Structure and functional impact of glycosaminoglycan modification of HSulf-2 endosulfatase revealed by atomic force microscopy and mass spectrometry
Abstract The human sulfatase HSulf-2 is one of only two known endosulfatases that play a decisive role in modulating the binding properties of heparan sulfate proteoglycans on the cell surface and in the extracellular matrix. Recently, HSulf-2 was shown to exhibit an unusual post-translational modif...
| Main Authors: | , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Nature Portfolio
2023-12-01
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| Series: | Scientific Reports |
| Online Access: | https://doi.org/10.1038/s41598-023-49147-5 |
| _version_ | 1827581580244156416 |
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| author | Ilham Seffouh Mélanie Bilong Cédric Przybylski Nesrine El Omrani Salomé Poyer Guillaume Lamour Marie-Jeanne Clément Rebecca-Joe Boustany Evelyne Gout Florence Gonnet Romain R. Vivès Régis Daniel |
| author_facet | Ilham Seffouh Mélanie Bilong Cédric Przybylski Nesrine El Omrani Salomé Poyer Guillaume Lamour Marie-Jeanne Clément Rebecca-Joe Boustany Evelyne Gout Florence Gonnet Romain R. Vivès Régis Daniel |
| author_sort | Ilham Seffouh |
| collection | DOAJ |
| description | Abstract The human sulfatase HSulf-2 is one of only two known endosulfatases that play a decisive role in modulating the binding properties of heparan sulfate proteoglycans on the cell surface and in the extracellular matrix. Recently, HSulf-2 was shown to exhibit an unusual post-translational modification consisting of a sulfated glycosaminoglycan chain. This study describes the structural characterization of this glycosaminoglycan (GAG) and provides new data on its impact on the catalytic properties of HSulf-2. The unrevealed nature of this GAG chain is identified as a chondroitin/dermatan sulfate (CS/DS) mixed chain, as shown by mass spectrometry combined with NMR analysis. It consists primarily of 6-O and 4-O monosulfated disaccharide units, with a slight predominance of the 4-O-sulfation. Using atomic force microscopy, we show that this unique post-translational modification dramatically impacts the enzyme hydrodynamic volume. We identified human hyaluronidase-4 as a secreted hydrolase that can digest HSulf-2 GAG chain. We also showed that HSulf-2 is able to efficiently 6-O-desulfate antithrombin III binding pentasaccharide motif, and that this activity was enhanced upon removal of the GAG chain. Finally, we identified five N-glycosylation sites on the protein and showed that, although required, reduced N-glycosylation profiles were sufficient to sustain HSulf-2 integrity. |
| first_indexed | 2024-03-08T22:38:26Z |
| format | Article |
| id | doaj.art-74ccc652ff974500a6d2da80b5385266 |
| institution | Directory Open Access Journal |
| issn | 2045-2322 |
| language | English |
| last_indexed | 2024-03-08T22:38:26Z |
| publishDate | 2023-12-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj.art-74ccc652ff974500a6d2da80b53852662023-12-17T12:18:39ZengNature PortfolioScientific Reports2045-23222023-12-0113111410.1038/s41598-023-49147-5Structure and functional impact of glycosaminoglycan modification of HSulf-2 endosulfatase revealed by atomic force microscopy and mass spectrometryIlham Seffouh0Mélanie Bilong1Cédric Przybylski2Nesrine El Omrani3Salomé Poyer4Guillaume Lamour5Marie-Jeanne Clément6Rebecca-Joe Boustany7Evelyne Gout8Florence Gonnet9Romain R. Vivès10Régis Daniel11Université Paris-Saclay, Univ Evry, CY Cergy Paris Université, CNRS, LAMBEUniversité Paris-Saclay, Univ Evry, CY Cergy Paris Université, CNRS, LAMBEUniversité Paris-Saclay, Univ Evry, CY Cergy Paris Université, CNRS, LAMBEUniversité Paris-Saclay, Univ Evry, CY Cergy Paris Université, CNRS, LAMBEUniversité Paris-Saclay, Univ Evry, CY Cergy Paris Université, CNRS, LAMBEUniversité Paris-Saclay, Univ Evry, CY Cergy Paris Université, CNRS, LAMBEUniversité Paris-Saclay, Univ Evry, INSERM, SABNPUniv. Grenoble Alpes, CNRS, CEA, IBSUniv. Grenoble Alpes, CNRS, CEA, IBSUniversité Paris-Saclay, Univ Evry, CY Cergy Paris Université, CNRS, LAMBEUniv. Grenoble Alpes, CNRS, CEA, IBSUniversité Paris-Saclay, Univ Evry, CY Cergy Paris Université, CNRS, LAMBEAbstract The human sulfatase HSulf-2 is one of only two known endosulfatases that play a decisive role in modulating the binding properties of heparan sulfate proteoglycans on the cell surface and in the extracellular matrix. Recently, HSulf-2 was shown to exhibit an unusual post-translational modification consisting of a sulfated glycosaminoglycan chain. This study describes the structural characterization of this glycosaminoglycan (GAG) and provides new data on its impact on the catalytic properties of HSulf-2. The unrevealed nature of this GAG chain is identified as a chondroitin/dermatan sulfate (CS/DS) mixed chain, as shown by mass spectrometry combined with NMR analysis. It consists primarily of 6-O and 4-O monosulfated disaccharide units, with a slight predominance of the 4-O-sulfation. Using atomic force microscopy, we show that this unique post-translational modification dramatically impacts the enzyme hydrodynamic volume. We identified human hyaluronidase-4 as a secreted hydrolase that can digest HSulf-2 GAG chain. We also showed that HSulf-2 is able to efficiently 6-O-desulfate antithrombin III binding pentasaccharide motif, and that this activity was enhanced upon removal of the GAG chain. Finally, we identified five N-glycosylation sites on the protein and showed that, although required, reduced N-glycosylation profiles were sufficient to sustain HSulf-2 integrity.https://doi.org/10.1038/s41598-023-49147-5 |
| spellingShingle | Ilham Seffouh Mélanie Bilong Cédric Przybylski Nesrine El Omrani Salomé Poyer Guillaume Lamour Marie-Jeanne Clément Rebecca-Joe Boustany Evelyne Gout Florence Gonnet Romain R. Vivès Régis Daniel Structure and functional impact of glycosaminoglycan modification of HSulf-2 endosulfatase revealed by atomic force microscopy and mass spectrometry Scientific Reports |
| title | Structure and functional impact of glycosaminoglycan modification of HSulf-2 endosulfatase revealed by atomic force microscopy and mass spectrometry |
| title_full | Structure and functional impact of glycosaminoglycan modification of HSulf-2 endosulfatase revealed by atomic force microscopy and mass spectrometry |
| title_fullStr | Structure and functional impact of glycosaminoglycan modification of HSulf-2 endosulfatase revealed by atomic force microscopy and mass spectrometry |
| title_full_unstemmed | Structure and functional impact of glycosaminoglycan modification of HSulf-2 endosulfatase revealed by atomic force microscopy and mass spectrometry |
| title_short | Structure and functional impact of glycosaminoglycan modification of HSulf-2 endosulfatase revealed by atomic force microscopy and mass spectrometry |
| title_sort | structure and functional impact of glycosaminoglycan modification of hsulf 2 endosulfatase revealed by atomic force microscopy and mass spectrometry |
| url | https://doi.org/10.1038/s41598-023-49147-5 |
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