The GlpT molecule has the shape (as described in the literature) of a "Mayan temple", as it has a flat, rectangular top and base. 

-Click on the surface button to see a rendering of the protein with a solid surface

Surface

-Use your mouse to rotate the protein until you can see the top and bottom, where the bottom is the larger of the two ends.

*Note that if you rotate the protein correctly, a near line of symmetry can be observed running vertically up the molecule.

This line of symmetry between the two pseudo domains is in agreement with the suggestion that MFS proteins arose by gene duplication, and the arrangement of the helix bundle in each domain suggests that gene insertion preceded the duplication.

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Turn on the a-helices.

Display a-helices

Color code:

H1 and H7

H2 and H8

H3 and H9

H4 and H10

H5 and H11

H6 and H12

The protein has 12 transmembrane a-helices  which can be divided into two similar domains by a pseudo two-fold symmetry.  This pseudo symmetry extends to all helices with H1 related to H7, H2 to H8, et cetera.

Display a-helices with fill

This space filling representation shows the helices in a space filled way.  By rotating the molecule with your mouse, you can see that the molecular pore (which extends through the center of the protein) is closed on one side and open on the other.

Display Periplasmic Residues

The periplasmic side of the molecule is flat and protrudes only slightly into the periplasm and the pore is closed.  This barrier is created by portions of H1 and H7.  The space between H1 and H7, is filled with nine aromatic side chains which help close the pore completely.

Display H1 and H7 Closing Region

Display Cytoplasmic Residues

The cytoplasmic residues include the N- and C- termini which also means that the opening of the pore is within the cytoplasm. 

Display All Helices Except H1 and H7

The pore opening can be seen by displaying all helices except H1 and H7 and rotating the molecule such that you are looking directly at the base of the molecule.  A small opening can be seen.

Display Lys₃₇₈ and Lys₃₇₉

It is believed that the cytoplasmic loop from H10-H11 plays an integral role in helix alignment when the protein is inserted into the membrane.  This is because it contains two lysine residues (Lys₃₇₈ and Lys₃₇₉) which have been shown to aid in insertion of the Glut1 protein (Sato et al.  J. Biol. Chem. 274m 24721 (1999)).

Display Both Cytoplasmic and Periplasmic Regions

With this representation, it is clearly shown how the molecule inserts itself into the membrane.  The periplasmic and cytoplasmic residues encompass the region of the protein which is membrane bound and the pore is closed.

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Now that we have a basic understanding of the GlpT Transporter structure it is important to see how the protein might facilitate the exchange of inorganic phosphate for glycerol-3-phosphate.

Display Molecular Electrostatic Surface Potential

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Because substrate binding to GlpT is mediated by the phosphate moiety, which is dibasic at physiological pH, the substrate-binding site is expected to have a positive surface electrostatic potential.  The only area in the central pore with this characteristic is at the closed end of the pore, in the middle of the membrane.  Thus, this area most likely represents the substrate binding site.

Looking at the electrostatic surface potential, it is noticed that at the innermost end of the protein the surface potential is also positive (blue color), this would be to attract the negatively charged phosphates to the transporter site.  However, one would imagine that this potential would also attract water, among other negatively charged molecules.  This is true, however the hydrophobicity of the interior funnel ensures that water and ions do not adhere to the funnel surface. 

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In other phosphate binding proteins, it has been observed that arginine residues often participate in phosphate recognition by engaging in hydrogen bonding with the oxygen atoms of phosphate.  In GlpT Arg₄₅ and Arg₂₆₉ were discovered to perform the equivalent task and GlpT shows no transport activity when Arg₄₅ and Arg₂₆₉ are replaced by Lys.  Additionally, the arginine molecules are located at approximately the same height in the membrane. 

Display Arg₄₅ and Arg₂₆₉

It is noted that Arg₄₅ and Arg₂₆₉ are embedded in H1 and H7 and by X-ray crystallography, the literature reports that the distance between then is 9.9A.  The optimal hydrogen bond length for the phosphate and Arg residues would be 2.9A, this means that the phosphate ligand must induce a conformation change in the protein to bring the Arg-Arg distance to the optimal dimension.  When this happens, the periplasmic ends of H1 and H7 separate and the cytoplasmic ends come together to close the cytoplasmic side.  This allows for the inorganic phosphate to be released and G3P to be bound and brought back into the cell.

Display Arg₄₅ and Arg₂₆₉ as situated in H1 and H7

The substrate binding sites as located on H1 and H7.

Conclusions

The GlpT protein works in a "rocker-switch" fashion which exposes the binding site, Arg₄₅ and Arg₂₆₉, to the periplasm, yielding the outward opening conformation of the molecule.  Cytoplasmic binding and periplasmic release of inorganic phosphate would allow its replacement in the substrate-binding site by G3P, which is then transferred into the cytoplasm.  Substrate binding is proposed to lower the energy barrier between the inward- and outward-facing conformations of GlpT, facilitating their interconversion and allowing the inorganic phosphate gradient to drive G3P transport.

Reference

1.  Huang, Yafei.  Lemieux, M. Joanne.  Song, Jinmei.  Auer, Manfred and Wang, Da-Neng.  Structure and Mechanism of the Glycerol-3-Phosphate Transporter from Escherichia coli.  Science.  301.  616-620  (2003).