Osmolyte Trimethylamine-N-Oxide Does Not Affect the Strength of Hydrophobic Interactions: Origin of Osmolyte Compatibility

Athawale, Manoj V.; Dordick, Jonathan S.; Garde, Shekhar

doi:10.1529/biophysj.104.056671

Cited by 143 publications

(218 citation statements)

References 56 publications

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“…In particular, this red shift is not caused by the effect of the cosolvents on the solution pH. Moreover, as previously observed by Gai and coworkers (22,23), our MD simulations indicate that the osmolytes all disrupt rather than strengthen the water tetrahedral H-bonding network, although TMAO seems the least disruptive to the water network (19,26). Consequently, we conclude that the observed red shifts primarily reflect direct interactions between water and the osmolytes.…”

Section: Resultssupporting

confidence: 87%

“…First of all, protein folding generally sequesters the peptide backbone from the solvent. Because TMAO interacts unfavorably with the peptide backbone (9,10), it will provide a stabilizing effect for any protein (19). Second, although it is generally accepted that hydrophobic forces provide the dominant driving force for protein folding (53), folded proteins do not completely sequester all hydrophobic groups from the solvent.…”

Section: Resultsmentioning

confidence: 99%

“…Garde and coworkers (19) concluded that TMAO has minimal impact on hydrophobic interactions, but Paul and Patey (20) suggested that TMAO actually weakens hydrophobic forces. Other studies suggested that osmolytes stabilize folded states of proteins indirectly by altering the bulk water structure.…”

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confidence: 99%

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Trimethylamine N -oxide stabilizes proteins via a distinct mechanism compared with betaine and glycine

Liao

Manson

DeLyser

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

165

197

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We report experimental and computational studies investigating the effects of three osmolytes, trimethylamine N-oxide (TMAO), betaine, and glycine, on the hydrophobic collapse of an elastin-like polypeptide (ELP). All three osmolytes stabilize collapsed conformations of the ELP and reduce the lower critical solution temperature (LSCT) linearly with osmolyte concentration. As expected from conventional preferential solvation arguments, betaine and glycine both increase the surface tension at the air-water interface. TMAO, however, reduces the surface tension. Atomically detailed molecular dynamics (MD) simulations suggest that TMAO also slightly accumulates at the polymer-water interface, whereas glycine and betaine are strongly depleted. To investigate alternative mechanisms for osmolyte effects, we performed FTIR experiments that characterized the impact of each cosolvent on the bulk water structure. These experiments showed that TMAO red-shifts the OH stretch of the IR spectrum via a mechanism that was very sensitive to the protonation state of the NO moiety. Glycine also caused a red shift in the OH stretch region, whereas betaine minimally impacted this region. Thus, the effects of osmolytes on the OH spectrum appear uncorrelated with their effects upon hydrophobic collapse. Similarly, MD simulations suggested that TMAO disrupts the water structure to the least extent, whereas glycine exerts the greatest influence on the water structure. These results suggest that TMAO stabilizes collapsed conformations via a mechanism that is distinct from glycine and betaine. In particular, we propose that TMAO stabilizes proteins by acting as a surfactant for the heterogeneous surfaces of folded proteins.osmolytes | protein folding | mechanism | spectroscopy | MD simulations M any organisms use small organic osmolytes to stabilize proteins in harsh environments, such as when the salinity is highly variable (1). In particular, trimethylamine N-oxide (TMAO) is known to counteract the denaturing effects of urea as well as salts, and it is present at high concentrations in some aquatic organisms (2). Its effects are often compared with the ions on the left side of the Hofmeister series, which help stabilize the native, folded structures of proteins (Fig. 1).Because of their fundamental biophysical importance, many studies have investigated the behavior and effects of osmolytes. In particular, Timasheff and coworkers (3, 4) proposed that osmolyte effects result from the relative partitioning of these molecules between the bulk solution and the protein-water interface. Stabilization should occur when osmolytes are depleted from the protein-water interface, but proteins will unfold when osmolytes accumulate at this interface. Accordingly, osmolyte effects are often interpreted in terms of an effective protein-water "surface tension." In fact, despite the significant differences between protein surfaces and air-water interfaces, osmolyte effects are often, although not always, consistent with their effect on the air-water interfa...

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Section: Resultssupporting

confidence: 87%

Section: Resultsmentioning

confidence: 99%

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Trimethylamine N -oxide stabilizes proteins via a distinct mechanism compared with betaine and glycine

Liao

Manson

DeLyser

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

165

197

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“…For longer repeats of the backbone model the preferential interaction parameter is usually negative indicating a significant decrease in the overall amount of TMAO as compared to bulk solution, but this depends on conformation and is quite noisy. The high relative error seen is common behavior for the preferential interaction parameter as evidenced by the calculations of others, [36][37][38] because the nature of the calculation depends upon the local number fluctuations at increasing distances. The inherent nosiness of the parameter, and the subsequent lack in statistical confidence of the interpretation, caused us to look at shorter ranged events that are less susceptible to artifacts caused by large local number fluctuations.…”

Section: Resultsmentioning

confidence: 84%

Backbone additivity in the transfer model of protein solvation

et al. 2010

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The transfer model implying additivity of the peptide backbone free energy of transfer is computationally tested. Molecular dynamics simulations are used to determine the extent of change in transfer free energy (DG tr ) with increase in chain length of oligoglycine with capped end groups. Solvation free energies of oligoglycine models of varying lengths in pure water and in the osmolyte solutions, 2M urea and 2M trimethylamine N-oxide (TMAO), were calculated from simulations of all atom models, and DG tr values for peptide backbone transfer from water to the osmolyte solutions were determined. The results show that the transfer free energies change linearly with increasing chain length, demonstrating the principle of additivity, and provide values in reasonable agreement with experiment. The peptide backbone transfer free energy contributions arise from van der Waals interactions in the case of transfer to urea, but from electrostatics on transfer to TMAO solution. The simulations used here allow for the calculation of the solvation and transfer free energy of longer oligoglycine models to be evaluated than is currently possible through experiment. The peptide backbone unit computed transfer free energy of 254 cal/mol/M compares quite favorably with 243 cal/mol/M determined experimentally.

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“…1, Inset) adopts a skewed tetrahedral structure with a charged oxygen capable of accepting hydrogen bonds (H bonds) and three hydrophobic (methyl) groups. This amphiphilic structural arrangement makes TMAO a rather special cosolvent, because it can form H bonds with water, self-associate in a manner similar to surfactants, and show preferential interactions with or exclusion (4)(5)(6)(7)(8)(9)(10)(11)(12) from certain protein functional groups (13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23). Indeed, these molecular properties of TMAO have been used, either individually or in combination, to rationalize its biological activities.…”

mentioning

confidence: 99%

Microscopic insights into the protein-stabilizing effect of trimethylamine N-oxide (TMAO)

Pazos

Gai

2014

Proc. Natl. Acad. Sci. U.S.A.

229

237

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Although it is widely known that trimethylamine N-oxide (TMAO), an osmolyte used by nature, stabilizes the folded state of proteins, the underlying mechanism of action is not entirely understood. To gain further insight into this important biological phenomenon, we use the C≡N stretching vibration of an unnatural amino acid, p-cyano-phenylalanine, to directly probe how TMAO affects the hydration and conformational dynamics of a model peptide and a small protein. By assessing how the lineshape and spectral diffusion properties of this vibration change with cosolvent conditions, we are able to show that TMAO achieves its protein-stabilizing ability through the combination of (at least) two mechanisms: (i) It decreases the hydrogen bonding ability of water and hence the stability of the unfolded state, and (ii) it acts as a molecular crowder, as suggested by a recent computational study, that can increase the stability of the folded state via the excluded volume effect.N ature employs a variety of small organic molecules to cope with osmotic stress. Trimethylamine N-oxide (TMAO) is one such naturally occurring osmolyte that protects intracellular components against disruptive stress conditions (1). In particular, previous studies have shown that TMAO is able to enhance protein stability and to counteract the denaturing effect of urea (2, 3). TMAO (Fig. 1, Inset) adopts a skewed tetrahedral structure with a charged oxygen capable of accepting hydrogen bonds (H bonds) and three hydrophobic (methyl) groups. This amphiphilic structural arrangement makes TMAO a rather special cosolvent, because it can form H bonds with water, self-associate in a manner similar to surfactants, and show preferential interactions with or exclusion (4-12) from certain protein functional groups (13-23). Indeed, these molecular properties of TMAO have been used, either individually or in combination, to rationalize its biological activities. For example, a prevailing view about TMAO is that its osmotic and protective role is caused by the molecule's tendency to be preferentially depleted from protein surfaces, as suggested by physicochemical measurements (24-28). This thermodynamic picture, as described by Bolen and coworkers (29), implies that the protein is preferentially hydrated. Although this notion is consistent with TMAO's being a protecting osmolyte, to the best of our knowledge no experimental studies have been carried out to directly examine the effect of TMAO on protein hydration dynamics, an aspect that is also important to protein function. Herein, we use twodimensional infrared (2D IR) spectroscopy and a site-specific IR probe to explore this critical issue and gain insight toward achieving a microscopic understanding of the molecular mechanism of the protecting action of TMAO.2D IR spectroscopy is capable of assessing the frequencyfrequency correlation function (FFCF) of a given IR probe, which reports on the underlying dynamics of events that lead to fluctuations of its vibrational frequency (30). For an IR probe that is able ...

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Osmolyte Trimethylamine-N-Oxide Does Not Affect the Strength of Hydrophobic Interactions: Origin of Osmolyte Compatibility

Cited by 143 publications

References 56 publications

Trimethylamine N -oxide stabilizes proteins via a distinct mechanism compared with betaine and glycine

Trimethylamine N -oxide stabilizes proteins via a distinct mechanism compared with betaine and glycine

Backbone additivity in the transfer model of protein solvation

Microscopic insights into the protein-stabilizing effect of trimethylamine N-oxide (TMAO)

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