Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2016 Oct 7:7:13083.
doi: 10.1038/ncomms13083.

Non-enzymatic pyridine ring formation in the biosynthesis of the rubrolone tropolone alkaloids

Yijun Yan  1 , Jing Yang  1 , Zhiyin Yu  1 , Mingming Yu  1 , Ya-Tuan Ma  1 , Li Wang  1 , Can Su  1 , Jianying Luo  1 , Geoffrey P Horsman  2 , Sheng-Xiong Huang  1
Affiliations

Non-enzymatic pyridine ring formation in the biosynthesis of the rubrolone tropolone alkaloids

Yijun Yan et al. Nat Commun. .

Abstract

The pyridine ring is a potent pharmacophore in alkaloid natural products. Nonetheless, its biosynthetic pathways are poorly understood. Rubrolones A and B are tropolone alkaloid natural products possessing a unique tetra-substituted pyridine moiety. Here, we report the gene cluster and propose a biosynthetic pathway for rubrolones, identifying a key intermediate that accumulates upon inactivation of sugar biosynthetic genes. Critically, this intermediate was converted to the aglycones of rubrolones by non-enzymatic condensation and cyclization with either ammonia or anthranilic acid to generate the respective pyridine rings. We propose that this non-enzymatic reaction occurs via hydrolysis of the key intermediate, which possesses a 1,5-dione moiety as an amine acceptor capable of cyclization. This study suggests that 1,5-dione moieties may represent a general strategy for pyridine ring biosynthesis, and more broadly highlights the utility of non-enzymatic diversification for exploring and expanding natural product chemical space.

PubMed Disclaimer

Figures

Figure 1
Figure 1. Chemical structures of rubrolones A (1) and B (2) and selected pyridine alkaloids.
Pyridine rings are highlighted in blue.
Figure 2
Figure 2. HPLC profiles of the fermentation extracts of engineered heterologous expression strains.
I: S. albus 9B10; II: S. albus pJTU2554; III: S. albus 9B10-ΔS1; IV: S. albus 9B10-ΔE9; V: S. albus 9B10-ΔB; VI: S. albus 9B10-ΔC; VII: S. albus 9B10-ΔE9 feeding with 3; VIII: S. albus 9B10-ΔE9 feeding with 4; IX: S. albus 9B10 feeding with anthranilic acid; and X: S. albus 9B10-ΔS1 feeding with anthranilic acid.
Figure 3
Figure 3. Biosynthetic gene cluster and proposed biosynthetic pathway of rubrolones.
(a) Organization of the rub biosynthetic gene cluster, with functional assignment of genes including PKS (black), oxygenases (orange), deoxysugar synthases (pink), regulation (green), unknown (cyan) and genes outside the cluster (white). (b) Proposed biosynthetic pathway for PKS and post-PKS modifications, with dashed arrows indicating the pathway generating the shunt metabolites R1128A (5), 6 and 7. (c) Proposed biosynthetic pathway for the deoxysugar dTDP-2-keto-D-fucose.
Figure 4
Figure 4. Chemical structures of rubrolone analogues and related metabolites produced by mutants.
Compounds 3 and 4 were isolated from mutant S. albus 9B10-ΔS1; 5–7 were isolated from mutant S. albus 9B10-ΔB; 8 was obtained from S. albus 9B10-ΔS1 feeding with anthranilic acid; and compounds 9 and 10 were obtained from S. albus 9B10 feeding with 2-amino-5-fluorobenzoic acid and 2-amino-5-chlorobenzoic acid, respectively.
Figure 5
Figure 5. Different possible biosynthetic relationships between 1 and 2.
(i) 2 being generated by the oxidative N–C coupling of benzoic acid and 1, (ii) 1 generated by the reductive N–C cleavage of 2 and (iii) both 1 and 2 arising from divergent amination of 4.
Figure 6
Figure 6. HPLC analysis of in vitro chemical conversions |:
4 was incubated with buffer only; II: 4 was incubated with ammonium acetate; and III: 4 was incubated with anthranilic acid (black diamond).
See this image and copyright information in PMC

References

    1. Palleroni N. J. et al.. Production of a novel red pigment, rubrolone, by Streptomyces echinoruber sp. nov. I. Taxonomy, fermentation and partial purification. J. Antibiot. 31, 1218–1225 (1978). - PubMed
    1. Schüep W., Blount J. F., Williams T. H. & Stempel A. Production of a novel red pigment, rubrolone, by Streptomyces echinoruber sp. nov. II. Chemistry and structure elucidation. J. Antibiot. 31, 1226–1232 (1978). - PubMed
    1. Kelly T. R., Echavairen A., Whiting A., Weibel F. R. & Miki Y. Synthesis of the chromophore of rubrolone. Tetrahedron Lett. 27, 6049–6050 (1986).
    1. Boger D. L., Ichikawa S. & Jiang H. Total synthesis of the rubrolone aglycon. J. Am. Chem. Soc. 122, 12169–12173 (2000).
    1. Yan Y. et al.. Tropolone ring construction in the biosynthesis of rubrolone B, a cationic tropolone alkaloid from endophytic Streptomyces. Org. Lett. 18, 1254–1257 (2016). - PubMed

Publication types