Catalytic residues and a predicted structure of tetrahydrobiopterin- dependent alkylglycerol mono-oxygenase

Katrin Watschinger, Julian E. Fuchs, Vladimir Yarov-Yarovoy, Markus A. Keller, Georg Golderer, Albin Hermetter, Gabriele Werner-Felmayer, Nicolas Hulo, Ernst R. Werner

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

Alkylglycerol mono-oxygenase (EC 1.14.16.5) forms a third, distinct, class among tetrahydrobiopterin-dependent enzymes in addition to aromatic amino acid hydroxylases and nitric oxide synthases. Its protein sequence contains the fatty acid hydroxylase motif, a signature indicative of a di-iron centre, which contains eight conserved histidine residues. Membrane enzymes containing this motif, including alkylglycerol mono-oxygenase, are especially labile and so far have not been purified to homogeneity in active form. To obtain a first insight into structure-function relationships of this enzyme, we performed site-directed mutagenesis of 26 selected amino acid residues and expressed wild-type and mutant proteins containing a C-terminal Myc tag together with fatty aldehyde dehydrogenase in Chinese-hamster ovary cells. Among all of the acidic residues within the eight-histidinemotif, onlymutation of Glu 137 to alanine led to an 18-fold increase in the Michaelis-Menten constant for tetrahydrobiopterin, suggesting a role in tetrahydrobiopterin interaction. A ninth additional histidine residue essential for activity was also identified. Nine membrane domains were predicted by four programs: ESKW, TMHMM, MEMSAT and Phobius. Prediction of a part of the structure using the Rosetta membrane ab initio method led to a plausible suggestion for a structure of the catalytic site of alkylglycerol mono-oxygenase.

Original languageEnglish (US)
Pages (from-to)279-286
Number of pages8
JournalBiochemical Journal
Volume443
Issue number1
DOIs
StatePublished - Apr 1 2012

Fingerprint

Oxygenases
long-chain-aldehyde dehydrogenase
glyceryl-ether monooxygenase
Mixed Function Oxygenases
Membranes
Histidine
Enzymes
Aromatic Amino Acids
Mutagenesis
Mutant Proteins
Site-Directed Mutagenesis
Cricetulus
Nitric Oxide Synthase
Alanine
Ovary
Catalytic Domain
Fatty Acids
Iron
Cells
Amino Acids

Keywords

  • Alkylglycerol mono-oxygenase
  • Fatty acid hydroxylase motif
  • Phenylalanine hydroxylase
  • Tetrahydrobiopterin
  • Transmembrane domain

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Watschinger, K., Fuchs, J. E., Yarov-Yarovoy, V., Keller, M. A., Golderer, G., Hermetter, A., ... Werner, E. R. (2012). Catalytic residues and a predicted structure of tetrahydrobiopterin- dependent alkylglycerol mono-oxygenase. Biochemical Journal, 443(1), 279-286. https://doi.org/10.1042/BJ20111509

Catalytic residues and a predicted structure of tetrahydrobiopterin- dependent alkylglycerol mono-oxygenase. / Watschinger, Katrin; Fuchs, Julian E.; Yarov-Yarovoy, Vladimir; Keller, Markus A.; Golderer, Georg; Hermetter, Albin; Werner-Felmayer, Gabriele; Hulo, Nicolas; Werner, Ernst R.

In: Biochemical Journal, Vol. 443, No. 1, 01.04.2012, p. 279-286.

Research output: Contribution to journalArticle

Watschinger, K, Fuchs, JE, Yarov-Yarovoy, V, Keller, MA, Golderer, G, Hermetter, A, Werner-Felmayer, G, Hulo, N & Werner, ER 2012, 'Catalytic residues and a predicted structure of tetrahydrobiopterin- dependent alkylglycerol mono-oxygenase', Biochemical Journal, vol. 443, no. 1, pp. 279-286. https://doi.org/10.1042/BJ20111509
Watschinger, Katrin ; Fuchs, Julian E. ; Yarov-Yarovoy, Vladimir ; Keller, Markus A. ; Golderer, Georg ; Hermetter, Albin ; Werner-Felmayer, Gabriele ; Hulo, Nicolas ; Werner, Ernst R. / Catalytic residues and a predicted structure of tetrahydrobiopterin- dependent alkylglycerol mono-oxygenase. In: Biochemical Journal. 2012 ; Vol. 443, No. 1. pp. 279-286.
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