Hydrogen Transfer in SAM‐Mediated Enzymatic Radical Reactions

Hioe, Johnny; Zipse, Hendrik

doi:10.1002/chem.201202869

Cited by 29 publications

(36 citation statements)

References 56 publications

(136 reference statements)

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“…97d Computational studies have provided further thermodynamic rationale for invoking intermediary amino acid radicals, as this would provide an SPL mechanism that avoids any strongly endothermic or exothermic steps. 478,479 …”

Section: Using Radical Sam Chemistry To Cleave C–x (X = C N P) Bondsmentioning

confidence: 99%

RadicalS-Adenosylmethionine Enzymes

et al. 2014

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Section: Using Radical Sam Chemistry To Cleave C–x (X = C N P) Bondsmentioning

confidence: 99%

RadicalS-Adenosylmethionine Enzymes

et al. 2014

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“…[1][2][3][4] Together with BDE data for other dipeptide radicals this was taken to reflect steric interactions between the C a substituents and the amide groups present on the N-and C-terminal side of the central amino acid radicals. …”

Section: Introductionmentioning

confidence: 99%

Dissociation energies of Cα–H bonds in amino acids – a re-examination

Hioe

Mosch

Smith³

et al. 2013

RSC Adv.

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aThe C a -H bond dissociation energies (BDE) in glycine and alanine peptide models have been assessed using selected theoretical methods from the G3 and, in part, G4 family. The BDE values (and thus the stability of the respective C a peptide radicals) are shown to depend significantly on the level of theory, the size of the model system and the coverage of conformational space. For the largest dipeptide models chosen here, BDE(C a -H) values of +363.8 kJ mol 21 (glycine) and +372.3 kJ mol 21 (alanine) have been obtained at G3B3 level. This reconfirms earlier findings that glycyl peptide radicals are more stable than radicals derived from alanine or any other amino acid carrying substituents at the C a position.

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“…While this manuscript was under review, Hioe and Zipse reported theoretical data of potential thermochemical profiles. 19 They also suggest that the SPL enzyme can circumvent the endothermic formation of the adenosyl radical when the radical chain reaction is propagated by an intermediate tyrosyl radical. Although formation of the Tyr98 radical is based on our data very likely, this conclusion needs further substantiation particularly based on EPR spectroscopic data.…”

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confidence: 99%

The radical SAM enzyme spore photoproduct lyase employs a tyrosyl radical for DNA repair

et al. 2013

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The spore photoproduct lyase is a radical SAM enzyme, which repairs 5-(a-thyminyl)-5,6-dihydrothymidine. Here we show that the enzyme establishes a complex radical transfer cascade and creates a cysteine and a tyrosyl radical dyade to establish repair. This allows the enzyme to solve topological and energetic problems associated with the radical based repair reaction.UV irradiation causes the formation of a variety of dinucleotide lesions, which are typically formed between two pyrimidines. 1,2In cellular DNA, UV irradiation causes the formation of the well studied cyclobutane pyrimidine dimers (CPDs), pyrimidine (6-4)-pyrimidone photoproducts (6-4PPs), and their Dewar-valence isomers.1 In bacterial endospores, in contrast, these bipyrimidine photoproducts are only formed in low amounts. Due to the unusual packing of the DNA in spores, UV irradiation was found to create selectively the (5R)-5-(a-thyminyl)-5,6-dihydrothymidine lesion (spore photoproduct or SP-lesion). [3][4][5] This dimeric thymidine-photoproduct is repaired during germination by the spore photoproduct lyase (SPL), which is a radical SAM enzyme. 6The SPL catalyzes the direct repair of the SP-lesion, utilizing a radical mediated mechanism (Fig. 1A) ( Fig. 1B) shows the topological problem associated with closing the catalytic cycle. While the allyl radical is situated and reduced at the 3 0 -side, transfer of the radical center back to the 5 0 -dAdoH requires moving the radical back to the 5 0 -part in the active side over a distance of roughly 10 Å. This creates next to a topological problem also an energetic obstacle, because regeneration of the adenosyl radical by the thiyl radical would be endothermic by 62 kJ mol À1. 16Here we provide first evidence that the enzyme uses a further tyrosyl radical intermediate to solve the energetic and topological problem. In the crystal structure the tyrosine bridges the conserved cysteine and the 5 0 -dAdoH. The structure shows that Tyr98 is located 3.6 Å and 5.1 Å away from both centers, respectively. In order to clarify the role of Cys140 and Tyr98 we prepared two mutants C140A and Y98F ( Fig. 2A) and determined their catalytic activity. To this end we quantified product formation after 3 h and after reaction overnight. The measured K m and V max values are listed in Table 1. The data show that both mutants have a strongly reduced catalytic activity. The Y98F mutant shows in addition a reduced substrate binding affinity, which indicates that the phenolic hydroxyl group is important to organize the substrate in the active site. Direct interactions with the lesion are, however, not observed in the crystal structure. For the catalytically competent wild type (wt) enzyme we measured a ratio of product/5 0 -dAdoH of greater than two indicating that the cleavage of one SAM molecule repairs more than one lesion (turnover). The mutants in contrast use more than one SAM per product showing that here the turnover is smaller than one product molecule (0.7 and 0.25, Fig. 2B) formed per SAM. This result shows that Cys1...

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Hydrogen Transfer in SAM‐Mediated Enzymatic Radical Reactions

Cited by 29 publications

References 56 publications

RadicalS-Adenosylmethionine Enzymes

RadicalS-Adenosylmethionine Enzymes

Dissociation energies of Cα–H bonds in amino acids – a re-examination

The radical SAM enzyme spore photoproduct lyase employs a tyrosyl radical for DNA repair

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