Insertions and Deletions (Indels): A Missing Piece of the Protein Engineering Jigsaw

Miton, C.M.; Tokuriki, Nobuhiko

doi:10.1021/acs.biochem.2c00188

Cited by 25 publications

(23 citation statements)

References 134 publications

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“…The role of insertions and deletions (indels) of single amino acids, small motifs, or whole domains in potential engineering campaigns should not be underestimated. , In nature, indels play an important role by facilitating phenotypic jumps in evolutionary trajectories. The importance of indels in the functional diversification of protein families has been described in the literature before, for example in the context of loops close to the active site responsible for different substrate spectra or altered ligand binding . In engineering campaigns, indels are often overlooked due to the strong focus on substituting amino acids.…”

Section: Enzyme Engineering Strategiesmentioning

confidence: 99%

“…In nature, the reduction of stability and structural integrity of proteins caused by indels seem to be compensated by substitutions. The apparent coevolution of indels and substitutions could reveal compensatory epistatic interactions and constraints on evolutionary trajectories mediated by epistatic effects …”

Section: Residue Coevolution and Its Effects On Protein Sequence Spac...mentioning

confidence: 99%

“…The importance of indels in the functional diversification of protein families has been described in the literature before, for example in the context of loops close to the active site responsible for different substrate spectra or altered ligand binding. 41 In engineering campaigns, indels are often overlooked due to the strong focus on substituting amino acids. Readers are referred to Miton et al 41 for a comprehensive summary of the importance of indels in functional diversification and to Savino et al 42 for an overview of recent methodological advances in directed-evolution campaigns, semirational approaches, and computational protein engineering pipelines.…”

Section: Directed Evolutionmentioning

confidence: 99%

“…41 In engineering campaigns, indels are often overlooked due to the strong focus on substituting amino acids. Readers are referred to Miton et al 41 for a comprehensive summary of the importance of indels in functional diversification and to Savino et al 42 for an overview of recent methodological advances in directed-evolution campaigns, semirational approaches, and computational protein engineering pipelines.…”

Section: Directed Evolutionmentioning

confidence: 99%

“…The apparent coevolution of indels and substitutions could reveal compensatory epistatic interactions and constraints on evolutionary trajectories mediated by epistatic effects. 41…”

Section: Epistasismentioning

confidence: 99%

See 4 more Smart Citations

Learning Epistasis and Residue Coevolution Patterns: Current Trends and Future Perspectives for Advancing Enzyme Engineering

2022

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Engineering proteins and enzymes with the desired functionality has broad applications in molecular biology, biotechnology, biomedical sciences, health, and medicine. The vastness of protein sequence space and all the possible proteins it represents can pose a considerable barrier for enzyme engineering campaigns through directed evolution and rational design. The nonlinear effects of coevolution between amino acids in protein sequences complicate this further. Data-driven models increasingly provide scientists with the computational tools to navigate through the largely undiscovered forest of protein variants and catch a glimpse of the rules and effects underlying the topology of sequence space. In this review, we outline a complete theoretical journey through the processes of protein engineering methods such as directed evolution and rational design and reflect on these strategies and data-driven hybrid strategies in the context of sequence space. We discuss crucial phenomena of residue coevolution, such as epistasis, and review the history of models created over the past decade, aiming to infer rules of protein evolution from data and use this knowledge to improve the prediction of the structure− function relationship of proteins. Data-driven models based on deep learning algorithms are among the most promising methods that can account for the nonlinear phenomena of sequence space to some degree. We also critically discuss the available models to predict evolutionary coupling and epistatic effects (classical and deep learning) in terms of their capabilities and limitations. Finally, we present our perspective on possible future directions for developing data-driven approaches and provide key orientation points and necessities for the future of the fast-evolving field of enzyme engineering.

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Section: Enzyme Engineering Strategiesmentioning

confidence: 99%

Section: Residue Coevolution and Its Effects On Protein Sequence Spac...mentioning

confidence: 99%

Section: Directed Evolutionmentioning

confidence: 99%

Section: Directed Evolutionmentioning

confidence: 99%

“…The apparent coevolution of indels and substitutions could reveal compensatory epistatic interactions and constraints on evolutionary trajectories mediated by epistatic effects. 41…”

Section: Epistasismentioning

confidence: 99%

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Learning Epistasis and Residue Coevolution Patterns: Current Trends and Future Perspectives for Advancing Enzyme Engineering

2022

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A cellular selection identifies elongated flavodoxins that support electron transfer to sulfite reductase

et al. 2023

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Flavodoxins (Flds) mediate the flux of electrons between oxidoreductases in diverse metabolic pathways. To investigate whether Flds can support electron transfer to a sulfite reductase (SIR) that evolved to couple with a ferredoxin, we evaluated the ability of Flds to transfer electrons from a ferredoxin‐NADP reductase (FNR) to a ferredoxin‐dependent SIR using growth complementation of an Escherichia coli strain with a sulfur metabolism defect. We show that Flds from cyanobacteria complement this growth defect when coexpressed with an FNR and an SIR that evolved to couple with a plant ferredoxin. When we evaluated the effect of peptide insertion on Fld‐mediated electron transfer, we observed a sensitivity to insertions within regions predicted to be proximal to the cofactor and partner binding sites, while a high insertion tolerance was detected within loops distal from the cofactor and within regions of helices and sheets that are proximal to those loops. Bioinformatic analysis showed that natural Fld sequence variability predicts a large fraction of the motifs that tolerate insertion of the octapeptide SGRPGSLS. These results represent the first evidence that Flds can support electron transfer to assimilatory SIRs, and they suggest that the pattern of insertion tolerance is influenced by interactions with oxidoreductase partners.This article is protected by copyright. All rights reserved.

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Creation and optimization of artificial metalloenzymes: Harnessing the power of directed evolution and beyond

Zou,

Higginson,

Ward

2024

Chem

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Insertions and Deletions (Indels): A Missing Piece of the Protein Engineering Jigsaw

Cited by 25 publications

References 134 publications

Learning Epistasis and Residue Coevolution Patterns: Current Trends and Future Perspectives for Advancing Enzyme Engineering

Learning Epistasis and Residue Coevolution Patterns: Current Trends and Future Perspectives for Advancing Enzyme Engineering

A cellular selection identifies elongated flavodoxins that support electron transfer to sulfite reductase

Creation and optimization of artificial metalloenzymes: Harnessing the power of directed evolution and beyond

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