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Nature. 2014 Mar 6;507(7490):73-7. doi: 10.1038/nature13074. Epub 2014 Feb 26.

Crystal structure of the plant dual-affinity nitrate transporter NRT1.1.

Author information

1
Department of Pharmacology, Box 357280, University of Washington, Seattle, Washington 98195, USA.
2
Department of Physiology and Biophysics, Box 357290, University of Washington, Seattle, Washington 98195, USA.
3
1] Department of Pharmacology, Box 357280, University of Washington, Seattle, Washington 98195, USA [2] Department of Structural Biology, Genentech Inc., South San Francisco, California 94080, USA.
4
1] Department of Pharmacology, Box 357280, University of Washington, Seattle, Washington 98195, USA [2] Howard Hughes Medical Institute, Box 357280, University of Washington, Seattle, Washington 98195, USA.

Abstract

Nitrate is a primary nutrient for plant growth, but its levels in soil can fluctuate by several orders of magnitude. Previous studies have identified Arabidopsis NRT1.1 as a dual-affinity nitrate transporter that can take up nitrate over a wide range of concentrations. The mode of action of NRT1.1 is controlled by phosphorylation of a key residue, Thr 101; however, how this post-translational modification switches the transporter between two affinity states remains unclear. Here we report the crystal structure of unphosphorylated NRT1.1, which reveals an unexpected homodimer in the inward-facing conformation. In this low-affinity state, the Thr 101 phosphorylation site is embedded in a pocket immediately adjacent to the dimer interface, linking the phosphorylation status of the transporter to its oligomeric state. Using a cell-based fluorescence resonance energy transfer assay, we show that functional NRT1.1 dimerizes in the cell membrane and that the phosphomimetic mutation of Thr 101 converts the protein into a monophasic high-affinity transporter by structurally decoupling the dimer. Together with analyses of the substrate transport tunnel, our results establish a phosphorylation-controlled dimerization switch that allows NRT1.1 to uptake nitrate with two distinct affinity modes.

PMID:
24572362
PMCID:
PMC3968801
DOI:
10.1038/nature13074
[Indexed for MEDLINE]
Free PMC Article

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