title: Eukaryotic formylglycine-generating enzyme catalyses a monooxygenase type of reaction.
abstract: C α-formylglycine (FGly) is the catalytic residue of sulfatases in eukaryotes. It is generated by a unique post-translational modification catalysed by the FGly-generating enzyme (FGE) in the endoplasmic reticulum. FGE oxidizes a cysteine residue within the conserved CxPxR sequence motif of nascent sulfatase polypeptides to FGly. Here we show that this oxidation is strictly dependent on molecular oxygen (O2) and consumes 1 mol O2 per mol FGly formed. For maximal activity FGE requires an O2 concentration of 9% (105 μM). Sustained FGE activity further requires the presence of a thiol-based reductant such as DTT. FGly is also formed in the absence of DTT, but its formation ceases rapidly. Thus inactivated FGE accumulates in which the cysteine pair Cys336/Cys341 in the catalytic site is oxidized to form disulfide bridges between either Cys336 and Cys341 or Cys341 and the CxPxR cysteine of the sulfatase. These results strongly suggest that the Cys336/Cys341 pair is directly involved in the O2 -dependent conversion of the CxPxR cysteine to FGly. The available data characterize eukaryotic FGE as a monooxygenase, in which Cys336/Cys341 disulfide bridge formation donates the electrons required to reduce one oxygen atom of O2 to water while the other oxygen atom oxidizes the CxPxR cysteine to FGly. Regeneration of a reduced Cys336/Cys341 pair is accomplished in vivo by a yet unknown reductant of the endoplasmic reticulum or in vitro by DTT. Remarkably, this monooxygenase reaction utilizes O2 without involvement of any activating cofactor.
authors.affiliation: Department of Cellular Biochemistry, University of Göttingen, Germany.
authors.forename: Jianhe
authors.initials: J
authors.lastname: Peng
authors.affiliation: Department of Cellular Biochemistry, University of Göttingen, Germany.
authors.forename: Sarfaraz
authors.initials: S
authors.lastname: Alam
authors.affiliation: Department of Cellular Biochemistry, University of Göttingen, Germany.|Department of Chemistry, Biochemistry I, Bielefeld University, Germany.
authors.forename: Karthikeyan
authors.initials: K
authors.lastname: Radhakrishnan
authors.affiliation: Department of Cellular Biochemistry, University of Göttingen, Germany.
authors.forename: Malaiyalam
authors.initials: M
authors.lastname: Mariappan
authors.affiliation: Discovery Technology, F. Hoffmann La Roche, Basel, Switzerland.
authors.forename: Markus Georg
authors.initials: MG
authors.lastname: Rudolph
authors.affiliation: Department of Medical Proteomics/Bioanalytics, Medizinisches Proteom-Center, Ruhr-University Bochum, Germany.
authors.forename: Caroline
authors.initials: C
authors.lastname: May
authors.affiliation: Department of Chemistry, Biochemistry I, Bielefeld University, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Dierks
authors.affiliation: Department of Cellular Biochemistry, University of Göttingen, Germany.
authors.forename: Kurt
authors.initials: K
authors.lastname: von Figura
authors.affiliation: Department of Cellular Biochemistry, University of Göttingen, Germany.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Schmidt
mesh_terms: D000409:Alanine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D055162:Biocatalysis; D020134:Catalytic Domain; D003545:Cysteine / Q000737:chemistry / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry; D004229:Dithiothreitol / Q000737:chemistry; D057075:Enzyme Assays; D015870:Gene Expression; D005998:Glycine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction; D050862:Oxidoreductases Acting on Sulfur Group Donors; D010100:Oxygen / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D061987:Sf9 Cells; D018411:Spodoptera; D013429:Sulfatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D004220:Disulfides; D011994:Recombinant Proteins; C524739:C(alpha)-formylglycine; D006899:Mixed Function Oxygenases; D050862:Oxidoreductases Acting on Sulfur Group Donors; C480483:SUMF1 protein, human; D013429:Sulfatases; D003545:Cysteine; D000409:Alanine; D010100:Oxygen; D004229:Dithiothreitol; D005998:Glycine
keywords: catalysis; endoplasmic reticulum; formylglycine-generating enzyme; monooxygenase; multiple sulfatase deficiency