title: Origin, diversification and substrate specificity in the family of NCS1/FUR transporters.
abstract: NCS1 proteins are H(+)/Na(+) symporters specific for the uptake of purines, pyrimidines and related metabolites. In this article, we study the origin, diversification and substrate specificity of fungal NCS1 transporters. We show that the two fungal NCS1 sub-families, Fur and Fcy, and plant homologues originate through independent horizontal transfers from prokaryotes and that expansion by gene duplication led to the functional diversification of fungal NCS1. We characterised all Fur proteins of the model fungus Aspergillus nidulans and discovered novel functions and specificities. Homology modelling, substrate docking, molecular dynamics and systematic mutational analysis in three Fur transporters with distinct specificities identified residues critical for function and specificity, located within a major substrate binding site, in transmembrane segments TMS1, TMS3, TMS6 and TMS8. Most importantly, we predict and confirm that residues determining substrate specificity are located not only in the major substrate binding site, but also in a putative outward-facing selective gate. Our evolutionary and structure-function analysis contributes in the understanding of the molecular mechanisms underlying the functional diversification of eukaryotic NCS1 transporters, and in particular, forward the concept that selective channel-like gates might contribute to substrate specificity.
authors.affiliation: Faculty of Biology, University of Athens, Panepistimioupolis, Athens, 15784, Greece.
authors.forename: Emilia
authors.initials: E
authors.lastname: Krypotou
authors.affiliation: Faculty of Pharmacy, University of Athens, Panepistimioupolis, Athens, 15771, Greece.
authors.forename: Thomas
authors.initials: T
authors.lastname: Evangelidis
authors.affiliation: Faculty of Biology, University of Athens, Panepistimioupolis, Athens, 15784, Greece.
authors.forename: Jacob
authors.initials: J
authors.lastname: Bobonis
authors.affiliation: Bioinformatics and Genomics Programme, Centre for Genomic Regulation (CRG), Dr. Aiguader, 88, Barcelona, 08003, Spain.|Department of Experimental and Health Sciences, Universitat Pompeu Fabra (UPF), Barcelona, 08003, Spain.
authors.forename: Alexandros A
authors.initials: AA
authors.lastname: Pittis
authors.affiliation: Bioinformatics and Genomics Programme, Centre for Genomic Regulation (CRG), Dr. Aiguader, 88, Barcelona, 08003, Spain.|Department of Experimental and Health Sciences, Universitat Pompeu Fabra (UPF), Barcelona, 08003, Spain.|Institució Catalana de Recerca i Estudis Avançats (ICREA), Pg. Lluís Companys 23, Barcelona, 08010, Spain.
authors.forename: Toni
authors.initials: T
authors.lastname: Gabaldón
authors.affiliation: Department of Microbiology, Imperial College, London, SW7 2AZ, UK.|Institut de Génétique et Microbiologie, Université Paris-Sud, France.
authors.forename: Claudio
authors.initials: C
authors.lastname: Scazzocchio
authors.affiliation: Faculty of Pharmacy, University of Athens, Panepistimioupolis, Athens, 15771, Greece.
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Mikros
authors.affiliation: Faculty of Biology, University of Athens, Panepistimioupolis, Athens, 15784, Greece.
authors.forename: George
authors.initials: G
authors.lastname: Diallinas
mesh_terms: D000595:Amino Acid Sequence; D001233:Aspergillus nidulans / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D020440:Gene Duplication; D022761:Gene Transfer, Horizontal; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000145:classification / Q000235:genetics* / Q000378:metabolism*; D062105:Molecular Docking Simulation; D056004:Molecular Dynamics Simulation; D009154:Mutation; D010802:Phylogeny; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011544:Pseudogenes; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D027981:Symporters / Q000235:genetics
chemical_list: D005656:Fungal Proteins; D026901:Membrane Transport Proteins; D027981:Symporters