A genetic system based on split-ubiquitin for the analysis of interactions between membrane proteins in vivo.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 95 ·No. 9 ·1998-04-28 ·Pages 5187-92

Stagljar I, Korostensky C, Johnsson N, te Heesen S

Abstract

A detection system for interactions between membrane proteins in vivo is described. The system is based on split-ubiquitin [Johnsson, N. & Varshavsky, A. (1994) Proc. Natl. Acad. Sci. USA 91, 10340-10344]. Interaction between two membrane proteins is detected by proteolytic cleavage of a protein fusion. The cleavage releases a transcription factor, which activates reporter genes in the nucleus. As a result, interaction between membrane proteins can be analyzed by the means of a colorimetric assay. We use membrane proteins of the endoplasmic reticulum as a model system. Wbp1p and Ost1p are both subunits of the oligosaccharyl transferase membrane protein complex. The Alg5 protein also localizes to the membrane of the endoplasmic reticulum, but does not interact with the oligosaccharyltransferase. Specific interactions are detected between Wbp1p and Ost1p, but not between Wbp1p and Alg5p. The new system might be useful as a genetic and biochemical tool for the analysis of interactions between membrane proteins in vivo.

MeSH Terms

Cloning, Molecular/methods Endopeptidases/metabolism Genetic Vectors Herpes Simplex Virus Protein Vmw65/metabolism Hexosyltransferases Membrane Proteins/chemistry Protein Binding Recombinant Fusion Proteins/metabolism Saccharomyces cerevisiae Transferases/metabolism Ubiquitin-Specific Proteases Ubiquitins/chemistry

Chemicals

Herpes Simplex Virus Protein Vmw65 Membrane Proteins Recombinant Fusion Proteins Ubiquitins Transferases Hexosyltransferases dolichyl-diphosphooligosaccharide - protein glycotransferase Endopeptidases Ubiquitin-Specific Proteases

Authors & Affiliations

4 authors, click to expand affiliations / ORCID

Stagljar I

Institute of Veterinary Biochemistry, University Zürich-Irchel, Winterthurerstrasse 190, CH-8057 Zurich, Switzerland.

Korostensky C

Johnsson N

te Heesen S

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21 references, click to expand

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Article Info

Journal

Proceedings of the National Academy of Sciences of the United States of America

Abbr.

Proc Natl Acad Sci U S A

ISSN

0027-8424

DOI

10.1073/pnas.95.9.5187

Published

1998-04-28

Pages

5187-92

Language

English

Region

United States

NLM ID

7505876

PMCID

PMC20236

Subset

External Links

PubMed source DOI: 10.1073/pnas.95.9.5187 PMC Full Text View journal details

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A genetic system based on split-ubiquitin for the analysis of interactions between membrane proteins in vivo.

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