Applications of Protein Thermodynamic Database for Understanding Protein Mutant Stability and Designing Stable Mutants

Gromiha, M. Michael; Paruchuri, Anoosha; Huang, Liang-Tsung

doi:10.1007/978-1-4939-3572-7_4

Cited by 21 publications

(9 citation statements)

References 74 publications

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“…The algorithms are better at predicting deletion mutations in the hydrophobic core, than mutations that increase the size of the side chain, mutations on the protein surface and mutations where electrostatic interactions contribute to the stabilization. This is partly a result of the data available for training the algorithms that mainly consist of deletion mutations in the hydrophobic core 26 , A particular challenge in predicting stabilizing protein variants is that among the few single substitutions that are actually stabilizing the effects are often small, so that multiple substitutions may be needed to create a substantial stabilizing effect 17 . As the effects of the mutations are not always independent, and non-additivity may result in both positive or negative epistasis 27 , 28 , it can be difficult to predict the stability of proteins with multiple substitutions.…”

Section: Introductionmentioning

confidence: 99%

Synergistic stabilization of a double mutant in chymotrypsin inhibitor 2 from a library screen in E. coli

et al. 2021

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Most single point mutations destabilize folded proteins. Mutations that stabilize a protein typically only have a small effect and multiple mutations are often needed to substantially increase the stability. Multiple point mutations may act synergistically on the stability, and it is often not straightforward to predict their combined effect from the individual contributions. Here, we have applied an efficient in-cell assay in E. coli to select variants of the barley chymotrypsin inhibitor 2 with increased stability. We find two variants that are more than 3.8 kJ mol−1 more stable than the wild-type. In one case, the increased stability is the effect of the single substitution D55G. The other case is a double mutant, L49I/I57V, which is 5.1 kJ mol−1 more stable than the sum of the effects of the individual mutations. In addition to demonstrating the strength of our selection system for finding stabilizing mutations, our work also demonstrate how subtle conformational effects may modulate stability.

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Section: Introductionmentioning

confidence: 99%

Synergistic stabilization of a double mutant in chymotrypsin inhibitor 2 from a library screen in E. coli

et al. 2021

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“…The algorithms are better at predicting deletion mutations in the hydrophobic core, than mutations that increase the size of the side chain, mutations on the protein surface and mutations where electrostatic interactions contribute to the stabilization. This is partly a result of the data available for training the algorithms that mainly consist of deletion mutations in the hydrophobic core (Gromiha et al, 2016). A particular challenge in predicting stabilizing protein variants is that among the few single substitutions that are actually stabilizing the effects are often small, so that multiple substitutions may be needed to create a substantial stabilizing effect (Goldenzweig et al, 2018).…”

Section: Introductionmentioning

confidence: 99%

Synergistic stabilization of a double mutant in CI2 from anin-celllibrary screen

Hamborg

Granata

Olsen

et al. 2020

Preprint

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Most single point mutations destabilize folded proteins. Mutations that stabilize a protein typically only have a small effect and multiple mutations are often needed to substantially increase the stability. Multiple point mutations may act synergistically on the stability, and it is not straightforward to predict their combined effect from the individual contributions. Here, we have applied an efficient in-cell assay to select variants of the barley chymotrypsin inhibitor 2 with increased stability. We find two variants that are more than 3.8 kJ/mol more stable than the wild-type. In one case the increased stability is the effect of the single substitution D55G. The other case is a double mutant, L49I/I57V, which is 5.1 kJ/mol more stable than the sum of the effects of the individual mutations. In addition to demonstrating the strength of our selection system for finding stabilizing mutations, our work also demonstrate how subtle conformational effects may modulate stability.

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“… 7 , 30 , 31 Let us provide some pieces of evidence that support this important conjecture. The proteins’ free energy of unfolding (Δ G U ) spans a wide range of variations, viz., between 5 and 25 kcal/mol, 32 revealing the complexity of the “protein folding problem”. 14 , 33 − 35 However, its range of variation upon point mutations (ΔΔ G U ) is small and well-defined, revealing the validity of the thermodynamic hypothesis or Anfinsen dogma, 14 as explained next.…”

Section: Introductionmentioning

confidence: 99%

Proteins’ Evolution upon Point Mutations

Vila

2022

ACS Omega

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As the reader must be already aware, state-of-the-art protein folding prediction methods have reached a smashing success in their goal of accurately determining the three-dimensional structures of proteins. Yet, a solution to simple problems such as the effects of protein point mutations on their (i) native conformation; (ii) marginal stability; (iii) ensemble of high-energy nativelike conformations; and (iv) metamorphism propensity and, hence, their evolvability, remains as an unsolved problem. As a plausible solution to the latter, some properties of the amide hydrogen-deuterium exchange, a highly sensitive probe of the structure, stability, and folding of proteins, are assessed from a new perspective. The preliminary results indicate that the protein marginal stability change upon point mutations provides the necessary and sufficient information to estimate, through a Boltzmann factor, the evolution of the amide hydrogen exchange protection factors and, consequently, that of the ensemble of folded conformations coexisting with the native state. This work contributes to our general understanding of the effects of point mutations on proteins and may spur significant progress in our efforts to develop methods to determine the appearance of new folds and functions accurately.

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Applications of Protein Thermodynamic Database for Understanding Protein Mutant Stability and Designing Stable Mutants

Cited by 21 publications

References 74 publications

Synergistic stabilization of a double mutant in chymotrypsin inhibitor 2 from a library screen in E. coli

Synergistic stabilization of a double mutant in chymotrypsin inhibitor 2 from a library screen in E. coli

Synergistic stabilization of a double mutant in CI2 from anin-celllibrary screen

Proteins’ Evolution upon Point Mutations

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