UN2A undergoes a conformational change with S100A1 binding. The UN2A region was cloned in between CFP and YFP to create a FRET reporter system to observe conformational changes in UN2A. Samples were excited at 433 nm and fluorescence emission spectra were measured from 445 to 600 nm. (A and B) Binding experiments were conducted in both low Ca²⁺ (pCa 10, A) and high Ca²⁺ (pCa 3, B) conditions. No significant differences were observed in low Ca²⁺ conditions. Under high Ca²⁺ conditions, there was an increase in the CFP (465 nm) peak and a decrease in the YFP (525 nm) peak, consistent with lower FRET due to an increase in the distance between the two fluorophores. This suggests that the UN2A adopts a more elongated conformation when S100A1 binds.