Mouse WIF1 Is Only Modified with O-Fucose in Its EGF-like Domain III Despite Two Evolutionarily Conserved Consensus Sites
Abstract
:1. Introduction
2. Materials and Methods
2.1. Phylogenetic Reconstruction and Sequence Conservation Analyses
2.2. Plasmid Constructs
2.3. Protein Expression and Purification
2.4. Glycosyltransferase Reactions
2.5. Click Chemistry Reactions
2.6. SDS-PAGE and Blotting Techniques
2.7. Automatic Modeling and Superimposition with X-ray Structures
2.8. Protein Digestions, Mass Spectrometry Analyses and Data Processing
2.9. Statistical Analysis
3. Results
3.1. Wif1 Appeared in Bilaterian Ancestor
3.2. Predicted O-Fucosylation Sites Are Evolutionarily Conserved in Deuterostomes
3.3. Only WIF1 EGF-LD III Can Be In Vitro Modified by O-Fucose
3.4. Full-Length Recombinant WIF1 Carried O-Fucose Only on Its EGF-LD III
3.5. Full-Length Recombinant WIF1 Carried a Non-Extended O-Fucose on Its EGF-LD III
3.6. The O-Fucose Carried by T255 Was Required for Optimal Secretion of Recombinant WIF1
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Pennarubia, F.; Pinault, E.; Al Jaam, B.; Brun, C.E.; Maftah, A.; Germot, A.; Legardinier, S. Mouse WIF1 Is Only Modified with O-Fucose in Its EGF-like Domain III Despite Two Evolutionarily Conserved Consensus Sites. Biomolecules 2020, 10, 1250. https://doi.org/10.3390/biom10091250
Pennarubia F, Pinault E, Al Jaam B, Brun CE, Maftah A, Germot A, Legardinier S. Mouse WIF1 Is Only Modified with O-Fucose in Its EGF-like Domain III Despite Two Evolutionarily Conserved Consensus Sites. Biomolecules. 2020; 10(9):1250. https://doi.org/10.3390/biom10091250
Chicago/Turabian StylePennarubia, Florian, Emilie Pinault, Bilal Al Jaam, Caroline E. Brun, Abderrahman Maftah, Agnès Germot, and Sébastien Legardinier. 2020. "Mouse WIF1 Is Only Modified with O-Fucose in Its EGF-like Domain III Despite Two Evolutionarily Conserved Consensus Sites" Biomolecules 10, no. 9: 1250. https://doi.org/10.3390/biom10091250