Metrics that Differentiate the Origins of Osmolyte Effects on Protein Stability: A Test of the Surface Tension Proposal

Auton, Matthew; Ferreon, Allan Chris M.; Bolen, D. Wayne

doi:10.1016/j.jmb.2006.07.003

Cited by 80 publications

(93 citation statements)

References 61 publications

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“…Such a result suggests that TMAO is actually enriched rather than depleted at hydrophobic interfaces. This finding is consistent with previous measurements (3,17,31), although it is perhaps puzzling given the fact that TMAO is nearly as effective as glycine at depressing the LCST of the ELP (Fig. 3A).…”

Section: Resultssupporting

confidence: 92%

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.

166

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: 92%

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.

166

197

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“…The nonzero peak observed in this histogram is clearly shifted relative to that observed for the 0 M TMAO data, showing a higher E FRET of approximately 0.86, corresponding to a more compact protein dimension. This observation is consistent with the known ability of TMAO to stabilize compact folded states of proteins (25,26,(28)(29)(30)49) and induce folding of unfolded proteins (30,50). To further probe the α-synuclein structural landscape in the presence of TMAO, we performed a series of smFRET experiments in varying TMAO concentrations.…”

Section: Resultssupporting

confidence: 80%

“…The interaction between TMAO and the protein backbone is highly unfavorable (62). Consequently, the protein shifts to collapsed forms with increasing concentrations of the protecting osmolyte TMAO (50). On the contrary, the denaturing osmolyte urea interacts favorably with the protein backbone (62).…”

Section: Resultsmentioning

confidence: 99%

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

Ferreon

Moosa

Gambin

et al. 2012

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

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Protein structure and function depend on a close interplay between intrinsic folding energy landscapes and the chemistry of the protein environment. Osmolytes are small-molecule compounds that can act as chemical chaperones by altering the environment in a cellular context. Despite their importance, detailed studies on the role of these chemical chaperones in modulating structure and dimensions of intrinsically disordered proteins have been limited. Here, we used single-molecule Förster resonance energy transfer to test the counteraction hypothesis of counterbalancing effects between the protecting osmolyte trimethylamine-N-oxide (TMAO) and denaturing osmolyte urea for the case of α-synuclein, a Parkinson's disease-linked protein whose monomer exhibits significant disorder. The single-molecule experiments, which avoid complications from protein aggregation, do not exhibit clear solvent-induced cooperative protein transitions for these osmolytes, unlike results from previous studies on globular proteins. Our data demonstrate the ability of TMAO and urea to shift α-synuclein structures towards either more compact or expanded average dimensions. Strikingly, the experiments directly reveal that a 2∶1 ½urea∶½TMAO ratio has a net neutral effect on the protein's dimensions, a result that holds regardless of the absolute osmolyte concentrations. Our findings shed light on a surprisingly simple aspect of the interplay between urea and TMAO on α-synuclein in the context of intrinsically disordered proteins, with potential implications for the biological roles of such chemical chaperones. The results also highlight the strengths of single-molecule experiments in directly probing the chemical physics of protein structure and disorder in more chemically complex environments.Parkinson's disease | protein folding | urea-TMAO counteraction | smFRET P roteins are dynamic entities that are in constant interaction with their environment. Several components of the protein environment can affect the folding landscape (1) and function, including solvents (2), osmolytes (3), crowding agents (4), and small-molecule and macromolecular ligands (5-7). Osmolytes are naturally occurring low-molecular weight compounds that are utilized by biological systems as chemical chaperones that counteract deleterious effects of extreme physical conditions such as high osmotic and hydrostatic pressures (8, 9), dehydration (10), and high or low temperatures (11, 12). Urea, a major metabolic byproduct, is known to be used as a balancing osmolyte by several marine vertebrates (8), air-breathing teleosts (13) and some amphibians (14, 15) to deal with osmotic stress. In mammalian kidneys, urea plays an important role in balancing the medullary osmotic gradient (16). However, even at physiologically relevant concentrations, urea shows a strong denaturing effect on proteins (8,17). This apparent paradox is solved by the activity of several protecting osmolytes (e.g., methylamines and polyhydric alcohols) found in these urea-rich biological systems (8,18,19).T...

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“…The effect of proline can be rationalized by its solvophobic destabilization of partially folded states and early aggregates (both of which expose excess hydrophobic surface) and its favorable effect on solubilization on the native state. 15,[22][23][24][25] In addition to its utility for monitoring aggregation in vivo, FlAsH can be advantageously used as a read-out of aggregation in vitro. We found that the aggregation kinetics monitored by FlAsH fluorescence faithfully recapitulates the kinetics of monomer depletion in the time course of aggregation.…”

Section: Review Of Our In Vivo Folding and Aggregation Studiesmentioning

confidence: 99%

From the test tube to the cell: Exploring the folding and aggregation of a β‐clam protein

Ignatova¹,

Krishnan²,

Bombardier³

et al. 2007

Biopolymers

107

126

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A crucial challenge in present biomedical research is the elucidation of how fundamental processes like protein folding and aggregation occur in the complex environment of the cell. Many new physico‐chemical factors like crowding and confinement must be considered, and immense technical hurdles must be overcome in order to explore these processes in vivo. Understanding protein misfolding and aggregation diseases and developing therapeutic strategies to these diseases demand that we gain mechanistic insight into behaviors and misbehaviors of proteins as they fold in vivo. We have developed a fluorescence approach using FlAsH labeling to study the thermodynamics of folding of a model β‐rich protein, cellular retinoic acid binding protein (CRABP) in Escherichia coli cells. The labeling approach has also enabled us to follow aggregation of a modified version of CRABP and chimeras between CRABP and huntingtin exon 1 with its glutamine repeat tract. In this article, we review our recent results using FlAsH labeling to study in‐vivo folding and present new observations that hint at fundamental differences between the thermodynamics and kinetics of protein folding in vivo and in vitro. © 2007 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 88: 157–163, 2007. This article was originally published online as an accepted preprint. The ‘Published Online’ date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com

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Metrics that Differentiate the Origins of Osmolyte Effects on Protein Stability: A Test of the Surface Tension Proposal

Cited by 80 publications

References 61 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

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

From the test tube to the cell: Exploring the folding and aggregation of a β‐clam protein

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