TY - JOUR
T1 - Gaugement of the inner space of the apomyoglobin's heme binding site by a single free diffusing proton. II. Interaction with a bulk proton
AU - Shimoni, E.
AU - Nachliel, E.
AU - Gutman, M.
N1 - Funding Information:
This research is supported by the United States-Israel Binational Science Foundation, grant No. 870035, and the Office of Naval Re- search, United States Navy grant N00014-89-J 1622.
PY - 1993
Y1 - 1993
N2 - The reaction mechanism and the dynamic aspects of protonation of a defined moiety located inside an aqueous cavity in a protein were monitored by time resolved spectroscopy using the pyranine apomyoglobin complex as a model (Shimoni, Tsfadia, Nachliel, and Gutman, 1993, Biophys. J. 64:472–479). The reaction was synchronized by a short laser pulse and the reprotonation of the ground state pyranine anion (phi O-) was monitored, in the microsecond time scale, by its transient absorption at 457 nm. The observed signal was reconstructed by a numeric solution of nonlinear, coupled differential equations which account for the direct reaction of phi O- with bulk proton and by proton transfer from the nearby amino acids: His 64, Asp 44, Asp 60, and Glu 59. A unique combination of rate constant was obtained which quantitates the contribution of each pathway to the overall relaxation process. In the first phase of the dynamics phi O- abstracts a proton from the nearby protonated histidine. The bulk proton interacts preferentially with the cluster of three carboxylates and immediately shuttled to the deprotonated histidine. The high proximity of the reactive groups and the strong electrostatic forces operating inside the heme binding cavity render the rate of proton transfer in the site ultrafast.
AB - The reaction mechanism and the dynamic aspects of protonation of a defined moiety located inside an aqueous cavity in a protein were monitored by time resolved spectroscopy using the pyranine apomyoglobin complex as a model (Shimoni, Tsfadia, Nachliel, and Gutman, 1993, Biophys. J. 64:472–479). The reaction was synchronized by a short laser pulse and the reprotonation of the ground state pyranine anion (phi O-) was monitored, in the microsecond time scale, by its transient absorption at 457 nm. The observed signal was reconstructed by a numeric solution of nonlinear, coupled differential equations which account for the direct reaction of phi O- with bulk proton and by proton transfer from the nearby amino acids: His 64, Asp 44, Asp 60, and Glu 59. A unique combination of rate constant was obtained which quantitates the contribution of each pathway to the overall relaxation process. In the first phase of the dynamics phi O- abstracts a proton from the nearby protonated histidine. The bulk proton interacts preferentially with the cluster of three carboxylates and immediately shuttled to the deprotonated histidine. The high proximity of the reactive groups and the strong electrostatic forces operating inside the heme binding cavity render the rate of proton transfer in the site ultrafast.
UR - http://www.scopus.com/inward/record.url?scp=0027464736&partnerID=8YFLogxK
U2 - 10.1016/S0006-3495(93)81390-0
DO - 10.1016/S0006-3495(93)81390-0
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AN - SCOPUS:0027464736
SN - 0006-3495
VL - 64
SP - 480
EP - 483
JO - Biophysical Journal
JF - Biophysical Journal
IS - 2
ER -