I have been on travel and caught up with numerous other things for several weeks, which not only means I have been unable to post but also that I’ve doubtless missed some interesting things over the recent period. If so, apologies for that – and feel free to let me know! But note also that my email address at Nature won’t be active for much longer, as I am very shortly leaving my role there as a writer – I’m no longer able to keep that going alongside the other demands on my time. I will no doubt keep writing occasionally for Nature nonetheless, but will be henceforth reachable at p.ball@btinternet.com.
To business, and in no particular order from the stack of papers in my pile…
Martina Havenith and her coworkers at Bochum have continued their highly revealing work on hydration structures using terahertz spectroscopy. In a new paper (B. Born et al., JACS 131, 3752 (2009) – paper here) they look at the hydration networks around various small peptides at low hydration levels, and find that the collective motions of peptide + solvent that seem to characterize protein hydration shells disappear below a minimum number of hydration waters, which appears to be well below that required for monolayer coverage.
This issue of solvent-protein dynamical correlations is also studied by Nigel Scrutton and colleagues at the University of Manchester (D.J. Heyes et al., Angew. Chem. Int. Ed. 48, 3850; 2009 – paper here). They find that proton tunnelling in protochlorophyllide oxireductase, a light-driven enzyme involved in chlorophyll biosynthesis, requires protein motions that appear to be slaved to the solvent, whereas hydride transfer in this species does not. It seems significant that proton, but not hydride, transfer occurs only close to the protein’s glass transition temperature, when the protein-solvent coupling is likely to be most acute.
In my recent ChemPhysChem paper (here), I referred to some recent work on denaturants by Jeremy England and coworkers, inadvertently describing it as though it came out of Vijay Pande’s lab (J. L. England, V. S. Pande & G. Haran, JACS 130, 11854; 2008). Actually I gather it was done mostly at the lab of Gilad Haran at the Weizmann Institute in Israel. Apologies for that oversight. Gilad has told me of his recent work on protein collapse using single-molecule FRET (G. Ziv & G. Haran, JACS 131, 2942; 2009 – paper here; and G. Ziv et al., Phys. Chem. Chem. Phys. 11, 83; 2009 – paper here). The former paper argues that the action of denaturants operates primarily by modulating the collapse of the denatured state, rather than acting on the transition from molten globule to folded state.
A recent special issue of J. Phys. Chem. B on aqueous solutions and interfaces (113(13)) has given me a lot to catch up with. Sotiris Xantheas and Greg Voth give a nice overview here. Among the interesting papers therein, Greg profiles the case for the amphiphilic nature of the hydrated proton at the interface with hydrophobic media (S. Iuchi et al., J. Phys. Chem. B 113, 4017; 2009 – paper here); Janamejaya Chowdhary and Branka Ladanyi use MD to investigate the dynamics of hydrogen-bond making and breaking (113, 4045; 2009 – paper here), and Yves Rezus and Huib Bakker use femtosecond IR spectroscopy to look at how various amphiliphilic small molecules alter water structure in the bulk (113, 4038; 2009 – paper here). Rezus and Bakker find that trimethylamine-N-oxide seems to be special in the latter regard, increasing the rate of reorientation of mobile water, which might be interpreted as a sign that it increases the number of defects in the H-bonded network.
On much the same topic, Greg Voth and colleagues have used MD to look at the interactions of hydrophobic, nonpolar solvents in aqueous salt (NaCl) and acid (HCl) solution (H. Chen et al., J. Phys. Chem. B ASAP; paper here). They find unusual solvated-proton structures in the acid which are bound to the hydrophobes, again supporting the idea that the hydrated protons act as amphiphiles. This might explain why protons seem anomalous in the Hofmeister series. And Michael Brindza and Robert Walker at the University of Maryland use SHG to look at solvation mechanisms of small molecules (p-nitroanisole, indoline) at the interface of polar solids and various liquids, offering a broader context for understanding what water in particular does in such cases (JACS 131, 6207; 2009 – paper here). Alenka Luzar and her coworkers have used MD to look at the solvation of monosodium glutamate, making a comparison to experimental data on this system (C. D. Daub et al., J. Phys. Chem. B ASAP; paper here). On the whole the agreement is good with the neutron-diffraction data, but there are some important differences – for example, the simulations couldn’t reproduce the experimentally observed reduction in water-water correlations – that presumably point to inadequacies of using classical potentials. (Incidentally, my spellchecker tirelessly insists that ‘solvation’ should be ‘salvation’ – forgive me if I don’t fail to catch them all…)
Alenka’s comparison with experiment here relies heavily on Alan Soper’s neutron data and its analysis using empirical potential structure refinement, which was conducted for those data by Alan with Sylvia McLain and Anthony Watts. Sylvia has sent me a paper in which she, with Soper and Watts alongside Jeremy Smith and Isabella Daidone, use this same technique to deduce the nature of hydration and interaction between various amino acid dimmers (S. E. McLain et al., Angew. Chem. Int. Ed. 47, 9059; 2008 – paper here). This is extremely interesting, as it challenges the conventional view that the main driving force for association is the interaction of hydrophobic regions. In contrast, this study finds that it is the charged sites on the peptides that dominate the association, and that interaction decreases as hydrophobicity increases. Could the same apply in protein folding itself?
Sason Shaik and his collaborators have now published the full analysis of the role on internal waters in the action of cytochrome P450 StaP (Y. Wang et al., JACS ASAP; 2009 – paper here), the preliminary account of which I discussed in my CPC paper.
Anders Nilsson, Lars Pettersson and their coworkers have now published the work that I referred to in a Nature article last year, in which they challenge the view that X-ray and neutron diffraction data uniquely support the conventional tetrahedral-coordinate picture of water structure (K. T. Wikfeldt et al., J. Phys. Chem. B 113, 6246; 2009 – paper here). Needless to say, this remains highly controversial stuff.
Pedro de Pablo and colleagues at Madrid have reported some very striking results on the desiccation of viruses. They say that drying of two different viruses causes the ejection of DNA and, in one case, collapse of the remaining capsid owing to capillary forces of the water menisci inside (C. Carrasco et al., PNAS 106, 5475; 2009 – paper here).
A couple more papers on nanoconfined water, in hydrophobic but less explicitly biological environments. Sow-Hsin Chen and colleagues say that supercooled water, which shows a density minimum under confinement in hydrophilic mesoporous materials, has none such in a hydrophobic material (Y. Zhang et al., J. Phys. Chem. B ASAP: paper here). And Gene Stanley and colleagues look at the H-bond dynamics of TIP5P water in a hydrophobic nanopore slit (S. Han et al., Phys. Rev. E 79, 041202; 2009 – paper here). They say that the H-bonds are shorter-lived in this case than in the bulk, and the relaxation time is smaller, but the general qualitative behaviour is much the same (e.g. the temperature-dependence of the average H-bond lifetime, and non-exponential lifetime distributions).
Ahmed Zewail and Ding-Shyue Yang recently reported ultrafast electron crystallography of water at low temperatures (c. 150 K) at the surface of graphite (PNAS 106, 4122; 2009 – paper here). It’s not strictly relevant to water in biology, perhaps, but an issue that I’ve started to follow increasingly and which touches on water’s general ability to order at interfaces. The results imply that, perhaps contrary to expectation (especially when compared with hydrophobic H-terminated silicon), this hydrophobic surface doesn’t disrupt a highly ordered interfacial structure. It seems this may be because the graphite surface is stepped, which apparently allows it to template a cubic-ice structure.
I recently wrote a column for Nature Materials (8, 250; 2009 – see here) on hydrophobicity at larger scales than the molecular, and in particular on the discussions of wetting by water of nano- and microstructured surfaces via Wenzel or Cassie states. This is relevant to biology insofar as it bears on the question of wettability of, e.g. insect legs and lotus leaves. There is a nice recent simulation paper on the topic by Takahiro Koishi at the University of Fukui and colleagues (PNAS doi:10.1073/pnas.0902027106 – not yet online), which describes the different conditions (of surface topology and so forth) under which Wenzel and Cassie wetting exist, and the possibility of their coexistence. And Abraham Marmur has flagged up his paper from last year (Langmuir 24, 7573; 2008 – paper here) in which he too looks at general geometric considerations that might promote such high-contact-angle states.
Here’s an intriguing thing that had escaped my notice until now: it seems that voltages may be generated in carbon nanotubes along the tube axis when water flows through them (see S. Ghosh et al., Science 299, 1042; 2003 – paper here). That’s suggestive from the perspective that views nanotubes as simple analogues of hydrophobic protein channels. Already it seems that this idea has been explored for power conversion (Y. C. Zhao et al., Adv. Mater. 20, 1772; 2008). Now Quanzi Yuan and Ya-Pu Zhao of the Institute of Mechanics in Beijing offer an explanation for the phenomenon in terms of the alignment of water molecules in the hydrogen-bonded chain within the nanotube (JACS ASAP; paper here).
More on this general topic from Bo Liu at the Graduate University of the Chinese Academy of Sciences in Beijing and coworkers, who describe simulations of a model system in which end-functionalized short carbon nanotubes are embedded in a phospholipid bilayer as mimics of aquaporin (B. Liu et al., Nano Lett. 9, 1386; 2009 – paper here). Two charges are placed near the tube midpoints, to simulate the NPA region of aquaporin where water selectivity and proton gating is thought to happen. But proton conduction couldn’t be studied explicitly in these classical MD simulations. That aside, the device seems to work more or less as planned, in theory. But can we make it?
Well, that’s not the end of it, but my desk looks a whole lot better.
Wednesday, May 6, 2009
Friday, March 13, 2009
There’s more to life than sequence
I have been meaning for some time to write about an interesting paper in JACS by Naoki Sugimoto’s group in Kobe. It found its way into an article that I wrote this week for Nature’s online news. So I’ve decided to simply post this article here – it’s not all strictly relevant to water in biology, but hopefully is interesting stuff anyway. This is the version before editing, which has more detail.
Shape might be one of the key factors in the function of mysterious ‘non-coding’ DNA.
Everyone knows what DNA looks like. Its double helix decorates countless articles on genetics, has been celebrated in sculpture, and was even engraved on the Golden Record, our message to the cosmos on board the Voyager spacecraft.
The entwined strands, whose form was deduced in 1953 by James Watson and Francis Crick, are admired as much for their beauty as for the light they shed on the mechanism of inheritance: the complementarity between juxtaposed chemical building blocks on the two strands, held together by weak ‘hydrogen’ bonds like a zipper, immediately suggested to Crick and Watson how information encoded in the sequence of blocks could be transmitted to a new strand assembled on the template of an existing one.
With the structure of DNA ‘solved’, genetics switched its focus to the sequence of the four constituent units (called nucleotide bases). By using biotechnological methods to deduce this sequence, they claimed to be ‘reading the book of life’, with the implication that all the information needed to build an organism was held within this abstract linear code.
But beauty has a tendency to inhibit critical thinking. There is now increasing evidence that the molecular structure of DNA is not a delightfully ordered epiphenomenon of its function as a digital data bank but a crucial – and mutable – aspect of the way genomes work. A new study in Science [1] underlines that notion by showing that the precise shape of some genomic DNA has been determined by evolution. In other words, genetics is not simply about sequence, but about structure too.
The standard view – indeed, part of biology’s ‘central dogma’ – is that in its sequence of the four fundamental building blocks (called nucleotide bases) DNA encodes corresponding sequences of amino-acid units that are strung together to make a protein enzyme, with the protein’s compact folded shape (and thus its function) being uniquely determined by that sequence.
This is basically true enough. Yet as the human genome was unpicked nucleotide base by base, it became clear that most of the DNA doesn’t ‘code for’ proteins at all. Fully 98 percent of the human genome is non-coding. So what does it do?
We don’t really know, except to say that it’s clearly not all ‘junk’, as was once suspected – the detritus of evolution, like obsolete files clogging up a computer. Much of the non-coding DNA evidently has a role in cell function, since mutations (changes in nucleotide sequence) in some of these regions have observable (phenotypic) consequences for the organism. We don’t know, however, how the former leads to the latter.
This is the question that Elliott Margulies of the National Institutes of Health in Bethesda, Maryland, Tom Tullius of Boston University, and their coworkers set out to investigate. According to the standard picture, the function of non-coding regions, whatever it is, should be determined by their sequence. Indeed, one way of identifying important non-coding regions is to look for ones that are sensitive to sequence, with the implication that the sequence has been finely tuned by evolution.
But Margulies and colleagues wondered if the shape of non-coding DNA might also be important. As they point out, DNA isn’t simply a uniform double helix: it can be bent or kinked, and may have a helical pitch of varying width, for example. These differences depend on the sequence, but not in any straightforward manner. Two near-identical sequences can adopt quite different shapes, or two very different sequences can have a similar shape.
The researchers used a chemical method to deduce the relationship between sequence and shape. They then searched for shape similarities between analogous non-coding regions in the genomes of 36 different species. Such similarity implies that the shapes have been selected and preserved by evolution – in other words, that shape, rather than sequence per se, is what is important. They found twice as many evolutionarily constrained (and thus functionally important) parts of the non-coding genome than were evident from trans-species correspondences using only sequence data.
So in these non-coding regions, at least, sequence appears to be important only insofar as it specifies a certain molecular shape and not because if its intrinsic information content – a different sequence with the same shape might do just as well.
That doesn’t answer why shape matters to DNA. But it suggests that we are wrong to imagine that the double helix is the beginning and end of the story.
There are plenty of other good reasons to suspect that is true. For example, DNA can adopt structures quite different from Watson and Crick’s helix, called the B-form. It can, under particular conditions of saltiness or temperature, switch to at least two other double-helical structures, called the A and Z forms. It may also from triple- and quadruple-stranded variants, linked by different types of hydrogen-bonding matches between nucleotides. One such is called Hoogsteen base-pairing.
Biochemist Naoki Sugimoto and colleagues at Konan University in Kobe, Japan, have recently shown that, when DNA in solution is surrounded by large polymer molecules, mimicking the crowded conditions of a real cell, Watson-Crick base pairing seems to be less stable than it is in pure, dilute solution, while Hoogsteen base-pairing, which favours the formation of triple and quadruple helices, becomes more stable [2-4].
The researchers think that this is linked to the way water molecules surround the DNA in a ‘hydration shell’. Hoogsteen pairing demands less water in this shell, and so is promoted when molecular crowding makes water scarce.
Changes to the hydration shell, for example induced by ions, may alter DNA shape in a sequence-dependent manner, perhaps being responsible for the sequence-structure relationships studied by Margulies and his colleagues. After all, says Tullius, the method they use to probe structure is a measure of “the local exposure of the surface of DNA to the solvent.”
The importance of DNA’s water sheath on its structure and function is also revealed in work that uses small synthetic molecules as drugs that bind to DNA and alter its behaviour, perhaps switching certain genes on or off. It is conventionally assumed that these molecules must fit snugly into the screw-like groove of the double helix. But some small molecules seem able to bind and show useful therapeutic activity even without such a fit, apparently because they can exploit water molecules in the hydration shell as ‘bridges’ to the DNA itself [5]. So here there is a subtle and irreducible interplay between sequence, shape and ‘environment’.
Then there are mechanical effects too. Some proteins bend and deform DNA significantly when they dock, making the molecule’s stiffness (and its dependence on sequence) a central factor in that process. And the shape and mechanics of DNA can influence gene function at larger scales. For example, the packaging of DNA and associated proteins into a compact form, called chromatin, in cells can affect whether particular genes are active or not. Special ‘chromatin-remodelling’ enzymes are needed to manipulate its structure and enable processes such as gene expression of DNA repair.
None of this is yet well understood. But it feels reminiscent of the way early work on protein structure in the 1930s and 40s grasped for dimly sensed principles before an understanding of the factors governing shape and function transformed our view of life’s molecular machinery. Are studies like these, then, a hint at some forthcoming insight that will reveal gene sequence to be just one element in the logic of life?
References
1. Parker, S. C. J. et al., Science Express doi:10.1126/science.1169050 (2009). Paper here.
2. Miyoshi, D., Karimata, H. & Sugimoto, N. J. Am. Chem. Soc. 128, 7957-7963 (2006). Paper here.
3. Nakano, S. et al., J. Am. Chem. Soc. 126, 14330-14331 (2004). Paper here.
4. Miyoshi, D. et al., J. Am. Chem. Soc. doi:10.1021/ja805972a (2009). Paper here.
5. Nguyen, B., Neidle, S. & Wilson, W. D. Acc. Chem. Res. 42, 11-21 (2009). Paper here.
Shape might be one of the key factors in the function of mysterious ‘non-coding’ DNA.
Everyone knows what DNA looks like. Its double helix decorates countless articles on genetics, has been celebrated in sculpture, and was even engraved on the Golden Record, our message to the cosmos on board the Voyager spacecraft.
The entwined strands, whose form was deduced in 1953 by James Watson and Francis Crick, are admired as much for their beauty as for the light they shed on the mechanism of inheritance: the complementarity between juxtaposed chemical building blocks on the two strands, held together by weak ‘hydrogen’ bonds like a zipper, immediately suggested to Crick and Watson how information encoded in the sequence of blocks could be transmitted to a new strand assembled on the template of an existing one.
With the structure of DNA ‘solved’, genetics switched its focus to the sequence of the four constituent units (called nucleotide bases). By using biotechnological methods to deduce this sequence, they claimed to be ‘reading the book of life’, with the implication that all the information needed to build an organism was held within this abstract linear code.
But beauty has a tendency to inhibit critical thinking. There is now increasing evidence that the molecular structure of DNA is not a delightfully ordered epiphenomenon of its function as a digital data bank but a crucial – and mutable – aspect of the way genomes work. A new study in Science [1] underlines that notion by showing that the precise shape of some genomic DNA has been determined by evolution. In other words, genetics is not simply about sequence, but about structure too.
The standard view – indeed, part of biology’s ‘central dogma’ – is that in its sequence of the four fundamental building blocks (called nucleotide bases) DNA encodes corresponding sequences of amino-acid units that are strung together to make a protein enzyme, with the protein’s compact folded shape (and thus its function) being uniquely determined by that sequence.
This is basically true enough. Yet as the human genome was unpicked nucleotide base by base, it became clear that most of the DNA doesn’t ‘code for’ proteins at all. Fully 98 percent of the human genome is non-coding. So what does it do?
We don’t really know, except to say that it’s clearly not all ‘junk’, as was once suspected – the detritus of evolution, like obsolete files clogging up a computer. Much of the non-coding DNA evidently has a role in cell function, since mutations (changes in nucleotide sequence) in some of these regions have observable (phenotypic) consequences for the organism. We don’t know, however, how the former leads to the latter.
This is the question that Elliott Margulies of the National Institutes of Health in Bethesda, Maryland, Tom Tullius of Boston University, and their coworkers set out to investigate. According to the standard picture, the function of non-coding regions, whatever it is, should be determined by their sequence. Indeed, one way of identifying important non-coding regions is to look for ones that are sensitive to sequence, with the implication that the sequence has been finely tuned by evolution.
But Margulies and colleagues wondered if the shape of non-coding DNA might also be important. As they point out, DNA isn’t simply a uniform double helix: it can be bent or kinked, and may have a helical pitch of varying width, for example. These differences depend on the sequence, but not in any straightforward manner. Two near-identical sequences can adopt quite different shapes, or two very different sequences can have a similar shape.
The researchers used a chemical method to deduce the relationship between sequence and shape. They then searched for shape similarities between analogous non-coding regions in the genomes of 36 different species. Such similarity implies that the shapes have been selected and preserved by evolution – in other words, that shape, rather than sequence per se, is what is important. They found twice as many evolutionarily constrained (and thus functionally important) parts of the non-coding genome than were evident from trans-species correspondences using only sequence data.
So in these non-coding regions, at least, sequence appears to be important only insofar as it specifies a certain molecular shape and not because if its intrinsic information content – a different sequence with the same shape might do just as well.
That doesn’t answer why shape matters to DNA. But it suggests that we are wrong to imagine that the double helix is the beginning and end of the story.
There are plenty of other good reasons to suspect that is true. For example, DNA can adopt structures quite different from Watson and Crick’s helix, called the B-form. It can, under particular conditions of saltiness or temperature, switch to at least two other double-helical structures, called the A and Z forms. It may also from triple- and quadruple-stranded variants, linked by different types of hydrogen-bonding matches between nucleotides. One such is called Hoogsteen base-pairing.
Biochemist Naoki Sugimoto and colleagues at Konan University in Kobe, Japan, have recently shown that, when DNA in solution is surrounded by large polymer molecules, mimicking the crowded conditions of a real cell, Watson-Crick base pairing seems to be less stable than it is in pure, dilute solution, while Hoogsteen base-pairing, which favours the formation of triple and quadruple helices, becomes more stable [2-4].
The researchers think that this is linked to the way water molecules surround the DNA in a ‘hydration shell’. Hoogsteen pairing demands less water in this shell, and so is promoted when molecular crowding makes water scarce.
Changes to the hydration shell, for example induced by ions, may alter DNA shape in a sequence-dependent manner, perhaps being responsible for the sequence-structure relationships studied by Margulies and his colleagues. After all, says Tullius, the method they use to probe structure is a measure of “the local exposure of the surface of DNA to the solvent.”
The importance of DNA’s water sheath on its structure and function is also revealed in work that uses small synthetic molecules as drugs that bind to DNA and alter its behaviour, perhaps switching certain genes on or off. It is conventionally assumed that these molecules must fit snugly into the screw-like groove of the double helix. But some small molecules seem able to bind and show useful therapeutic activity even without such a fit, apparently because they can exploit water molecules in the hydration shell as ‘bridges’ to the DNA itself [5]. So here there is a subtle and irreducible interplay between sequence, shape and ‘environment’.
Then there are mechanical effects too. Some proteins bend and deform DNA significantly when they dock, making the molecule’s stiffness (and its dependence on sequence) a central factor in that process. And the shape and mechanics of DNA can influence gene function at larger scales. For example, the packaging of DNA and associated proteins into a compact form, called chromatin, in cells can affect whether particular genes are active or not. Special ‘chromatin-remodelling’ enzymes are needed to manipulate its structure and enable processes such as gene expression of DNA repair.
None of this is yet well understood. But it feels reminiscent of the way early work on protein structure in the 1930s and 40s grasped for dimly sensed principles before an understanding of the factors governing shape and function transformed our view of life’s molecular machinery. Are studies like these, then, a hint at some forthcoming insight that will reveal gene sequence to be just one element in the logic of life?
References
1. Parker, S. C. J. et al., Science Express doi:10.1126/science.1169050 (2009). Paper here.
2. Miyoshi, D., Karimata, H. & Sugimoto, N. J. Am. Chem. Soc. 128, 7957-7963 (2006). Paper here.
3. Nakano, S. et al., J. Am. Chem. Soc. 126, 14330-14331 (2004). Paper here.
4. Miyoshi, D. et al., J. Am. Chem. Soc. doi:10.1021/ja805972a (2009). Paper here.
5. Nguyen, B., Neidle, S. & Wilson, W. D. Acc. Chem. Res. 42, 11-21 (2009). Paper here.
Tuesday, March 3, 2009
Making sense of solvent slaving
In my previous post I mentioned work by Pablo Debenedetti on ‘toy models’ of water. The places to look are: Buldyrev et al., PNAS 104, 20177 (2007) (here) for the solvation thermodynamics of ‘spherical’ water; and Patel et al., Biophys. J. 93, 4116 (2007) (here) and J. Chem. Phys. 128, 175102 (2008) (here) for water-explicit lattice models of proteins.
And in discussing recent work on the mechanism of urea-induced protein denaturation, I neglected to mention Bruce Berne’s PNAS paper from late last year with Ruhong Zhou, Dave Thirumalai and Lan Hua (105, 16928; paper here). That paper on MD simulations for lysozyme anticipated the more recent work showing that denaturation seems to be caused by direct urea-protein interactions: the urea displaced water from the first hydration shell and penetrates into the hydrophobic core to give a ‘dry globule’.
The notion that protein dynamics are ‘slaved’ to those of the hydration shell has been floating around for some time now. Hans Frauenfelder and colleagues have now brought considerable focus to the idea (PNAS doi:10.1073/pnas.0900336106; paper here) with dynamical measurements using dielectric spectroscopy, Mossbauer and neutron scattering. They find that large-scale protein motions follow the fluctuations of the solvent and are dependent on solvent viscosity. There are two classes of fluctuation in the solvent, alpha and beta, with different timescales. It seems that the former are ‘structural’ in nature and control protein shape; the latter are those to which the protein’s internal motions are slaved.
Jianxing Song at the National University of Singapore, who I met in Hangzhou, has sent me three papers on the intriguing solubilisation of ‘water-insoluble’ proteins in pure water. He and his coworkers found this effect in 2006 for a range of diverse proteins (M. Li et al., Protein Science 15, 1835 (2006) – paper here; M. Li et al., Biophys. J. 91, 4201 (2006) – paper here). They attributed it to the tendency of the ‘insoluble’ proteins to form partially folded states with many exposed hydrophobic residues, such that only a very low ionic strength is sufficient to screen out repulsive interactions and cause aggregation. In pure water, however, those electrostatic interactions remain sufficiently strong to suppress aggregation and precipitation. Jianxing has now provided an overview of this work, expanding on the importance of pH for this effect, in FEBS Letters (doi:10.1016/j.febslet.2009.02.022; paper here).
Roberto Righini at the University of Florence and colleagues have used IR spectroscopy to identify and quantify the various aqueous species that solvate the polar heads of phospholipids in bilayers (V. V. Volkov et al., J. Phys. Chem. B doi:10.1021/jp806650c; paper here). And Davide Donadio and coworkers in California have shown how electronic charge fluctuations show up in the IR spectra of water close to nonpolar surfaces (here graphite) (D. Donadio et al., J. Phys. Chem. B doi:10.1021/jp807709z; paper here). Still in that neck of the woods, Chuan-Shan Tian and Ron Shen have used sum-frequency-IR spectroscopy to sort out the nature of hydrogen-bonding at the air-water interface (JACS 131, 2791 (2009) – paper here). And Heather Allen and colleagues at Ohio State University have used this and other spectroscopic techniques to study hydration structure of the air-water interface for various divalent nitrates (M. Xu et al., J. Phys. Chem. B doi:10.1021/jp806565a; paper here).
Haiping Fang in Shanghai continues his exploration of how water and solutes can be manipulated within the confinement of carbon nanotubes. He and colleagues now show, using MD simulations, how a single charge outside a nanotube can be used to move a hydrated peptide inside it, regardless of whether the peptide itself is charged (P. Xiu et al., JACS 131, 2840 (2009) – paper here). This is due to the dipole-orientational ordering of the water molecules caused by confinement and interaction with the external charge.
There’s more, but later.
And in discussing recent work on the mechanism of urea-induced protein denaturation, I neglected to mention Bruce Berne’s PNAS paper from late last year with Ruhong Zhou, Dave Thirumalai and Lan Hua (105, 16928; paper here). That paper on MD simulations for lysozyme anticipated the more recent work showing that denaturation seems to be caused by direct urea-protein interactions: the urea displaced water from the first hydration shell and penetrates into the hydrophobic core to give a ‘dry globule’.
The notion that protein dynamics are ‘slaved’ to those of the hydration shell has been floating around for some time now. Hans Frauenfelder and colleagues have now brought considerable focus to the idea (PNAS doi:10.1073/pnas.0900336106; paper here) with dynamical measurements using dielectric spectroscopy, Mossbauer and neutron scattering. They find that large-scale protein motions follow the fluctuations of the solvent and are dependent on solvent viscosity. There are two classes of fluctuation in the solvent, alpha and beta, with different timescales. It seems that the former are ‘structural’ in nature and control protein shape; the latter are those to which the protein’s internal motions are slaved.
Jianxing Song at the National University of Singapore, who I met in Hangzhou, has sent me three papers on the intriguing solubilisation of ‘water-insoluble’ proteins in pure water. He and his coworkers found this effect in 2006 for a range of diverse proteins (M. Li et al., Protein Science 15, 1835 (2006) – paper here; M. Li et al., Biophys. J. 91, 4201 (2006) – paper here). They attributed it to the tendency of the ‘insoluble’ proteins to form partially folded states with many exposed hydrophobic residues, such that only a very low ionic strength is sufficient to screen out repulsive interactions and cause aggregation. In pure water, however, those electrostatic interactions remain sufficiently strong to suppress aggregation and precipitation. Jianxing has now provided an overview of this work, expanding on the importance of pH for this effect, in FEBS Letters (doi:10.1016/j.febslet.2009.02.022; paper here).
Roberto Righini at the University of Florence and colleagues have used IR spectroscopy to identify and quantify the various aqueous species that solvate the polar heads of phospholipids in bilayers (V. V. Volkov et al., J. Phys. Chem. B doi:10.1021/jp806650c; paper here). And Davide Donadio and coworkers in California have shown how electronic charge fluctuations show up in the IR spectra of water close to nonpolar surfaces (here graphite) (D. Donadio et al., J. Phys. Chem. B doi:10.1021/jp807709z; paper here). Still in that neck of the woods, Chuan-Shan Tian and Ron Shen have used sum-frequency-IR spectroscopy to sort out the nature of hydrogen-bonding at the air-water interface (JACS 131, 2791 (2009) – paper here). And Heather Allen and colleagues at Ohio State University have used this and other spectroscopic techniques to study hydration structure of the air-water interface for various divalent nitrates (M. Xu et al., J. Phys. Chem. B doi:10.1021/jp806565a; paper here).
Haiping Fang in Shanghai continues his exploration of how water and solutes can be manipulated within the confinement of carbon nanotubes. He and colleagues now show, using MD simulations, how a single charge outside a nanotube can be used to move a hydrated peptide inside it, regardless of whether the peptide itself is charged (P. Xiu et al., JACS 131, 2840 (2009) – paper here). This is due to the dipole-orientational ordering of the water molecules caused by confinement and interaction with the external charge.
There’s more, but later.
Thursday, February 5, 2009
What does denaturation mean?
At a recent symposium in honour of John Finney, who ‘retires’ (quote marks sure to be apt) this year, I heard Bertil Halle talk about the work described in a new paper (M. Davidovic et al., JACS 10.1021/ja8056419 – paper here), in which he and his colleagues argue that – if I am not being too liberal with the message here – cold-induced protein denaturation is not really denaturation at all, but rather the formation of a solvent-pentrated compact state. They report here that the hydration dynamics of BPTI, ubiquitin, apomyoglobin and beta-lactoglobulin, as monitored with NMR spin relaxation, shows that only apomyoglobin – that is, the only ‘modified’ protein here – truly denatures in picolitre emulsion droplets cooled to minus 35 C. Thus, they argue that cold denaturation is not equivalent to heat denaturation, and is perhaps not a well-defined state at all.
I have the abstract only of a simulation study of (partially) denaturation of human alpha-lactalbumin by Doug Tobias and colleagues (N. Sengupta et al., Biophys. J. 95, 5257; 2008 – paper here). So I can’t say too much about it, except that I assume this to be heat-induced, and that the results seem to show quite subtle effects on solvation dynamics. It seems that the partially denatured structure is rather patchily and ‘imperfectly’ solvated, with the solvent influx not keeping pace with the exposure of new surface in the protein.
Pablo Debenedetti was also at John’s symposium, and reported some most intriguing work on ‘toy’ models of water – spherical with two length scales, and four-coordinate on a square lattice – that offered insights into the possible origins of some behaviours both in the bulk and as a solvent. Forgive me, I will try to track that down when I have a free moment. Meanwhile, Pablo’s latest paper in J. Phys. Chem. B. (S. Romero-Vargas Castrillon et al., 10.1021/jp809032n – paper here) involves rather more ‘realistic’ water. They look at how water behaves in confinement between the silica surfaces of beta-cristobalite, primarily as a function of the parametrized surface Coulombic charge (k). When the surfaces are wholly apolar – no surface charge – they template an ice-like layer with slow dynamics. When the surfaces are strongly hydrophilic (high charge), the dynamics are also slow, but for a different reason: a dense, disordered water layer forms, with strong H-bonds. Both the diffusion coefficient and the rotational relaxation are thus non-monotonic with k.
Jan Engberts has brought my attention to a deeply interesting paper in Acc. Chem. Res. by Stephen Neidle of UCL and coworkers (Acc. Chem. Res. 42, 11-21; 2009; paper here) on the water mediation of DNA-ligand interactions. They describe how the binding of various small molecules in the minor groove can’t be easily understood on the basis of shape complementarity, but can be rationalized in terms of water-bridging in the hydration shell. This concept has already been used to identify drug candidates with high binding affinity and biological activity, which are now in clinical trials. This is probably the most striking example I have come across of how hydration structures can be used to inform drug development, something that I said in my Chem. Rev. paper was very much under-utilized owing to a lack of understanding. Perhaps that statement is now too strong. In any event, this looks like very important work.
Takashi Hayashi of Osaka University and coworkers report X-ray structures of cytochrome P450cam (which catalyses hydroxylation of camphor), showing how a propionate side chain acts as a gate that helps expel water from the active site (T. Hayashi et al., JACS 10.1021/ja807420k; paper here).
Mark Berg and colleagues have simulated the dynamics of water and ions near DNA in order to explain Stokes-shift data (S. Sen et al., JACS 131, 1724 (2009); paper here). It seems that the anomalous power-law dynamics seen experimentally can be accounted for by the water motions alone, the bottom line being that ‘water near DNA is strongly perturbed and is quite unlike bulk water.’
Damien Laage, Guillaume Stirnemann and Casey Hynes add to the overwhelming body of argument for dispensing with any notion of ‘iceberg’ hydration around hydrophobic groups (J. Phys. Chem. B 10.1021/jp809521t; paper here). They show that, contrary to what Rezus and Bakker recently claimed (Phys. Rev. Lett. 99, 148301; 2007), no water molecules are actually immobilized by hydrophobic solutes. Their model suggests that only a moderate degree of reorientational slowing, owing to slower hydrogen-bond exchange, is sufficient to explain both the ultrafast spectroscopic and the NMR data.
Mauricio Alcolea Palafox and coworkers in Madrid have simulated the hydration shells of thymidine and its derivative D4T (an alternative substrate for HIV-1 reverse transcriptase) from first principles (J. Phys. Chem. B 10.1021/jp806684v; paper here). And Dor Ben-Amotz and colleagues at Purdue look at the hydration shells of halide ions using Raman spectroscopy to probe the OH stretch (J. Phys. Chem. B 10.1021/jp808732s; paper here). They say the results support earlier work showing that the H-bonds between the halide ions and water are weaker than those in water.
I have the abstract only of a simulation study of (partially) denaturation of human alpha-lactalbumin by Doug Tobias and colleagues (N. Sengupta et al., Biophys. J. 95, 5257; 2008 – paper here). So I can’t say too much about it, except that I assume this to be heat-induced, and that the results seem to show quite subtle effects on solvation dynamics. It seems that the partially denatured structure is rather patchily and ‘imperfectly’ solvated, with the solvent influx not keeping pace with the exposure of new surface in the protein.
Pablo Debenedetti was also at John’s symposium, and reported some most intriguing work on ‘toy’ models of water – spherical with two length scales, and four-coordinate on a square lattice – that offered insights into the possible origins of some behaviours both in the bulk and as a solvent. Forgive me, I will try to track that down when I have a free moment. Meanwhile, Pablo’s latest paper in J. Phys. Chem. B. (S. Romero-Vargas Castrillon et al., 10.1021/jp809032n – paper here) involves rather more ‘realistic’ water. They look at how water behaves in confinement between the silica surfaces of beta-cristobalite, primarily as a function of the parametrized surface Coulombic charge (k). When the surfaces are wholly apolar – no surface charge – they template an ice-like layer with slow dynamics. When the surfaces are strongly hydrophilic (high charge), the dynamics are also slow, but for a different reason: a dense, disordered water layer forms, with strong H-bonds. Both the diffusion coefficient and the rotational relaxation are thus non-monotonic with k.
Jan Engberts has brought my attention to a deeply interesting paper in Acc. Chem. Res. by Stephen Neidle of UCL and coworkers (Acc. Chem. Res. 42, 11-21; 2009; paper here) on the water mediation of DNA-ligand interactions. They describe how the binding of various small molecules in the minor groove can’t be easily understood on the basis of shape complementarity, but can be rationalized in terms of water-bridging in the hydration shell. This concept has already been used to identify drug candidates with high binding affinity and biological activity, which are now in clinical trials. This is probably the most striking example I have come across of how hydration structures can be used to inform drug development, something that I said in my Chem. Rev. paper was very much under-utilized owing to a lack of understanding. Perhaps that statement is now too strong. In any event, this looks like very important work.
Takashi Hayashi of Osaka University and coworkers report X-ray structures of cytochrome P450cam (which catalyses hydroxylation of camphor), showing how a propionate side chain acts as a gate that helps expel water from the active site (T. Hayashi et al., JACS 10.1021/ja807420k; paper here).
Mark Berg and colleagues have simulated the dynamics of water and ions near DNA in order to explain Stokes-shift data (S. Sen et al., JACS 131, 1724 (2009); paper here). It seems that the anomalous power-law dynamics seen experimentally can be accounted for by the water motions alone, the bottom line being that ‘water near DNA is strongly perturbed and is quite unlike bulk water.’
Damien Laage, Guillaume Stirnemann and Casey Hynes add to the overwhelming body of argument for dispensing with any notion of ‘iceberg’ hydration around hydrophobic groups (J. Phys. Chem. B 10.1021/jp809521t; paper here). They show that, contrary to what Rezus and Bakker recently claimed (Phys. Rev. Lett. 99, 148301; 2007), no water molecules are actually immobilized by hydrophobic solutes. Their model suggests that only a moderate degree of reorientational slowing, owing to slower hydrogen-bond exchange, is sufficient to explain both the ultrafast spectroscopic and the NMR data.
Mauricio Alcolea Palafox and coworkers in Madrid have simulated the hydration shells of thymidine and its derivative D4T (an alternative substrate for HIV-1 reverse transcriptase) from first principles (J. Phys. Chem. B 10.1021/jp806684v; paper here). And Dor Ben-Amotz and colleagues at Purdue look at the hydration shells of halide ions using Raman spectroscopy to probe the OH stretch (J. Phys. Chem. B 10.1021/jp808732s; paper here). They say the results support earlier work showing that the H-bonds between the halide ions and water are weaker than those in water.
Tuesday, January 6, 2009
How do protons get through bacteriorhodopsin?
I’ve just received a copy of the special issue of ChemPhysChem (here) on water at molecular interfaces (Vol. 9, 2635-2879), containing presentations from the DFG Forschergruppe 436 meeting in Dortmund last July. I won’t list everything in it – there is too much that is all worth reading.
One of the contributions, from Klaus Gerwert and colleagues, looks at how vectorial proton transport is achieved in bacteriorhodopsin via a network of water molecules (p.2772). That is also the topic of a recent paper from Qiang Cui of the University of Wisconsin and colleagues (P. Phatak et al., PNAS 105, 19672; 2008 – paper here), who look specifically at the much-debated question of what the proton storage site in bR is. They argue, against the conclusions of Gerwert and coworkers (e.g. Nature 439, 109; 2006), that the proton is kept on a pair of glutamate residues (Glu 194/204), not on a nearby water cluster. I daresay the debate will continue.
Jeremy Smith and colleagues have looked at another aspect of the problem – the possible role of a bound water molecule on the cytoplasmic side of the retinal Schiff base chromophore in the initial transfer of a proton from this chromophore to Asp85, the first step in its motion to the extracellular side (A.-N. Bondar et al., J. Phys. Chem. B 112, 14729; 2008 – paper here). They report calculations which suggest that a water molecule bound to the ‘back’ of retinal in this way helps to direct proton transfer to Asp85 rather than towards Asp212 on the other side of the channel. A surprisingly subtle and indirect form of ‘water-tuning’.
Feng Gai and colleagues at the University of Pennsylvania have added to the unfolding (forgive me) story of how hydration influences amyloid aggregation (S. Mukherjee et al., J. Phys. Chem. B 10.1021/jp809817s – paper here). They have manipulated the degree of hydration of two amyloid-forming peptides by encapsulating them in reverse micelles, and find that aggregation is enhanced when hydration is lessened.
Ronen Zangi, Ruhong Zhou and Bruce Berne report simulations that support what seems to be a growing view that urea’s denaturing action results from direct interaction with hydrophobic surfaces and not any kind of ‘chaotropic’ effect on ‘water structure’ (R. Zangi et al., JACS 10.1021/ja807887g – paper here). They find that urea weakens hydrophobic interactions both in a hydrophobic model polymer and between hydrophobic (and graphene) plates, owing to its binding to the surface and acting as a kind of surfactant.
One of the contributions, from Klaus Gerwert and colleagues, looks at how vectorial proton transport is achieved in bacteriorhodopsin via a network of water molecules (p.2772). That is also the topic of a recent paper from Qiang Cui of the University of Wisconsin and colleagues (P. Phatak et al., PNAS 105, 19672; 2008 – paper here), who look specifically at the much-debated question of what the proton storage site in bR is. They argue, against the conclusions of Gerwert and coworkers (e.g. Nature 439, 109; 2006), that the proton is kept on a pair of glutamate residues (Glu 194/204), not on a nearby water cluster. I daresay the debate will continue.
Jeremy Smith and colleagues have looked at another aspect of the problem – the possible role of a bound water molecule on the cytoplasmic side of the retinal Schiff base chromophore in the initial transfer of a proton from this chromophore to Asp85, the first step in its motion to the extracellular side (A.-N. Bondar et al., J. Phys. Chem. B 112, 14729; 2008 – paper here). They report calculations which suggest that a water molecule bound to the ‘back’ of retinal in this way helps to direct proton transfer to Asp85 rather than towards Asp212 on the other side of the channel. A surprisingly subtle and indirect form of ‘water-tuning’.
Feng Gai and colleagues at the University of Pennsylvania have added to the unfolding (forgive me) story of how hydration influences amyloid aggregation (S. Mukherjee et al., J. Phys. Chem. B 10.1021/jp809817s – paper here). They have manipulated the degree of hydration of two amyloid-forming peptides by encapsulating them in reverse micelles, and find that aggregation is enhanced when hydration is lessened.
Ronen Zangi, Ruhong Zhou and Bruce Berne report simulations that support what seems to be a growing view that urea’s denaturing action results from direct interaction with hydrophobic surfaces and not any kind of ‘chaotropic’ effect on ‘water structure’ (R. Zangi et al., JACS 10.1021/ja807887g – paper here). They find that urea weakens hydrophobic interactions both in a hydrophobic model polymer and between hydrophobic (and graphene) plates, owing to its binding to the surface and acting as a kind of surfactant.
Thursday, December 18, 2008
A nice Christmas package
There are some important and provocative papers in this batch…
Teresa Head-Gordon and her coworkers have extended their recent work on quasi-elastic neutron scattering in peptide hydration shells (e.g. Russo et al., J. Phys. Chem. B 108, 19885 and 109, 12966 (2005); Russo et al., Biophys. J. 86, 1852 (2004)) by using MD simulations to explore the way in which the hydration dynamics are affected by the heterogeneous, amphiphilic nature of most protein surfaces (M. E. Johnson et al., J. Phys. Chem. B doi:10.1021/jp806183v – paper here). The notion they proposed earlier is that the dynamics are most perturbed at the interfaces of hydrophobic and hydrophilic patches, due to the frustration created by different styles of hydration in the adjacent regimes. This now seems to be borne out by the simulations, where the water dynamics seen experimentally are reproduced for an amphiphilic peptide but not a hydrophilic one. The strongest dynamical perturbations are found for the first hydration shell of hydrophobic residues.
Jeetain Mittal and Gerhard Hummer have used simulations to try to clarify exactly what goes on at the interface of a hydrophobic surface and water (PNAS doi:10.1073/pnas.0809029105 – paper here). They are in particular examining the vexed question of whether there is a depletion layer in water density close to the surface, as proposed first by Frank Stillinger and invoked in the Lum-Chandler-Weeks model of dewetting-induced hydrophobic collapse (K. Lum, D. Chandler & J. D. Weeks, J. Phys. Chem. B 103, 4570; 1999). Experiments have now shown some evidence for a depletion layer perhaps 1-2 Å thick. But is there a sharp transition between a liquid-like and vapour-like phase, or a gradual thinning? In the former case, capillary waves are expected to blur the interface, so it’s hard to tell the difference. Mittal and Hummer find, for a purely repulsive spherical solute particle, that the interface is indeed rather sharp, but broadened by capillary waves in line with what theory predicts for a free air-water interface. The ‘dry’ layer looks to be instantaneously about 2 Å or so thick. The result is a flickering interface with patches that are intermittently dry and wet (in proportions that depend on the solute size), and transitions between them that are slow on a molecular timescale. This is all very illuminating, but I’m hard to satisfy – what happens when van der Waals forces between solvent and surface are included, I wonder?
Roland Netz at the TU Munich and his colleagues have also explored the depletion-layer problem from a very different angle. They have used MD simulations to examine how the slip length for water flow past a hydrophobic surface depends on the contact angle (D. M. Huang et al., Phys. Rev. Lett. 101, 226101; 2008 – paper here). Experimental studies in this area have given confusing and conflicting results, with slip lengths orders of magnitude different for surfaces that seem very similar. But the simulations show a rather systematic (though nonlinear) dependence of slip length on static contact angle. Moreover, they see depletion layers of molecular dimensions, whose average width varies with the ¼ power of the slip length. Thus, anything that influences the width of the depletion layer (dissolved gases) should have a marked effect on the slip length.
I referred recently to a study that challenged the notion of a dynamical transition for protein hydration water at 220 K and its interpretation as a fragile-to-strong crossover (S. Khodadadi et al., J. Phys. Chem. B 112, 14273-14280; 2008). Now here comes another one, from Michael Vogel at the Technical University of Damstadt (Phys. Rev. Lett. 101, 225701; 2008 – paper here). He has used deuterium NMR to study reorientational dynamics of hydration water for elastin and collagen, and sees no sign of a transition at 225 K. There is one at 200 K, but Vogel says that it corresponds to the onset, at lower temperatures, of thermally activated jumps in tetrahedral coordination, perhaps related to defect motion in the hydrogen-bonded network.
Fabio Sterpone and colleagues in Rome argue that the thermostability of proteins is primarily determined by protein-water interactions, with the intra-chain interactions between packed portions of the polypeptide being of only secondary importance (F. Sterpone et al., J. Phys. Chem. B doi:10.1021/jp805199c – paper here). They looked, using simulations, at the thermal stability and flexibility of three homologous proteins – one mesophilic, one thermophilic, and one hyperthermophilic. As thermal stability increases, so the proteins seem to be encased in an increasingly persistent hydration shell linked by hydrogen bonds. The idea, crudely speaking, seems to be that this shell supplies an increasingly robust protective coat against the penetration of water into the folded protein.
At the recent Hangzhou workshop I heard about the work of Shengfu Chen of Zhejiang University and colleagues on anti-fouling films that incorporate heterogeneously charged peptides. The idea is that the ability of these films to resist non-specific protein adsorption is linked to the nature of hydration of the surface chemical groups: the ‘more’ hydration there is, the stronger the disrupting influence of an incoming adsorbate and thus the more its attachment is inhibited. Shengfu and his workers in Washington and Taiwan develop this idea in a paper here (J. Phys. Chem. B doi:10.1021/jp8065713). They introduce a method for deducing the number of water molecules hydrating a given solute, and find that the greater the ‘hydration capacity’ of a solute, the greater its ability to resist protein adsorption in anti-fouling films.
Haiping Fang, my co-chair at that meeting, has an intriguing paper on the effect on water flux through a nanotube on the nature of the ‘outside structure’, in this case meaning whether the nanotube threads through a single, double or multiple sheets of graphene (X. Gong et al., Phys. Rev. Lett. 101, 257801; 2008 - paper here). In simulations, they find that the flux of water can be quite different in the various cases. For example, with two graphene sheets separated by a vacuum, the flux and flow both increases as the separation increases. And if water surrounds the nanotube in the space(s) between sheets, the flux is lowered. They deduce that interactions between water molecules inside the nanotube and the species outside the tube are responsible for the differences, emphasizing how sensitive, in this confined geometry with more or less single-file molecular traffic (where molecular motions are strongly correlated), the water transport is to the internal configurations of water molecules.
It seems clear that nanobubbles can form on hydrophobic surfaces, and very likely that these play a key role in the long-ranged hydrophobic interaction that is sometimes observed between such surfaces. The question has remained of how such bubbles, with a very high radius of curvature, can be stable when that curvature creates a large Laplace pressure which should lead to rapid diffusive efflux of gas out of the bubble. Michael Brenner and Detlef Lohse have considered this question (Phys. Rev. Lett. 101, 214505; 2008 – paper here). They say that the outflux can be balanced by an enhanced influx of gas at the contact line of bubble and surface, owing to the attraction of dissolved gas to the hydrophobic surface. They acknowledge that this is a non-equilibrium situation which suggests that in the long term the bubbles should disappear. But there haven’t yet been any long-term studies of closed systems to see whether that is the case.
Apparently sobering news from Michael Levitt and colleagues: MD simulations for protein structure refinement perform worse in explicit solvent than implicit solvent (G. Chopra et al., PNAS doi:10.1073/pnas.0810818105 – paper here). This seems to be because the potential in explicit solvent is more rugged, and so there is more chance of getting stuck in local minima unless the simulation is very long. So there are some situations in which it is still best not to consider the hydration shell molecule by molecule.
Angel Garcia at RPI and coworkers have calculated the stability diagram of the well-studied Trp-cage miniprotein (D. Paschek et al., PNAS 105, 17754; 2008 – paper here). They derive some insights into the role of hydration in pressure-induced denaturation, which they link in part to tighter packing of water around nonpolar atoms as pressure increases.
The debate over the ‘pH’ of the air-water interface continues. First-principles empirical-valence-bond calculations by Greg Voth and colleagues seem to indicate that the preference of hydrated protons for the surface (as claimed in their earlier work) is energetically (rather than entropically) promoted, due to the amphiphilic nature of the hydrated proton (S. Iuchi et al., J. Phys. Chem. B doi:10.1021/jp805304j – paper here). They say that much the same applies for a water-hydrophobe interface too.
Teresa Head-Gordon and her coworkers have extended their recent work on quasi-elastic neutron scattering in peptide hydration shells (e.g. Russo et al., J. Phys. Chem. B 108, 19885 and 109, 12966 (2005); Russo et al., Biophys. J. 86, 1852 (2004)) by using MD simulations to explore the way in which the hydration dynamics are affected by the heterogeneous, amphiphilic nature of most protein surfaces (M. E. Johnson et al., J. Phys. Chem. B doi:10.1021/jp806183v – paper here). The notion they proposed earlier is that the dynamics are most perturbed at the interfaces of hydrophobic and hydrophilic patches, due to the frustration created by different styles of hydration in the adjacent regimes. This now seems to be borne out by the simulations, where the water dynamics seen experimentally are reproduced for an amphiphilic peptide but not a hydrophilic one. The strongest dynamical perturbations are found for the first hydration shell of hydrophobic residues.
Jeetain Mittal and Gerhard Hummer have used simulations to try to clarify exactly what goes on at the interface of a hydrophobic surface and water (PNAS doi:10.1073/pnas.0809029105 – paper here). They are in particular examining the vexed question of whether there is a depletion layer in water density close to the surface, as proposed first by Frank Stillinger and invoked in the Lum-Chandler-Weeks model of dewetting-induced hydrophobic collapse (K. Lum, D. Chandler & J. D. Weeks, J. Phys. Chem. B 103, 4570; 1999). Experiments have now shown some evidence for a depletion layer perhaps 1-2 Å thick. But is there a sharp transition between a liquid-like and vapour-like phase, or a gradual thinning? In the former case, capillary waves are expected to blur the interface, so it’s hard to tell the difference. Mittal and Hummer find, for a purely repulsive spherical solute particle, that the interface is indeed rather sharp, but broadened by capillary waves in line with what theory predicts for a free air-water interface. The ‘dry’ layer looks to be instantaneously about 2 Å or so thick. The result is a flickering interface with patches that are intermittently dry and wet (in proportions that depend on the solute size), and transitions between them that are slow on a molecular timescale. This is all very illuminating, but I’m hard to satisfy – what happens when van der Waals forces between solvent and surface are included, I wonder?
Roland Netz at the TU Munich and his colleagues have also explored the depletion-layer problem from a very different angle. They have used MD simulations to examine how the slip length for water flow past a hydrophobic surface depends on the contact angle (D. M. Huang et al., Phys. Rev. Lett. 101, 226101; 2008 – paper here). Experimental studies in this area have given confusing and conflicting results, with slip lengths orders of magnitude different for surfaces that seem very similar. But the simulations show a rather systematic (though nonlinear) dependence of slip length on static contact angle. Moreover, they see depletion layers of molecular dimensions, whose average width varies with the ¼ power of the slip length. Thus, anything that influences the width of the depletion layer (dissolved gases) should have a marked effect on the slip length.
I referred recently to a study that challenged the notion of a dynamical transition for protein hydration water at 220 K and its interpretation as a fragile-to-strong crossover (S. Khodadadi et al., J. Phys. Chem. B 112, 14273-14280; 2008). Now here comes another one, from Michael Vogel at the Technical University of Damstadt (Phys. Rev. Lett. 101, 225701; 2008 – paper here). He has used deuterium NMR to study reorientational dynamics of hydration water for elastin and collagen, and sees no sign of a transition at 225 K. There is one at 200 K, but Vogel says that it corresponds to the onset, at lower temperatures, of thermally activated jumps in tetrahedral coordination, perhaps related to defect motion in the hydrogen-bonded network.
Fabio Sterpone and colleagues in Rome argue that the thermostability of proteins is primarily determined by protein-water interactions, with the intra-chain interactions between packed portions of the polypeptide being of only secondary importance (F. Sterpone et al., J. Phys. Chem. B doi:10.1021/jp805199c – paper here). They looked, using simulations, at the thermal stability and flexibility of three homologous proteins – one mesophilic, one thermophilic, and one hyperthermophilic. As thermal stability increases, so the proteins seem to be encased in an increasingly persistent hydration shell linked by hydrogen bonds. The idea, crudely speaking, seems to be that this shell supplies an increasingly robust protective coat against the penetration of water into the folded protein.
At the recent Hangzhou workshop I heard about the work of Shengfu Chen of Zhejiang University and colleagues on anti-fouling films that incorporate heterogeneously charged peptides. The idea is that the ability of these films to resist non-specific protein adsorption is linked to the nature of hydration of the surface chemical groups: the ‘more’ hydration there is, the stronger the disrupting influence of an incoming adsorbate and thus the more its attachment is inhibited. Shengfu and his workers in Washington and Taiwan develop this idea in a paper here (J. Phys. Chem. B doi:10.1021/jp8065713). They introduce a method for deducing the number of water molecules hydrating a given solute, and find that the greater the ‘hydration capacity’ of a solute, the greater its ability to resist protein adsorption in anti-fouling films.
Haiping Fang, my co-chair at that meeting, has an intriguing paper on the effect on water flux through a nanotube on the nature of the ‘outside structure’, in this case meaning whether the nanotube threads through a single, double or multiple sheets of graphene (X. Gong et al., Phys. Rev. Lett. 101, 257801; 2008 - paper here). In simulations, they find that the flux of water can be quite different in the various cases. For example, with two graphene sheets separated by a vacuum, the flux and flow both increases as the separation increases. And if water surrounds the nanotube in the space(s) between sheets, the flux is lowered. They deduce that interactions between water molecules inside the nanotube and the species outside the tube are responsible for the differences, emphasizing how sensitive, in this confined geometry with more or less single-file molecular traffic (where molecular motions are strongly correlated), the water transport is to the internal configurations of water molecules.
It seems clear that nanobubbles can form on hydrophobic surfaces, and very likely that these play a key role in the long-ranged hydrophobic interaction that is sometimes observed between such surfaces. The question has remained of how such bubbles, with a very high radius of curvature, can be stable when that curvature creates a large Laplace pressure which should lead to rapid diffusive efflux of gas out of the bubble. Michael Brenner and Detlef Lohse have considered this question (Phys. Rev. Lett. 101, 214505; 2008 – paper here). They say that the outflux can be balanced by an enhanced influx of gas at the contact line of bubble and surface, owing to the attraction of dissolved gas to the hydrophobic surface. They acknowledge that this is a non-equilibrium situation which suggests that in the long term the bubbles should disappear. But there haven’t yet been any long-term studies of closed systems to see whether that is the case.
Apparently sobering news from Michael Levitt and colleagues: MD simulations for protein structure refinement perform worse in explicit solvent than implicit solvent (G. Chopra et al., PNAS doi:10.1073/pnas.0810818105 – paper here). This seems to be because the potential in explicit solvent is more rugged, and so there is more chance of getting stuck in local minima unless the simulation is very long. So there are some situations in which it is still best not to consider the hydration shell molecule by molecule.
Angel Garcia at RPI and coworkers have calculated the stability diagram of the well-studied Trp-cage miniprotein (D. Paschek et al., PNAS 105, 17754; 2008 – paper here). They derive some insights into the role of hydration in pressure-induced denaturation, which they link in part to tighter packing of water around nonpolar atoms as pressure increases.
The debate over the ‘pH’ of the air-water interface continues. First-principles empirical-valence-bond calculations by Greg Voth and colleagues seem to indicate that the preference of hydrated protons for the surface (as claimed in their earlier work) is energetically (rather than entropically) promoted, due to the amphiphilic nature of the hydrated proton (S. Iuchi et al., J. Phys. Chem. B doi:10.1021/jp805304j – paper here). They say that much the same applies for a water-hydrophobe interface too.
Tuesday, December 9, 2008
A lot about interfaces
Janamejaya Chowdhary and Branka Ladanyi at Colorado State have used MD simulations to look at the dynamics of H-bonds at a water-hydrocarbon interface (J. Phys. Chem. B ASAP doi:10.1021/jp; paper here). They find that the reorientation of the O-H bond is anisotropic, and quantify the effects of cooperativity in the dynamics.
Robert Woods and colleagues at the University of Georgia study how bound water mediates the binding of concanavalin A to its target carbohydrate ligand (R. Kadirvelraj et al., JACS ASAP; paper here). Or rather, they look at a modified ligand of the natural trisaccharide, with a hydroxylethyl side chain that may or may not displace a conserved water in binding of the natural ligand. The crystal structure reported here shows that this water is retained, though its position is distorted. This helps to explain the previous thermodynamic data on ligand specificity for Con A, showing that there is no entropic component for the synthetic ligand arising from water displacement.
Roger Tam and colleagues in Ottawa have looked at the inhibition of ice recrystallization by mono- and disaccharides (JACS ASAP; paper here). Specifically, they look for correlates of ice-growth inhibition in the degree of hydration of the sugars, and find that, rather than using the total number of tightly bound water molecules, a better predictor of inhibiting ability is a hydration index in which the hydration number is divided by the molar volume. The researchers conclude that the inhibition arises from a disruption of water ‘pre-ordering’ at the ice-water interface.
Joe Zaccai and colleagues have measured water dynamics in human red blood cells using quasielastic incoherent neutron scattering (A. M. Stadler et al., JACS ASAP; paper here). In line with their previous work on E. coli, they find that most (90%) of the cell water has similar translational diffusion to the bulk, while about 10% is slower, this presumably being the water hydrating haemoglobin.
Sherwin Singer and colleagues at Ohio State have looked at the hydration dynamics of myoglobin using MD simulations (T. Li et al., J. Phys. Chem. B 10.1021/jp803042u; paper here). Specifically, they look at the time-dependent fluorescence Stokes shift after photoexcitation of the Trp-7 residue, a measure of the relaxation dynamics of the chromophore’s environment. The question is whether the water dynamics are due to constraint of the water by interactions with the protein, or whether they are controlled by the dynamics of the protein itself. This distinction should be revealed by arresting the protein in the simulations. Singer and colleagues find that doing so significantly changes the Stokes shift, suggesting that the intrinsic protein flexibility is important. They caution, however, that this does not necessarily imply that the water dynamics exhibit no intrinsic slow component of relaxation; rather, the protein and water dynamics are so intimately coupled that either slow water dynamics or slow protein dynamics (or both) could alter the Stokes shift.
Shekhar Garde and colleagues at RPI have conducted simulations of hydrophobically induced polymer collapse near to the interface with air or a hydrophobic wall (S. N. Jamadagni et al., J. Phys. Chem. B 10.1021/jp806528m – paper here here). They find that the driving force for collapse is smaller at the water-alkane interface, and all but vanishes at the air-vapour interface, where the polymer remains unfolded. They think that both the weaker hydration of the polymer and the enhanced density fluctuations of water at the interface produce faster conformational switches in the folded chain. The results throws up lots of interesting questions, most obviously of course what this implies for the conformational flexibility of two peptide chains approaching one another via the hydrophobic interaction.
Hangjun Lu and colleagues at Zhejiang Normal Univerity have looked at how an external charge of +1e near a carbon nanotube will affect the filling and emptying by water (H. Lu et al., J. Phys. Chem. B 10.1021/jp802263v – paper here here). It seems that the charge stabilizes the water-filled state when it is at the midpoint of the nanotube, but much less so if it is moved towards the ends. The implication is that this is a method that might be exploited by protein channels to control water transport via the positioning of ionized residues.
The freezing-point depression of water that hydrates phospholipid membranes has been studied using NMR by Dong-Kuk Lee at Seoul National University of Technology and coworkers (D.-K. Lee et al., Langmuir 24, 13598 (2008) – paper here). They find that water molecules still show liquid-like signatures below -20 C in bilayers, and that the freezing behaviour is depressed still further by cholesterol, a known cryoprotectant.
I have a kind of follow-up to my Chem. Rev. article in a forthcoming issue of ChemPhysChem, which has now appeared online (here). This will form part of a special issue on the subject of water at interfaces, stemming from a meeting of the DFG Forschergruppe 436 in Dortmund last summer.
Robert Woods and colleagues at the University of Georgia study how bound water mediates the binding of concanavalin A to its target carbohydrate ligand (R. Kadirvelraj et al., JACS ASAP; paper here). Or rather, they look at a modified ligand of the natural trisaccharide, with a hydroxylethyl side chain that may or may not displace a conserved water in binding of the natural ligand. The crystal structure reported here shows that this water is retained, though its position is distorted. This helps to explain the previous thermodynamic data on ligand specificity for Con A, showing that there is no entropic component for the synthetic ligand arising from water displacement.
Roger Tam and colleagues in Ottawa have looked at the inhibition of ice recrystallization by mono- and disaccharides (JACS ASAP; paper here). Specifically, they look for correlates of ice-growth inhibition in the degree of hydration of the sugars, and find that, rather than using the total number of tightly bound water molecules, a better predictor of inhibiting ability is a hydration index in which the hydration number is divided by the molar volume. The researchers conclude that the inhibition arises from a disruption of water ‘pre-ordering’ at the ice-water interface.
Joe Zaccai and colleagues have measured water dynamics in human red blood cells using quasielastic incoherent neutron scattering (A. M. Stadler et al., JACS ASAP; paper here). In line with their previous work on E. coli, they find that most (90%) of the cell water has similar translational diffusion to the bulk, while about 10% is slower, this presumably being the water hydrating haemoglobin.
Sherwin Singer and colleagues at Ohio State have looked at the hydration dynamics of myoglobin using MD simulations (T. Li et al., J. Phys. Chem. B 10.1021/jp803042u; paper here). Specifically, they look at the time-dependent fluorescence Stokes shift after photoexcitation of the Trp-7 residue, a measure of the relaxation dynamics of the chromophore’s environment. The question is whether the water dynamics are due to constraint of the water by interactions with the protein, or whether they are controlled by the dynamics of the protein itself. This distinction should be revealed by arresting the protein in the simulations. Singer and colleagues find that doing so significantly changes the Stokes shift, suggesting that the intrinsic protein flexibility is important. They caution, however, that this does not necessarily imply that the water dynamics exhibit no intrinsic slow component of relaxation; rather, the protein and water dynamics are so intimately coupled that either slow water dynamics or slow protein dynamics (or both) could alter the Stokes shift.
Shekhar Garde and colleagues at RPI have conducted simulations of hydrophobically induced polymer collapse near to the interface with air or a hydrophobic wall (S. N. Jamadagni et al., J. Phys. Chem. B 10.1021/jp806528m – paper here here). They find that the driving force for collapse is smaller at the water-alkane interface, and all but vanishes at the air-vapour interface, where the polymer remains unfolded. They think that both the weaker hydration of the polymer and the enhanced density fluctuations of water at the interface produce faster conformational switches in the folded chain. The results throws up lots of interesting questions, most obviously of course what this implies for the conformational flexibility of two peptide chains approaching one another via the hydrophobic interaction.
Hangjun Lu and colleagues at Zhejiang Normal Univerity have looked at how an external charge of +1e near a carbon nanotube will affect the filling and emptying by water (H. Lu et al., J. Phys. Chem. B 10.1021/jp802263v – paper here here). It seems that the charge stabilizes the water-filled state when it is at the midpoint of the nanotube, but much less so if it is moved towards the ends. The implication is that this is a method that might be exploited by protein channels to control water transport via the positioning of ionized residues.
The freezing-point depression of water that hydrates phospholipid membranes has been studied using NMR by Dong-Kuk Lee at Seoul National University of Technology and coworkers (D.-K. Lee et al., Langmuir 24, 13598 (2008) – paper here). They find that water molecules still show liquid-like signatures below -20 C in bilayers, and that the freezing behaviour is depressed still further by cholesterol, a known cryoprotectant.
I have a kind of follow-up to my Chem. Rev. article in a forthcoming issue of ChemPhysChem, which has now appeared online (here). This will form part of a special issue on the subject of water at interfaces, stemming from a meeting of the DFG Forschergruppe 436 in Dortmund last summer.
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