Real-time tracking of CO migration and binding in the α and β subunits of human hemoglobin via 150-ps time-resolved Laue crystallography.
We have developed the method of picosecond Laue crystallography and used this capability to probe ligand dynamics in tetrameric R-state hemoglobin (Hb). Time-resolved, 2 Å-resolution electron density maps of photolyzed HbCO reveal the time-dependent population of CO in the binding (A) and primary docking (B) sites of both α and β subunits from 100 ps to 10 μs. The proximity of the B site in the β subunit is about 0.25 Å closer to its A binding site, and itskBArebinding rate (~300 μs-1) is six times faster, suggesting distal control of the rebinding dynamics. Geminate rebinding in the β subunit exhibits both prompt and delayed geminate phases. We developed a microscopic model to quantitatively explain the observed kinetics, with three states for the α subunit and four states for the β subunit. This model provides a consistent framework for interpreting rebinding kinetics reported in prior studies of both HbCO and HbO2.
Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520, USA.
Friedrich
F
Schotte
Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520, USA.
Hyun Sun
HS
Cho
Department of Biochemistry and Cell Biology, and W.M. Keck Center for Computational Biology, Rice University, Houston, TX 77251-1892, USA.
Jayashree
J
Soman
European Synchrotron Radiation Facility, BP 220, 38043 Grenoble Cedex, France.
Michael
M
Wulff
Department of Biochemistry and Cell Biology, and W.M. Keck Center for Computational Biology, Rice University, Houston, TX 77251-1892, USA.
John S
JS
Olson
Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520, USA.
Philip A
PA
Anfinrud
Geminate rebinding; Hemoglobin; Ligand migration; Time-resolved Laue crystallography