Cover Feature: Modeling‐Assisted Design of Thermostable Benzaldehyde Lyases from Rhodococcus erythropolis for Continuous Production of α‐Hydroxy Ketones (ChemBioChem 7/2022)

Martin Peng,Martin K M Engqvist,Kersten S Rabe,Dominik L Siebert,Christof M Niemeyer

doi:10.1002/cbic.202100668

Martin Peng, Martin K M Engqvist + Show 3 more

Open Access

https://doi.org/10.1002/cbic.202100668

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Abstract

The right tool can do the trick: A novel thermostable benzaldehyde lyase from Rhodococcus erythropolis was identified using the computer-assisted prediction tool Tome, and two variants with altered active sites were constructed based on homology models. All new enzyme variants showed significantly increased thermostability compared with the only experimentally confirmed benzaldehyde lyase from Pseudomonas fluorescens to date. The enzyme variants showed different substrate specificities and different activities in the presence of a range of co-solvents. The most stable variant was then immobilized and used for continuous production of enantiopure benzoin. More information can be found in the Full Paper by K. S. Rabe et al.

Highlights

The production of active pharmaceutical ingredients (APIs) and natural compounds with multiple stereocenters is a highly topical field in research and development
Our work emphasizes that modeling-assisted rational design of ReBAL enzymes in combination with robust immobilization strategies is a powerful approach to create novel biocatalytic processes for continuous production of valued-added molecules
We demonstrated the immobilization of a thermostable benzaldehyde lyase in a continuous packed-bed microreactor for the production of αhydroxy ketones with constant activity over days

Summary

Introduction

The production of active pharmaceutical ingredients (APIs) and natural compounds with multiple stereocenters is a highly topical field in research and development. The BAL from Pseudomonas fluorescens[4,5] (PfBAL) is the only experimentally confirmed representative in this enzyme class recorded in the BRENDA[6] and TEED[7] database to date (for more detailed analysis of TEED database see supporting information page 20) It has an broad substrate spectrum[8,9] and can catalyze CÀ N bond formations,[10] cyclizations[11] and intramolecular Stetter reactions.[12] via rational engineering of its binding site, PfBAL was successfully altered to perform benzoylformate decarboxylase[13] or formolase reactions.[14] current applications are limited by the relatively low stability of this enzyme. Several strategies for the direct, site-selective immobilization of PfBAL on solid supports by employment of genetically-encoded tag systems, like the hexahistine (His6)-tag,[19,20] HaloTag[21] or aggregation-inducing tags,[22] did not show the desired performance and only the His6-tag variant could be used in a fluidic setup.[19]

Results and Discussion

Conclusion

Conflict of Interest