Figure 5. His-330 and His-362 are key residues in the interaction of Pb²⁺ with Tau_244–372.

10 µM single mutants H330A (A) and H362A (B) or double mutant H330A/H362A (C) of Tau_244–372 were incubated with 0–40 µM Pb²⁺ (black: 0 µM, red: 5 µM, blue: 10 µM, yellow: 20 µM, green: 30 µM, and magenta: 40 µM) in 10 mM HEPES buffer (pH 7.4) containing 100 mM NaCl, 1 mM DTT, 2.5 µM heparin, and 20 µM ThT, and ThT binding assays were carried out at 37°C. Data are expressed as mean ± S.D. (n = 3–4). (D) Effect of Pb²⁺ on the relative change in the β-sheet content of single mutants H330A (square) and H362A (circle) or double mutant H330A/H362A (triangle) during fibril formation, studied by monitoring the CD signal at 218 nm. 10 µM Tau_244–372 was incubated with 0–10 µM Pb²⁺ (black: 0 µM, and blue: 10 µM) in 30 mM phosphate buffer (pH 7.4) containing 1 mM DTT and 2.5 µM heparin.