Oceanic cyanobacteria are the most abundant oxygen-generating phototrophs on our planet and are therefore important to life. These organisms are infected by viruses called cyanophages, which have recently shown to encode metabolic genes that modulate host photosynthesis, phosphorus cycling and nucleotide metabolism. Herein we report the characterization of a wild-type flavin-dependent viral halogenase (VirX1) from a cyanophage. Notably, halogenases have been previously associated with secondary metabolism, tailoring natural products. Exploration of this viral halogenase reveals it capable of regioselective halogenation of a diverse range of substrates with a preference for forming aryl iodide species; this has potential implications for the metabolism of the infected host. Until recently, a flavin-dependent halogenase that is capable of iodination in vitro had not been reported. VirX1 is interesting from a biocatalytic perspective as it shows strikingly broad substrate flexibility and a clear preference for iodination, as illustrated by kinetic analysis. These factors together render it an attractive tool for synthesis.
Subscribe to Journal
Get full journal access for 1 year
only $13.33 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Rent or Buy article
Get time limited or full article access on ReadCube.
All prices are NET prices.
Agarwal, V. et al. Enzymatic halogenation and dehalogenation reactions: pervasive and mechanistically diverse. Chem. Rev. 117, 5619–5674 (2017).
Gkotsi, D. S., Dhaliwal, J., McLachlan, M. M., Mulholand, K. R. & Goss, R. J. M. Halogenases: powerful tools for biocatalysis (mechanisms applications and scope). Curr. Opin. Chem. Biol. 43, 119–126 (2018).
Weichold, V., Milbredt, D. & van Pée, K.-H. Specific enzymatic halogenation-from the discovery of halogenated enzymes to their applications in vitro and in vivo. Angew. Chem. Int. Ed. 55, 6374–6389 (2016).
Dong, C. et al. Tryptophan 7-halogenase (PrnA) structure suggests a mechanism for regioselective chlorination. Science 309, 2216–2219 (2005).
Keller, S. et al. Purification and partial characterization of tryptophan 7-halogenase (PrnA) from Pseudomonas fluorescence. Angew. Chem. Int. Ed. 39, 2300–2302 (2000).
Dong, C. J. et al. Crystal structure and mechanism of a bacterial fluorinating enzyme. Nature 427, 561–565 (2004).
Mori, S., Pang, A. H., Chandrika, N. T., Garneau-Tsodikova, S. & Tsodikov, O. V. Unusual substrate and halide versatility of phenolic halogenase PltM. Nat. Commun. 10, 1255 (2019).
Zeng, J. & Zhan, J. A novel fungal flavin-dependent halogenase for natural product biosynthesis. ChemBioChem. 11, 2119–2123 (2010).
Neumann, C. S., Walsh, C. T. & Kay, R. R. A flavin-dependent halogenase catalyzes the chlorination step in the biosynthesis of Dictyostelium differentiation-inducing factor 1. Proc. Natl Acad. Sci. USA 107, 5798–5803 (2010).
Lang, A. et al. Changing the regioselectivity of the tryptophan 7-halogenase PrnA by site directed mutagenesis. Angew. Chem. Int. Ed. 50, 2951–2951 (2011).
Glenn, W. S., Nims, E. & O’Connor, S. E. Reengineering a tryptophan halogenase to preferentially chlorinate a direct alkaloid precursor. J. Am. Chem. Soc. 133, 19348–19349 (2011).
Payne, J. T., Poor, C. B. & Lewis, J. C. Directed evolution of RebH for site selective halogenation of large biologically active molecules. Angew. Chem. Int. Ed. 54, 4226–4230 (2015).
Menon, B. R. K. et al. RadH: a versatile halogenase for integration into synthetic pathways. Angew. Chem. Int. Ed. 56, 11841–11845 (2017).
Schnepel, C., Mignes, H., Frese, M. & Sewald, N. A high-throughput fluorescence assay to determine the activity of tryptophan halogenases. Angew. Chem. Int. Ed. 55, 14159–14163 (2017).
Goss, R. J. M. & Gkotsi, D. S. Discovery and utilisation of wildly different halogenases, powerful new tools for medicinal chemistry. UK patent GB1803491.8 (2018).
Agarwal, V. et al. Biosynthesis of polybrominated aromatic organic compounds by marine bacteria. Nat. Chem. Bio. 10, 640–647 (2014).
Sullivan, M. B., Waterbury, J. B. & Chisholm, S. W. Cyanophage infecting the oceanic cyanobacterium Prochlorococcus. Nature 424, 1047–1051 (2003).
Rost, B. Twilight zone of protein sequence alignments. Protein Engineering 12, 85–94 (1999).
Kelley, L. A., Mezulis, S., Yates, C. M., Wass, M. N. & Sternberg, M. J. The Phyre2 web portal for protein modeling, prediction and analysis. Nat. Protoc. 10, 845–858 (2015).
James, M. J., Cuthbertson, J. D., O’Brien, P., Taylor, R. J. K. & Unsworth, W. P. Silver(i) or copper(ii)-mediated dearomatisation of aromatic ynones: direct access to spirocyclic scaffolds. Angew. Chem. Int. Ed. 54, 7640–7643 (2015).
Chambers, S. J. et al. Heteroaromatic acids and imines to azaspirocycles: stereoselective synthesis and 3D shape analysis. Chem. Eur. J. 22, 6496–6500 (2016).
Yeh, E., Blasiak, L. C., Koglin, A., Drennan, C. L. & Walsh, C. T. Chlorination by a long-lived intermediate in the mechanism of flavin-dependent halogenases. Biochem. 46, 1284–1292 (2007).
Holm, L. & Laakso, L. M. Dali server update. Nucl. Acids Res. 44, W351–W355 (2016).
Krissinel, E. Stock-based detection of protein oligomeric states in jsPISA. Nucl. Acids Res. 43, W314–W319 (2015).
Neubauer, P. R. et al. A flavin-dependent halogenase from metagenomic analysis prefers bromination over chlorination. PLoS ONE 13, e0196797 (2018).
Sharma, S. V. et al. Living GenoChemetics: hyphenating synthetic biology and synthetic chemistry in vivo. Nat. Commun. 8, 229 (2017).
Breitbart, M., Bonnain, C., Malki, K. & Sawaya, N. A. Phage puppet masters of the marine microbial relm. Nat. Microbiol. 3, 754–766 (2018).
Amachi, S. Microbial contribution to global iodine cycling: volatilization, accumulation, reduction, oxidation and sorption of iodine. Microbes. Environ. 23, 269–276 (2008).
Crockford, S. J. Evolutionary roots of iodine and thyroid hormones in cell-cell signalling. Integr. Com. Biol. 49, 155–166 (2009).
We thank the European Research Council under the European Union’s Seventh Framework Programme (FP7/2007–2013/ERC grant agreement no. 614779 GenoChemetics to R.J.M.G.), Syngenta and Wellcome ISSF (grant no. 204821/Z/16/Z to D.S.G.) for generous financial support. We thank G. Harris and M. Weckener (Harwell) for size-excluion chromatography multiangle light scattering and analytical ultracentrifugation analysis. We thank all of our colleagues, in particular, T. Smith and co-workers in the School of Chemistry and the Biomedical Sciences Research Complex at the University of St Andrews for all of the help that they have afforded us in the aftermath of the BMS fire. We thank I. M. Wilson for assistance with graphics.
The authors declare no competing interests.
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
About this article
Cite this article
Gkotsi, D.S., Ludewig, H., Sharma, S.V. et al. A marine viral halogenase that iodinates diverse substrates. Nat. Chem. 11, 1091–1097 (2019) doi:10.1038/s41557-019-0349-z
Nature Chemistry (2019)
ACS Central Science (2019)