Identification of Specific Effect of Chloride on the Spectral Properties and Structural Stability of Multiple Extracellular Glutamic Acid Mutants of Bacteriorhodopsin
Fig 2
Alkaline titration of the Schiff base of multiple EC Glu mutants and WT.
(A) Representative difference spectra (pHi–pH6) of 3Glu in 1 M KCl (for pH values i = 9 to 12.5), at room temperature. The cross over point at 515 nm reflects the transition from the basic (purple in 1 M KCl) to red spectral form. (B) Representative difference spectra (pHi–pH6) of 2Glu in 150 M KCl (for pH values i = 6 to 13.2), at room temperature. Insets in A and B represent plots of the absorption changes at 470 nm and 370 nm, as a function of the pH. The apparent pKa of SB, at 12.8 and 12.3 for 2Glu (150 mM KCl) and 3Glu (1 M KCl), respectively, were obtained from the best fit of ΔA at 370 nm vs pH using the Henderson-Hasselbach equation. (C) Plots of the absorption changes at 570 nm as a function of pH,for WT (■) and 2Glu () in 150 mM KCl, and 3Glu (
,
) and 4Glu (
,
) in 150 mM and 1 M KCl, respectively, at room temperature. Continuous lines are empirically drawn for a visual help and do not represent a fit of the data.