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. 2021 Jan 15;11(6):3510-3515.
doi: 10.1039/d0ra10878b. eCollection 2021 Jan 14.

Characterization of NucPNP and NucV involved in the early steps of nucleocidin biosynthesis in Streptomyces calvus

Utumporn Ngivprom  1 , Surayut Kluaiphanngam  1 , Wenjuan Ji  2 , Siriwalee Siriwibool  1 , Anyanee Kamkaew  1 , James R Ketudat Cairns  1   3 , Qi Zhang  2 , Rung-Yi Lai  1   3
Affiliations

Characterization of NucPNP and NucV involved in the early steps of nucleocidin biosynthesis in Streptomyces calvus

Utumporn Ngivprom et al. RSC Adv. .

Abstract

Nucleocidin 1 produced by Streptomyces calvus is one of five characterized natural products containing fluorine. It was discovered in 1956, but its biosynthesis is not yet completely resolved. Recently, the biosynthetic gene cluster of 1 was identified. The nucPNP gene, which was initially annotated as orf206 and encodes a putative purine nucleoside phosphorylase, is essential for nucleocidin production. In this study, we performed in vitro assays and showed NucPNP produced adenine 3 from methylthioadenosine (MTA) 2 and adenosine 4. We also showed the downstream enzyme, NucV annotated as adenine phosphoribosyltransferase (APRT), catalyzes AMP formation from adenine 3 and 5-phospho-α-d-ribose-1-diphosphate (PRPP) 5. However, the catalytic efficiency of NucV was much slower than its homolog ScAPRT involved in the biosynthesis of canonical purine nucleoside in the same strain. These results provide new insights in nucleocidin biosynthesis and could guide future research on organofluorine formation.

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Conflict of interest statement

There are no conflicts to declare.

Figures

Fig. 1
Fig. 1. The five characterized natural products containing fluorine.
Scheme 1
Scheme 1. Proposed biosynthetic pathway of nucleocidin 1. The reactions (solid arrow) are studied in this article.
Fig. 2
Fig. 2. The HPLC analysis of MBP-NucPNP assays to convert MTA 2 to adenine 3.
Fig. 3
Fig. 3. The HPLC analysis of MBP-NucPNP assays to convert adenosine 4 to adenine 3.
Fig. 4
Fig. 4. MBP-NucPNP catalyzed the phosphorylation of adenosine 4 to generate adenine 3 faster than MTA 2.
Fig. 5
Fig. 5. NucV catalyzed the reaction slower than ScAPRT under the same condition. The relative rate of AMP 6 formation was analyzed by HPLC (A). PPi formation and PRPP 5 consumption by ScAPRT (B) and NucV (C) were monitored by 31P-NMR.
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