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J Biol Chem. 2016 May 20;291(21):11385-93. doi: 10.1074/jbc.M115.704015. Epub 2016 Mar 15.

Catalytically Active Guanylyl Cyclase B Requires Endoplasmic Reticulum-mediated Glycosylation, and Mutations That Inhibit This Process Cause Dwarfism.

Author information

1
From the Departments of Biochemistry, Molecular Biology, and Biophysics and.
2
From the Departments of Biochemistry, Molecular Biology, and Biophysics and Pharmacology, University of Minnesota, Minneapolis, Minnesota 55455 potter@umn.edu.

Abstract

C-type natriuretic peptide activation of guanylyl cyclase B (GC-B), also known as natriuretic peptide receptor B or NPR2, stimulates long bone growth, and missense mutations in GC-B cause dwarfism. Four such mutants (L658F, Y708C, R776W, and G959A) bound (125)I-C-type natriuretic peptide on the surface of cells but failed to synthesize cGMP in membrane GC assays. Immunofluorescence microscopy also indicated that the mutant receptors were on the cell surface. All mutant proteins were dephosphorylated and incompletely glycosylated, but dephosphorylation did not explain the inactivation because the mutations inactivated a "constitutively phosphorylated" enzyme. Tunicamycin inhibition of glycosylation in the endoplasmic reticulum or mutation of the Asn-24 glycosylation site decreased GC activity, but neither inhibition of glycosylation in the Golgi by N-acetylglucosaminyltransferase I gene inactivation nor PNGase F deglycosylation of fully processed GC-B reduced GC activity. We conclude that endoplasmic reticulum-mediated glycosylation is required for the formation of an active catalytic, but not ligand-binding domain, and that mutations that inhibit this process cause dwarfism.

KEYWORDS:

N-linked glycosylation; cGMP; glycosylation; glycosylation inhibitor; guanylate cyclase (guanylyl cyclase); natriuretic peptide

PMID:
26980729
PMCID:
PMC4900282
DOI:
10.1074/jbc.M115.704015
[Indexed for MEDLINE]
Free PMC Article

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