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Repositioning the Leader Peptide in Graspetide Biosynthesis.

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|March 6, 2026
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Summary
This summary is machine-generated.

Synthetic biology enables new peptide products by repositioning leader sequences in ribosomally synthesized and post-translationally modified peptides (RiPPs). This strategy generates novel, mechanically interlocked RiPPs with unique modifications.

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Area of Science:

  • Synthetic biology
  • Biochemistry
  • Natural product synthesis

Background:

  • Ribosomally synthesized and post-translationally modified peptides (RiPPs) exhibit diverse structures, including mechanically interlocked architectures.
  • Tailoring enzymes modify precursor peptides guided by N-terminal leader sequences to produce mature RiPPs.

Purpose of the Study:

  • To investigate the effect of repositioning the leader sequence of fuscimiditide precursor peptide from N-terminal to C-terminal.
  • To explore the generation of novel RiPP structures using leader peptide engineering.

Main Methods:

  • Enzymatic modification and cyclization of precursor peptides with native and C-terminal leader sequences.
  • In cellulo and in vitro assays using graspetide synthetase ThfB.
  • Generation of chimeric RiPPs using engineered precursor peptides.

Main Results:

  • Substrate-selective post-translational modification by ThfB was retained with the C-terminal leader sequence.
  • A modest 2-fold reduction in modification rate was observed upon leader sequence repositioning.
  • Chimeric RiPPs were successfully generated, incorporating modifications from both lasso peptide and graspetide families.

Conclusions:

  • Repositioning RiPP leader sequences is a viable strategy for generating novel peptide products.
  • This approach expands the structural diversity of RiPPs, enabling access to new mechanically interlocked molecules.
  • Leader peptide engineering offers a powerful tool for creating new-to-nature peptide architectures.