Engineering the stambomycin modular polyketide synthase yields 37-membered mini-stambomycins

Area of Science:

• Biochemistry
• Synthetic Biology
• Metabolic Engineering

Background:

• Type I polyketide synthases (PKSs) possess a modular structure, theoretically enabling rational engineering for novel analogs.
• Despite this potential, engineering PKSs has historically been inefficient, limiting the production of desired compounds.

Purpose of the Study:

• To reprogram the stambomycin PKS by deleting seven internal modules to create truncated polyketide metabolites.
• To investigate the efficiency of engineered PKS assembly lines and understand limitations in production.

Main Methods:

• Utilized state-of-the-art approaches to engineer the stambomycin PKS.
• Performed module deletion experiments within the PKS gene cluster.
• Analyzed the resulting metabolites and characterized production yields.

Main Results:

• Successfully produced 37-membered mini-stambomycin metabolites, a significant reduction in chain length.
• Observed substantial production of shunt metabolites alongside the target compounds.
• Identified an unprecedented off-loading mechanism involving the C-terminal thioesterase domain for stalled intermediates.
• Mini-stambomycin yields were lower than wild-type, indicating issues with module tolerance to non-native substrates.

Conclusions:

• Engineering modular PKSs presents challenges, including module tolerance to altered substrates.
• The study identified key factors influencing the productivity of engineered PKS assembly lines.
• Further research is necessary to enhance production titers for engineered polyketide compounds.