title: Comparative kinetic analysis of AzgA and Fcy21p, prototypes of the two major fungal hypoxanthine-adenine-guanine transporter families.
abstract: In fungi, uptake of salvageable purines is carried out by members of two evolutionarily distinct protein families, the Purine-Related Transporters (PRT/NCS1) and the AzgA-like Transporters. We carried out a comparative kinetic analysis of two prototypes of these transporter families. The first was Fcy21p, a herein characterized protein of Candida albicans, and the second was AzgA, a transporter of Aspergillus nidulans. Our results showed that: (i) AzgA and Fcy21p are equally efficient high-affinity, high-capacity, purine transporters, (ii) Fcy21p, but not AzgA, is an efficient cytosine and 5-fluorocytosine transporter, interacting with =O2 and C4-NH2 of the pyrimidine ring, (iii) the major interactions of AzgA and Fcy21p with the purine ring are similar, but not identical, involving in all cases positions 6 and 7, and for some substrates, positions 1 and 9 as well, and (iv) in AzgA, bulky groups at position N3 have a detrimental steric effect on substrate binding, while similar substitutions at C2 or N9 are fully or partially tolerated. In contrast, in Fcy21p, C2 and N9 bulky substitutions abolish substrate binding, while similar substitutions in N3 are fully tolerated. These results suggest that all fungal purine transporters might have evolved from a single ancestral protein, and show that fungal transporters use different substrate interactions compared to the analogous protozoan or mammalian proteins. Finally, results are also discussed in respect of the possibility of using fungal purine transporters as specific gateways for the development of targeted antifungal pharmacological therapies.
authors.affiliation: Faculty of Biology, Department of Botany, University of Athens, Panepistimioupolis, Athens, Greece.
authors.forename: Sophia
authors.initials: S
authors.lastname: Goudela
authors.forename: Helen
authors.initials: H
authors.lastname: Tsilivi
authors.forename: George
authors.initials: G
authors.lastname: Diallinas
mesh_terms: D000225:Adenine / Q000378:metabolism; D001233:Aspergillus nidulans / Q000378:metabolism*; D001692:Biological Transport; D002176:Candida albicans / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006147:Guanine / Q000378:metabolism; D019271:Hypoxanthine / Q000378:metabolism; D007700:Kinetics; D011687:Purines / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D002352:Carrier Proteins; D005656:Fungal Proteins; D011687:Purines; D019271:Hypoxanthine; D006147:Guanine; D000225:Adenine