The Pseudomonas aeruginosa nirE gene encodes the S-adenosyl-L-methionine- dependent uroporphyrinogen III methyltransferase required for heme d1 biosynthesis

Sonja Storbeck, Johannes Walther, Judith Müller, Vina Parmar, Hans Martin Schiebel, Dorit Kemken, Thomas Dülcks, Martin J. Warren, Gunhild Layer

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26 Citations (Scopus)

Abstract

Biosynthesis of heme d1, the essential prosthetic group of the dissimilatory nitrite reductase cytochrome cd1, requires the methylation of the tetrapyrrole precursor uroporphyrinogen III at positions C-2 and C-7. We produced Pseudomonas aeruginosa NirE, a putative S-adenosyl-l-methionine (SAM)-dependent uroporphyrinogen III methyltransferase, as a recombinant protein in Escherichia coli and purified it to apparent homogeneity by metal chelate and gel filtration chromatography. Analytical gel filtration of purified NirE indicated that the recombinant protein is a homodimer. NirE was shown to be a SAM-dependent uroporphyrinogen III methyltransferase that catalyzes the conversion of uroporphyrinogen III into precorrin-2 in vivo and in vitro. A specific activity of 316.8 nmol of precorrin-2 h-1·mg-1 of NirE was found for the conversion of uroporphyrinogen III to precorrin-2. At high enzyme concentrations NirE catalyzed an overmethylation of uroporphyrinogen III, resulting in the formation of trimethylpyrrocorphin. Substrate inhibition was observed at uroporphyrinogen III concentrations above 17 m. The protein did bind SAM, although not with the same avidity as reported for other SAM-dependent uroporphyrinogen III methyltransferases involved in siroheme and cobalamin biosynthesis. A P. aeruginosa nirE transposon mutant was not complemented by native cobA encoding the SAM-dependent uroporphyrinogen III methyltransferase involved in cobalamin formation. However, bacterial growth of the nirE mutant was observed when cobA was constitutively expressed by a complementing plasmid, underscoring the special requirement of NirE for heme d1 biosynthesis.

Original languageEnglish
Pages (from-to)5973-5982
Number of pages10
JournalFEBS Journal
Volume276
Issue number20
DOIs
Publication statusPublished - Oct 2009

Keywords

  • Heme d biosynthesis
  • Precorrin-2
  • Pseudomonas aeruginosa
  • SAM-dependent uroporphyrinogen III methyltransferase
  • Uroporphyrinogen III

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