Mutation and functional analysis of the Aspergillus nidulans ammonium permease MeaA and evidence for interaction with itself and MepA.
The movement of ammonium across biological membranes is mediated in both prokaryotic and eukaryotic systems by ammonium transport proteins which constitute a family of related sequences (called the AMT/MEP family). Interestingly, recent evidence suggests that human and mouse Rhesus proteins which display significant relatedness to AMT/MEP sequences may function as ammonium transporters. To add to the functional understanding of ammonium transport proteins, the sequence changes in 37 loss-of-function mutations within the Aspergillus nidulans ammonium permease gene, meaA, were characterized. Together with the identification of conserved AMT/MEP residues and regions, the mutational analysis predicted regions important for uptake activity. Specifically, a major facilitator superfamily like motif (161-GAVAERGR-168 in MeaA) may be important for the translocation of ammonium across the membrane as may the conserved Pro186 residue. A specific Gly447 to Asp mutation was introduced into MeaA and this mutant protein was found to trans-inhibit the activity of endogenous MeaA and the other A. nidulans ammonium transporter, MepA. These results suggest that MeaA may interact with itself and with MepA, although any hetero-interaction is not required for ammonium transport function. In addition, cross-feeding studies showed that MeaA and to a lesser extent MepA are also required for the retention of intracellular ammonium.
Monahan
Brendon J
BJ
Department of Genetics, University of Melbourne, Parkville, Victoria 3010, Australia.
Unkles
Shiela E
SE
Tsing I
Tchuc
T
Kinghorn
James R
JR
Hynes
Michael J
MJ
Davis
Meryl A
MA
D000483:Alleles; D000595:Amino Acid Sequence; D001233:Aspergillus nidulans / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D027682:Cation Transport Proteins; D004271:DNA, Fungal / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D005800:Genes, Fungal; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D017433:Protein Structure, Secondary; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D002352:Carrier Proteins; D027682:Cation Transport Proteins; D004271:DNA, Fungal; D005656:Fungal Proteins; C464582:MeaA protein, Aspergillus nidulans; D026901:Membrane Transport Proteins; C464583:MepA protein, Aspergillus nidulans; D000644:Quaternary Ammonium Compounds