Functional characterization of a H+/nucleoside co-transporter (CaCNT) from Candida albicans, a fungal member of the concentrative nucleoside transporter (CNT) family of membrane proteins.
Human and other mammalian concentrative (Na(+)-linked) nucleoside transport proteins belong to a membrane protein family (CNT, TC 2.A.41) that also includes Escherichia coli H(+)-dependent nucleoside transport protein NupC. Here, we report the cDNA cloning and functional characterization of a CNT family member from the pathogenic yeast Candida albicans. This 608 amino acid residue H(+)/nucleoside symporter, designated CaCNT, contains 13 predicted transmembrane domains (TMs), but lacks the exofacial, glycosylated carboxyl-terminus of its mammalian counterparts. When produced in Xenopus oocytes, CaCNT exhibited transport activity for adenosine, uridine, inosine and guanosine but not cytidine, thymidine or the nucleobase hypoxanthine. Apparent K(m) values were in the range 16-64 micro M, with V(max) : K(m) ratios of 0.58-1.31. CaCNT also accepted purine and uridine analogue nucleoside drugs as permeants, including cordycepin (3'-deoxyadenosine), a nucleoside analogue with anti-fungal activity. Electrophysiological measurements under voltage clamp conditions gave a H(+) to [(14)C]uridine coupling ratio of 1 : 1. CaCNT, obtained from logarithmically growing cells, is the first described cation-coupled nucleoside transporter in yeast, and the first member of the CNT family of proteins to be characterized from a unicellular eukaryotic organism.
Loewen
Shaun K
SK
Department of Physiology, University of Alberta, Edmonton, Alberta T6G 2H7, Canada.
Ng
Amy M L
AM
Mohabir
Nadira N
NN
Baldwin
Stephen A
SA
Cass
Carol E
CE
Young
James D
JD
D000595:Amino Acid Sequence; D000818:Animals; D000935:Antifungal Agents / Q000378:metabolism; D001483:Base Sequence; D002176:Candida albicans / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D003839:Deoxyadenosines / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008564:Membrane Potentials / Q000502:physiology; D026901:Membrane Transport Proteins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D033703:Nucleoside Transport Proteins / Q000235:genetics* / Q000378:metabolism; D009705:Nucleosides / Q000378:metabolism; D018408:Patch-Clamp Techniques; D016133:Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D014981:Xenopus / Q000378:metabolism
D000935:Antifungal Agents; D004271:DNA, Fungal; D003839:Deoxyadenosines; D005656:Fungal Proteins; D026901:Membrane Transport Proteins; D033703:Nucleoside Transport Proteins; D009705:Nucleosides; C065171:cif nucleoside transporter; C058120:cordycepin