Binding of 5′-GTP to the C-terminal FeS cluster of the radical
            <i>S</i>
            -adenosylmethionine enzyme MoaA provides insights into its mechanism

Hänzelmann, Petra; Schindelin, Hermann

doi:10.1073/pnas.0510711103

Cited by 148 publications

(252 citation statements)

References 36 publications

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“…It lacks both the CX 2 CX 5 CX 3 C motif and the second auxiliary cluster that are common in SPASM family members. With only these three protein ligands to its auxiliary cluster, MoaA uses the available coordination site to bind substrate (24,25). In anSMEcpe, the final cysteine of the CX 2 CX 5 CX 3 C motif is the fourth ligand to this cluster ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…In the case of MoaA, where substrate does directly ligate cluster, the ligation site (the N1 position of the guanine base) is on the opposite end of the GTP substrate than the hydrogen abstraction site (the 3′ hydrogen atom of the ribose; Fig. S3C, gray) (24,25,39). A similar mode of binding would be required for any SPASM members that use cluster ligation for substrate binding.…”

Section: Discussionmentioning

confidence: 96%

“…While the function of these auxiliary clusters is unknown, the 7-cysteine motif prompted speculation that members of the SPASM subfamily, including anSMEs, use an available ligation site on one of the [4Fe-4S] clusters for substrate binding (10). Direct binding of substrate to an auxiliary [4Fe-4S] cluster has been observed in the molybdenum cofactor biosynthetic enzyme MoaA, another AdoMet radical enzyme (24,25). Other possible functions specific to anSMEs include the second oxidation of substrate or substrate deprotonation (Fig.…”

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confidence: 99%

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X-ray structure of an AdoMet radical activase reveals an anaerobic solution for formylglycine posttranslational modification

Goldman¹,

Grove²,

Sites³

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

112

197

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Arylsulfatases require a maturating enzyme to perform a co-or posttranslational modification to form a catalytically essential formylglycine (FGly) residue. In organisms that live aerobically, molecular oxygen is used enzymatically to oxidize cysteine to FGly. Under anaerobic conditions, S-adenosylmethionine (AdoMet) radical chemistry is used. Here we present the structures of an anaerobic sulfatase maturating enzyme (anSME), both with and without peptidyl-substrates, at 1.6-1.8 Å resolution. We find that anSMEs differ from their aerobic counterparts in using backbone-based hydrogen-bonding patterns to interact with their peptidylsubstrates, leading to decreased sequence specificity. These anSME structures from Clostridium perfringens are also the first of an AdoMet radical enzyme that performs dehydrogenase chemistry. Together with accompanying mutagenesis data, a mechanistic proposal is put forth for how AdoMet radical chemistry is coopted to perform a dehydrogenation reaction. In the oxidation of cysteine or serine to FGly by anSME, we identify D277 and an auxiliary [4Fe-4S] cluster as the likely acceptor of the final proton and electron, respectively. D277 and both auxiliary clusters are housed in a cysteinerich C-terminal domain, termed SPASM domain, that contains homology to ∼1,400 other unique AdoMet radical enzymes proposed to use [4Fe-4S] clusters to ligate peptidyl-substrates for subsequent modification. In contrast to this proposal, we find that neither auxiliary cluster in anSME bind substrate, and both are fully ligated by cysteine residues. Instead, our structural data suggest that the placement of these auxiliary clusters creates a conduit for electrons to travel from the buried substrate to the protein surface.iron-sulfur cluster fold | radical SAM dehydrogenase

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 96%

mentioning

confidence: 99%

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X-ray structure of an AdoMet radical activase reveals an anaerobic solution for formylglycine posttranslational modification

Goldman¹,

Grove²,

Sites³

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

112

197

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“…Given that BtrN contains these sequence elements (CX 9-15 GX 4 CX n ), it was hypothesized to contain its Aux cluster in a twitch domain similar to MoaA (20). Interestingly, the Aux cluster in MoaA has an available iron ligation site that, unlike in anSMEcpe, is used to bind substrate (27,28). The structures of BtrN, presented below, address both the role and the structure of the Aux clusterbinding region and provide the structure of an AdoMet radical dehydrogenase that acts on a nonpeptide substrate.…”

mentioning

confidence: 99%

X-ray analysis of butirosin biosynthetic enzyme BtrN redefines structural motifs for AdoMet radical chemistry

Goldman

Grove²,

Booker

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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The 2-deoxy-scyllo-inosamine (DOIA) dehydrogenases are key enzymes in the biosynthesis of 2-deoxystreptamine-containing aminoglycoside antibiotics. In contrast to most DOIA dehydrogenases, which are NAD-dependent, the DOIA dehydrogenase from Bacillus circulans (BtrN) is an S-adenosyl-L-methionine (AdoMet) radical enzyme. To examine how BtrN employs AdoMet radical chemistry, we have determined its structure with AdoMet and substrate to 1.56 Å resolution. We find a previously undescribed modification to the core AdoMet radical fold: instead of the canonical (β/α) 6 architecture, BtrN displays a (β 5 /α 4 ) motif. We further find that an auxiliary [4Fe-4S] cluster in BtrN, thought to bind substrate, is instead implicated in substrate-radical oxidation. High structural homology in the auxiliary cluster binding region between BtrN, fellow AdoMet radical dehydrogenase anSME, and molybdenum cofactor biosynthetic enzyme MoaA provides support for the establishment of an AdoMet radical structural motif that is likely common to ∼6,400 uncharacterized AdoMet radical enzymes.radical SAM enzyme | iron-sulfur cluster fold | twitch domain D ue to mounting antibiotic resistance, the discovery and/or modification of antibiotic compounds to fight bacterial infection is a vital task of our time (1). Understanding antibiotic biosynthesis is an important first step in being able to engineer a new wave of therapeutic molecules. Aminoglycosides are a class of antibiotics that inhibit protein synthesis by interacting with the 30S subunit of the bacterial ribosome and have had considerable success in the clinical setting (2). Most FDA-approved aminoglycosides, including neomycin, gentamicin, and kanamycin, share a common glucose-6-phosphate-derived 2-deoxystreptamine (DOS) structural core (3) (blue in Scheme 1). Although the majority of aminoglycoside-producing bacteria use similar reaction sequences to biosynthesize this DOS core, including the penultimate two-electron oxidation of 2-deoxy-scyllo-inosamine (DOIA) to amino-dideoxy-scyllo-inosose (amino-DOI) (Scheme 1, A), identification of the enzymes responsible has not always been straightforward. For example, the btrE gene product in the butirosin B producing Bacillus circulans was proposed to be an NADdependent enzyme responsible for the generation of amino-DOI, as in other aminoglycoside pathways (4). This hypothesis proved incorrect upon further sequence and biochemical analysis (5, 6). Instead, Yokoyama et al. found that production of amino-DOI required another enzyme, BtrN, an S-adenosyl-L-methionine (AdoMet, SAM) radical dehydrogenase (5). Here, we report structures of catalytic and noncatalytic forms of BtrN, providing a structural basis for the unique reaction it performs.The AdoMet radical enzyme family catalyzes a diverse array of radical-based reactions, including sulfur insertions, complex chemical transformations and rearrangements, DNA and RNA modifications, and, in the case of BtrN, dehydrogenation (7). BtrN is one of two known members of the dehydrogenase subfamily of ...

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“…A common mechanism is shared for generation of a powerful reagent 5Ј-deoxyadenosyl radical, with the [4Fe-4S] cluster serving as an electron donor to reductively cleave the carbon-sulfur bond of AdoMet, thereby initiating highly diverse transformations. One fascinating aspect of radical AdoMet proteins is their potential to catalyze highly complex structural rearrangements, as exemplified by ThiC in thiamine biosynthesis (12) and MoaA in molybdenum cofactor biosynthesis (13,14).…”

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confidence: 99%

Characterization of NocL Involved in Thiopeptide Nocathiacin I Biosynthesis

Zhang

Chen

Lin

et al. 2011

Journal of Biological Chemistry

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The radical S-adenosylmethionine (AdoMet) enzyme superfamily is remarkable at catalyzing chemically diverse and complex reactions. We have previously shown that NosL, which is involved in forming the indole side ring of the thiopeptide nosiheptide, is a radical AdoMet enzyme that processes L-Trp to afford 3-methyl-2-indolic acid (MIA) via an unusual fragmentation-recombination mechanism. We now report the expansion of the MIA synthase family by characterization of NocL, which is involved in nocathiacin I biosynthesis.

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Binding of 5′-GTP to the C-terminal FeS cluster of the radical S -adenosylmethionine enzyme MoaA provides insights into its mechanism

Abstract: The first step in molybdenum cofactor biosynthesis, the conversion of 5-GTP to precursor Z, an oxygen-sensitive tetrahydropyranopterin is catalyzed by the S-adenosylmethionine ( iron-sulfur ͉ radicals ͉ molybdenum cofactor ͉ GTP cyclohydrolase

Cited by 148 publications

References 36 publications

X-ray structure of an AdoMet radical activase reveals an anaerobic solution for formylglycine posttranslational modification

X-ray structure of an AdoMet radical activase reveals an anaerobic solution for formylglycine posttranslational modification

X-ray analysis of butirosin biosynthetic enzyme BtrN redefines structural motifs for AdoMet radical chemistry

Characterization of NocL Involved in Thiopeptide Nocathiacin I Biosynthesis

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