The Glut 1 Glucose Transporter Interacts with Calnexin and Calreticulin*
- From the School of Biological Sciences, University of Manchester, 2.205 Stopford Building, Oxford Road, Manchester M13 9PT, United Kingdom and the
- § Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110-1031
- ∥ BBSRC Advanced Research Fellow. To whom correspondence should be addressed. Tel.: 161 275 5070; Fax: 161 275 5082.
Abstract
Calnexin is an integral membrane protein that acts as a chaperone during glycoprotein folding in the endoplasmic reticulum. Cross-linking studies were carried out with the aim of investigating the interactions of calnexin with glycoproteins in vitro. A truncated version of the integral membrane glycoprotein Glut 1 (GT155) was synthesized in a rabbit reticulocyte translation system in the presence of canine pancreatic microsomes. Following immunoprecipitation with an anticalnexin antiserum, a cross-linker-independent association was observed between GT155 and calnexin. In addition, the anti-calnexin antiserum immunoprecipitated a UV-dependent cross-linking product consisting of GT155 and a protein of approximately 60 kDa designated CAP-60 (calnexin-associated protein of 60 kDa).
Both the GT155-calnexin and the GT155-CAP-60 interactions were dependent on the presence of a correctly modified oligosaccharide group on GT155, a characteristic of many calnexin interactions. A GT155 mutant that was not glycosylated (AGGT155) did not associate with calnexin or CAP-60.
Calreticulin, the soluble homologue of calnexin, was also shown to interact with GT155 only when the protein bore a correctly modified oligosaccharide group. Thus, our data show that both calnexin and calreticulin with Glut 1 in a glycosylation-dependent manner.
Footnotes
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↵‡ Supported by grants from the Biotechnology and Biological Sciences Research Council (BBSRC) and Human Frontier Science Program.
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↵¶ Supported by NIH Grant (DK43695).
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↵* The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵1 The abbreviations used are:
- ER
-
endoplasmic reticulum
- AGGT155
-
155-amino acid truncation of glucose transporter lacking N-linked glycosylation site
- BASED
-
bis[β-(4-azidosalicylamido)ethyl] disulfide
- CAP-60
-
calnexin-associated protein of 60 kDa
- DADE
-
4-4′-diazidodiphenyl ether
- dNM
-
1-deoxynojirimycin
- Endo H
-
endoglycosidase H
- Glut 1
-
glucose transporter (human erythrocyte)
- GT155
-
155-amino acid truncation of glucose transporter
- MHC
-
major histocompatibility complex
- PAGE
-
polyacrylamide gel electrophoresis
- IP
-
immunoprecipitation.
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- Received October 2, 1995.
- Revision received March 15, 1996.
- © 1996 by The American Society for Biochemistry and Molecular Biology, Inc.
