4-Hydroxyphenylacetate decarboxylase activating enzyme catalyses a classical S-adenosylmethionine reductive cleavage reaction.
J Biol Inorg Chem
; 18(6): 633-43, 2013 Aug.
Article
em En
| MEDLINE
| ID: mdl-23716017
ABSTRACT
4-Hydroxyphenylacetate decarboxylase (4Hpad) is an Fe/S cluster containing glycyl radical enzyme (GRE), which catalyses the last step of tyrosine fermentation in clostridia, generating the bacteriostatic p-cresol. The respective activating enzyme (4Hpad-AE) displays two cysteine-rich motifs in addition to the classical S-adenosylmethionine (SAM) binding cluster (RS cluster) motif. These additional motifs are also present in other glycyl radical activating enzymes (GR-AE) and it has been postulated that these orthologues may use an alternative SAM homolytic cleavage mechanism, generating a putative 3-amino-3-carboxypropyl radical and 5'-deoxy-5'-(methylthio)adenosine but not a 5'-deoxyadenosyl radical and methionine. 4Hpad-AE produced from a codon-optimized synthetic gene binds a maximum of two [4Fe-4S](2+/+) clusters as revealed by EPR and Mössbauer spectroscopy. The enzyme only catalyses the turnover of SAM under reducing conditions, and the reaction products were identified as 5'-deoxyadenosine (quenched form of 5'-deoxyadenosyl radical) and methionine. We demonstrate that the 5'-deoxyadenosyl radical is the activating agent for 4Hpad through p-cresol formation and correlation between the production of 5'-deoxyadenosine and the generation of glycyl radical in 4Hpad. Therefore, we conclude that 4Hpad-AE catalyses a classical SAM-dependent glycyl radical formation as reported for GR-AE without auxiliary clusters. Our observation casts doubt on the suggestion that GR-AE containing auxiliary clusters catalyse the alternative cleavage reaction detected for glycerol dehydratase activating enzyme.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
S-Adenosilmetionina
/
Carboxiliases
/
Biocatálise
Idioma:
En
Revista:
J Biol Inorg Chem
Assunto da revista:
BIOQUIMICA
Ano de publicação:
2013
Tipo de documento:
Article
País de afiliação:
Alemanha