrolysis. These changes also stabilize the charge on the phosphate and accelerate the deprotonation of the catalytic water involved in hydrolysis. In short, the bound water helps to organize the active-site geometry.

Another nail in the coffin of ‘structure-making and –breaking’: Fabio Bruni and colleagues in Rome have looked at how local solvent structure around ions affects their influence on viscosity – specifically, on how changes in ionic concentration affect the viscosity of the solution (T. Corridoni et al., J. Phys. Chem. B jp202755u – paper here). Classically, the viscosity is found to be (almost) linearly related to the concentration. It has been asserted that the magnitude of the coefficient of proportionality, denoted B, depends on how the ions perturb the water structure. Fabio et al. use neutron scattering and simulations to look for some structural parameter that can be correlated with B. They find that the nature of the (univalent) ions is all but irrelevant to the size of the percolating water clusters in solution. The change in viscosity seems to be unrelated to any structural changes in the bulk liquid, but instead pertain to changes in the local hydration shells of the ions. As a result, they say, “the particular effect of solutes ranked in the Hofmeister series must be looked at in terms of specific ion interactions with hydrophilic or hydrophobic surfaces”, and not in terms of any generalized propensity for structure-making or –breaking.

In an intriguing preprint, Jampa Maruthi Pradeep Kanth and Ramesh Anishetty at the Institute of Mathematical Sciences in Chennai propose that the hydrophobic interaction should be understood as an effect analogous to the Casimir effect (the attraction of two surfaces separated by a vacuum due to the suppression of long-wavelength electromagnetic fluctuations of the vacuum in the gap) (preprint here). Their molecular mean-field analytical method suggests that confinement alters the allowed fluctuations of the hydrogen-bond network, specifically the long-ranged correlations between water molecular orientations. Now what I’d like to know is whether an explicit connection can be made to the alleged role of fluctuations in the Lum-Chandler-Weeks model. But that seems to argue in the opposite direction, namely that fluctuations are actually enhanced in the gap owing to the destabilizing influence of the hydrophobic surfaces on the intervening water layer. It’s not clear to me whether in Kanth and Anishetty the water in the gap is, aside from the suppression of fluctuations, any different from bulk water, except perhaps for the monolayer adjacent to the surfaces…? And why would this not work for hydrophilic confinement too?

Alenka Luzar and her colleagues Christopher Daub and Dusan Bratko have just published a review of how electric fields at interfaces can modify their wettability (Top. Curr. Chem. 10.1007/128_2011_188; 2011 – paper here). Effects of this nature may play a role in the behaviour of voltage-sensitive ion channels.

Still more on denaturants. Thomas Record and colleagues at Wisconsin ask why urea is a denaturant while glycine betaine is a protein stabilizer (E. J. Guinn et al., PNAS 108, 16932-16937; 2011 – paper here). They use osmomentry and solubility measurements to look at the interactions of these molecules with 45 model proteins, and conclude that the explanation for the different behaviours lies with the details of how and where the molecules interact with the peptide surfaces. For example, urea accumulates at amide O groups, and to a lesser extent at aliphatic carbon atoms, whereas glycine betaine is excluded from them. This adds further weight to the notion that such osmolytes exert their effects via direct interactions with proteins rather than any generalized influence on ‘water structure’.

Phosphate groups turn out to be a sensitive probe of electric fields, including those that can be induced by hydration. As Steven Boxer and colleagues at Stanford show (N. M. Levinson et al., JACS 133, 13236; 2011 – paper here), electric fields perturb the vibrational spectra of organophosphates in a way that can reveal changes in hydration within partially hydrated environments, such as the active sites of enzymes.

Is there a liquid-liquid transition in confined water? That question is investigated via MD simulations by Limei Xu and Valeria Molinero at Utah (J. Phys. Chem. B jp205045k – paper here). The possibility has been raised by simulations of water in slit-like pores 2.4 nm wide (Brovchenko & Oleinikova, J. Chem. Phys. 126, 214701; 2007). Valeria and Limei use the mW water model to look at water’s behaviour in 1.5-nm hydrophilic pores at a range of temperatures and pressures up to 4000 atm. They find that at high pressures there is a signature of a somewhat abrupt but nonetheless continuous phase transition in the supercooled regime which could be interpreted as a ‘shadow’ of a L-L transition in the bulk phase which cannot itself be accessed.

A new and unusual view of the protein dynamical transition at c.200-220 K is presented in a preprint by Andrei Krokhotin and Antti Niemi at Uppsala University (paper here). They say that it this transition can be regarded as an analogue of the transition of a high-temperature superconductor to a non-superconducting pseudo-gap state. In other words, proteins can be assigned an order parameter formally equivalent to the quasiparticle wave function of superconductors. It’s not clear to me how/if this description modifies what is known already about the transition (on which, and on the role that hydration plays, there seems still to be no real consensus), but it is an original idea.