Abstract
The recently discovered FeII/α-ketoglutarate-dependent dioxygenase AsqJ from Aspergillus nidulans stereoselectively catalyzes a multistep synthesis of quinolone alkaloids, natural products with significant biomedical applications. To probe molecular mechanisms of this elusive catalytic process, we combine here multi-scale quantum and classical molecular simulations with X-ray crystallography, and in vitro biochemical activity studies. We discover that methylation of the substrate is essential for the activity of AsqJ, establishing molecular strain that fine-tunes π-stacking interactions within the active site. To rationally engineer AsqJ for modified substrates, we amplify dispersive interactions within the active site. We demonstrate that the engineered enzyme has a drastically enhanced catalytic activity for non-methylated surrogates, confirming our computational data and resolved high-resolution X-ray structures at 1.55 Å resolution. Our combined findings provide crucial mechanistic understanding of the function of AsqJ and showcase how combination of computational and experimental data enables to rationally engineer enzymes.
| Original language | English (US) |
|---|---|
| Journal | Nature Communications |
| Volume | 9 |
| Issue number | 1 |
| DOIs | |
| State | Published - Mar 21 2018 |
| Externally published | Yes |
Bibliographical note
KAUST Repository Item: Exported on 2020-10-01Acknowledgements: This work was supported by the German Research Foundation (SFB1035 project B12 to V.R.I.K., SFB749 to M.G.) and by the 1974-01 TUM-KAUST agreement on selective C–H bond activation (A.B.). The authors are thankful for the computing time provided by the Leibniz Rechenzentrum. We thank the staff of the beamline X06SA at the Paul Scherrer Institute, SLS, Villigen (Switzerland) for assistance during data collection.
This publication acknowledges KAUST support, but has no KAUST affiliated authors.