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Pore mutations in ammonium transporter AMT1 with increased electrogenic ammonium transport activity.

ABSTRACT: AMT/Mep ammonium transporters mediate high affinity ammonium/ammonia uptake in bacteria, fungi, and plants. The Arabidopsis AMT1 proteins mediate uptake of the ionic form of ammonium. AMT transport activity is controlled allosterically via a highly conserved cytosolic C terminus that interacts with neighboring subunits in a trimer. The C terminus is thus capable of modulating the conductivity of the pore. To gain insight into the underlying mechanism, pore mutants suppressing the inhibitory effect of mutations in the C-terminal trans-activation domain were characterized. AMT1;1 carrying the mutation Q57H in transmembrane helix I (TMH I) showed increased ammonium uptake but reduced capacity to take up methylammonium. To explore whether the transport mechanism was altered, the AMT1;1-Q57H mutant was expressed in Xenopus oocytes and analyzed electrophysiologically. AMT1;1-Q57H was characterized by increased ammonium-induced and reduced methylammonium-induced currents. AMT1;1-Q57H possesses a 100x lower affinity for ammonium (K(m)) and a 10-fold higher V(max) as compared with the wild type form. To test whether the trans-regulatory mechanism is conserved in archaeal homologs, AfAmt-2 from Archaeoglobus fulgidus was expressed in yeast. The transport function of AfAmt-2 also depends on trans-activation by the C terminus, and mutations in pore-residues corresponding to Q57H of AMT1;1 suppress nonfunctional AfAmt-2 mutants lacking the activating C terminus. Altogether, our data suggest that bacterial and plant AMTs use a conserved allosteric mechanism to control ammonium flux, potentially using a gating mechanism that limits flux to protect against ammonium toxicity.

SUBMITTER: Loque D

PROVIDER: S-EPMC2757203 | biostudies-literature | 2009 Sep

REPOSITORIES: biostudies-literature

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Pore mutations in ammonium transporter AMT1 with increased electrogenic ammonium transport activity.

Loqué Dominique D Mora Silvia I SI Andrade Susana L A SL Pantoja Omar O Frommer Wolf B WB

The Journal of biological chemistry 20090706 37

AMT/Mep ammonium transporters mediate high affinity ammonium/ammonia uptake in bacteria, fungi, and plants. The Arabidopsis AMT1 proteins mediate uptake of the ionic form of ammonium. AMT transport activity is controlled allosterically via a highly conserved cytosolic C terminus that interacts with neighboring subunits in a trimer. The C terminus is thus capable of modulating the conductivity of the pore. To gain insight into the underlying mechanism, pore mutants suppressing the inhibitory effe ...[more]

PMID: 19581303

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Project description:BACKGROUND:Cyanobacteria account for 20-30% of Earth's primary photosynthetic productivity and convert solar energy into biomass-stored chemical energy at the rate of approximately 450 TW [1]. These single-cell microorganisms are resilient predecessors of all higher oxygenic phototrophs and can be found in self-sustaining, nitrogen-fixing communities the world over, from Antarctic glaciers to the Sahara desert [2]. METHODOLOGY/PRINCIPAL FINDINGS:Here we show that diverse genera of cyanobacteria including biofilm-forming and pelagic strains have a conserved light-dependent electrogenic activity, i.e. the ability to transfer electrons to their surroundings in response to illumination. Naturally-growing biofilm-forming photosynthetic consortia also displayed light-dependent electrogenic activity, demonstrating that this phenomenon is not limited to individual cultures. Treatment with site-specific inhibitors revealed the electrons originate at the photosynthetic electron transfer chain (P-ETC). Moreover, electrogenic activity was observed upon illumination only with blue or red but not green light confirming that P-ETC is the source of electrons. The yield of electrons harvested by extracellular electron acceptor to photons available for photosynthesis ranged from 0.05% to 0.3%, although the efficiency of electron harvesting likely varies depending on terminal electron acceptor. CONCLUSIONS/SIGNIFICANCE:The current study illustrates that cyanobacterial electrogenic activity is an important microbiological conduit of solar energy into the biosphere. The mechanism responsible for electrogenic activity in cyanobacteria appears to be fundamentally different from the one exploited in previously discovered electrogenic bacteria, such as Geobacter, where electrons are derived from oxidation of organic compounds and transported via a respiratory electron transfer chain (R-ETC) [3], [4]. The electrogenic pathway of cyanobacteria might be exploited to develop light-sensitive devices or future technologies that convert solar energy into limited amounts of electricity in a self-sustainable, CO(2)-free manner.

Dataset Information

Pore mutations in ammonium transporter AMT1 with increased electrogenic ammonium transport activity.

Publications

Pore mutations in ammonium transporter AMT1 with increased electrogenic ammonium transport activity.

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