In vivo characterization of NcsB3 to establish the complete biosynthesis of the naphthoic acid moiety of the neocarzinostatin chromophore

Abstract

Neocarzinostatin (NCS) is an enediyne antibiotic produced by Streptomyces carzinostaticus. The NCS chromophore consists of an enediyne core, a sugar moiety, and a naphthoic acid (NA) moiety. The latter plays a key role in binding the NCS chromophore to its apoprotein to protect and stabilize the bioactive NCS chromophore. In this study, we expressed three genes: ncsB (naphthoic acid synthase), ncsB3 (P450 hydroxylase), and ncsB1 (O-methyltransferase), in Streptomyces lividans TK24. The three genes were sufficient to produce 2-hydroxy-7-methoxy-5-methyl-1-naphthoic acid. Production was analyzed and confirmed by LC-MS and nuclear magnetic resonance. Here, we report the functional characterization of ncsB3 and thereby elucidate the complete biosynthetic pathway of NA moiety of the NCS chromophore.

Deciphering the cytochrome P450 genes in the microbiome of a chronically polluted soil with history of agricultural activities

“…In the Streptomyces CYPome, the CYP107 family constituted 12% even as the majority were CYP105 (24%) and a host of standalone CYPs involved in the biosynthesis off the enediyne antitumor antibiotic neocarzinostatin (ncsB3), azinomycin B (aziB1), as well as natamycin (Hang et al 2010;Ding et al 2010;Liu et al 2015). Even though CYP107 were not associated with Mycobacterium, also a member of the Actinobacteria, it is interesting Salam et al Bulletin of the National Research Centre (2022) 46:256 that four CYP107 subfamilies (CYP107A1, CYP107B1, CYP107BR1, and CYP107E1) associated with other Actinobacteria genera Saccharopolyspora, Micromonospora, and Pseudonocardia and found to be involved in catabolism of xenobiotics, biosynthesis of erythromycin and mycinamicin, and hydroxylation of vitamin D 3 to its active form were annotated (McLean et al 2015;Mnguni et al 2020).…”

Biocatalytic role of cytochrome P450s to produce antibiotics: A review

“…NCS, an enediyne antibiotic, produced by Streptomyces carzinostaticus , ncsB3 (P450) functions as 7‐hydroxylase, which is involved in the biosynthesis of napthoic acid (NA) moiety of NCS chromophore (Hang et al, 2010) (Figure 7n). In the calicheamicin 68 BGC, CalO2 is involved in modifying bacterial iterative type I polyketide products (Figure 7o) (McCoy et al, 2009).…”

Aromatic Polyketides Produced by Bacterial Iterative Type I Polyketide Synthases

“…The modified NPA moiety is then incorporated into a nonribosomal peptide synthase (NRPS) system by AziA, which consists of an adenylation domain and an ACP domain (Figure B) . In neocarzinostatin biosynthesis, the C-7 regiospecific hydroxylation on 2-hydroxyl-5-methyl-1-NPA and its subsequent methylation on the nascent C-7 hydroxyl group are catalyzed by the P450 hydroxylase NcsB3 and O -methyltransferase NcsB1, respectively. − The resulting 2-hydroxyl-7-methoxy-5-methyl-NPA is then activated by NscB2 as a CoA thioester and transferred to the enediyne core by the putative AT NcsB4 (Figure C) …”