An NMR Confirmation for Increased Folded State Entropy Following Loop Truncation

Gavrilov, Yulian; Dagan, Shlomi; Reich, Ziv; Scherf, Tali; Levy, Yaakov

doi:10.1021/acs.jpcb.8b09658

Cited by 6 publications

(6 citation statements)

References 56 publications

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“…In order to assess the convergence of the calculations, we focused on the side-chain S 2 axis,sim values since they, due to the higher flexibility of the side chains, may converge more slowly than the backbone order parameters. 55−57 We averaged the 240 original blocks of 10 ns into 10 blocks of 240 ns and performed a leave-N-out cross-validation with N = 1,..., 5 evaluate the convergence of the side-chain order parameters. 58 Specifically, for each N, we split the 10 blocks in a reference set of 10-N and a test set of N blocks (the left-out blocks), averaged the S 2 axis,sim values within the two sets, and computed the RMSD between the resulting average values of the order parameters.…”

Section: ■ Materials and Methodsmentioning

confidence: 99%

“…The conformational dynamics of the native state of a protein contributes to its conformational stability. If the dynamics of the native state is increased without significantly affecting the intramolecular interactions, it will lead to a more stable protein. , Typically, however, the increased dynamics is counteracted by fewer stabilizing interactions through entropy-enthalpy compensation in the more dynamic molecule. , In some cases, evolution has circumvented this problem. Thus, in hyperthermophilic proteins, Lys is preferred over Arg compared to the mesophilic homologs.…”

Section: Introductionmentioning

confidence: 99%

“…If the dynamics of the native state is increased without significantly affecting the intramolecular interactions, it will lead to a more stable protein. 5,6 Typically, however, the increased dynamics is counteracted by fewer stabilizing interactions through entropy-enthalpy compensation in the more dynamic molecule. 7,8 In some cases, evolution has circumvented this problem.…”

Section: ■ Introductionmentioning

confidence: 99%

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Double Mutant of Chymotrypsin Inhibitor 2 Stabilized through Increased Conformational Entropy

et al. 2022

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The conformational heterogeneity of a folded protein can affect not only its function but also stability and folding. We recently discovered and characterized a stabilized double mutant (L49I/I57V) of the protein CI2 and showed that state-of-the-art prediction methods could not predict the increased stability relative to the wild-type protein. Here, we have examined whether changed native-state dynamics, and resulting entropy changes, can explain the stability changes in the double mutant protein, as well as the two single mutant forms. We have combined NMR relaxation measurements of the ps–ns dynamics of amide groups in the backbone and the methyl groups in the side chains with molecular dynamics simulations to quantify the native-state dynamics. The NMR experiments reveal that the mutations have different effects on the conformational flexibility of CI2: a reduction in conformational dynamics (and entropy estimated from this) of the native state of the L49I variant correlates with its decreased stability, while increased dynamics of the I57V and L49I/I57V variants correlates with their increased stability. These findings suggest that explicitly accounting for changes in native-state entropy might be needed to improve the predictions of the effect of mutations on protein stability.

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Section: ■ Materials and Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: ■ Introductionmentioning

confidence: 99%

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Double Mutant of Chymotrypsin Inhibitor 2 Stabilized through Increased Conformational Entropy

et al. 2022

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“…Previous studies investigating the functional impact of indels generally fall into two categories ( Savino et al 2022 ). First, protein engineering studies have shown that indels can impact a protein's function, especially if they overlap important secondary structures ( Simm et al 2007 ; Arpino et al 2014 ; Tóth-Petróczy and Tawfik 2014 ; Gavrilov et al 2015 , 2018 ; Grocholski et al 2015 ; Liu et al 2015 , 2016 ; Jackson et al 2017 ; Halliwell et al 2018 ; Gonzalez et al 2019 ; Woods et al 2023 ). For example, Liu et al (2016) found that experimentally deleting amino acids in beta strands and alpha helices of green fluorescent protein tended to reduce fluorescence, while deletions outside such regions were relatively neutral.…”

Section: Introductionmentioning

confidence: 99%

Insertion–Deletion Events Are Depleted in Protein Regions with Predicted Secondary Structure

Yang,

Braga,

Dean

2024

Genome Biology and Evolution

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A fundamental goal in evolutionary biology and population genetics is to understand how selection shapes the fate of new mutations. Here we test the null hypothesis that insertion-deletion events (indels) in protein coding regions occur randomly with respect to secondary structures. We identified indels across 11,444 sequence alignments in mouse, rat, human, chimp, and dog genomes, then quantified their overlap with four different types of secondary structure – alpha helices, beta strands, protein bends, and protein turns – predicted by deep-learning methods of AlphaFold2. Indels overlapped secondary structures 54% as much as expected, and were especially under-represented over beta strands, which tend to form internal, stable regions of proteins. In contrast, indels were enriched by 155% over regions without any predicted secondary structures. These skews were stronger in the rodent lineages compared to the primate lineages, consistent with population genetic theory predicting that natural selection will be more efficient in species with larger effective population sizes. Nonsynonymous substitutions were also less common in regions of protein secondary structure, although not as strongly reduced as in indels. In a complementary analysis of thousands of human genomes, we showed that indels overlapping secondary structure segregated at significantly lower frequency than indels outside of secondary structure. Taken together, our study shows that indels are selected against if they overlap secondary structure, presumably because they disrupt the tertiary structure and function of a protein.

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“…If the dynamics of the native state is increased without significantly affecting the intermolecular interactions it will lead to a more stable protein. 5,6 Typically, however, the increased dynamics is counteracted by fewer stabilizing interactions through entropy-enthalpy compensation in the more dynamic molecule. 7,8 In some cases evolution has circumvented this problem.…”

Section: Introductionmentioning

confidence: 99%

Double mutant of chymotrypsin inhibitor 2 stabilized through increased conformational entropy

Gavrilov¹,

Kümmerer²,

Orioli³

et al. 2021

Preprint

Self Cite

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The conformational heterogeneity of a folded protein can affect both its function but also stability and folding. We recently discovered and characterized a stabilized double mutant (L49I/I57V) of the protein CI2 and showed that state-of-the-art prediction methods could not predict the increased stability relative to the wild-type protein. Here we have examined whether changed native state dynamics, and resulting entropy changes, can explain the stability changes in the double mutant protein, as well as the two single mutant forms. We have combined NMR relaxation measurements of the ps-ns dynamics of amide groups in the backbone and the methyl groups in the side-chains with molecular dynamics simulations to quantify the native state dynamics. The NMR experiments reveal that the mutations have different effects on the conformational flexibility of CI2: A reduction in conformational dynamics (and entropy) of the native state of L49I variant correlates with its decreased stability, while increased dynamics of the I57V and L49I/I57V variants correlates with their increased stability. These findings suggest that explicitly accounting for changes in native state entropy might be needed to improve the predictions of the effect of mutations on protein stability.

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An NMR Confirmation for Increased Folded State Entropy Following Loop Truncation

Cited by 6 publications

References 56 publications

Double Mutant of Chymotrypsin Inhibitor 2 Stabilized through Increased Conformational Entropy

Double Mutant of Chymotrypsin Inhibitor 2 Stabilized through Increased Conformational Entropy

Insertion–Deletion Events Are Depleted in Protein Regions with Predicted Secondary Structure

Double mutant of chymotrypsin inhibitor 2 stabilized through increased conformational entropy

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