Fig. 4: SAXS and cryo-EM demonstrates that human TG2 adopts the open state when bound to Ca²⁺ and forms a dimer at high enzyme concentrations.

a The 3.4 Å cryo-EM structure of the TG2 catalytic domain bound to Ca²⁺ ions suggests that TG2 adopts the open state. b The three Ca²⁺ binding sites (lime) within the TG2 catalytic domain. The conserved residues involved in Ca²⁺ binding is shown in the inset for each site. c Ca²⁺ bound TG2 can be described as a open state dimer in solution, as demonstrated using DAMMIF to construct a SAXS envelope (light blue) and CRYSOL to fit to the open state dimer (yellow, see Fig. 3c). The CRYSOL fit of the open state monomer is shown in black for comparison. The Guinier R_g and the model Χ² are shown in the legend. d TG2 incubated with 2 mM Ca²⁺ and then treated with 5 mM GTP (gray) can be described as a closed state monomer in solution, as demonstrated using DAMMIF to construct a SAXS envelope (light blue) and CRYSOL to fit to the closed state monomer (black). The Guinier R_g and the model Χ² are shown in the legend. e The Guinier R_g of TG2 pre-incubated with 2 mM GTP increases in response to treatment with 1 mM to 5 mM Ca²⁺, supporting the formation of the open state. The experimental molecular weight calculated from the Porod Volume is shown on the secondary Y-axis (red). f The Guinier R_g of TG2 R580K pre-incubated with 2 mM GTP increases in response to treatment with 1 mM to 5 mM Ca²⁺, indicating it can promote oligomerization of the R580K open state dimer. The experimental molecular weight calculated from the Porod Volume is shown on the secondary Y-axis (red).