Characterization of calcium transport by basolateral membrane vesicles of human small intestine.
The present studies investigated the mechanism of Ca2+ transport across basolateral membrane vesicles (BLMVs) prepared from human small intestine. Ca2+ uptake represented transport into the intravesicular space as evident by osmolality study and by the demonstration of Ca2+ efflux from the intravesicular space by Ca2+ ionophore A23187. Ca2+ uptake was stimulated by Mg2+-ATP. Kinetic parameters for ATP-dependent Ca2+ uptake revealed a Michaelis constant (Km) of 0.02 +/- 0.01 microM and a maximum rate of uptake (Vmax) of 1.00 +/- 0.03 nmol.mg protein-1.min-1.Ca2+ uptake in the absence of Mg2+ was inhibited by 75%. The Km of ATP concentration required for half-maximal Ca2+ uptake was 0.50 +/- 0.1 mM. Calmodulin (10 micrograms/ml) increased Vmax to 1.62 +/- 0.02 nmol.mg protein-1.min-1 (P less than 0.001). Km values were 0.017 +/- 0.001 microM, which was not significantly different from control values. Basolateral membranes depleted of calmodulin by EDTA osmotic shock decreased ATP-dependent Ca2+ uptake by 65%. Trifluoperazine, an anticalmodulin drug, inhibited ATP-dependent Ca2+ uptake by 50%, while no inhibition was noted in calmodulin-depleted membranes. Efflux of Ca2+ in the BLMVs was stimulated by trans-Na+. Na+-dependent Ca2+ uptake was saturable with respect to Ca2+ concentration and exhibited a Km of 0.09 +/- 0.03 microM and a Vmax of 1.08 +/- 0.01 nmol.mg protein-1.min-1. These results are consistent with the existence of a Na+-Ca2+ exchange system and ATP and Mg2+-dependent, calmodulin-regulated Ca2+, transport mechanism in BLMVs of human enterocytes.
D000255:Adenosine Triphosphate / Q000378:metabolism; D001693:Biological Transport, Active / Q000187:drug effects; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D004492:Edetic Acid / Q000494:pharmacology; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D007421:Intestine, Small / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008875:Middle Aged; D011188:Potassium / Q000494:pharmacology; D012964:Sodium / Q000494:pharmacology; D014268:Trifluoperazine / Q000494:pharmacology
D002147:Calmodulin; D014268:Trifluoperazine; D000001:Calcimycin; D000255:Adenosine Triphosphate; D004492:Edetic Acid; D012964:Sodium; D011188:Potassium; D002118:Calcium