Abstract
Protein O-mannosylation is initiated at the endoplasmic reticulum (ER) by dolichyl phosphate-mannose: protein O-mannosyltransferases (PMTs). PMTs are members of the glycosyltransferase (GT) C superfamily. They are large polytopic integral membrane proteins located in the ER membrane. PMTs utilize dolichyl phosphate-Âactivated mannose as sugar donor. Glycosyltransfer of mannose to serine and threonine residues of nascent polypeptides leads to an inversion of the stereochemistry of the glycosidic bond. Here, we describe photoaffinity labeling of yeast Pmt1p using a photo-reactive probe that is based on the artificial mannosyl acceptor peptide YATAV. Due to the high homology of PMTs, this method can also be applied to study PMT1 and PMT2 subfamily members from fungi other than baker’s yeast.
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Acknowledgments
We thank V. Hofmann for excellent technical advice. We thank A. Schott and M. Buettner for critical reading of the manuscript. This work was partially supported by the Deutsche Forschungs Gemeinschaft (SFB638). S. Strahl is a member of CellNetworks-Cluster of Excellence (EXC81).
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Loibl, M., Strahl, S. (2013). Photoaffinity Labeling of Protein O-Mannosyltransferases of the PMT1/PMT2 Subfamily. In: Brockhausen, I. (eds) Glycosyltransferases. Methods in Molecular Biology, vol 1022. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-465-4_9
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DOI: https://doi.org/10.1007/978-1-62703-465-4_9
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