Chemical Interactions of Polyethylene Glycols (PEGs) and Glycerol with Protein Functional Groups: Applications to Effects of PEG and Glycerol on Protein Processes

Knowles, D.B.; Shkel, Irina A.; Phan, Noel; Sternke, Matt; Lingeman, Emily; Cheng, Xian; Cheng, Lixue; O’Connor, Kevin; Record, M. Thomas

doi:10.1021/acs.biochem.5b00246

Cited by 102 publications

(194 citation statements)

References 60 publications

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“…The free energy of transfer depends upon the effective potential of interaction between the probe and crowder molecules in solution, the nature of which may be deduced from the composition dependence of various solution properties (see for example 18–21 ).…”

Section: What Have We Learned From Theory and In Vitro Experiment?mentioning

confidence: 99%

“…Nonspecific electrostatic attraction between probe and crowder species was found to counteract the effects of volume exclusion to a varying extent, and in certain instances to dominate the overall behavior of individual tracer proteins, peptides, and nucleotides 19, 38–40 . Several approaches to the generalization of crowding theory to allow for enthalpic as well as entropic intermolecular interactions have been published 21, 36, 38, 41–46 . While these generalized theories can be used to rationalize experimental results, they are not yet capable of quantitatively predicting the effect of a specific macromolecular crowding species upon a specific macromolecular reaction.…”

Section: What Have We Learned From Theory and In Vitro Experiment?mentioning

confidence: 99%

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Macromolecular Crowding In Vitro , In Vivo , and In Between

Rivas

Minton

2016

Trends in Biochemical Sciences

403

370

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Biochemical processes take place in heterogeneous and highly volume occupied or crowded environments that can considerably influence the reactivity and distribution of participating macromolecules. Here we summarize the thermodynamic consequences of excluded volume and longer-ranged nonspecific intermolecular interactions for macromolecular reactions in volume-occupied media. In addition, we summarize and compare the information content of studies of crowding in vitro and in vivo. We emphasize the importance of characterizing the behavior not only of labeled tracer macromolecules, but also the composition and behavior of unlabeled macromolecules in the immediate vicinity of the tracer. Finally, we propose strategies for extending quantitative analyses of crowding in simple model systems to increasingly complex media up to and including intact cells.

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Section: What Have We Learned From Theory and In Vitro Experiment?mentioning

confidence: 99%

Section: What Have We Learned From Theory and In Vitro Experiment?mentioning

confidence: 99%

Macromolecular Crowding In Vitro , In Vivo , and In Between

Rivas

Minton

2016

Trends in Biochemical Sciences

403

370

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“…5,6 Often, the addition of sugars, polymers, and other osmolytes to protein solutions alters the chemical potential of the protein in its native and unfolded states. 3,7,8 Folding/unfolding equilibria are altered when addition of a given osmolyte shifts the chemical potential of the native state to a different extent than for the unfolded state. 9,10 Historically, measurements of protein solubility and/ or protein unfolding thermodynamics have often been used to infer differences between protein−water and protein−osmolyte interactions.…”

Section: ■ Introductionmentioning

confidence: 99%

Osmolyte Effects on Monoclonal Antibody Stability and Concentration-Dependent Protein Interactions with Water and Common Osmolytes

et al. 2016

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Preferential interactions of proteins with water and osmolytes play a major role in controlling the thermodynamics of protein solutions. While changes in protein stability and shifts in phase behavior are often reported with the addition of osmolytes, the underlying protein interactions with water and/or osmolytes are typically inferred rather than measured directly. In this work, Kirkwood−Buff integrals for protein−water interactions (G 12 ) and protein−osmolyte interactions (G 23 ) were determined as a function of osmolyte concentration from density measurements of antistreptavidin immunoglobulin gamma-1 (AS-IgG1) in ternary aqueous solutions for a set of common neutral osmolytes: sucrose, trehalose, sorbitol, and poly(ethylene glycol) (PEG). For sucrose and PEG solutions, both protein−water and protein−osmolyte interactions depend strongly on osmolyte concentrations (c 3 ). Strikingly, both osmolytes change from being preferentially excluded to preferentially accumulated with increasing c 3 . In contrast, sorbitol and trehalose solutions do not show large enough preferential interactions to be detected by densimetry. G 12 and G 23 values are used to estimate the transfer free energy for native AS-IgG1 (Δμ 2 N ) and compared with existing models. AS-IgG1 unfolding via calorimetry shows a linear increase in midpoint temperatures as a function of trehalose, sucrose, and sorbitol concentrations, but the opposite behavior for PEG. Together, the results highlight limitations of existing models and common assumptions regarding the mechanisms of protein stabilization by osmolytes. Finally, PEG preferential interactions destabilize the Fab regions of AS-IgG1 more so than the C H 2 or C H 3 domains, illustrating preferential interactions can be specific to different protein domains.

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“…19−22 This approach uses multiple linear regressions (based on the number of compounds used), which describe all the surface types present in the molecules. We added our 15 compounds to the list of 27 molecules published by Capp et al 19 As our model compounds were mostly nitrogen-containing aromatic heterocycles, we aimed to calculate α values for aromatic N surface types in addition to the surface types analyzed by the Record lab.…”

Section: Resultsmentioning

confidence: 99%

“…MATLAB (version R2016A) was used for fitting (a sample Excel sheet along with the MATLAB code is provided in the Supporting Information). Errors were calculated using eq S17 from the supplement of Knowles et al 22 …”

Section: Methodsmentioning

confidence: 99%

Aspects of Weak Interactions between Folate and Glycine Betaine

et al. 2016

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Folate, or vitamin B9, is an important compound in one-carbon metabolism. Previous studies have found weaker binding of dihydrofolate to dihydrofolate reductase in the presence of osmolytes. In other words, osmolytes are more difficult to remove from the dihydrofolate solvation shell than water; this shifts the equilibrium toward the free ligand and protein species. This study uses vapor-pressure osmometry to explore the interaction of folate with the model osmolyte, glycine betaine. This method yields a preferential interaction potential (μ23/RT value). This value is concentration-dependent as folate dimerizes. The μ23/RT value also tracks the deprotonation of folate’s N3–O4 keto–enol group, yielding a pKa of 8.1. To determine which folate atoms interact most strongly with betaine, the interaction of heterocyclic aromatic compounds (as well as other small molecules) with betaine was monitored. Using an accessible surface area approach coupled with osmometry measurements, deconvolution of the μ23/RT values into α values for atom types was achieved. This allows prediction of μ23/RT values for larger molecules such as folate. Molecular dynamics simulations of folate show a variety of structures from extended to L-shaped. These conformers possess μ23/RT values from −0.18 to 0.09 m–1, where a negative value indicates a preference for solvation by betaine and a positive value indicates a preference for water. This range of values is consistent with values observed in osmometry and solubility experiments. As the average predicted folate μ23/RT value is near zero, this indicates folate interacts almost equally well with betaine and water. Specifically, the glutamate tail prefers to interact with water, while the aromatic rings prefer betaine. In general, the more protonated species in our small molecule survey interact better with betaine as they provide a source of hydrogens (betaine is not a hydrogen bond donor). Upon deprotonation of the small molecule, the preference swings toward water interaction because of its hydrogen bond donating capacities.

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Chemical Interactions of Polyethylene Glycols (PEGs) and Glycerol with Protein Functional Groups: Applications to Effects of PEG and Glycerol on Protein Processes

Cited by 102 publications

References 60 publications

Macromolecular Crowding In Vitro , In Vivo , and In Between

Macromolecular Crowding In Vitro , In Vivo , and In Between

Osmolyte Effects on Monoclonal Antibody Stability and Concentration-Dependent Protein Interactions with Water and Common Osmolytes

Aspects of Weak Interactions between Folate and Glycine Betaine

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