Oral Presentation The 16th Australian Peptide Conference 2025

Chemical Synthesis and Characterisation of the Structurally and Functionally Unique Peptide Natural Product Recifin A (130787)

K. Johan Rosengren 1 , Christina I Schroeder 2 , Richard J Clark 1 , Taylor B Smallwood 1 , Lauren RH Krumpe 3 , Barry R O'Keefe 3
  1. The University of Queensland, Brisbane, QLD, Australia
  2. Genentech Inc, South San Francisco, CA 94080, USA
  3. National Cancer Institute, National Institute of Health, Frederick, MD 21702, USA

Recifin A is a 42 amino acid cysteine-rich peptide isolated from the marine sponge Axinella sp. It represents the inaugural member of a newly identified structural class termed the "tyrosine-lock".1 This unique fold features a central four-stranded β-sheet stabilized by three disulfide bonds. The disulfides are arranged such that they together with connecting backbone segments form an embedded ring that wraps around one of the β-strands, encapsulating a buried tyrosine side chain, and resulting in a tightly constrained molecular architecture.

Functionally, recifin A acts as a selective allosteric inhibitor of tyrosyl-DNA phosphodiesterase 1 (TDP1), a DNA repair enzyme implicated in the repair of topoisomerase I-induced DNA damage.1 By binding to a site distinct from the active site, recifin A modulates TDP1 activity, thereby enhancing the cytotoxic effects of topoisomerase inhibitors such as topotecan and irinotecan. This mechanism positions recifin A as a promising candidate for combination cancer therapies aimed at overcoming resistance to these chemotherapeutic agents.

The total chemical synthesis of recifin A was achieved using native chemical ligation chemistry and solid-phase peptide synthesis (SPPS), followed by oxidative folding under controlled redox conditions to promote correct disulfide bond formation.2 Despite the complex fold the native structure is remarkably favoured during folding, and synthetic recifin A and analogues were structurally validated via NMR spectroscopy. Pharmacological studies revealed initial structure activity relationships that suggest that the intrinsically disordered regulatory domain of TDP-1 wraps around recifin A, creating a large contact area.

The discovery of recifin A underscores the potential of marine-derived peptides in drug development and introduces the tyrosine-lock fold as a novel scaffold for therapeutic intervention. Its unique structural features and functional properties make it a valuable subject for further research in the design of targeted cancer therapies.

  1. 1. Krumpe LRH, Wilson BAP, Marchand C, Sunassee SN, Bermingham A, Wang W, Price E, Guszczynski T, Kelley JA, Gustafson KR, Pommier Y, Rosengren KJ, Schroeder CI, O'Keefe BR. Recifin A, Initial Example of the Tyr-Lock Peptide Structural Family, Is a Selective Allosteric Inhibitor of Tyrosyl-DNA Phosphodiesterase I. J Am Chem Soc. 2020 142(50):21178-21188.
  2. 2. Smallwood TB, Krumpe LRH, Payne CD, Klein VG, O'Keefe BR, Clark RJ, Schroeder CI, Rosengren KJ. Picking the tyrosine-lock: chemical synthesis of the tyrosyl-DNA phosphodiesterase I inhibitor recifin A and analogues. Chem Sci. 2024 15(33):13227-13233.