Recombinant Mouse Anti-S. lividans KcsA ( T75C) Antibody (CAT#: PABW-084)

Figure 1 Formation of a dimer of the T75C mutant of KcsA catalyzed by Cu 2+ .

(A) SDS-PAGE analysis of dimer formation. Identical samples of T75C protein (0.5 µg) were incubated in 10 mM Hepes-Tris (pH 7.4), 100 mM choline Cl, and the following concentrations of CuCl 2 corresponding to individual lanes of the gel proceeding from left to right: 0, 1, 10, 50, 100, 250, 500, and 750 µM and 1, 2.5, 5, 7.5, and 100 mM. Samples were subjected to SDS-PAGE after incubation for 1 h at 22 °C. (B) Fraction of tetramer (b) normalized to tetramer band measured in the absence of Cu 2+ and fraction of dimer (O) normalized to dimer band measured in the presence of 5 mM Cu 2+ . Solid lines correspond to fits of data to eq 1 with an N of 1.5 and a K 0.5 of 630 µM Cu 2+ for tetramer and an N of 1.6 and a K 0.5 of 580 µM Cu 2+ for dimer.

Krishnan, M. N., Trombley, P., & Moczydlowski, E. G. (2008). Thermal stability of the K+ channel tetramer: cation interactions and the conserved threonine residue at the innermost site (S4) of the KcsA selectivity filter. Biochemistry, 47(19), 5354-5367.

Figure 2 Effect of thiophilic cations on the thermal stability of KcsA and the T75C mutant.

KcsA (A) and T75C (B) proteins were purified and exchanged into buffer containing 10 mM Hepes-Tris, 100 mM Tris-NO 3 (pH 7.4), and ∼4 mM C 12 M. Protein samples (∼0.5 µg) were equilibrated at room temperature for 1 h in this same buffer with either no addition (lanes U and H) or buffer containing 1 mM KCl (lane K + ), 1 mM Hg(NO 3 ) 2 (lane Hg 2+ ), 1 mM Ag(NO 3 ) (lane Ag + ), or 1 mM Pb(NO 3 ) 2 (lane Pb 2+ ). Samples were analyzed at room temperature by SDS-PAGE either without heating (lane U) or after heating at 65 °C for 10 min (all other lanes).

Krishnan, M. N., Trombley, P., & Moczydlowski, E. G. (2008). Thermal stability of the K+ channel tetramer: cation interactions and the conserved threonine residue at the innermost site (S4) of the KcsA selectivity filter. Biochemistry, 47(19), 5354-5367.