Complex oligosaccharides are N-linked to Kv3 voltage-gated K+ channels in rat brain.
Neuronal Kv3 voltage-gated K(+) channels have two absolutely conserved N-glycosylation sites. Here, it is shown that Kv3.1, 3.3, and 3.4 channels are N-glycosylated in rat brain. Digestion of total brain membranes with peptide N glycosidase F (PNGase F) produced faster migrating immunobands than those of undigested membranes. Additionally, partial PNGase F digests showed that both sites are occupied by oligosaccharides. Neuraminidase treatment produced a smaller immunoband shift relative to PNGase F treatment. These results indicate that both sites are highly available and occupied by N-linked oligosaccharides for Kv3.1, 3.3, and 3.4 in rat brain, and furthermore that at least one oligosaccharide is of complex type. Additionally, these results point to an extracytoplasmic S1-S2 linker in Kv3 proteins expressed in native membranes. We suggest that N-glycosylation processing of Kv3 channels is critical for the expression of K(+) currents at the surface of neurons, and perhaps contributes to the pathophysiology of congenital disorders of glycosylation.
Cartwright
Tara A
TA
Department of Biochemistry and Molecular Biology, Brody School of Medicine at East Carolina University, 600 Moye Boulevard, Greenville, NC 27834, USA.
Corey
Melissa J
MJ
Schwalbe
Ruth A
RA
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D002236:Carbohydrate Conformation; D002462:Cell Membrane / Q000378:metabolism; D005260:Female; D006031:Glycosylation; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase / Q000378:metabolism; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009439:Neuraminidase / Q000378:metabolism; D009474:Neurons / Q000378:metabolism; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism*; D024642:Potassium Channels, Voltage-Gated / Q000378:metabolism; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D051381:Rats; D051670:Shaw Potassium Channels / Q000737:chemistry / Q000378:metabolism*
C098254:Kcnc1 protein, rat; C484184:Kcnc3 protein, rat; C492014:Kcnc4 protein, rat; D009419:Nerve Tissue Proteins; D009844:Oligosaccharides; D024642:Potassium Channels, Voltage-Gated; D051670:Shaw Potassium Channels; D009439:Neuraminidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase