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- Title
Membrane topology of the Mep/Amt family of ammonium transporters.
- Authors
Thomas, G H; Mullins, J G; Merrick, M
- Abstract
The Mep/Amt proteins constitute a new family of transport proteins that are ubiquitous in nature. Members from bacteria, yeast and plants have been identified experimentally as high-affinity ammonium transporters. We have determined the topology of AmtB, a Mep/Amt protein from Escherichia coli, as a representative protein for the complete family. This was established using a minimal set of AmtB-PhoA fusion proteins with a complementary set of AmtB-LacZ fusions. These data, accompanied by an in silico analysis, indicate that the majority of the Mep/Amt proteins contain 11 membrane-spanning helices, with the N-terminus on the exterior face of the membrane and the C-terminus on the interior. A small subset, including E. coli AmtB, probably have an additional twelfth membrane-spanning region at the N-terminus. Addition of PhoA or LacZ alpha-peptide to the C-terminus of E. coli AmtB resulted in complete loss of transport activity, as judged by measurements of [14C]-methylammonium uptake. This C-terminal region, along with four membrane-spanning helices, contains multiple residues that are conserved within the Mep/Amt protein family. Structural modelling of the E. coli AmtB protein suggests a number of secondary structural features that might contribute to function, including a putative ammonium binding site on the periplasmic face of the membrane at residue Asp-182. The implications of these results are discussed in relation to the structure and function of the related human Rhesus proteins.
- Publication
Molecular microbiology, 2000, Vol 37, Issue 2, p331
- ISSN
0950-382X
- Publication type
Journal Article
- DOI
10.1046/j.1365-2958.2000.01994.x