When programmed with yeast prepro-α-factor mRNA, the heterologous reticulocyte/dog pancreas translation system synthesizes two pheromone related polypeptides, a cytosolically located primary translation product (pp-α-Fcyt, 21 kDa) and a membrane-specific and multiply glycosylated e-factor precursor (pp-α-F3, 27.5 kDa). Glycosylation of the membrane specific pp-α-F3 species is competitively inhibited by synthetic peptides containing the consensus sequence Asn-Xaa-Thr as indicated by a shift of its molecular mass from 27.5 kDa to about 19.5 kDa (pp-α-F0), whereas the primary translation product pp-α-F cyt is not affected. Likewise, only the glycosylated pp-α-F3 structure is digested by Endo H yielding a polypeptide with a molecular mass between PP-α-F0 and pp-α-F cyt. These observations strongly suggest that the primary translation product is proteolytically processed during/on its translocation into the lumen of the microsomal vesicles. We believe that this proteolytic processing is due to the cleavage of a signal sequence from the pp-α-F cyt species, although this interpretation contradicts previous data from other groups. The distinct effect exerted by various glycosidase inhibitors (e.g. 1-deoxynojirimycin, N-methyl-dNM, 1-deoxymannojirimycin) on the electrophoretic mobility of the pp-α-F3 polypeptide indicates that its oligosaccharide chains are processed to presumbly Man9-GlcNAc2 structures under the in vitro conditions of translation. This oligosaccharide processing is most likely to involve the action of glucosidase I and glucosidase II as follows from the specificity of the glycosidase inhibitors applied and the differences of the molecular mass observed in their presence. In addition, several arguments suggest that both trimming enzymes are located in the lumen of the microsomal vesicles derived from endoplasmic reticulum membranes.
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September 01 1986
In vitro studies on the subcellular location of glucosidase I and glucosidase II in dog pancreas
Ernst Bause;
Ernst Bause
1Institut für Biochemie, Universität zu Köln, Zülpicher Str. 47, 5000 Köln
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Roland Günther;
Roland Günther
1Institut für Biochemie, Universität zu Köln, Zülpicher Str. 47, 5000 Köln
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Jürgen Schweden;
Jürgen Schweden
1Institut für Biochemie, Universität zu Köln, Zülpicher Str. 47, 5000 Köln
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Ulrich Tillmann
Ulrich Tillmann
2Institut für Botanik, Bonn, Meckenheimer Allee 170
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Publisher: Portland Press Ltd
Received:
September 12 1986
Online ISSN: 1573-4935
Print ISSN: 0144-8463
© 1986 Plenum Publishing Corporation
1986
Biosci Rep (1986) 6 (9): 827–834.
Article history
Received:
September 12 1986
Citation
Ernst Bause, Roland Günther, Jürgen Schweden, Ulrich Tillmann; In vitro studies on the subcellular location of glucosidase I and glucosidase II in dog pancreas. Biosci Rep 1 September 1986; 6 (9): 827–834. doi: https://doi.org/10.1007/BF01117106
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