Molecular Characterization of the Human Cα-formylglycine-generating Enzyme

Preusser-Kunze, Andrea; Mariappan, Malaiyalam; Schmidt, Bernhard; Gande, S.L.; Mutenda, Kudzai E.; Wenzel, Dirk; Figura, Kurt Von; Dierks, Thomas

doi:10.1074/jbc.m413383200

Cited by 75 publications

(150 citation statements)

References 32 publications

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“…The highest residual activity (16% of wt) was exhibited by FGE G263V and the lowest by FGE S155P (1.6%). These data, obtained by measuring directly the FGly formation in a sulfatase substrate peptide, 23,29,40 are in accordance with observations made by Annunziata et al, 25 who expressed different MSD causing SUMF1 mutations together with reporter sulfatases in FGE-deficient cells and observed detectable, although reduced sulfatase activities in all cases. Thus, all MSD causing mutations analyzed so far are hypomorphic -at least one per patient.…”

Section: Molecular Phenotype Of Msd: Differential Functional Impairmesupporting

confidence: 78%

“…36,37 Importantly, the four studied FGE variants with single substitutions were more (p.G247R, p.G263V and p.R345C) or less (p.S155P) efficient in passing this quality control mechanism, as these proteins were also secreted -just like wt FGE (Figures 1, 2 and 4), which is secreted even at normal expression levels. 29,38,39 In fact, all FGE variants except p.R327X and p.A149_A173del (see below) were able to catalyze a significant substrate turnover in vitro, as determined after isolation from the medium or cell lysates. The highest residual activity (16% of wt) was exhibited by FGE G263V and the lowest by FGE S155P (1.6%).…”

Section: Molecular Phenotype Of Msd: Differential Functional Impairmementioning

confidence: 99%

“…wt FGE is secreted and processed in the Golgi leading to an N-terminally truncated form in the medium. 29,33 Only three of the FGE variants under study also showed secretion into the cell-culture medium (p.G247R, p.G263V and p.R345C). All other variants, despite their comparable intracellular expression, were not secreted ( Figure 1, lower panel).…”

Section: Expression Of Recombinant Fge Mutant Proteinsmentioning

confidence: 99%

“…13,15,23,20,29 Our approach was substantiated by mapping the amino acid substitutions on the crystal structure of wt FGE (see Supplementary Figure S1), which allowed a solid prediction of structural consequences. Without exception, all missense mutations affected strictly conserved residues and were expected to impair FGE protein stability because of unfavorable side chain interactions resulting in clashes with other residues (p.S155P, p.G247R, p.G263V) and/or losses of specific hydrogen bonds (p.R345C).…”

Section: Molecular Phenotype Of Msd: Differential Functional Impairmementioning

confidence: 99%

“…13,20,29,21 ECL signals were quantified using the AIDA software package (Raytest, Straubenhardt, Germany). Endogenous FGE in MSD fibroblasts was detected using ECL femto substrate and enhancer diluted 1:10 in ECL solution (Pierce, Bonn, Germany).…”

Section: Western Blot Analysismentioning

confidence: 99%

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SUMF1 mutations affecting stability and activity of formylglycine generating enzyme predict clinical outcome in multiple sulfatase deficiency

et al. 2011

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Multiple Sulfatase Deficiency (MSD) is caused by mutations in the sulfatase-modifying factor 1 gene encoding the formylglycine-generating enzyme (FGE). FGE post translationally activates all newly synthesized sulfatases by generating the catalytic residue formylglycine. Impaired FGE function leads to reduced sulfatase activities. Patients display combined clinical symptoms of single sulfatase deficiencies. For ten MSD patients, we determined the clinical phenotype, FGE expression, localization and stability, as well as residual FGE and sulfatase activities. A neonatal, very severe clinical phenotype resulted from a combination of two nonsense mutations leading to almost fully abrogated FGE activity, highly unstable FGE protein and nearly undetectable sulfatase activities. A late infantile mild phenotype resulted from FGE G263V leading to unstable protein but high residual FGE activity. Other missense mutations resulted in a late infantile severe phenotype because of unstable protein with low residual FGE activity. Patients with identical mutations displayed comparable clinical phenotypes. These data confirm the hypothesis that the phenotypic outcome in MSD depends on both residual FGE activity as well as protein stability. Predicting the clinical course in case of molecularly characterized mutations seems feasible, which will be helpful for genetic counseling and developing therapeutic strategies aiming at enhancement of FGE.

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Section: Molecular Phenotype Of Msd: Differential Functional Impairmesupporting

confidence: 78%

Section: Molecular Phenotype Of Msd: Differential Functional Impairmementioning

confidence: 99%

Section: Expression Of Recombinant Fge Mutant Proteinsmentioning

confidence: 99%

Section: Molecular Phenotype Of Msd: Differential Functional Impairmementioning

confidence: 99%

Section: Western Blot Analysismentioning

confidence: 99%

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SUMF1 mutations affecting stability and activity of formylglycine generating enzyme predict clinical outcome in multiple sulfatase deficiency

et al. 2011

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Incorporation of desmocollin‐2 into the plasma membrane requires N‐glycosylation at multiple sites

et al. 2019

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Desmocollin‐2 (DSC2) is a desmosomal protein of the cadherin family. Desmosomes are multiprotein complexes, which are involved in cell adhesion of cardiomyocytes and of keratinocytes. The molecular structure of the complete extracellular domain (ECD) of DSC2 was recently described, revealing three disulfide bridges, four N ‐glycosylation sites, and four O ‐mannosylation sites. However, the functional relevance of these post‐translational modifications for the protein trafficking of DSC2 to the plasma membrane is still unknown. Here, we generated a set of DSC2 mutants, in which we systematically exchanged all N ‐glycosylation sites, O ‐mannosylation sites, and disulfide bridges within the ECD and investigated the resulting subcellular localization by confocal laser scanning microscopy. Of note, all single and double N ‐glycosylation‐ deficient mutants were efficiently incorporated into the plasma membrane, indicating that the absence of these glycosylation sites has a minor effect on the protein trafficking of DSC2. However, the exchange of multiple N ‐glycosylation sites resulted in intracellular accumulation. Colocalization analysis using cell compartment trackers revealed that N ‐glycosylation‐ deficient DSC2 mutants were retained within the Golgi apparatus. In contrast, elimination of the four O ‐mannosylation sites or the disulfide bridges in the ECD has no obvious effect on the intracellular protein processing of DSC2. These experiments underscore the importance of N ‐glycosylation at multiple sites of DSC2 for efficient intracellular transport to the plasma membrane.

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