Poster Presentation The 44th Lorne Conference on Protein Structure and Function 2019

A revised biosynthetic pathway for cofactor F420 in bacteria (#234)

James Antoney 1 2 , Ghader Bashiri 3 , Ehab NM Jirgis 3 , Mihir V Shah 2 , Blair Ney 1 2 , Janine Copp 4 , Stephanie M Stuteley 3 , Sreevalsan Sreebhavan 5 , Brian Palmer 5 , Martin Middleditch 3 , Nobuhiko Tokuriki 4 , Chris Greening 6 , Edward N Baker 3 , Colin Scott 2 , Colin J Jackson 1 2
  1. Research School of Chemistry, Australian National University, Canberra, ACT, Australia
  2. Synthetic Biology Future Science Platform, CSIRO Land & Water Flagship, Canberra, ACT, Australia
  3. School of Biological Sciences and Maurice Wilkins Centre for Molecular Biodiscovery, The University of Auckland, Auckland, New Zealand
  4. Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia, Canada
  5. Auckland Cancer Society Research Centre, Faculty of Medical and Health Sciences, The University of Auckland, Auckland, New Zealand
  6. School of Biological Sciences, Monash University, Clayton, Victoria, Australia

Cofactor F420 plays critical roles in primary and secondary metabolism in a range of bacteria and archaea as a low-potential hydride transfer agent. It mediates a variety of important redox transformations involved in bacterial persistence, antibiotic biosynthesis, pro-drug activation and methanogenesis. However, the biosynthetic pathway for F420 has not been fully elucidated: neither the enzyme that generates the putative intermediate 2-phospho-L-lactate, nor the function of the FMN-binding C-terminal domain of the γ-glutamyl ligase (FbiB) in bacteria are known. Here we show that the guanylyltransferases FbiD and CofC accept phosphoenolpyruvate, rather than 2-phospho-L-lactate, as their substrate, leading to the formation of the previously uncharacterised intermediate, dehydro-F420-0. The C-terminal domain of FbiB then utilises FMNH2 to reduce dehydro-F420-0, which produces mature F420 species when combined with the γ-glutamyl ligase activity of the N-terminal domain. This new insight has allowed the heterologous expression F420 from a recombinant F420 biosynthetic pathway in Escherichia coli.

  1. Bashiri, G., Antoney, J. et al. A revised biosynthetic pathway for the cofactor F420 in bacteria. Preprint at https://www.biorxiv.org/content/biorxiv/early/2018/11/27/470336.full.pdf (2018).