Identification of a broad-specificity nucleoside transporter with affinity for the sugar moiety in Giardia intestinalis trophozoites.
A broad-specificity nucleoside transporter has been identified in Giardia intestinalis trophozoites, using a rapid sampling assay to measure influx of [3H]deoxycytidine, [3H]adenosine and [3H]guanosine at 0 degrees C. The influx of each labelled nucleoside was inhibited strongly by all common, naturally-occurring nucleosides but only poorly or not at all by nucleobases, indicating that the transporter recognizes structural features on the furanosyl moiety of ribo- and 2'-deoxyribonucleosides. Both 2'- and 5'-deoxyadenosine were potent inhibitors of influx (greater than 95% inhibition at 2 mM), whereas 3'-deoxyadenosine was significantly less effective (approx. 70% inhibition), and 2',3'-dideoxycytidine and cytosine arabinoside were virtually inactive (0-20% inhibition). The data reveal that the 2'- and 5'-hydroxyl groups are not necessary for the recognition of nucleosides by this transporter. However, the 3'-hydroxyl appears to be important. Michaelis-Menten constants (Km) were calculated for the influx at 0 degrees C of deoxycytidine (220 +/- 116 microM) and adenosine (45 +/- 24 microM), with respective Vmax values of 13 +/- 4 and 11 +/- 2 pmol min-1 (10(6) cells)-1. Only 12-26% of [3H]thymidine influx occurred through this transporter, the remainder entering the cells through a thymine/uracil-specific transporter described previously. Thymidine exhibited a Ki of 205 +/- 90 microM against [3H]deoxycytidine influx.
Davey
R A
RA
Department of Microbiology and Immunology, University of Adelaide, Australia.
Mayrhofer
G
G
Ey
P L
PL
D000241:Adenosine / Q000378:metabolism; D000818:Animals; D001667:Binding, Competitive; D001692:Biological Transport; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003841:Deoxycytidine / Q000378:metabolism; D005663:Furans / Q000378:metabolism; D016829:Giardia lamblia / Q000378:metabolism*; D006151:Guanosine / Q000378:metabolism; D007700:Kinetics; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D033703:Nucleoside Transport Proteins; D009705:Nucleosides / Q000378:metabolism*; D013936:Thymidine / Q000378:metabolism / Q000494:pharmacology
D002352:Carrier Proteins; D005663:Furans; D008565:Membrane Proteins; D033703:Nucleoside Transport Proteins; D009705:Nucleosides; D003841:Deoxycytidine; D006151:Guanosine; D000241:Adenosine; D013936:Thymidine