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Scientists developed a microfluidic platform to detect gramicidin S production by nonribosomal peptide synthetases (NRPSs). This tool accelerates antibiotic discovery by enabling faster screening of NRPS enzyme activity.

Keywords:
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Area of Science:

  • Biochemistry
  • Synthetic Biology
  • Microfluidics

Background:

  • Nonribosomal peptide synthetases (NRPSs) are crucial for producing valuable natural products like antibiotics.
  • Developing new antibiotics necessitates efficient methods for redesigning NRPS pathways.
  • Current screening platforms lack the robustness and speed required for NRPS engineering.

Purpose of the Study:

  • To establish a microfluidic platform for detecting nonribosomal peptide production.
  • To enable rapid sorting and screening of NRPS mutants.
  • To facilitate the engineering of NRPS pathways for novel antibiotic discovery.

Main Methods:

  • A microfluidic platform was developed utilizing calcein-filled sensor liposomes.
  • Liposome fluorescence increases upon permeabilization, indicating gramicidin S production.
  • The platform was used to sort a library of NRPS mutants.

Main Results:

  • The platform reliably detected gramicidin S production.
  • Enrichment of gramicidin S producers by 14.5-fold was achieved.
  • A 250-fold decrease in internal stop codons and correlation with enzyme activity were observed.

Conclusions:

  • The microfluidic platform provides a robust and rapid method for screening NRPS activity.
  • This technology can accelerate the discovery of new antibiotics and facilitate NRPS engineering.
  • Future applications include advanced structure-activity relationship studies and pathway engineering.