Validation of response function construction and probing heterogeneous protein hydration by intrinsic tryptophan.
Protein solvation dynamics usually occur on multiple time scales with site specificity, and the characterization of such heterogeneous dynamics requires a convenient optical probe. We proposed a tryptophan methodology, and with site-directed mutagenesis we can use a tryptophan scan to probe any desirable position around protein surfaces. Here, we report our extended solvation model for construction of response functions for probes such as tryptophan with multiple emission peaks and lifetimes. We show our systematic construction procedure and careful analyses of the possible missing percentage of an initial ultrafast component with the established zero-time emission spectrum and limited temporal resolution through two methods of the direct mapping of femtosecond-resolved fluorescence spectra (3D FRES) and the constructed FRES (2D) from the fluorescence transients. We unambiguously validate our extended model with reexamination of solvation dynamics (methanol, water, and proteins) using conventional dye coumarin, intrinsic tryptophan, and cofactor flavin. Using mutant proteins of GB1, we show again the generality of the powerful probe tryptophan for protein hydration (solvation) and the slowdown of the hydration layer dynamics especially at the water-protein interface. These results justify the necessity of our extended solvation model, clarify the confusion of protein hydration in the recent literature, and establish the universal optical probe of tryptophan for heterogeneous protein dynamics.
Department of Physics, and Program of Biophysics, 191 West Woodruff Avenue, The Ohio State University, Columbus, Ohio 43210, USA.
Yangzhong
Y
Qin
Chih-Wei
CW
Chang
Lijuan
L
Wang
Dongping
D
Zhong
D011506:Proteins / Q000737:chemistry*; D014364:Tryptophan / Q000737:chemistry*; D014867:Water / Q000737:chemistry*
D011506:Proteins; D014867:Water; D014364:Tryptophan