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Recognition sequences and substrate evolution in cyanobactin biosynthesis.

Debosmita Sardar1, Elizabeth Pierce, John A McIntosh

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Summary
This summary is machine-generated.

Ribosomally synthesized and posttranslationally modified peptide (RiPP) natural products show remarkable enzyme promiscuity due to short recognition sequences (RSs) on substrates. These portable RSs can be swapped to control modifications, enabling synthetic biology applications.

Keywords:
cyanobactinsposttranslational modificationsrecognition sequencesribosomally synthesized and posttranslationally modified peptides (RiPP)substrate evolution

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Area of Science:

  • Biochemistry
  • Synthetic Biology
  • Molecular Biology

Background:

  • Ribosomally synthesized and posttranslationally modified peptides (RiPPs) are valuable natural products with diverse bioactivities.
  • RiPP pathways like cyanobactins exhibit substrate evolution, where enzymes are constant but substrates vary, suggesting underlying mechanisms of enzyme promiscuity.

Purpose of the Study:

  • To elucidate the mechanism behind the substrate promiscuity observed in RiPP cyanobactin pathways.
  • To identify and characterize the recognition elements responsible for directing enzymatic modifications in RiPP biosynthesis.

Main Methods:

  • Analysis of enzyme-substrate interactions across five cyanobactin gene clusters.
  • Engineering of synthetic substrates to probe recognition sequences (RSs).
  • In vivo experiments in E. coli to test the exchangeability of RSs and production of modified peptides.

Main Results:

  • Defined short, discrete recognition sequences (RSs) within substrates that direct enzymatic modifications.
  • Demonstrated that these RSs are portable and can be interchanged to alter functional group additions.
  • Identified the RS for heterocyclization modification, in addition to previously known N- and C-terminal proteolysis RSs.
  • Successfully produced modified natural products in E. coli by swapping substrate elements.

Conclusions:

  • The promiscuity of RiPP cyanobactin pathways is governed by portable recognition sequences (RSs) on the substrates.
  • These RSs can be manipulated to control post-translational modifications, offering a powerful tool for synthetic biology.
  • The exchangeability of RSs facilitates the tailoring of peptide products in vitro and in vivo, expanding possibilities for natural product synthesis.