title: Determination of the Fe-CO bond energy in myoglobin using heterodyne-detected transient thermal phase grating spectroscopy.
abstract: The bond energies at active sites of proteins are intimately coupled to the structure-function relationship in biological systems. Due to the unknown nature of the protein relaxation along a reaction coordinate, it has not been possible to directly determine bond energies relevant to protein function. By embedding proteins in trehalose glasses, it is possible to freeze out protein relaxation on short time scales and determine the bond energies of photolabile ligands using photothermal spectroscopies. As a prototypical example, the photodissociation dynamics and energetics of carboxy-myoglobin (MbCO) in a trehalose glass matrix at room temperature were studied by transient absorption (or pump-probe) and transient thermal phase grating spectroscopy to determine the CO recombination dynamics and associated energetics, respectively. Both the initial energetics of the bond breaking and the energy released upon bond reformation could be used, on a time scale faster than significant protein relaxation, to determine the Fe-CO bond energy as 34 +/- 4 kcal/mol. This bond energy is significantly larger than that typically cited (25 kcal/mol) on the basis of indirect measurements but is in good agreement with recent theoretical predictions (35 kcal/mol) (Rovira, C.; Parrinello, M. Int. J. Quantum Chem. 2000, 80, 1172). This result in combination with the theoretical study suggests that protein structure plays a significant role in the bond energies at active sites which in turn provides a tuning element of the effective barrier heights independent to the transition state region.
authors.affiliation: Department of Chemistry, University of Toronto, 80 St. George Street, Toronto, Ontario M5S 3H6, Canada.
authors.forename: Markus
authors.initials: M
authors.lastname: Walther
authors.forename: Valerica
authors.initials: V
authors.lastname: Raicu
authors.forename: Jennifer P
authors.initials: JP
authors.lastname: Ogilvie
authors.forename: Ralph
authors.initials: R
authors.lastname: Phillips
authors.forename: Ronald
authors.initials: R
authors.lastname: Kluger
authors.forename: R J Dwayne
authors.initials: RJ
authors.lastname: Miller
mesh_terms: D000818:Animals; D002248:Carbon Monoxide; D006736:Horses; D007501:Iron; D007700:Kinetics; D009206:Myocardium / Q000378:metabolism; D009211:Myoglobin / Q000737:chemistry*; D013053:Spectrophotometry; D013816:Thermodynamics
chemical_list: D009211:Myoglobin; C018013:carboxymyoglobin; D002248:Carbon Monoxide; D007501:Iron