TY - JOUR
T1 - The yeast Na+/H+ exchanger Nhx1 is an N-linked glycoprotein. Topological implications
AU - Wells, Karen M.
AU - Rao, Rajini
PY - 2001/2/2
Y1 - 2001/2/2
N2 - Nhx1, the endosomal Na+/H+ exchanger of Saccharomyces cerevisiae represents the founding member of a newly emerging subfamily of intracellular Na+/H+ exchangers. These proteins share significantly greater sequence homology to one another than to members of the mammalian Na+/H+ exchanger (NHE) family encoding plasma membrane Na+/H+ exchangers. Members of both subtypes are predicted to share a common organization, with an N-terminal transporter domain of transmembrane helices followed by a C-terminal hydrophilic tail. In the present study, we show that Nhx1 is an asparagine-linked glycoprotein and that the sites of glycosylation map to two residues within the C-terminal stretch of the polypeptide. This is the first evidence, to date, for glycosylation of the C-terminal region of any known NHE isoform. Importantly, the mapping of N-linked glycosylation to the C-terminal domain of Nhx1 is indicative of an unexpected membrane topology, particularly with regard to the orientation of the tail region. Although one recent study demonstrated that certain epitopes in the C-terminal domain of NHE3 were accessible from the exoplasmic side of the plasma membrane (Biemesderfer, D., DeGray, B., and Aronson, P. S. (1998) J. Biol. Chem. 273, 12391-12396), numerous other studies implicate a cytosolic disposition for the hydrophilic C-terminal tail of plasma membrane NHE isoforms. Our analysis of the glycosylation of Nhx1 is strongly indicative of residence of at least some portion of the hydrophilic tail domain within the endosomal lumen. These findings imply that the organization of the tail domain may be more complex than previously assumed.
AB - Nhx1, the endosomal Na+/H+ exchanger of Saccharomyces cerevisiae represents the founding member of a newly emerging subfamily of intracellular Na+/H+ exchangers. These proteins share significantly greater sequence homology to one another than to members of the mammalian Na+/H+ exchanger (NHE) family encoding plasma membrane Na+/H+ exchangers. Members of both subtypes are predicted to share a common organization, with an N-terminal transporter domain of transmembrane helices followed by a C-terminal hydrophilic tail. In the present study, we show that Nhx1 is an asparagine-linked glycoprotein and that the sites of glycosylation map to two residues within the C-terminal stretch of the polypeptide. This is the first evidence, to date, for glycosylation of the C-terminal region of any known NHE isoform. Importantly, the mapping of N-linked glycosylation to the C-terminal domain of Nhx1 is indicative of an unexpected membrane topology, particularly with regard to the orientation of the tail region. Although one recent study demonstrated that certain epitopes in the C-terminal domain of NHE3 were accessible from the exoplasmic side of the plasma membrane (Biemesderfer, D., DeGray, B., and Aronson, P. S. (1998) J. Biol. Chem. 273, 12391-12396), numerous other studies implicate a cytosolic disposition for the hydrophilic C-terminal tail of plasma membrane NHE isoforms. Our analysis of the glycosylation of Nhx1 is strongly indicative of residence of at least some portion of the hydrophilic tail domain within the endosomal lumen. These findings imply that the organization of the tail domain may be more complex than previously assumed.
UR - http://www.scopus.com/inward/record.url?scp=0035793559&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0035793559&partnerID=8YFLogxK
U2 - 10.1074/jbc.M001688200
DO - 10.1074/jbc.M001688200
M3 - Article
C2 - 11036065
AN - SCOPUS:0035793559
SN - 0021-9258
VL - 276
SP - 3401
EP - 3407
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 5
ER -