Further Characterization of Fungal Halogenase RadH and Its Homologs

Biomolecules. 2023 Jul 6;13(7):1081. doi: 10.3390/biom13071081.

Abstract

RadH is one of the flavin-dependent halogenases that has previously exhibited promising catalytic activity towards hydroxycoumarin, hydroxyisoquinoline, and phenolic derivatives. Here, we evaluated new functional homologs of RadH and expanded its specificities for the halogenation of non-tryptophan-derived, heterocyclic scaffolds. Our investigation revealed that RadH could effectively halogenate hydroxyquinoline and hydroxybenzothiophene. Assay optimization studies revealed the need to balance the various co-factor concentrations and where a GDHi co-factor recycling system most significantly improves the conversion and efficiency of the reaction. A crystal structure of RadH was also obtained with a resolution of 2.4 Å, and docking studies were conducted to pinpoint the binding and catalytic sites for substrates.

Keywords: X-ray crystallography; flavin-dependent halogenases; halogenation; heteroaromatic.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Catalytic Domain
  • Flavins / chemistry
  • Flavins / metabolism
  • Halogenation*
  • Oxidoreductases* / metabolism

Substances

  • Oxidoreductases
  • Flavins

Grants and funding

This research was funded by the National Research Foundation, Singapore (NRF-CRP19-2017-05-00), and the Agency for Science, Technology, and Research (A*STAR), Singapore (C211917006 and C211917010). G.A.G. acknowledges funding from A*STAR Graduate Academy (A*GA) for his PhD scholarship.