Bacterial Enzymes Catalyzing the Synthesis of 1,8-Dihydroxynaphthalene, a Key Precursor of Dihydroxynaphthalene Melanin, from Sorangium cellulosum

• Published in: Applied and environmental microbiology Vol. 84; no. 9; p. e00258-18
• Main Authors: Sone, Yusuke, Nakamura, Shuto, Sasaki, Makoto, Hasebe, Fumihito, Kim, Seung-Young, Funa, Nobutaka
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 01.05.2018
• Subjects: Aldo-keto reductase; Bacteria; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Biocatalysis; Biosynthesis; Chemical synthesis; Chromatography; Coenzyme A; Dehydration; Enzymes; Enzymology and Protein Engineering; Fungi; High performance liquid chromatography; Incubation; Liquid chromatography; Melanin; Melanins - biosynthesis; Myxococcales - enzymology; NADP; Naphthol; Naphthols - metabolism; Operon - genetics; Polyketide synthase; Proteins; Reductases; Reduction; Scytalone dehydratase; Secretion; Sorangium cellulosum; Spotlight; Stereoselectivity; Vermelone; 1,8-dihydroxynaphthalene; melanin; biosynthesis
• ISSN: 0099-2240; 1098-5336
• DOI: 10.1128/aem.00258-18

1,8-Dihydroxynaphthalene (1,8-DHN) is a key intermediate in the biosynthesis of DHN melanin, which is specific to fungi. In this study, we characterized the enzymatic properties of the gene products of an operon consisting of , , and from the Gram-negative bacterium Heterologous expression of , , and in caused secretion of a dark-brown pigment into the broth. High-performance liquid chromatography (HPLC) analysis of the broth revealed that the recombinant strain produced 1,8-DHN, indicating that the operon encoded a novel enzymatic system for the synthesis of 1,8-DHN. Simultaneous incubation of the recombinant SoceCHS1, BdsA, and BdsB with malonyl-coenzyme A (malonyl-CoA) and NADPH resulted in the synthesis of 1,8-DHN. SoceCHS1, a type III polyketide synthase (PKS), catalyzed the synthesis of 1,3,6,8-tetrahydroxynaphthalene (T HN) T HN was in turn converted to 1,8-DHN by successive steps of reduction and dehydration, which were catalyzed by BdsA and BdsB. BdsA, which is a member of the aldo-keto reductase (AKR) superfamily, catalyzed the reduction of T HN and 1,3,8-tetrahydroxynaphthalene (T HN) to scytalone and vermelone, respectively. The stereoselectivity of T HN reduction by BdsA occurred on the -face to give ( )-scytalone with more than 99% optical purity. BdsB, a SnoaL2-like protein, catalyzed the dehydration of scytalone and vermelone to T HN and 1,8-DHN, respectively. The fungal pathway for the synthesis of 1,8-DHN is composed of a type I PKS, naphthol reductases of the short-chain dehydrogenase/reductase (SDR) superfamily, and scytalone dehydratase (SD). These findings demonstrated 1,8-DHN synthesis by novel enzymes of bacterial origin. Although the DHN biosynthetic pathway was thought to be specific to fungi, we discovered novel DHN synthesis enzymes of bacterial origin. The biosynthesis of bacterial DHN utilized a type III PKS for polyketide synthesis, an AKR superfamily for reduction, and a SnoaL2-like NTF2 superfamily for dehydration, whereas the biosynthesis of fungal DHN utilized a type I PKS, SDR superfamily enzyme, and SD-like NTF2 superfamily. Surprisingly, the enzyme systems comprising the pathway were significantly different from each other, suggesting independent, parallel evolution leading to the same biosynthesis. DHN melanin plays roles in host invasion and adaptation to stress in pathogenic fungi and is therefore important to study. However, it is unclear whether DHN biosynthesis occurs in bacteria. Importantly, we did find that bacterial DHN biosynthetic enzymes were conserved among pathogenic bacteria.