High-affinity phlorizin binding in Mytilus gill.
The gill of the marine mussel, Mytilus, contains a high affinity, Na-dependent D-glucose transporter capable of accumulating glucose directly from sea water. We examined the ability of the beta-glucoside, phlorizin, to act as a high-affinity ligand of this process in intact gills and isolated brush border membrane vesicles (BBMV). The time course of association of nanomolar [3H]phlorizin to gills and BBMV was slow, with t50 values between 10 and 30 min, and a half-time for dissociation of approx. 30 min. 1 mM D-glucose reduced equilibrium binding of 1 nM phlorizin by 90-95%, indicating that there was little non-specific binding of this ligand to the gill. In addition, there was little, if any, hydrolysis by the gill of phlorizin to its constituents, glucose and phloretin. Phlorizin binding to gills and BBMV was significantly inhibited by the addition of 50 microM concentrations of D-glucose and alpha-methyl-D-glucose, and unaffected by the addition of L-glucose and fructose. Binding to gills and BBMV was reduced by greater than 90% when Na+ was replaced by K+. Replacement of Na+ by Li+ effectively blocked binding to the intact gill, although Li+ did support a limited amount of glucose-specific phlorizin binding in BBMV. The Kd values for glucose-specific phlorizin binding in intact gills and BBMV were 0.5 nM and 6 nM, respectively. We conclude that phlorizin binds with extremely high affinity to the Na-dependent glucose transporter of Mytilus gill, which may be useful in future efforts to isolate and purify the protein(s) involved in integumental glucose transport.
Wright
S H
SH
Department of Physiology, College of Medicine, University of Arizona, Tucson 85724.
Pajor
A M
AM
Moon
D A
DA
Wunz
T M
TM
D000818:Animals; D049872:Bivalvia; D050260:Carbohydrate Metabolism; D005880:Gills / Q000378:metabolism* / Q000648:ultrastructure; D005947:Glucose / Q000378:metabolism; D007700:Kinetics; D008094:Lithium / Q000378:metabolism; D008871:Microvilli / Q000378:metabolism; D010695:Phlorhizin / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011188:Potassium / Q000378:metabolism; D012964:Sodium / Q000378:metabolism
D008094:Lithium; D012964:Sodium; D010695:Phlorhizin; D005947:Glucose; D011188:Potassium