Mutant glycosidases for labeling sialoglycans with high specificity and affinity
Abstract Affinity labeling of biomacromolecules is vital for bioimaging and functional studies. However, affinity probes recognizing glycans with high specificity remain scarce. Here we report the development of glycan recombinant affinity binders (GRABs) based on mutant bacterial sialidases, which...
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| Format: | Article |
| Language: | English |
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Nature Portfolio
2025-02-01
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| Series: | Nature Communications |
| Online Access: | https://doi.org/10.1038/s41467-025-56629-9 |
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| author | Shuyu Liang Qi Tang Xunzi Guo Zi’an Li Yilan Guo Jinghan Chang Bo Cheng Qitao Song Jiayu Sun Peng Dai Xing Chen |
| author_facet | Shuyu Liang Qi Tang Xunzi Guo Zi’an Li Yilan Guo Jinghan Chang Bo Cheng Qitao Song Jiayu Sun Peng Dai Xing Chen |
| author_sort | Shuyu Liang |
| collection | DOAJ |
| description | Abstract Affinity labeling of biomacromolecules is vital for bioimaging and functional studies. However, affinity probes recognizing glycans with high specificity remain scarce. Here we report the development of glycan recombinant affinity binders (GRABs) based on mutant bacterial sialidases, which are enzymatically inactive but preserve stringent specificity for sialoglycan substrates. By mutating a key catalytic residue of Streptococcus pneumoniae neuraminidase A (SpNanA) and Ruminococcus gnavus neuraminidase H (RgNanH), we develop GRAB-Sia and GRAB-Sia3 recognizing total sialoglycans and α2,3-sialosides, respectively. The GRABs exhibit strict substrate and linkage specificity, and tetramerization with streptavidin substantially increases their avidity. The GRABs and tetrameric GRABs (tetra-GRABs) are effective tools for probing sialoglycans in immunoblotting, flow cytometry, immunoprecipitation, and fluorescence imaging. Furthermore, multiplex analysis with tetra-GRABs uncovers spatially distinct sialoglycans in the various mouse organs. This work provides a versatile toolkit for labeling and analyzing sialoglycans with high specificity, sensitivity, and convenience. |
| format | Article |
| id | doaj-art-10c004a2696146138e8acde4679da277 |
| institution | Kabale University |
| issn | 2041-1723 |
| language | English |
| publishDate | 2025-02-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Nature Communications |
| spelling | doaj-art-10c004a2696146138e8acde4679da2772025-02-09T12:44:10ZengNature PortfolioNature Communications2041-17232025-02-0116111410.1038/s41467-025-56629-9Mutant glycosidases for labeling sialoglycans with high specificity and affinityShuyu Liang0Qi Tang1Xunzi Guo2Zi’an Li3Yilan Guo4Jinghan Chang5Bo Cheng6Qitao Song7Jiayu Sun8Peng Dai9Xing Chen10College of Chemistry and Molecular Engineering, Peking UniversityCollege of Chemistry and Molecular Engineering, Peking UniversityCollege of Chemistry and Molecular Engineering, Peking UniversityCollege of Chemistry and Molecular Engineering, Peking UniversityCollege of Chemistry and Molecular Engineering, Peking UniversityCollege of Chemistry and Molecular Engineering, Peking UniversityCollege of Chemistry and Molecular Engineering, Peking UniversityCollege of Chemistry and Molecular Engineering, Peking UniversityCollege of Chemistry and Molecular Engineering, Peking UniversityCollege of Chemistry and Molecular Engineering, Peking UniversityCollege of Chemistry and Molecular Engineering, Peking UniversityAbstract Affinity labeling of biomacromolecules is vital for bioimaging and functional studies. However, affinity probes recognizing glycans with high specificity remain scarce. Here we report the development of glycan recombinant affinity binders (GRABs) based on mutant bacterial sialidases, which are enzymatically inactive but preserve stringent specificity for sialoglycan substrates. By mutating a key catalytic residue of Streptococcus pneumoniae neuraminidase A (SpNanA) and Ruminococcus gnavus neuraminidase H (RgNanH), we develop GRAB-Sia and GRAB-Sia3 recognizing total sialoglycans and α2,3-sialosides, respectively. The GRABs exhibit strict substrate and linkage specificity, and tetramerization with streptavidin substantially increases their avidity. The GRABs and tetrameric GRABs (tetra-GRABs) are effective tools for probing sialoglycans in immunoblotting, flow cytometry, immunoprecipitation, and fluorescence imaging. Furthermore, multiplex analysis with tetra-GRABs uncovers spatially distinct sialoglycans in the various mouse organs. This work provides a versatile toolkit for labeling and analyzing sialoglycans with high specificity, sensitivity, and convenience.https://doi.org/10.1038/s41467-025-56629-9 |
| spellingShingle | Shuyu Liang Qi Tang Xunzi Guo Zi’an Li Yilan Guo Jinghan Chang Bo Cheng Qitao Song Jiayu Sun Peng Dai Xing Chen Mutant glycosidases for labeling sialoglycans with high specificity and affinity Nature Communications |
| title | Mutant glycosidases for labeling sialoglycans with high specificity and affinity |
| title_full | Mutant glycosidases for labeling sialoglycans with high specificity and affinity |
| title_fullStr | Mutant glycosidases for labeling sialoglycans with high specificity and affinity |
| title_full_unstemmed | Mutant glycosidases for labeling sialoglycans with high specificity and affinity |
| title_short | Mutant glycosidases for labeling sialoglycans with high specificity and affinity |
| title_sort | mutant glycosidases for labeling sialoglycans with high specificity and affinity |
| url | https://doi.org/10.1038/s41467-025-56629-9 |
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