The Enthalpy and Entropy of Reaction for Formation of P + Q A - from Excited Reaction Centers of Rhodobacter sphaeroides Gregory J. Edens, † M. R. Gunner, ‡ Qiang Xu, ‡ and David Mauzerall* ,† Contribution from Rockefeller UniVersity, 1230 York AVenue, New York, New York 10021, and Department of Physics, City UniVersity of New York, ConVent AVenue and 138th Street, New York, New York 10031 ReceiVed May 28, 1999. ReVised Manuscript ReceiVed NoVember 9, 1999 Abstract: The enthalpy and volume changes for the charge-transfer reaction between excited donor and ionized donor and acceptor in bacterial reaction centers were determined using pulsed photoacoustics. Excitation in the lowest absorption band of the centers at 860 nm minimized the thermal signal caused by degradation of excess energy. Knowing the free energy of this reaction, -0.86 eV, the determination of the enthalpy, -0.44 eV, fixes the entropy at 25 °C as about one-half (T∆S )+0.42 eV) of the free energy for the normal ubiquinone- 10 containing centers. This is a larger contribution than anticipated from previous estimates of the enthalpy. The unexpected sign of the entropy is assigned to the release of counterions from the reaction center surfaces when the charge transfer cancels the dominant opposite charges at the interfaces. The enthalpy and entropy of six reaction centers containing exchanged quinones did not correlate with their free energies. The volume contractions ranged from -28 to -42 Å 3 and roughly correlated with the size of the quinone as expected from electrostriction. Introduction The kinetics of the electron transfer steps in bacterial reaction centers have been thoroughly investigated from femtoseconds to seconds, 1,2 and thus the reaction sequence is well character- ized. However, the thermodynamic properties of these inter- mediates are far less well-known. The sequence begins, following excitation of the donor dimer bacteriochlorophyll (P), with a rapid (∼3 ps) electron transfer to the bacteriopheophytin (H) in the L branch of the reaction center and is followed by a slower (∼200 ps) step to the primary quinone (Q A ). The free energies of these steps have been obtained as the separate redox potentials of the donor and acceptor. 3-7 Arata and Parson 8 have measured the free energy (-0.86 eV) of the excited state to donor cation acceptor anion reaction from the ratio of delayed to prompt fluorescence and the enthalpy (-0.7 eV) by use of the temperature dependence of the kinetics of delayed light emission. These and other measures of ∆H will be discussed in the body of this paper. Photoacoustic (PA) methodology allows a direct measure of the enthalpy of reaction plus changes in the reaction volume in a photochemical sequence. 9 The two contributions can be separated by measurement at the temperature of maximum density of water where only changes in volume contribute to the PA signal. Our previous measurements on reaction centers gave a volume change of -22 Å 3 for the formation of P + Q A - . This contraction was assigned to electrostriction and was used to obtain an estimate of the effective dielectric coefficient of the protein. 10 Measurements by several workers 11-13 have reported volume changes ranging from -12 to -34 Å 3 and values of the enthalpy change varying from -0.44 to -1.33 eV. Photoacoustic data for Rhodobacter sphaeroides reaction centers have been obtained previously using 532 nm excitation. At that wavelength, ca. 30% of the photon energy is degraded to heat instantly and adds a large background to the measure- ment. With the tuning ability of optical parametric oscillator technology, we can now excite the reaction centers near their trap energy, avoiding this excess heat. We set out to obtain ∆V, ∆H°, and, using the literature value of ∆G°, ∆S° of this reaction. Experimental Section R. sphaeroides reaction centers (RC’s) were isolated following standard procedures 14 using lauryl dimethylamine oxide (LDAO) extraction and purified using ammonium sulfate and DEAE (dieth- ylaminoethyl) chromatography. These RC’s have QA occupancy >95% and ∼5% QB occupancy. Solutions of OD 860 ) 1 in 1 cm (∼7 µM RC) in 10 mM Tris pH 8 were deoxygenated by stirring a thin layer in † Rockefeller University. ‡ City University of New York. (1) Blankenship, R. E.; Madigan, M. T.; Bauer, C. E. Anoxygenic photosynthetic bacteria; Kluwer Academic Publishers: Dordrecht, 1995; Vol. 2. (2) Vos, M. H.; Breton, J.; Martin, J. L. J. Phys. Chem. B 1997, 101, 9820-9832. (3) Dutton, P. L.; Leigh, J. S.; Wraight, C. A. 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