Journal: Nature Communications

Article Title: Structural basis of phosphate export by human XPR1

doi: 10.1038/s41467-025-55995-8

Figure Lengend Snippet: a , b Electron density maps ( a ) and cartoon representations ( b ) of XPR1 in two views. c Expanded view of the dimer interface of XPR1 (the region boxed in b ), highlighting key residues involved in dimer formation. d Disruption of XPR1 dimer by mutation of single residues in the dimer interface as shown by SEC. SEC was repeated twice with similar results. Approximate elution volumes for protein standards with known molecular weight were marked according to the manual provided by the manufacturer. e Kinetics of Pi export in HCT116 XPR1 KO cells expressing XPR1 WT or dimer interface mutants (F235G, L239G). Pi released in culture medium was quantified by malachite green phosphate assay and normalized to the total amount of proteins. Data are means ± SD from a representative experiment ( n = 3). f Relative Pi export level in cells expressing dimer interface mutants as compared with Pi export level measured in XPR1 WT cells at time point of 30 min. Data are means ± SD from n = 6 independent experiments. One-way ANOVA with Dunnett’s multiple comparisons test; ****, p ≤ 0.0001.

Article Snippet: For endogenous XPR1 expression, anti XPR1 rabbit polyclonal antibody from proteintech (14174-1-AP) was used at 1:1000 dilution, followed by Horseradish Peroxidase (HRP)-conjugated anti rabbit antibody (1:2000).

Techniques: Disruption, Mutagenesis, Molecular Weight, Expressing

Journal: Nature Communications

Article Title: Structural basis of phosphate export by human XPR1

doi: 10.1038/s41467-025-55995-8

Figure Lengend Snippet: a Protomer structure of XPR1 shown in two views, which can be separated into a core domain and a scaffold domain, with the core domain showing similarity to ion-translocating rhodopsin. Bound lipid is shown as red sticks. b Structure of the core domain of XPR1 in two views. c Structure of Cl - -translocating rhodopsin (pdb# 5b2n) protomer in two views. d Overlay of structures of the core domain of XPR1 and Cl - -translocating rhodopsin in two views. Bound Pi in XPR1 and bound Cl - in Cl - -translocating rhodopsin are shown as red and magenta spheres respectively. The Root mean square deviation between Cα atoms of XPR1 core domain and Cl - -translocating rhodopsin is ~4 Å.

Article Snippet: For endogenous XPR1 expression, anti XPR1 rabbit polyclonal antibody from proteintech (14174-1-AP) was used at 1:1000 dilution, followed by Horseradish Peroxidase (HRP)-conjugated anti rabbit antibody (1:2000).

Techniques:

Journal: Nature Communications

Article Title: Structural basis of phosphate export by human XPR1

doi: 10.1038/s41467-025-55995-8

Figure Lengend Snippet: a , b Pi translocation pathway in the core domain of XPR1, as calculated by Caver, showing EV ( a ) and IV ( b ). Red asterisk indicates the position of Pi binding site. c Distribution of positively-charged residues (shown as sticks) along the Pi translocation pathway, with those positioned in the entrance of EV and IV colored green, and those surrounding Pi binding site colored yellow. d Expanded view of the bound Pi and its interaction with the surrounding residues. Two views are shown. In the lower panel, densities for the bound Pi and side chains of its coordinating residues were shown as green and blue meshes respectively (contoured at 3 Å). e Time-dependent r.m.s.d. of bound Pi during the 200-ns simulations. For two bound Pi in an XPR1 dimer (one in each protomer), the r.m.s.d. values of three replicates are overlaid and plotted separately. f Relative Pi export level in cells expressing Pi coordinating residues mutants compared with Pi export level measured in XPR1 WT cells at time point of 30 min. Data are means ± SD from n = 5 independent experiments. One-way ANOVA with Dunnett’s multiple comparisons test; ****, p ≤ 0.0001.

Article Snippet: For endogenous XPR1 expression, anti XPR1 rabbit polyclonal antibody from proteintech (14174-1-AP) was used at 1:1000 dilution, followed by Horseradish Peroxidase (HRP)-conjugated anti rabbit antibody (1:2000).

Techniques: Translocation Assay, Binding Assay, Expressing

Journal: Nature Communications

Article Title: Structural basis of phosphate export by human XPR1

doi: 10.1038/s41467-025-55995-8

Figure Lengend Snippet: a , b Comparison of surface electrostatic potential of XPR1 wt ( a ) and XPR1 3 mut ( b ), highlighting the changes in the EV. Two views are shown for each structure. c Superimposition of structures of XPR1 wt and XPR1 3 mut , highlighting the conformational changes occurred in extracellular half of TM9 (cyan for XPR1 wt and purple for XPR1 3 mut ), viewed from the angle indicated in ( a ) (eye symbol). d Expanded view of the region boxed in ( c ) in two views, highlighting the movement of sidechain of W573. e Schematics of proposed working model. Only one protomer is shown, with each domain represented by different shapes and colored differently in the same way as in Fig. .

Article Snippet: For endogenous XPR1 expression, anti XPR1 rabbit polyclonal antibody from proteintech (14174-1-AP) was used at 1:1000 dilution, followed by Horseradish Peroxidase (HRP)-conjugated anti rabbit antibody (1:2000).

Techniques: Comparison

Journal: Nature Communications

Article Title: Structural basis of phosphate export by human XPR1

doi: 10.1038/s41467-025-55995-8

Figure Lengend Snippet: a Location and structure of the blocking peptide in front and bottom views. b – c Difference in the IV between full-length XPR1 wt structures ( b ) and XPR1 wt model with the C-terminal peptide manually removed ( c ).

Article Snippet: For endogenous XPR1 expression, anti XPR1 rabbit polyclonal antibody from proteintech (14174-1-AP) was used at 1:1000 dilution, followed by Horseradish Peroxidase (HRP)-conjugated anti rabbit antibody (1:2000).

Techniques: Blocking Assay

Journal: Nature communications

Article Title: Structural basis of phosphate export by human XPR1.

doi: 10.1038/s41467-025-55995-8

Figure Lengend Snippet: Fig. 5 | A C-terminal peptide shapes IV of Pi translocation pathway in XPR1. a Location and structure of the blocking peptide in front and bottom views. b–c Difference in the IV between full-length XPR1wt structures (b) and XPR1wt model with the C-terminal peptide manually removed (c).

Article Snippet: For endogenous XPR1 expression, anti XPR1 rabbit polyclonal antibody from proteintech (14174-1-AP) was used at 1:1000 dilution, followed by Horseradish Peroxidase (HRP)-conjugated anti rabbit antibody (1:2000).

Techniques: Translocation Assay, Blocking Assay