Structural basis for the glycosyltransferase activity of the Salmonella effector SseK3
| Autor: | Regina A. Günster, Teresa L. M. Thurston, Kamel El Omari, Armin Wagner, Luigi Martino, Katrin Rittinger, Diego Esposito |
|---|---|
| Přispěvatelé: | Wellcome Trust |
| Rok vydání: | 2018 |
| Předmět: |
CONFORMATIONAL-CHANGES
0301 basic medicine HOST Arginine PROTEIN GT-A family glycosyltransferase Biochemistry Protein structure arginine modification GLCNACYLATION CRYSTAL-STRUCTURE III SECRETION SYSTEM chemistry.chemical_classification biology Effector Chemistry bacterial effectors Salmonella enterica 11 Medical And Health Sciences CLOSTRIDIUM-DIFFICILE TOXIN Ligand (biochemistry) SseK3 ESCHERICHIA-COLI 03 Chemical Sciences Life Sciences & Biomedicine Biochemistry & Molecular Biology GLUCOSYLTRANSFERASE ACTIVITY 03 medical and health sciences Glycosyltransferase bacterial toxin enzyme mechanism arginine-modification structural analysis protein structure Molecular Biology X-ray crystallography Science & Technology 030102 biochemistry & molecular biology Active site Isothermal titration calorimetry Cell Biology 06 Biological Sciences UDP-GlcNAc 030104 developmental biology Enzyme glycosyltransferase type-A glycosyltransgerase type-A biology.protein DEATH DOMAIN |
| Zdroj: | Journal of Biological Chemistry. 293:5064-5078 |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.ra118.001796 |
| Popis: | The Salmonella secreted effector SseK3 translocates into host cells, targeting innate immune responses including NF-κB activation. SseK3 is a glycosyltransferase that transfers an N-acetylglucosamine (GlcNAc) moiety onto the guanidino group of a target arginine, modulating host cell function. However, a lack of structural information has precluded elucidation of the molecular mechanisms in arginine and GlcNAc selection. We report here the crystal structure of SseK3 in its apo form and in complex with hydrolysed UDP-GlcNAc. SseK3 possesses the typical glycosyltransferase type-A (GT-A)-family fold and the metal-coordinating DXD motif essential for ligand binding and enzymatic activity. Several conserved residues were essential for arginine-GlcNAcylation and SseK3-mediated inhibition of NF-κB activation. Isothermal titration calorimetry revealed SseK3's preference for manganese coordination. The pattern of interactions in the substrate-bound SseK3 structure explained the selection of the primary ligand. Structural re-arrangement of the C-terminal residues upon ligand binding was crucial for SseK3's catalytic activity and NMR analysis indicated that SseK3 has limited UDP-GlcNAc hydrolysis activity. The release of free N-acetyl α-D-glucosamine, and the presence of the same molecule in the SseK3 active site, classified it as a retaining glycosyltransferase. A glutamate residue in the active site suggested a double-inversion mechanism for the arginine N-glycosylation reaction. Homology models of SseK1, SseK2, and the Escherichia coli orthologue NleB1, reveal differences in the surface electrostatic charge distribution possibly accounting for their diverse activities. This first structure of a retaining GT-A arginine N-glycosyltransferase provides an important step towards a better understanding of this enzyme class and their roles as bacterial effectors. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|