Characterization of amino acid transport systems in human placental brush-border membrane vesicles.
Brush-border microvillous plasma membrane vesicles were prepared from human full-term placental syncytiotrophoblasts and purified 33-fold from the homogenate with reference to a membrane marker enzyme, alkaline phosphatase (EC 3.1.3.1). Transport of alpha-(methylamino)isobutyrate by the membrane vesicles was stimulated in the presence of an Na+ gradient from the outside to the inside of the vesicles. The initial rate of uptake in a 10-s period was enhanced with increasing concentration of Na+ in the external medium. The level of alpha-(methylamino)isobutyrate transported into the vesicles reached a maximum 1 min after the start of incubation at 37 degrees C, and then decreased with time due to efflux. Extrapolation to infinite medium osmolarity showed no uptake, indicating transport of alpha-(methylamino)isobutyrate into membrane vesicles. The initial rate of uptake was dependent on temperature and pH: the highest rate occurred at 37 degrees C and the optimal pH was 8.0. When the alpha-(methylamino)isobutyrate concentration was varied, the initial rate of uptake dependent on an Na+ gradient (out greater than in) obeyed Michaelis-Menten kinetics with Km and Vmax values of 1.07 mM and 3.23 nmol/10 s per mg of protein, respectively. Cross-inhibition patterns indicated that at least three Na+-dependent and two Na+-independent carrier-mediated pathways existed in the human placental brush border. One Na+-dependent pathway interacted with all substrates tested. Another Na+-dependent route interacted with L-proline, alpha-(methylamino)isobutyrate, and L-methionine, while a third pathway was selective for L-methionine. One Na+-independent pathway was selective for L-cysteine, while the other pathway interacted with all substrates tested.
Kudo
Y
Y
Department of Biochemistry, Hiroshima University School of Medicine, Japan.
Yamada
K
K
Fujiwara
A
A
Kawasaki
T
T
D000596:Amino Acids / Q000378:metabolism* / Q000494:pharmacology; D000621:Aminoisobutyric Acids / Q000378:metabolism*; D000838:Anions; D001692:Biological Transport; D002412:Cations; D002458:Cell Fractionation; D005260:Female; D006801:Humans; D007700:Kinetics; D008871:Microvilli / Q000378:metabolism* / Q000648:ultrastructure; D008954:Models, Biological; D010920:Placenta / Q000378:metabolism*
D000596:Amino Acids; D000621:Aminoisobutyric Acids; D000838:Anions; D002412:Cations; C017911:2-(methylamino)isobutyric acid