The Mep/Amt proteins constitute a new family of transport proteins that are
ubiquitous in nature. Members from bacteria, yeast and plants have been id
entified experimentally as high-affinity ammonium transporters. We have det
ermined 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 Amt
B-LacZ fusions. These data, accompanied by an in silico analysis, indicate
that the majority of the Mep/Amt proteins contain 11 membrane-spanning heli
ces, with the N-terminus on the exterior face of the membrane and the C-ter
minus on the interior. A small subset, including E. coli AmtB, probably hav
e an additional twelfth membrane-spanning region at the N-terminus. Additio
n 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 [C-14]
-methylammonium uptake. This C-terminal region, along with four membrane-sp
anning helices, contains multiple residues that are conserved within the Me
p/Amt protein family. Structural modelling of the E. coli AmtB protein sugg
ests a number of secondary structural features that might contribute to fun
ction, including a putative ammonium binding site on the periplasmic face o
f the membrane at residue Asp-182. The implications of these results are di
scussed in relation to the structure and function of the related human Rhes
us proteins.