Conversion of cysteine to formylglycine in eukaryotic sulfatases occurs by a common mechanism in the endoplasmic reticulum

Dierks, Thomas; Lecca, M. Rita; Schmidt, Bernhard; Figura, Kurt Von

doi:10.1016/s0014-5793(98)00065-9

Cited by 58 publications

(73 citation statements)

References 15 publications

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“…In our case, it is probable that a percentage of the expressed hrIDS remain inactive as a result of the non-modification of the Cys 84 because of the saturation of the FGly generating system as was suggested by Dierks et al (1998b). Since it has been demonstrated that prokaryotic system for the modification of Cys presents a similar specificity as the eukaryotic cells (Dierks et al 1998a;Dierks et al 1998b;Dierks et al 1999;Marquordt et al 2003;Berteau et al 2006), it is possible that different E. coli proteins (possibly not identified) used this system, saturating it (Henderson and Milazzo, 1979). Any way the function or saturation of FGly generating system, and the percentage of inactive hrIDS will have to be experimentally proved.…”

Section: Discussionmentioning

confidence: 52%

“…Some authors has reported that depending of the type of sulfatase expressed, in some prokaryotes a percentage of the sulfatase produced, carry out the FGly residue at its active site; while another percentage did not; for example the 60% of the prokaryote arylsulfatase expressed under "strong induction conditions" carry out the FGly residue, while 40% remain as original amino acids coded in the DNA sequence (Dierks et al 1998a;Dierks et al 1998b). In our case, it is probable that a percentage of the expressed hrIDS remain inactive as a result of the non-modification of the Cys 84 because of the saturation of the FGly generating system as was suggested by Dierks et al (1998b).…”

Section: Discussionmentioning

confidence: 99%

“…In Linfoblastoid and COS cells it has been found that the mature forms presents a molecular weight between 45-55 kDa. As other members of the sulfatases family, the hIDS undergo a common posttranslational modification in which a Cys or Ser residue at the active enzyme core is converted into FGly (Schmidt et al 1995;Dierks et al 1998a;Dierks et al 1998b;Knaust et al 1998;Dierks et al 1999;Waldow et al 1999). The Cys at position 84 in hIDS is co-translationally modified to formylglycine (FGly) within the endoplasmic reticulum (ER) to produce an active enzyme (von ).…”

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confidence: 99%

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Human sulfatase transiently and functionally active expressed in E. coli K12

Poutou-Piñales

Niño

Landázuri

et al. 2010

Electron. J. Biotechnol.

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Section: Discussionmentioning

confidence: 52%

Section: Discussionmentioning

confidence: 99%

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confidence: 99%

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Human sulfatase transiently and functionally active expressed in E. coli K12

Poutou-Piñales

Niño

Landázuri

et al. 2010

Electron. J. Biotechnol.

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“…active site (1)(2)(3)(4)(5)(6)(7)(8)(9). In the native enzyme FGly is present as an aldehyde hydrate carrying two geminal hydroxyls at its ␤-carbon that both participate in catalysis (10 -12).…”

mentioning

confidence: 99%

Molecular Characterization of the Human Cα-formylglycine-generating Enzyme

Preusser-Kunze¹,

Mariappan²,

Schmidt³

et al. 2005

Journal of Biological Chemistry

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139

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C␣-formylglycine (FGly) is the catalytic residue in the active site of sulfatases. In eukaryotes, it is generated in the endoplasmic reticulum by post-translational modification of a conserved cysteine residue. The FGly-generating enzyme (FGE), performing this modification, is an endoplasmic reticulum-resident enzyme that upon overexpression is secreted. Recombinant FGE was purified from cells and secretions to homogeneity. Intracellular FGE contains a high mannose type N-glycan, which is processed to the complex type in secreted FGE. Secreted FGE shows partial N-terminal trimming up to residue 73 without loosing catalytic activity. FGE is a calciumbinding protein containing an N-terminal (residues 86 -168) and a C-terminal (residues 178 -374) protease-resistant domain. The latter is stabilized by three disulfide bridges arranged in a clamp-like manner, which links the third to the eighth, the fourth to the seventh, and the fifth to the sixth cysteine residue. The innermost cysteine pair is partially reduced. The first two cysteine residues are located in the sequence preceding the Nterminal protease-resistant domain. They can form intramolecular or intermolecular disulfide bonds, the latter stabilizing homodimers. The C-terminal domain comprises the substrate binding site, as evidenced by yeast two-hybrid interaction assays and photocrosslinking of a substrate peptide to proline 182. Peptides derived from all known human sulfatases served as substrates for purified FGE indicating that FGE is sufficient to modify all sulfatases of the same species.

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“…During synthesis of sulfatases this cysteine is incorporated into the nascent polypeptide (16). It is converted to FGly during or shortly after protein translocation into the endoplasmic reticulum (ER), as could be shown in an in vitro translation/translocation system comprising import-competent dog pancreas microsomes (16,17). The FGly modification is directed by an autonomous linear sequence motif (CXPSRXXX(L/M)TG(R/K/L)) that is located within the N-terminal 80 -120 amino acid residues of the sulfatases, i.e.…”

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confidence: 99%

Characterization of Posttranslational Formylglycine Formation by Luminal Components of the Endoplasmic Reticulum

Fey¹,

Balleininger²,

Borissenko³

et al. 2001

Journal of Biological Chemistry

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C ␣ -formylglycine is the key catalytic residue in the active site of sulfatases. In eukaryotes formylglycine is generated during or immediately after sulfatase translocation into the endoplasmic reticulum by oxidation of a specific cysteine residue. We established an in vitro assay that allowed us to measure formylglycine modification independent of protein translocation. The modifying enzyme was recovered in a microsomal detergent extract. As a substrate we used ribosome-associated nascent chain complexes comprising in vitro synthesized sulfatase fragments that were released from the ribosomes by puromycin. Formylglycine modification was highly efficient and did not require a signal sequence in the substrate polypeptide. Ribosome association helped to maintain the modification competence of nascent chains but only after their release efficient modification occurred. The modifying machinery consists of soluble components of the endoplasmic reticulum lumen, as shown by differential extraction of microsomes. The in vitro assay can be performed under kinetically controlled conditions. The activation energy for formylglycine formation is 61 kJ/mol, and the pH optimum is Ϸ10. The activity is sensitive to the SH/SS equilibrium and is stimulated by Ca 2؉ . Formylglycine formation is efficiently inhibited by a synthetic sulfatase peptide representing the sequence directing formylglycine modification. The established assay system should make possible the biochemical identification of the modifying enzyme.

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Conversion of cysteine to formylglycine in eukaryotic sulfatases occurs by a common mechanism in the endoplasmic reticulum

Cited by 58 publications

References 15 publications

Human sulfatase transiently and functionally active expressed in E. coli K12

Human sulfatase transiently and functionally active expressed in E. coli K12

Molecular Characterization of the Human Cα-formylglycine-generating Enzyme

Characterization of Posttranslational Formylglycine Formation by Luminal Components of the Endoplasmic Reticulum

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