Computational Protein Engineering: Bridging the Gap between Rational Design and Laboratory Evolution

Barrozo, Alexandre; Borštnar, Rok; Marloie, Gaël; Kamerlin, Shina Caroline Lynn

doi:10.3390/ijms131012428

Cited by 39 publications

(32 citation statements)

References 153 publications

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“…Another important aspect of the design front is the issue of controlling the enantioselectivity of enzyme active sites (e.g., see [49,72–74]). Our studies in this direction focused on the ability to use the EVB in an analysis that considers the protein relaxation consistently.…”

Section: Computer Aided Enzyme Designmentioning

confidence: 99%

Computer aided enzyme design and catalytic concepts

Frushicheva

Mills

Schöpf

et al. 2014

Current Opinion in Chemical Biology

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Gaining a deeper understanding of enzyme catalysis is of great practical and fundamental importance. Over the years it has become clear that despite advances made in experimental mutational studies, a quantitative understanding of enzyme catalysis will not be possible without the use of computer modeling approaches. While we believe that electrostatic preorganization is by far the most important catalytic factor, convincing the wider scientific community of this may require the demonstration of effective rational enzyme design. Here we make the point that the main current advances in enzyme design are basically advances in directed evolution and that computer aided enzyme design must involve approaches that can reproduce catalysis in well-defined test cases. Such an approach is provided by the empirical valence bond method.

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Section: Computer Aided Enzyme Designmentioning

confidence: 99%

Computer aided enzyme design and catalytic concepts

Frushicheva

Mills

Schöpf

et al. 2014

Current Opinion in Chemical Biology

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“…Detailed control of the effect of each mutation at the atomic level can be provided by in silico rational protein-engineering techniques (111–115). Recently, Haidar et al (116) engineered the human A6 TCR for enhanced affinity toward the Tax peptide/HLA-A2 MHC complex.…”

Section: Computer-aided Protein Engineeringmentioning

confidence: 99%

Structure-Based, Rational Design of T Cell Receptors

Zoete¹,

Irving²,

Ferber³

et al. 2013

Front. Immunol.

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Adoptive cell transfer using engineered T cells is emerging as a promising treatment for metastatic melanoma. Such an approach allows one to introduce T cell receptor (TCR) modifications that, while maintaining the specificity for the targeted antigen, can enhance the binding and kinetic parameters for the interaction with peptides (p) bound to major histocompatibility complexes (MHC). Using the well-characterized 2C TCR/SIYR/H-2K(b) structure as a model system, we demonstrated that a binding free energy decomposition based on the MM-GBSA approach provides a detailed and reliable description of the TCR/pMHC interactions at the structural and thermodynamic levels. Starting from this result, we developed a new structure-based approach, to rationally design new TCR sequences, and applied it to the BC1 TCR targeting the HLA-A2 restricted NY-ESO-1157–165 cancer-testis epitope. Fifty-four percent of the designed sequence replacements exhibited improved pMHC binding as compared to the native TCR, with up to 150-fold increase in affinity, while preserving specificity. Genetically engineered CD8+ T cells expressing these modified TCRs showed an improved functional activity compared to those expressing BC1 TCR. We measured maximum levels of activities for TCRs within the upper limit of natural affinity, KD = ∼1 − 5 μM. Beyond the affinity threshold at KD < 1 μM we observed an attenuation in cellular function, in line with the “half-life” model of T cell activation. Our computer-aided protein-engineering approach requires the 3D-structure of the TCR-pMHC complex of interest, which can be obtained from X-ray crystallography. We have also developed a homology modeling-based approach, TCRep 3D, to obtain accurate structural models of any TCR-pMHC complexes when experimental data is not available. Since the accuracy of the models depends on the prediction of the TCR orientation over pMHC, we have complemented the approach with a simplified rigid method to predict this orientation and successfully assessed it using all non-redundant TCR-pMHC crystal structures available. These methods potentially extend the use of our TCR engineering method to entire TCR repertoires for which no X-ray structure is available. We have also performed a steered molecular dynamics study of the unbinding of the TCR-pMHC complex to get a better understanding of how TCRs interact with pMHCs. This entire rational TCR design pipeline is now being used to produce rationally optimized TCRs for adoptive cell therapies of stage IV melanoma.

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“…The substrate is shown in stick mode, while the residue 207 is in sphere mode (Rothlisberger et al 2008;Siegel et al 2010). The artificial catalysts are always significantly inferior to naturally occurring enzymes in the aspect of catalytic efficiency, partially attributed to the undesirable electronic preorganization in the active site of the enzyme (Barrozo et al 2012;Kiss et al 2013). However, with the intimate understanding of the precise molecular details of enzyme catalysis and parallel progress in computational technology, this problem might be solved in the near future.…”

Section: Discussionmentioning

confidence: 99%

Rational design of esterase BioH with enhanced enantioselectivity towards methyl (S)-o-chloromandelate

Guo

et al. 2014

Appl Microbiol Biotechnol

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Methyl (R)-o-chloromandelate (R-CMM) is an intermediate for the platelet aggregation inhibitor clopidogrel. Its preparation through enzymatic resolution of the corresponding ester has been hindered by the lack of an enzyme with satisfying enantioselectivity and activity. In the present work, we aimed to improve the enzymatic enantioselectivity towards methyl (S)-o-chloromandelate (S-CMM) by rational design, using esterase BioH as a model enzyme. Based on the differences in the binding mode of S- and R-enantiomers at the active cavity of the enzyme, the steric and electronic interactions between the key amino acids of BioH and the enantiomers were finely tuned. The enantioselectivity of esterase BioH towards S-CMM was improved from 3.3 (the wild type) to 73.4 (L123V/L181A/L207F). Synergistic interaction was observed between point mutations, and insight into the source of enzymatic enantioselectivity was gained by molecular dynamics simulations. The results can provide a reference for the enzyme design of other enzymes towards S-CMM for the enhancement of enantioselectivity.

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Computational Protein Engineering: Bridging the Gap between Rational Design and Laboratory Evolution

Cited by 39 publications

References 153 publications

Computer aided enzyme design and catalytic concepts

Computer aided enzyme design and catalytic concepts

Structure-Based, Rational Design of T Cell Receptors

Rational design of esterase BioH with enhanced enantioselectivity towards methyl (S)-o-chloromandelate

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