Publications

• p-Hydroxyphenylacetate decarboxylase from Clostridium difficile. A novel glycyl radical enzyme catalysing the formation of p-cresol.

  Selmer T., Andrei P.I.

  The human pathogenic bacterium Clostridium difficile is a versatile organism concerning its ability to ferment amino acids. The formation of p-cresol as the main fermentation product of tyrosine by C. difficile is unique among clostridial species. The enzyme responsible for p-cresol formation is p ... >> More

  The human pathogenic bacterium Clostridium difficile is a versatile organism concerning its ability to ferment amino acids. The formation of p-cresol as the main fermentation product of tyrosine by C. difficile is unique among clostridial species. The enzyme responsible for p-cresol formation is p-hydroxyphenylacetate decarboxylase. The enzyme was purified from C. difficile strain DMSZ 1296(T) and initially characterized. The N-terminal amino-acid sequence was 100% identical to an open reading frame in the unfinished genome of C. difficile strain 630. The ORF encoded a protein of the same size as the purified decarboxylase and was very similar to pyruvate formate-lyase-like proteins from Escherichia coli and Archaeoglobus fulgidus. The enzyme decarboxylated p-hydroxyphenylacetate (K(m) = 2.8 mM) and 3,4-dihydroxyphenylacetate (K(m) = 0.5 mM). It was competitively inhibited by the substrate analogues p-hydroxyphenylacetylamide and p-hydroxymandelate with K(i) values of 0.7 mM and 0.48 mM, respectively. The protein was readily and irreversibly inactivated by molecular oxygen. Although the purified enzyme was active in the presence of sodium sulfide, there are some indications for an as yet unidentified low molecular mass cofactor that is required for catalytic activity in vivo. Based on the identification of p-hydroxyphenylacetate decarboxylase as a novel glycyl radical enzyme and the substrate specificity of the enzyme, a catalytic mechanism involving ketyl radicals as intermediates is proposed. << Less

  Eur. J. Biochem. 268:1363-1372(2001) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• 4-Hydroxyphenylacetate decarboxylases: properties of a novel subclass of glycyl radical enzyme systems.

  Yu L., Blaser M., Andrei P.I., Pierik A.J., Selmer T.

  The 4-hydroxyphenylacetate decarboxylases from Clostridium difficile and Clostridium scatologenes, which catalyze the formation of p-cresol, form a distinct group of glycyl radical enzymes (GREs). Cresol formation provides metabolic toxicity, which allows an active suppression of other microbes an ... >> More

  The 4-hydroxyphenylacetate decarboxylases from Clostridium difficile and Clostridium scatologenes, which catalyze the formation of p-cresol, form a distinct group of glycyl radical enzymes (GREs). Cresol formation provides metabolic toxicity, which allows an active suppression of other microbes and may provide growth advantages for the producers in highly competitive environments. The GRE decarboxylases are characterized by a small subunit, which is not similar to any protein of known function in the databases, and provides unique properties that have not been observed in other GREs. Both decarboxylases are functional hetero-octamers (beta(4)gamma(4)), which contain iron-sulfur centers in addition to the glycyl radical prosthetic group. The small subunit is responsible for metal binding and is also involved in the regulation of the enzymes' oligomeric state and activity, which are triggered by reversible serine phosphorylation of the glycyl radical subunits. Biochemical data suggest that the iron-sulfur centers of the decarboxylases could be involved in the radical dissipation of previously activated enzymes in the absence of substrate. The cognate activating enzymes differ from their Pfl and Nrd counterparts in that up to two iron-sulfur centers, in addition to the characteristic SAM cluster, were found. Biochemical data suggested that these [4Fe-4S] centers are involved in the electron transfer to the SAM cluster but do not directly participate in the reductive cleavage of SAM. These data imply a tight regulation of p-cresol formation, which is necessary in order to avoid detrimental effects of the toxic product on the producers. << Less

  Biochemistry 45:9584-9592(2006) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.