Functional characterization of a novel violacein biosynthesis operon from Janthinobacterium sp. B9-8.

Violacein is a secondary metabolite mainly produced by Gram-negative bacteria that is formed from tryptophan by five enzymes encoded by a single operon. It is a broad-spectrum antibacterial pigment with various important biological activities such as anti-tumor, antiviral, and antioxidative effects. The newly discovered violacein operon vioABCDE was identified in the genome of the extremophile Janthinobacterium sp. B9-8. The key enzyme-encoding genes were cloned to construct the multigene coexpression plasmids pET-vioAB and pRSF-vioCDE. The violacein biosynthesis pathway was heterologously introduced into engineered Escherichia coli VioABCDE and VioABCDE-SD. The factors affecting violacein production, including temperature, pH, inoculum size, carbon and nitrogen source, precursor, and inducers were investigated. The violacein titer of VioABCDE-SD reached 107 mg/L in a two-stage fermentation process, representing a 454.4% increase over the original strain. The violacein operon from B9-8 provides a new microbial gene source for the analysis of the violacein synthesis mechanism, and the constructed engineering E. coli strains lay a foundation for the efficient and rapid synthesis of other natural products.Key points• The newly discovered violacein operon vioABCDE was identified in the genome of the extremophile Janthinobacterium sp. B9-8.• The violacein synthesis pathway was reconstructed in E. coli using two compatible plasmids.• A two-stage fermentation process was optimized for improved violacein accumulation.

Recent Advances on Key Enzymes of Microbial Origin in the Lycopene Biosynthesis Pathway.

Lycopene is a common carotenoid found mainly in ripe red fruits and vegetables that is widely used in the food industry due to its characteristic color and health benefits. Microbial synthesis of lycopene is gradually replacing the traditional methods of plant extraction and chemical synthesis as a more economical and productive manufacturing strategy. The biosynthesis of lycopene is a typical multienzyme cascade reaction, and it is important to understand the characteristics of each key enzyme involved and how they are regulated. In this paper, the catalytic characteristics of the key enzymes involved in the lycopene biosynthesis pathway and related studies are first discussed in detail. Then, the strategies applied to the key enzymes of lycopene synthesis, including fusion proteins, enzyme screening, combinatorial engineering, CRISPR/Cas9-based gene editing, DNA assembly, and scaffolding technologies are purposefully illustrated and compared in terms of both traditional and emerging multienzyme regulatory strategies. Finally, future developments and regulatory options for multienzyme synthesis of lycopene and similar secondary metabolites are also discussed.

Draft genome sequence of a new carotenoid-producing strain Brevibacterium sp. XU54, isolated from radioactive soil in Xinjiang, China.

Some species of the genus Brevibacterium are orange bacteria involved in cheese ripening, synthesis of odoriferous compounds, and carotenoids with aromatic end groups. Here, we report the genome sequence of Brevibacterium sp. XU54, isolated from radioactive soil in Xinjiang, China. The genome of XU54 consists of 4,899,099 base pairs with a GC content of 62.2%. The genome sequence was annotated with 4453 genes, encoding 4260 proteins, 13 rRNAs, and 49 tRNAs. 16S rRNA BLAST and comparative genomic analysis both indicated that XU54 may be a new species of Brevibacterium. In addition, compared to the type strains, some enzymes related to sulfur metabolism showed a low similarity of 66.85, 79.53 and 14.61%, respectively. The carotenoids biosynthesis gene cluster was identified and analyzed according to the genomic data, which revealed relatively low identity (5-85%) with existing strains. The optimum conditions for its growth and carotenoid production were then discussed. The whole-genome sequence of Brevibacterium sp. XU54 will be beneficial for utilizing these newly identified genes in carotenoid biosynthesis and regulation of sulfur metabolism pathway to promote the production of novel carotenoids and other structurally diverse compounds through combinatorial biosynthesis, which facilitates cheese ripening and coloration. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03366-1.