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Protein O-Mannosyltransferases Associate with the Translocon to Modify Translocating Polypeptide Chains
- Source :
- Journal of Biological Chemistry. 289:8599-8611
- Publication Year :
- 2014
- Publisher :
- Elsevier BV, 2014.
-
Abstract
- O-Mannosylation and N-glycosylation are essential protein modifications that are initiated in the endoplasmic reticulum (ER). Protein translocation across the ER membrane and N-glycosylation are highly coordinated processes that take place at the translocon-oligosaccharyltransferase (OST) complex. In analogy, it was assumed that protein O-mannosyltransferases (PMTs) also act at the translocon, however, in recent years it turned out that prolonged ER residence allows O-mannosylation of un-/misfolded proteins or slow folding intermediates by Pmt1-Pmt2 complexes. Here, we reinvestigate protein O-mannosylation in the context of protein translocation. We demonstrate the association of Pmt1-Pmt2 with the OST, the trimeric Sec61, and the tetrameric Sec63 complex in vivo by co-immunoprecipitation. The coordinated interplay between PMTs and OST in vivo is further shown by a comprehensive mass spectrometry-based analysis of N-glycosylation site occupancy in pmtĪ mutants. In addition, we established a microsomal translation/translocation/O-mannosylation system. Using the serine/threonine-rich cell wall protein Ccw5 as a model, we show that PMTs efficiently mannosylate proteins during their translocation into microsomes. This in vitro system will help to unravel mechanistic differences between co- and post-translocational O-mannosylation.
- Subjects :
- Sec61
Glycosylation
Saccharomyces cerevisiae Proteins
Endoplasmic reticulum
Glycobiology and Extracellular Matrices
Context (language use)
Translation (biology)
Saccharomyces cerevisiae
Cell Biology
Biology
Translocon
Mannosyltransferases
Biochemistry
Cell biology
Transport protein
carbohydrates (lipids)
Protein Transport
SEC63
Protein folding
Molecular Biology
Subjects
Details
- ISSN :
- 00219258
- Volume :
- 289
- Database :
- OpenAIRE
- Journal :
- Journal of Biological Chemistry
- Accession number :
- edsair.doi.dedup.....a9c73f75adcbf6583a2298a630de7968