Aqueous ionic liquids redistribute local enzyme stability via long-range perturbation pathways

Harrar, Till El; Frieg, Benedikt; Davari, Mehdi D.; Jaeger, Karl‐Erich; Schwaneberg, Ulrich; Gohlke, Holger

doi:10.1016/j.csbj.2021.07.001

Cited by 16 publications

(32 citation statements)

References 119 publications

(210 reference statements)

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“…From the RMSF we find that, in general, the fluctuations of the individual amino acids are dampened with increasing ionic strength. This is consistent with previous protein-IL simulation studies that have shown that the addition of ILs reduces protein dynamics …”

Section: Resultssupporting

confidence: 93%

“…This is consistent with previous protein-IL simulation studies that have shown that the addition of ILs reduces protein dynamics. 59 …”

Section: Resultsmentioning

confidence: 99%

“…This is consistent with previous protein-IL simulation studies that have shown that the addition of ILs reduces protein dynamics. 59 We note that the X-ray crystal structure (Figure 6A) presents an idealized Y-shaped conformer of the antibody.…”

Section: Simulations For Assisting In Evaluating Predictions and Inte...mentioning

confidence: 93%

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Ionic Liquid-Based Strategy for Predicting Protein Aggregation Propensity and Thermodynamic Stability

Shmool

Martin

Matthews

et al. 2022

JACS Au

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Novel drug candidates are continuously being developed to combat the most life-threatening diseases; however, many promising protein therapeutics are dropped from the pipeline. During biological and industrial processes, protein therapeutics are exposed to various stresses such as fluctuations in temperature, solvent pH, and ionic strength. These can lead to enhanced protein aggregation propensity, one of the greatest challenges in drug development. Recently, ionic liquids (ILs), in particular, biocompatible choline chloride ([Cho]Cl)-based ILs, have been used to hinder stress-induced protein conformational changes. Herein, we develop an IL-based strategy to predict protein aggregation propensity and thermodynamic stability. We examine three key variables influencing protein misfolding: pH, ionic strength, and temperature. Using dynamic light scattering, zeta potential, and variable temperature circular dichroism measurements, we systematically evaluate the structural, thermal, and thermodynamic stability of fresh immunoglobin G4 (IgG4) antibody in water and 10, 30, and 50 wt % [Cho]Cl. Additionally, we conduct molecular dynamics simulations to examine IgG4 aggregation propensity in each system and the relative favorability of different [Cho]Cl-IgG4 packing interactions. We re-evaluate each system following 365 days of storage at 4 °C and demonstrate how to predict the thermodynamic properties and protein aggregation propensity over extended storage, even under stress conditions. We find that increasing [Cho]Cl concentration reduced IgG4 aggregation propensity both fresh and following 365 days of storage and demonstrate the potential of using our predictive IL-based strategy and formulations to radically increase protein stability and storage.

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Section: Resultssupporting

confidence: 93%

“…This is consistent with previous protein-IL simulation studies that have shown that the addition of ILs reduces protein dynamics. 59 …”

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Ionic Liquid-Based Strategy for Predicting Protein Aggregation Propensity and Thermodynamic Stability

Shmool

Martin

Matthews

et al. 2022

JACS Au

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“…Also, some negatively charging approaches emphasized that negative enzyme surfaces favorably diminish the attraction of anions. − , Interestingly, our recent study revealed that the effects of cations and anions depend on their different combinations . In detail, BMIM + cations interacted more favorably with the BSLA surface in the case of halogenated-BMIM (i.e., [BMIM]Cl, [BMIM]Br, [BMIM]I), whereas BMIM + cation interactions were almost equal with TfO – anions (in case of [BMIM][TfO]) on the BSLA surface. , In addition, we found that identifying the perturbation pathways from IL indirect effects and specific IL ion–residue interactions enable the effective prediction of focused variant libraries with enhanced IL resistance …”

Section: Introductionmentioning

confidence: 96%

“…11,34 In addition, we found that identifying the perturbation pathways from IL indirect effects and specific IL ion−residue interactions enable the effective prediction of focused variant libraries with enhanced IL resistance. 35 Collectively, charge engineering of enzymes and its implication on IL ions is complicated and is far from fully understood, which hamper the protein engineers' ability to modify the enzymatic activity and stability in ILs efficiently. Generally, the global design principles of enzymes are difficult to find in random mutagenesis libraries.…”

Section: ■ Introductionmentioning

confidence: 99%

How Does Surface Charge Engineering of Bacillus subtilis Lipase A Improve Ionic Liquid Resistance? Lessons Learned from Molecular Dynamics Simulations

Pramanik

Cui

Dhoke

et al. 2022

ACS Sustainable Chem. Eng.

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Biocatalysis in ionic liquids (ILs) gained substantial interest due to solvent properties of the ILs, such as near-zero vapor pressure, high thermal stability, and wide tunability. Enzymes are often not catalytically active in ILs; therefore, understanding and improving enzyme resistance in ILs are essential to enable efficient biocatalysis in ionic liquids. Surface charge engineering has repeatedly been reported to enhance enzyme resistance toward ILs. However, the molecular knowledge about how substitutions to charged amino acids improve enzyme activity in an IL is far from being understood. Here, we report a comprehensive study to provide some principles of how surface charged amino acid substitutions (negatively and positively) strengthen the IL resistance of the Bacillus subtilis lipase A (BSLA) in [BMIM]Cl. Twenty typical BSLA substitutions (ten beneficial and ten nonbeneficial, obtained from the BSLA-SSM library) were studied by molecular dynamics (MD) simulations in the [BMIM]Cl system. The BSLA-IL interaction patterns were printed by analyzing several structural-and solvation-based observables. Lessons learned by analyzing the SSM library of BSLA comprise the following: (i) A general trend was found where both negatively and positively charged substitutions increased the essential water molecules locally at the substitution site, thereby contributing to the overall protein hydration shell. (ii) Electrostatic repulsion of both IL ions and the refined hydration shell are ultimately the two main drivers to enhanced IL resistance. The analysis of 20 BSLA substitutions and the identified common interactions reveals that surface charge engineering is very likely to be a general protein engineering strategy to enhance lipase/enzyme activity in ILs. Moreover, this study also suggests that MD is a valuable technique to screen for beneficial substitutions that repel/recruit surface solvation.

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Efficient asymmetric synthesis of ethyl (R)-3-hydroxybutyrate by recombinant Escherichia coli cells under high substrate loading using eco-friendly ionic liquids as cosolvent

Duan

Wang

Ouyang

et al. 2023

Bioprocess Biosyst Eng

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Aqueous ionic liquids redistribute local enzyme stability via long-range perturbation pathways

Abstract: Graphical abstract

Cited by 16 publications

References 119 publications

Ionic Liquid-Based Strategy for Predicting Protein Aggregation Propensity and Thermodynamic Stability

Ionic Liquid-Based Strategy for Predicting Protein Aggregation Propensity and Thermodynamic Stability

How Does Surface Charge Engineering of Bacillus subtilis Lipase A Improve Ionic Liquid Resistance? Lessons Learned from Molecular Dynamics Simulations

Efficient asymmetric synthesis of ethyl (R)-3-hydroxybutyrate by recombinant Escherichia coli cells under high substrate loading using eco-friendly ionic liquids as cosolvent

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