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Identification of Pyridine Synthase Recognition Sequences Allows a Modular Solid-Phase Route to Thiopeptide Variants
Walter J Wever1, Jonathan W Bogart1, Albert A Bowers1
1Division of Chemical Biology and Medicinal Chemistry, University of North Carolina at Chapel Hill, Eshelman School of Pharmacy , Chapel Hill, North Carolina 27599, United States.
Journal of the American Chemical Society
|August 31, 2016
Summary
Researchers identified a minimal leader peptide sequence essential for TclM enzyme activity in thiopeptide biosynthesis. This discovery enables efficient, modular synthesis of novel thiopeptide variants using solid-phase peptide synthesis (SPPS).
Area of Science:
- Natural Product Biosynthesis
- Enzymology
- Synthetic Biology
Background:
- Thiopeptides are complex bioactive molecules produced by ribosomal peptide synthesis and post-translational modifications.
- Pyridine synthases catalyze key steps in thiopeptide formation, including macrocyclization and leader peptide cleavage.
- These enzymes offer potential as biocatalysts for generating novel thiopeptide structures.
Purpose of the Study:
- To investigate the leader peptide requirements for the TclM enzyme involved in thiocillin biosynthesis.
- To establish an efficient and modular method for synthesizing thiopeptide variants.
- To explore the substrate specificity of TclM and TbtD pyridine synthases.
Main Methods:
- Truncation analysis of the TclM leader peptide to define the minimum recognition sequence (RS).
- Solid-phase peptide synthesis (SPPS) to prepare linear thiopeptide cores.
- Ligation of synthesized RS to thiopeptide cores for enzymatic processing by TclM.
- Comparative analysis of TclM and TbtD activity on various thiopeptide substrates.
Main Results:
- A minimum recognition sequence (RS) necessary and sufficient for TclM activity was identified.
- A modular strategy combining SPPS and enzymatic processing allows for efficient synthesis of thiopeptide variants.
- Differences in promiscuity between TclM and TbtD were observed, enabling access to new structural diversity.
Conclusions:
- The defined leader peptide RS is crucial for TclM-mediated thiopeptide maturation.
- This work provides a versatile platform for the rational design and synthesis of novel thiopeptide analogs.
- The study highlights the potential of pyridine synthases as biocatalysts for expanding thiopeptide chemical space.
