Journal
JOURNAL OF BIOLOGICAL CHEMISTRY
Volume 284, Issue 6, Pages 3433-3444Publisher
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M804866200
Keywords
-
Categories
Funding
- Canadian Institutes of Health Research
- Canadian Cancer Society
Ask authors/readers for more resources
Calnexin is a Ca2+-binding transmembrane chaperone of the endoplasmic reticulum that recognizes Glc(1)Man(5-9)GlcNAc(2) oligosaccharides on folding glycoproteins as well as non-native elements of the polypeptide backbone. This latter mode of recognition enables calnexin to suppress the aggregation of both glycosylated and nonglycosylated substrates. The luminal portion of calnexin (S-Cnx) consists of two domains, a globular lectin domain and an extended arm domain. To understand the function of these domains during the interaction of calnexin with non-native protein conformers, we tested deletion mutants of S-Cnx for their abilities to suppress the aggregation of nonglycosylated firefly luciferase. The arm domain alone exhibited no capacity to suppress aggregation. However, stepwise truncation of the arm domain in S-Cnx resulted in a progressive reduction in aggregation suppression potency to the point where the globular domain alone exhibited 25% potency. To characterize the polypeptide-binding site, we used hydrophobic peptides that were competitors of the ability of S-Cnx to suppress luciferase aggregation. Direct binding experiments revealed a single site of peptide binding in the globular domain (K-d = 0.9 mu M) at a location distinct from the lectin site. Progressive truncation of the arm domain in S-Cnx had no effect on the binding of small peptides but reduced the binding affinity of S-Cnx for large, non-native protein substrates. Because protein substrates exhibited no binding to the isolated arm domain, our findings support a model in which calnexin suppresses aggregation through a polypeptide-binding site in its globular domain, with the arm domain enhancing aggregation suppression by sterically constraining large substrates.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available