Poster Presentation The 16th Australian Peptide Conference 2025

Phage-encoded bismuth bicycles enable instant access to targeted bioactive peptides (#117)

Upamali Somathilake 1 , Sven Ullrich 1 2 , Minghao Shang 1 , Christoph Nitsche 1
  1. Research School of Chemistry, Australian National University, Canberra, Australian Capital Territory, Australia
  2. Department of chemistry, University of Tokyo, Bonkyo-ku Tokyo, Japan

Phage display is an efficient screening platform that allows the discovery of promising drug candidates, including cyclic and bicyclic peptide ligands.1 Constraining peptides around metal centres has emerged as an attractive strategy to synthesise bicyclic peptides.2,3 This study applies the metal-peptide bicyclisation strategy on phage, displaying bismuth(III) and arsenic(III) metal(loid) peptides. Applying this bismuth-bicyclisation chemistry on a protein fused to a three-cysteine peptide confirmed that the formation of the bismuth-bicycle conjugate on proteins is feasible under fully biocompatible conditions. To select bismuth-bicyclic peptide ligands that exhibit affinity towards our target; streptavidin, we used water-insoluble bismuth tribromide (BiBr3) as well as water-soluble bismuth tripotassium dicitrate (gastrodenol), avoiding the potential challenges associated with the use of organic co-solvents. Additionally, a parallel screening was conducted to select arsenic bicyclic peptides by modifying the phage library with water-soluble sodium arsenite (NaAsO2). No effect of these bicyclisation reagents on the phage infectivity was observed at screening concentrations. To favour the selection of peptides that bind to the ligand-binding site of streptavidin, competitive elution was conducted with biotin.4 Despite the use of two different bismuth-based reagents, both Bi(III) screenings resulted in similar enriched sequences. Several Bi(III)- and As(III)-bicyclic peptides identified through the screenings were synthesised in situ from their linear analogues. Subsequent evaluation through surface plasmon resonance revealed two orders of magnitude higher affinity for bicycles compared to their linear counterparts, with KD values in the lower micromolar range.5

  1. Sohrabi, C.; Foster, A.; Tavassoli, A., Methods for generating and screening libraries of genetically encoded cyclic peptides in drug discovery. Nat. Rev. Chem. 2020, 4, 90–101.
  2. Voss, S.; Rademann, J.; Nitsche, C., Peptide–bismuth bicycles: In situ access to stable constrained peptides with superior bioactivity. Angew. Chem. Int. Ed.; 2022, e202113857.
  3. Ullrich, S.; Ghosh, P.; Shang, M.; Siryer, S.; Kumaresan, S.; Panda, B.; Davies, L.J.; Somathilake, U.; Patel, A. P.; Nitsche C., Synthesis and stability studies of constrained peptide–antimony bicycles. Aust. J. Chem. 2024, 77, CH24094.
  4. Owens, A. E.; Iannuzzelli, J. A.; Gu, Y. Fasan, R., MOrPH-PhD: An integrated phage display platform for the discovery of functional genetically encoded peptide macrocycles. ACS Cent. Sci. 2020, 6, 368–381.
  5. Ullrich, S.; Somathilake, U.; Shang, M.; Nitsche, C., Phage-encoded bismuth bicycles enable instant access to targeted bioactive peptides. Commun. Chem. 2024, 7, 143.