Biosynthetic Pathway Engineering for Natural Product Development

Biosynthetic pathway engineering harnesses advances in genome mining, synthetic biology and molecular genetics to reprogramme microorganisms for the production and diversification of natural products. Central to this endeavour is the identification and manipulation of biosynthetic gene clusters (BGCs), which encode the enzymatic machinery for assembling complex molecules such as polyketides, non-ribosomal peptides and alkaloids. By refactoring regulatory elements, swapping enzyme domains and introducing heterologous clusters into optimised host strains, researchers can activate silent pathways, increase yields and generate novel analogues with improved pharmacological properties. This integrated approach has spurred the discovery of new antibiotics, anticancer agents and agrichemicals, addressing pressing global challenges such as drug resistance and sustainable crop protection. The field draws on high-throughput DNA assembly methods, precision genome editing tools and bespoke microbial chassis to accelerate the translation of genomic information into valuable bioactive compounds.

Research from Nature Portfolio

Recent studies have introduced a scalable direct-cloning strategy combining Cas12a-mediated DNA cleavage, a tailored exo + fill-in assembly and in vivo Cre-lox circularisation. This method achieves near-100% efficiency in capturing 10–113 kb BGCs from diverse Actinomycetes and Bacilli, leading to the discovery of previously uncharacterised antimicrobial compounds with unique cyclic scaffolds. Earlier work employed RNA-guided Cas9 nuclease to excise large genomic segments of up to 100 kb at designated loci, followed by Gibson assembly into expression vectors in a single step, thereby providing a rapid tool for targeted cluster cloning. Yeast-mediated transformation-associated recombination has also been adapted to integrate intact BGCs into Bacillus subtilis, enabling heterologous production of lipopeptide and pro-drug intermediates and revealing novel maturation processes crucial for bioactivity.

Biosynthetic Pathway Engineering for Natural Product Development publication trend

The graph below shows the total number of articles in biosynthetic pathway engineering for natural product development across all publications each year (not limited to Nature Index journals).

Technical terms

Biosynthetic gene cluster (BGC): A contiguous set of genes encoding enzymes and regulators responsible for the stepwise assembly of a specialised metabolite.

Heterologous expression: The introduction and functional expression of a gene or gene cluster in a non-native host organism to enable production of its encoded compound.

Transformation-associated recombination (TAR): A yeast-based technique that exploits homologous recombination to capture and assemble large DNA fragments in vivo.

Cas12a-assisted precise targeted cloning (CAPTURE): A method combining Cas12a cleavage, polymerase-based DNA assembly and Cre-lox recombination to efficiently clone large gene clusters.

Gibson assembly: A one-pot enzymatic method for joining multiple DNA fragments that have overlapping ends.

Cre-lox recombination: A site-specific recombination system used to circularise or excise DNA segments via Cre recombinase acting on lox sites.

References

1. Cas12a-assisted precise targeted cloning using in vivo Cre-lox recombination. Nature Communications (2021).
2. Cas9-Assisted Targeting of CHromosome segments CATCH enables one-step targeted cloning of large gene clusters. Nature Communications (2015).
3. Directed natural product biosynthesis gene cluster capture and expression in the model bacterium Bacillus subtilis. Scientific Reports (2015).
4. Parallelized gene cluster editing illuminates mechanisms of epoxyketone proteasome inhibitor biosynthesis. Nucleic Acids Research (2023).
5. Recent advances in the direct cloning of large natural product biosynthetic gene clusters. Engineering Microbiology (2023).
6. A CRISPR-Cas9-Mediated Large-Fragment Assembly Method for Cloning Genomes and Biosynthetic Gene Cluster. Microorganisms (2024).