Abstract
Calnexin is an endoplasmic reticulum protein that has a role in folding newly synthesized glycoproteins. In this study, we used site-specific mutagenesis to disrupt cysteine and histidine amino acid residues in the N- and P-domains of calnexin and determined whether these mutations impact the structure and function of calnexin. We identified that disruption of the N-domain cysteines resulted in significant loss of the chaperone activity of calnexin toward the glycosylated substrate, IgY, while disruption of the P-domain cysteines only had a small impact toward IgY. We observed that wild-type calnexin as well as the P-domain double cysteine mutant contained an intramolecular disulfide bond which is lost when the N-domain cysteines are mutated. Mutation to the N-domain histidine and N-domain cysteines resulted in increased binding of ERp57. Mutations to the P-domain cysteines further enhanced ERp57 binding to calnexin. Taken together, these observations indicated that the cysteine residues within calnexin were important for the structure and function of calnexin.
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Abbreviations
- ER:
-
Endoplasmic reticulum
- CNX:
-
Calnexin
- MDH:
-
Malate dehydrogenase
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Acknowledgments
This work was supported by a grant to M.M. from the Canadian Institutes of Health Research (MOP-53050). H.C. and J.J. are supported by a studentship from the Alberta Innovates-Health Solutions.
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These authors have no conflicting interest.
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Coe, H., Schneider, J.D., Dabrowska, M. et al. Role of cysteine amino acid residues in calnexin. Mol Cell Biochem 359, 271–281 (2012). https://doi.org/10.1007/s11010-011-1021-0
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DOI: https://doi.org/10.1007/s11010-011-1021-0