title: Viscosity-dependent energy barriers and equilibrium conformational fluctuations in oxygen recombination with hemerythrin.
abstract: The recombination kinetics of photo-dissociated oxyhemerythrin (Sipunculus nudus) have been investigated between 298 K and 90 K. Fast geminate recombinations compete with oxygen escape into the solvent, from which a subsequent slower bimolecular rebinding takes place. In phosphate buffer (pH 7.7) at 278 K, the fast and slow processes are exponential and have comparable amplitudes. Whereas the oxygen escape rate rapidly decreases upon increasing the viscosity, the inward rate from the solvent is found to be independent of viscosity, up to about 50 cP (50 mPa.s). The data suggest that a Brownian-motion-driven displacement of one or several side-chain residues is implied in oxygen escape from within the protein and also that hemerythrin undergoes a conformational change in the deoxy state. At higher viscosities and lower temperature only the geminate phase is observed and the kinetics progressively depart from an exponential. Below about 130 K, the kinetics resemble those reported in the literature for heme proteins. They are consistent with a temperature-independent non-equilibrium frozen distribution of conformational substates. However, between 190 K and 130 K, the profile of the kinetics is invariant on a log/log plot and the results simply differ by a translation along the log t axis. It is shown that this property is expected only for a temperature-dependent distribution of substates in a Boltzmann equilibrium. From room temperature, where rebinding is exponential, down to the 'freezing' temperature, the geminate recombinations display a variety of kinetic laws. It can be shown, however, that for a broad class of substate distributions, the initial slope of the kinetic plot follows an Arrhenius relationship. The activation energy is equal to that of the exponential rate constant measured at high temperature. This result establishes the conditions under which protein data obtained from low-temperature kinetics can be extrapolated to physiological temperature.
authors.affiliation: Unité 219, Institut National de la Santé et de la Recherche Médicale, Institut Curie, Orsay, France.
authors.forename: D
authors.initials: D
authors.lastname: Lavalette
authors.forename: C
authors.initials: C
authors.lastname: Tetreau
mesh_terms: D000818:Animals; D006422:Hemerythrin; D007700:Kinetics; D008433:Mathematics; D008667:Metalloproteins; D009348:Nematoda; D010084:Oxidation-Reduction; D010100:Oxygen; D010777:Photochemistry; D011485:Protein Binding; D011487:Protein Conformation; D012997:Solvents; D013696:Temperature; D014783:Viscosity
chemical_list: D006422:Hemerythrin; D008667:Metalloproteins; D012997:Solvents; D010100:Oxygen