title: High-affinity nitrate/nitrite transporters NrtA and NrtB of Aspergillus nidulans exhibit high specificity and different inhibitor sensitivity.
abstract: The NrtA and NrtB nitrate transporters are paralogous members of the major facilitator superfamily in Aspergillus nidulans. The availability of loss-of-function mutations allowed individual investigation of the specificity and inhibitor sensitivity of both NrtA and NrtB. In this study, growth response tests were carried out at a growth-limiting concentration of nitrate (1 mM) as the sole nitrogen source, in the presence of a number of potential nitrate analogues at various concentrations, to evaluate their effect on nitrate transport. Both chlorate and chlorite inhibited fungal growth, with chlorite exerting the greater inhibition. The main transporter of nitrate, NrtA, proved to be more sensitive to chlorate than the minor transporter, NrtB. Similarly, the cation caesium was shown to exert differential effects, strongly inhibiting the activity of NrtB, but not NrtA. In contrast, no inhibition of nitrate uptake by NrtA or NrtB transporters was observed in either growth tests or uptake assays in the presence of bicarbonate, formate, malonate or oxalate (sulphite could not be tested in uptake assays owing to its reaction with nitrate), indicating significant specificity of nitrate transport. Kinetic analyses of nitrate uptake revealed that both chlorate and chlorite inhibited NrtA competitively, while these same inhibitors inhibited NrtB in a non-competitive fashion. The caesium ion appeared to inhibit NrtA in a non-competitive fashion, while NrtB was inhibited uncompetitively. The results provide further evidence of the distinctly different characteristics as well as the high specificity of nitrate uptake by these two transporters.
authors: 1​School of Biology, University of St Andrews, St Andrews, Fife, KY16 9TH, UK.
authors: Naureen
authors: N
authors: Akhtar
authors: 2​Biochemistry Laboratory, Chemistry Department, University of Ioannina, Ioannina 45110, Greece.
authors: Eugenia
authors: E
authors: Karabika
authors: 1​School of Biology, University of St Andrews, St Andrews, Fife, KY16 9TH, UK.
authors: James R
authors: JR
authors: Kinghorn
authors: 3​Department of Botany, University of British Columbia, Vancouver, British Columbia, V6T 1Z4, Canada.
authors: Anthony D M
authors: AD
authors: Glass
authors: 1​School of Biology, University of St Andrews, St Andrews, Fife, KY16 9TH, UK.
authors: Shiela E
authors: SE
authors: Unkles
authors: 4​Biotechnology and Environmental Biology, RMIT University, Melbourne, Australia.
authors: Duncan A
authors: DA
authors: Rouch
mesh_terms: D027321:Anion Transport Proteins / Q000235:genetics / Q000378:metabolism*; D000935:Antifungal Agents / Q000378:metabolism; D001233:Aspergillus nidulans / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D002586:Cesium / Q000378:metabolism; D002704:Chlorates / Q000378:metabolism; D002712:Chlorides / Q000378:metabolism; D003470:Culture Media / Q000737:chemistry; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D008826:Microbial Sensitivity Tests; D009566:Nitrates / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism*; D009584:Nitrogen / Q000378:metabolism; D013379:Substrate Specificity; D013447:Sulfites / Q000378:metabolism
chemical_list: D027321:Anion Transport Proteins; D000935:Antifungal Agents; D002704:Chlorates; D002712:Chlorides; D003470:Culture Media; D005656:Fungal Proteins; D009566:Nitrates; D009573:Nitrites; C444100:NrtA protein, Aspergillus nidulans; C444102:NrtB protein, Aspergillus nidulans; D013447:Sulfites; D002586:Cesium; D009584:Nitrogen; C001599:chlorite