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Am J Physiol Cell Physiol 277: C1111-C1121, 1999;
0363-6143/99 $5.00
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Vol. 277, Issue 6, C1111-C1121, December 1999

EDITORIAL FOCUS
Na+/H+ exchangers (NHE1-3) have similar turnover numbers but different percentages on the cell surface

Megan E. Cavet, Shafinaz Akhter, Fermin Sanchez de Medina, Mark Donowitz, and Chung-Ming Tse

Gastrointestinal Division, Department of Medicine, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205-2195

NHE1, NHE2, and NHE3 are well-characterized cloned members of the mammalian Na+/H+ exchanger (NHE) gene family. Given the specialized function and regulation of NHE1, NHE2, and NHE3, we compared basal turnover numbers of NHE1, NHE2, and NHE3 measured in the same cell system: PS120 fibroblasts lacking endogenous NHEs. NHE1, NHE2, and NHE3 were epitope tagged with vesicular stomatitis virus glycoprotein (VSVG). The following characteristics were determined on the same passage of cells transfected with NHE1V, NHE2V, or NHE3V: 1) maximal reaction velocity (Vmax) by 22Na+ uptake and fluorometery, 2) total amount of NHE protein by quantitative Western analysis with internal standards of VSVG-tagged maltose-binding protein, and 3) cell surface expression by cell surface biotinylation. Cell surface expression (percentage of total NHE) was 88.8 ± 3.5, 64.6 ± 3.3, 20.0 ± 2.6, and 14.0 ± 1.3 for NHE1V, 85- and 75-kDa NHE2V, and NHE3V, respectively. Despite these divergent cell surface expression levels, turnover numbers for NHE1, NHE2, and NHE3 were similar (80.3 ± 9.6, 92.1 ± 8.6, and 99.2 ± 9.1 s-1, when Vmax was determined using 22Na uptake at 22°C and 742 ± 47, 459 ± 16, and 609 ± 39 s-1 when Vmax was determined using fluorometry at 37°C). These data indicate that, in the same cell system, intrinsic properties that determine turnover number are conserved among NHE1, NHE2, and NHE3.

sodium/hydrogen antiporter; quantitative Western analysis; cell surface biotinylation; PS120 fibroblasts


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