Ammonia transporter

Structure

The structure of the ammonia channel from E. coli, was, at the time of its publication, the highest resolution structure of any integral membrane protein. It shows a trimer of subunits, each made up of 11 transmembrane segments (TMSs) and containing a pseudo two-fold symmetry. Each monomer contains a hydrophobic ammonia conducting channel.

While prokaryotic ammonia channel proteins have an N-terminal region which acts as a signal sequence and is cleaved in the mature protein, the Rhesus glycoproteins retain this as a 12th transmembrane helix in the mature protein.

Substrate specificity

Most functionally characterized members of the family are ammonium uptake transporters. Some, but not other Amt proteins also transport methylammonium. Detailed phylogenetic analyses of plant homologues have been published. In E. coli, NH4+, rather than NH3, may be the substrate of AmtB, but controversy still exists. If NH4+ is transported, K+ possibly serves as a counter ion in an antiport process with K+, and that one histidine removes a proton off of NH4+ to yield NH3.

Transport reaction

The generalized transport reaction catalyzed by members of the Amt family are suggested to be: :NH4+ (out) ⇌ NH4+ (in)

Mechanism

The X-ray structures have revealed that the pore of the Amt and Rh proteins is characterized by a hydrophobic portion about 12 Å long, in which electronic density was observed in the crystallographic study of AmtB from Escherichia coli. This electronic density was initially only observed when crystals were grown in the presence of ammonium, and was thus attributed to ammonia molecules. The Amt/Rh protein mechanism might involve the single-file diffusion of NH3 molecules. However, the pore could also be filled with water molecules. The possible presence of water molecules in the pore lumen calls for a reassessment of the notion that Amt/Rh proteins work as plain NH3 channels. Indeed, functional experiments on plant ammonium transporters and Rh proteins suggest a variety of permeation mechanisms including the passive diffusion of NH3, the antiport of NH4+/H+, the transport of NH4+, or the cotransport of NH3/H+. Lamoureux et al. discuss these mechanisms in light of functional and simulation studies on the AmtB transporter.

Regulation

In E. coli the AmtB gene is expressed only under limiting nitrogen levels to yield the AmtB protein. It is co-expressed with the GlnK gene which encodes a PII protein. This protein is also trimeric and remains in the cytoplasm. It is covalently modified by a U/U deuridylylated group at Y51. The hydrolyzed product, adenosine 5'-diphosphate, orients the surface of GlnK for AmtB blockade. When nitrogen levels outside the cell rise, the ammonia channel must be deactivated to prevent excessive ammonia entering the cell (where ammonia would be combined with glutamate to make glutamine, utilizing ATP and thereby depleting the cell's ATP reserves). This deactivation is achieved by deuridylylation of the GlnK protein which then binds to the cytoplasmic face of AmtB and inserts a loop into the ammonia conducting pore. At the tip of this loop is an arginine residue which sterically blocks the channel.

Human ammonia transporter-related proteins

RHAG, RHBG, RHCE, RHCG, RHD

References

References

1. (Feb 2004). "Non-erythroid Rh glycoproteins: a putative new family of mammalian ammonium transporters". Pflügers Archiv.
2. (Sep 2004). "Mechanism of ammonia transport by Amt/MEP/Rh: structure of AmtB at 1.35 A". Science.
3. (Dec 2004). "The mechanism of ammonia transport based on the crystal structure of AmtB of Escherichia coli". Proceedings of the National Academy of Sciences of the United States of America.
4. (Dec 2006). "The Amt/MEP/Rh family: structure of AmtB and the mechanism of ammonia gas conduction". Physiology.
5. (November 2013}} that the [[erythrocyte]] Rh complex is a heterotrimer of RhAG, [[RHD (gene)). "Functional reconstitution into liposomes of purified human RhCG ammonia channel". PLOS ONE.
6. (Jan 2011). "The rhesus protein RhCG: a new perspective in ammonium transport and distal urinary acidification". Kidney International.
7. (Nov 2008). "A role for Rhesus factor Rhcg in renal ammonium excretion and male fertility". Nature.
8. (Dec 2004). "Electron and atomic force microscopy of the trimeric ammonium transporter AmtB". EMBO Reports.
9. (May 2006). "The ammonia channel protein AmtB from Escherichia coli is a polytopic membrane protein with a cleavable signal peptide". FEMS Microbiology Letters.
10. (2002-05-28). "Rhesus expression in a green alga is regulated by CO(2)". Proceedings of the National Academy of Sciences of the United States of America.
11. (2009-03-31). "Relative CO2/NH3 selectivities of AQP1, AQP4, AQP5, AmtB, and RhAG". Proceedings of the National Academy of Sciences of the United States of America.
12. (2007-12-01). "The Amt/Mep/Rh family of ammonium transport proteins". Molecular Membrane Biology.
13. (2014-01-01). "Evolutionary classification of ammonium, nitrate, and peptide transporters in land plants". BMC Evolutionary Biology.
14. (2007-11-20). "The W148L substitution in the Escherichia coli ammonium channel AmtB increases flux and indicates that the substrate is an ion". Proceedings of the National Academy of Sciences of the United States of America.
15. (2007-02-07). "Protonation states of ammonia/ammonium in the hydrophobic pore of ammonia transporter protein AmtB". Journal of the American Chemical Society.
16. (2006-12-22). "An unusual twin-his arrangement in the pore of ammonia channels is essential for substrate conductance". The Journal of Biological Chemistry.
17. (2010-09-01). "Transport mechanisms in the ammonium transporter family". Transfusion Clinique et Biologique.
18. (Oct 2006). "In vitro analysis of the Escherichia coli AmtB-GlnK complex reveals a stoichiometric interaction and sensitivity to ATP and 2-oxoglutarate". The Journal of Biological Chemistry.
19. (Jan 2007). "The crystal structure of the Escherichia coli AmtB-GlnK complex reveals how GlnK regulates the ammonia channel". Proceedings of the National Academy of Sciences of the United States of America.