Role of glycosylation in the renal electrogenic Na+-HCO3 - cotransporter (NBCe1)

Inyeong Choi, Lihui Hu, Jose Rojas, Bernhard M. Schmitt, Walter F. Boron

Research output: Contribution to journalArticle

34 Citations (Scopus)

Abstract

The electrogenic Na+-HCO3 - cotransporter NBCe1 is important for the regulation of intracellular pH (pHi) and for epithelial HCO3 - transport in many tissues, including kidney, pancreas, and brain. In the present study, we investigate glycosylation sites in NBCe1. Treatment of rat kidney membrane extracts with peptide N-glycosidase F (PNGase F) shifted the apparent molecular weight (MW) of NBCe1 from 130 to 116, the MW predicted from the deduced amino acid sequence. Treatment with endoglycosidase F2 or H or O-glycosidase did not affect the MW of NBCe1. Lectin-binding studies, together with the enzyme data, suggest that the N-linked carbohydrates are of tri- or tetra-antennary type. To localize glycosylation sites, we individually mutated the seven consensus N-glycosylation sites by replacing asparagine (N) with glutamine (Q) and assessing mutant transporters in Xenopus laevis oocytes. Immunoblotting of oocyte membrane extracts treated with PNGase F indicates that NBCe1 is normally glycosylated at N597 and N617 (both on the third extracellular loop). However, N592 (on the same loop) is glycosylated when the other two sites are mutated. The triple mutant (N592Q/N597Q/N617Q) is completely unglycosylated but, based on microelectrode measurements of membrane potential and pHi in oocytes, preserves the Na+ and HCO3 - dependence and electrogenicity of wild-type NBCe1.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Renal Physiology
Volume284
Issue number6 53-6
StatePublished - Jun 1 2003
Externally publishedYes

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Glycosylation
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase
Oocytes
Molecular Weight
Glycoside Hydrolases
Kidney
Membranes
Asparagine
Xenopus laevis
Microelectrodes
Glutamine
Immunoblotting
Lectins
Membrane Potentials
Pancreas
Amino Acid Sequence
Carbohydrates
Brain
Enzymes

Keywords

  • Acid-base mechanism
  • PH measurement
  • Transporter

ASJC Scopus subject areas

  • Physiology

Cite this

Role of glycosylation in the renal electrogenic Na+-HCO3 - cotransporter (NBCe1). / Choi, Inyeong; Hu, Lihui; Rojas, Jose; Schmitt, Bernhard M.; Boron, Walter F.

In: American Journal of Physiology - Renal Physiology, Vol. 284, No. 6 53-6, 01.06.2003.

Research output: Contribution to journalArticle

Choi, Inyeong ; Hu, Lihui ; Rojas, Jose ; Schmitt, Bernhard M. ; Boron, Walter F. / Role of glycosylation in the renal electrogenic Na+-HCO3 - cotransporter (NBCe1). In: American Journal of Physiology - Renal Physiology. 2003 ; Vol. 284, No. 6 53-6.
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