Mechanistic investigation of B12-independent glycerol dehydratase and its activating enzyme GD-AE

Li, Yaoyang; Yao, Yadi; Yu, Lu; Tian, Changlin; Dong, Min

doi:10.1039/d1cc06991h

Cited by 5 publications

(3 citation statements)

References 32 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Interestingly, the special SAM cleavage activity of Dph2 resulted from the distinct geometry of SAM binding with 4Fe-4S, which was conjugated by three cysteines from different domains of the protein instead of the CxxxCxxC motif. 25 The Broderick group 27 and our group 28 recently demonstrated that glycerol dehydrogenase-activating enzyme GD-AE, the only previously assumed noncanonical radical SAM enzyme with the CxxxCxxC motif, is a canonical radical SAM enzyme that generates the 5′-dA radical. Moreover, many enzymes, such as TWY2 29 , Tsr3 30 and AzeJ 31 , cleave the C γ,Met -S bond of SAM by a nucleophilic mechanism.…”

mentioning

confidence: 99%

Arsinothricin Biosynthesis Involving a Noncanonical Radical SAM Enzyme for C-As Bond Formation

Yao,

He,

Dong

2024

Preprint

Self Cite

View full text Add to dashboard Cite

Arsinothricin is a potent antibiotic secreted by soil bacteria. The biosynthesis of Arsinothricin was proposed to involve two steps. The first step is C–As bond formation between trivalent As and the 3–amino–3–carboxypropyl (ACP) group of S–adenosyl–L–methionine (SAM), which is catalyzed by the protein ArsL. However, the reaction has not been verified in vitro, and ArsL has not been characterized in detail. Interestingly, ArsL contains a CxxxCxxC motif and thus belongs to the radical SAM enzyme superfamily, the members of which cleave SAM and generate a 5′–deoxyadenosyl radical. Here, we found that ArsL cleaves the Cγ,Met–S bond of SAM and generates an ACP radical that resembles Dph2, a noncanonical radical SAM enzyme involved in diphthamid biosynthesis. As Dph2 does not contain the CxxxCxxC motif, ArsL is a unique noncanonical radical SAM enzyme that contains this motif but generates an ACP radical. Together with the methyltransferase ArsM, we successfully reconstituted arsinothricin biosynthesis in vitro. ArsL has a conserved RCCLKC motif in the C–terminal sequence and belongs to the RCCLKC–tail radical SAM protein subfamily. By truncation, we showed that this motif binds to the substrate arsenite and is highly important for its activity. Our results suggested that ArsL is a noncanonical radical SAM enzyme with a canonical radical SAM enzyme motif, implying that more noncanonical radical SAM chemistry may exist within the radical SAM enzyme superfamily

show abstract

mentioning

confidence: 99%

Arsinothricin Biosynthesis Involving a Noncanonical Radical SAM Enzyme for C-As Bond Formation

Yao,

He,

Dong

2024

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…Given not only the value of fluorine-containing compounds for drug discovery − but also an emerging interest in organosilanes for this purpose, silicon-substituted fluorohydrins could present a novel platform for designing conformationally restricted biologically active structures. Alternatively, oxidative desilylation would generate 2-fluoro-1,3-propanediols, which have proven to be useful for studying enzymatic reactions involving glycerol , as well as starting points to access fluorinated carbohydrate analogues of medicinal value . To that end, Tamao–Fleming oxidation of the 3-silyl fluorohydrin products was investigated.…”

Section: Resultsmentioning

confidence: 99%

Regioselective Fluorohydrin Synthesis from Allylsilanes and Evidence for a Silicon–Fluorine Gauche Effect

Clover,

Jones,

Berger

et al. 2024

J. Org. Chem.

View full text Add to dashboard Cite

Allylsilanes can be regioselectively transformed into the corresponding 3-silylfluorohydrin in good yield using a sequence of epoxidation followed by treatment with HF•Et 3 N with or without isolation of the intermediate epoxide. Various siliconsubstitutions are tolerated, resulting in a range of 2-fluoro-3-silylpropan-1-ol products from this method. Whereas other fluorohydrin syntheses by epoxide opening using HF•Et 3 N generally require more forcing conditions (e.g., higher reaction temperature), opening of allylsilane-derived epoxides with this reagent occurs at room temperature. We attribute this rate acceleration along with the observed regioselectivity to a β-silyl effect that stabilizes a proposed cationic intermediate. The use of enantioenriched epoxides indicates that both S N 1-and S N 2-type mechanisms may be operable depending on substitution at silicon. Conformational analysis by NMR and theory along with a crystal structure obtained by X-ray diffraction points to a preference for silicon and fluorine to be proximal to one another in the products, perhaps favored due to electrostatic interactions.

show abstract

“…Interestingly, the special SAM cleavage activity of Dph2 results from the distinct geometry of SAM binding with 4Fe-4S, which is ligated by three cysteines from different domains of the protein instead of the CxxxCxxC motif. 24 The Broderick group 26 and our group 27 recently demonstrated that glycerol dehydrogenase-activating enzyme GD-AE, the only previously assumed noncanonical RS enzyme with the CxxxCxxC motif, 28 is a canonical RS enzyme that generates the 5′-dA radical. Moreover, many enzymes, such as TWY2, 29 Tsr3 30 and AzeJ, 31 cleave the C γ,Met −S bond of SAM by a nucleophilic mechanism.…”

mentioning

confidence: 99%

Arsinothricin Biosynthesis Involving a Radical SAM Enzyme for Noncanonical SAM Cleavage and C–As Bond Formation

Yao,

He,

Chen

et al. 2024

J. Am. Chem. Soc.

Self Cite

View full text Add to dashboard Cite

Arsinothricin is a potent antibiotic secreted by soil bacteria. The biosynthesis of arsinothricin was proposed to involve a C−As bond formation between trivalent As and the 3-amino-3-carboxypropyl (ACP) group of S-adenosyl-L-methionine (SAM), which is catalyzed by the protein ArsL. However, ArsL has not been characterized in detail. Interestingly, ArsL contains a CxxxCxxC motif and thus belongs to the radical SAM enzyme superfamily, the members of which cleave SAM and generate a 5′-deoxyadenosyl radical. Here, we found that ArsL cleaves the C γ,Met −S bond of SAM and generates an ACP radical that resembles Dph2, a noncanonical radical SAM enzyme involved in diphthamid biosynthesis. As Dph2 does not contain the CxxxCxxC motif, ArsL is a unique radical SAM enzyme that contains this motif but generates a noncanonical ACP radical. Together with the methyltransferase ArsM, we successfully reconstituted arsinothricin biosynthesis in vitro. ArsL has a conserved RCCLKC motif in the C-terminal sequence and belongs to the RCCLKC-tail radical SAM protein subfamily. By truncation and mutagenesis, we showed that this motif plays an important role in binding to the substrate arsenite and is highly important for its activity. Our results suggested that ArsL has a canonical radical SAM enzyme motif but catalyzes a noncanonical radical SAM reaction, implying that more noncanonical radical SAM chemistry may exist within the radical SAM enzyme superfamily.

show abstract

Mechanistic investigation of B12-independent glycerol dehydratase and its activating enzyme GD-AE

Abstract: B12-independent glycerol dehydratase (GD) is a glycyl radical enzyme in the biotransformation of glycerol to 1,3-propanediol. GD requires the activating enzyme GD-AE to initiate the radical reaction. GD-AE belongs to...

Cited by 5 publications

References 32 publications

Arsinothricin Biosynthesis Involving a Noncanonical Radical SAM Enzyme for C-As Bond Formation

Arsinothricin Biosynthesis Involving a Noncanonical Radical SAM Enzyme for C-As Bond Formation

Regioselective Fluorohydrin Synthesis from Allylsilanes and Evidence for a Silicon–Fluorine Gauche Effect

Arsinothricin Biosynthesis Involving a Radical SAM Enzyme for Noncanonical SAM Cleavage and C–As Bond Formation

Contact Info

Product

Resources

About