An HD domain phosphohydrolase active site tailored for oxetanocin-A biosynthesis

Bridwell‐Rabb, Jennifer; Kang, Gyunghoon; Zhong, Aoshu; Liu, Hung Wen; Drennan, Catherine L.

doi:10.1073/pnas.1613610113

Cited by 27 publications

(48 citation statements)

References 56 publications

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“…This work confirms that oxsA and oxsB are the essential OXT-A biosynthetic genes; we show here that OxsB uses AdoMet radical chemistry to catalyze ring contraction of a deoxyadenosine phosphate and recently showed that OxsA catalyzes hydrolysis of the resulting phosphorylated OXT-A compounds 22 . Although we do not know the roles of the remaining two genes located within the OXT-A gene cluster, our genetic experiments confirm that they are not required for OXT-A production.…”

Section: Discussionsupporting

confidence: 88%

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A B12-dependent radical SAM enzyme involved in oxetanocin A biosynthesis

Bridwell‐Rabb

Zhong

Sun

et al. 2017

Nature

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209

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Summary Oxetanocin-A (OXT-A, 1) is a potent antitumor, antiviral, and antibacterial compound. Biosynthesis of OXT-A has been linked to a plasmid-borne, Bacillus megaterium gene cluster that contains four genes, oxsA, oxsB, oxrA, and oxrB. Here, we show that the oxsA and oxsB genes are both required for the production of OXT-A. Biochemical analysis of the encoded proteins, a cobalamin (Cbl)-dependent S-adenosylmethionine (AdoMet) radical enzyme, OxsB, and an HD-domain phosphohydrolase, OxsA, revealed that OXT-A is derived from 2′-deoxyadenosine phosphate in an OxsB-catalyzed ring contraction reaction initiated by H-atom abstraction from C2′. Hence, OxsB represents the first biochemically characterized non-methylating Cbl-dependent AdoMet radical enzyme. X-ray analysis of OxsB reveals the fold of a Cbl-dependent AdoMet radical enzyme for which there are an estimated 7000 members. Overall, this work provides a framework for understanding the interplay of AdoMet and Cbl cofactors and expands the catalytic repertoire of Cbl-dependent AdoMet radical enzymes.

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Section: Discussionsupporting

confidence: 88%

“…OxsA was recently shown to catalyze the sequential hydrolysis of mono-, di-, and tri-phosphorylated OXT-A compounds into OXT-A ( 3→1 , Fig. 1d) 22 , supporting the ability of the OxsB/OxsA pair to turnover dAMP, dADP and dATP.…”

Section: Oxsa B Are Oxt-a Biosynthetic Genesmentioning

confidence: 63%

A B12-dependent radical SAM enzyme involved in oxetanocin A biosynthesis

Bridwell‐Rabb

Zhong

Sun

et al. 2017

Nature

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209

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“…Recombinant OxsA was shown to catalyze hydrolysis of phosphorylated OXT-A compounds. 99 Recombinant OxsB harbored a single [4Fe–4S] cluster and hydroxocobalamin, consistent with its annotation as a cobalamin-dependent radical SAM enzyme. 81 Initially, no catalytic activity was observed in OxsB assays using various nucleosides and nucleotides.…”

Section: Radical Sam Enzymes With N-terminal Cofactor Binding Domainssupporting

confidence: 66%

“…20a). Since OxsA catalyzes the hydrolysis of dAMP, dADP and dATP, 99 the function of OxsB was proposed to be conversion of dAMP into dehydro-OXT-A phosphate (Fig. 20a).…”

Section: Radical Sam Enzymes With N-terminal Cofactor Binding Domainsmentioning

confidence: 99%

C–C bond forming radical SAM enzymes involved in the construction of carbon skeletons of cofactors and natural products

Yokoyama

Lilla

2018

Nat. Prod. Rep.

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C–C bond formations are frequently the key steps in cofactor and natural product biosynthesis. Historically, C–C bond formations were thought to proceed by two electron mechanisms, represented by Claisen condensation in fatty acids and polyketide biosynthesis. These types of mechanisms require activated substrates to create a nucleophile and an electrophile. More recently, increasing number of C–C bond formations catalyzed by radical SAM enzymes are being identified. These free radical mediated reactions can proceed between almost any sp3 and sp2 carbon centers, allowing introduction of C–C bonds at unconventional positions in metabolites. Therefore, free radical mediated C–C bond formations are frequently found in the construction of structurally unique and complex metabolites. This review discusses our current understanding of the functions and mechanisms of C–C bond forming radical SAM enzymes and highlights their important roles in the biosynthesis of structurally complex, naturally occurring organic molecules. Mechanistic consideration of C–C bond formation by radical SAM enzymes identifies the significance of three key mechanistic factors: radical initiation, acceptor substrate activation and radical quenching. Understanding the functions and mechanisms of these characteristic enzymes will be important not only in promoting our understanding of radical SAM enzymes, but also for understanding natural product and cofactor biosynthesis.

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“…dAMP, dADP and dATP were identified as direct oxetanocin precursors [24]. The product of oxsB catalyses the contraction of the deoxyribose ring, while the product of oxsA is responsible for the removal of one or multiple phosphates from a phosphorylated 2'-deoxyadenosine derivative [24,25]. Through the simultaneous actions of OxsA and OxsB, the phosphorylated 2'-deoxyadenosine is converted to the oxetanocin A precursor, its aldehyde form, which must be reduced to complete biosynthesis [24].…”

Section: Proposed Biosynthetic Pathway Of Albucidinmentioning

confidence: 99%

Identification and Heterologous Expression of the Albucidin Gene Cluster from the Marine Strain Streptomyces Albus Subsp. Chlorinus NRRL B-24108

et al. 2020

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Herbicides with new modes of action and safer toxicological and environmental profiles are needed to manage the evolution of weeds that are resistant to commercial herbicides. The unparalleled structural diversity of natural products makes these compounds a promising source for new herbicides. In 2009, a novel nucleoside phytotoxin, albucidin, with broad activity against grass and broadleaf weeds was isolated from a strain of Streptomyces albus subsp. chlorinus NRRL B-24108. Here, we report the identification and heterologous expression of the previously uncharacterized albucidin gene cluster. Through a series of gene inactivation experiments, a minimal set of albucidin biosynthetic genes was determined. Based on gene annotation and sequence homology, a model for albucidin biosynthesis was suggested. The presented results enable the construction of producer strains for a sustainable supply of albucidin for biological activity studies.

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An HD domain phosphohydrolase active site tailored for oxetanocin-A biosynthesis

Cited by 27 publications

References 56 publications

A B12-dependent radical SAM enzyme involved in oxetanocin A biosynthesis

A B12-dependent radical SAM enzyme involved in oxetanocin A biosynthesis

C–C bond forming radical SAM enzymes involved in the construction of carbon skeletons of cofactors and natural products

Identification and Heterologous Expression of the Albucidin Gene Cluster from the Marine Strain Streptomyces Albus Subsp. Chlorinus NRRL B-24108

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