Catalytic mechanism and molecular engineering of quinolone biosynthesis in dioxygenase AsqJ

Nat Commun. 2018 Mar 21;9(1):1168. doi: 10.1038/s41467-018-03442-2.


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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alkaloids / biosynthesis*
  • Alpha-Ketoglutarate-Dependent Dioxygenase FTO / chemistry*
  • Alpha-Ketoglutarate-Dependent Dioxygenase FTO / genetics
  • Alpha-Ketoglutarate-Dependent Dioxygenase FTO / metabolism
  • Aspergillus nidulans / chemistry
  • Aspergillus nidulans / enzymology*
  • Biocatalysis
  • Catalytic Domain
  • Crystallography, X-Ray
  • Fungal Proteins / chemistry*
  • Fungal Proteins / genetics
  • Fungal Proteins / metabolism
  • Gene Expression
  • Kinetics
  • Molecular Dynamics Simulation
  • Protein Binding
  • Protein Conformation, alpha-Helical
  • Protein Conformation, beta-Strand
  • Protein Engineering / methods
  • Protein Interaction Domains and Motifs
  • Quantum Theory
  • Quinolones / metabolism*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Substrate Specificity
  • Thermodynamics


  • Alkaloids
  • Fungal Proteins
  • Quinolones
  • Recombinant Proteins
  • Alpha-Ketoglutarate-Dependent Dioxygenase FTO