Characteristics of Na+-dependent hexose transport in OK, an established renal epithelial cell line.
The characteristics of Na+-dependent hexose uptake were determined for monolayers of OK, an established renal epithelial cell line derived from an opossum kidney. A comparison is made with other cultured cells, particularly LLC-PK1. The capacity to accumulate alpha-methyl D-glucoside (AMG) in OK cells develops with time, reaching a maximum level of 18 nmol/mg protein per h, 3 days after confluency. In contrast to LLC-PK1, this level is not influenced by the medium D-glucose concentration. AMG uptake in OK cells was characterized by an apparent Km of 2.9 mM and a Vmax of 17.1 nmol/mg protein per min. For Na+-dependent phlorisin binding, a KD of 0.025 microM and a Bmax of 1.5 pmol/mg protein were found. A turnover frequency of 158/s was derived from our data. The hexose carrier of OK shares with the carrier of LLC-PK1 a high level of expression, its substrate specificity and turnover frequency. It differs however with respect to the substrate binding site. The affinity for AMG and D-glucose is 3- and 10-fold lower, whereas the affinity for phlorizin is 3-times higher in OK than in LLC-PK1. The Na+ dependence of AMG uptake was also different for both cell lines and suggested for OK cells a 1:1, Na+:substrate stoichiometry. In OK cells, the phlorizin-sensitive uptake rate of D-glucose is much lower than the one for AMG. Nevertheless, D-glucose interacts with the AMG binding site in a competitive way and with an affinity similar to AMG. This could indicate a malfunction of the carrier with D-glucose as a substrate at the level of the translocation step.
Van den Bosch
L
L
Laboratorium voor Fysiologie, Katholieke Universiteit Leuven, Belgium.
De Smedt
H
H
Borghgraef
R
R
D000818:Animals; D001693:Biological Transport, Active; D002460:Cell Line; D004848:Epithelium / Q000378:metabolism; D006601:Hexoses / Q000378:metabolism*; D066298:In Vitro Techniques; D007668:Kidney / Q000378:metabolism*; D007700:Kinetics; D008757:Methylglucosides / Q000378:metabolism; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D009893:Opossums; D012964:Sodium / Q000502:physiology*; D013329:Structure-Activity Relationship
D006601:Hexoses; D008757:Methylglucosides; D009004:Monosaccharide Transport Proteins; D012964:Sodium