Multiplexed mobilization and expression of biosynthetic gene clusters

Area of Science:

• Microbiology
• Synthetic Biology
• Natural Product Discovery

Background:

• Bacterial genomes harbor numerous biosynthetic gene clusters (BGCs) with potential for novel natural product discovery.
• Heterologous expression of these BGCs is key to accessing new bioactive molecules but is often hampered by difficulties in large-scale manipulation.
• Current methods for BGC manipulation are cumbersome, limiting the systematic exploration of microbial natural product potential.

Purpose of the Study:

• To develop a method for parallelized identification, mobilization, and heterologous expression of bacterial BGCs.
• To enable the large-scale interrogation of cryptic BGCs within bacterial strain collections.
• To accelerate the discovery of novel therapeutic natural products.

Main Methods:

• Genomic DNA from a bacterial strain collection was cloned into a large-insert library.
• CONKAT-seq (co-occurrence network analysis of targeted sequences) was employed to efficiently identify clones containing intact BGCs.
• Parallelized heterologous expression of identified BGCs was performed.

Main Results:

• A scalable method for capturing and identifying numerous BGCs was established.
• Several novel natural products were discovered through heterologous expression.
• An antibiotic effective against multi-drug resistant Staphylococcus aureus was identified.

Conclusions:

• The developed method significantly enhances the efficiency of discovering natural products from bacterial BGCs.
• This approach facilitates the systematic exploration of cryptic BGCs, unlocking vast chemical diversity.
• Economies of scale in BGC manipulation can accelerate the discovery of urgently needed therapeutics.