Second-generation probes for biosynthetic intermediate capture: towards a comprehensive profiling of polyketide...

Area of Science:

• Biochemistry
• Organic Chemistry
• Molecular Biology

Background:

• Polyketide natural products are synthesized by large, modular enzyme assembly lines.
• Understanding the kinetics of intermediate processing is crucial for elucidating these complex biosynthetic pathways.
• Existing methods for capturing biosynthetic intermediates in vivo have limitations.

Purpose of the Study:

• To develop and utilize novel second-generation probes for capturing polyketide intermediates in vivo.
• To characterize the nearly complete set of biosynthetic intermediates in a modular polyketide assembly line.
• To provide the first kinetic overview of intermediate processing during complex natural product formation.

Main Methods:

• Synthesis of malonyl carba(dethia) N-decanoyl cysteamine methyl esters and acetoxymethyl esters as chemical probes.
• In vivo application of these second-generation probes in a relevant biological system.
• Characterization of captured intermediates using advanced analytical techniques.

Main Results:

• Successful in vivo capture of a near-complete set of polyketide biosynthetic intermediates.
• Identification of key intermediates and their processing order within the modular assembly line.
• Establishment of a kinetic framework for intermediate processing in polyketide biosynthesis.

Conclusions:

• Second-generation probes are effective tools for in vivo capture and characterization of polyketide intermediates.
• The study provides unprecedented kinetic insights into complex natural product biosynthesis.
• This work lays the foundation for future metabolic engineering and drug discovery efforts targeting polyketide pathways.