I. Wavelength-dependent photocycle activity of xanthorhodopsin in the visible region II. Solvent isotope effect on the dark adaptation of bacteriorhodopsin in purple membrane

• Main Authors: Chiang, Han-Kuei, 江函桂
• Other Authors: Chu, Li-Kang
• Format: Others
• Language: zh-TW
• Published: 2016
• Online Access: http://ndltd.ncl.edu.tw/handle/qkcp79

碩士 === 國立清華大學 === 化學系 === 105 === Part I. Xanthorhodopsin (xR) is a dual-chromophore proton-pump photosynthetic protein comprising one retinal Schiff base and one light-harvesting antenna salinixanthin (SX). The excitation wavelength-dependent transient population of the intermediate M demonstrates that the excitation of the retinal at 570 nm leads to the highest photocycle activity and the excitations of SX at 460 and 430 nm reduce the activity to ca. 37% relatively, suggesting an energy transfer pathway from the S2 state of the SX to the S1 state of the retinal and a quick internal vibrational relaxation in the S2 state of SX prior to the energy transfer from SX to retinal. Part II. The thermal retinal isomerization from all-trans, 15-anti to 13-cis, 15-syn of bacteriorhodopsin in purple membrane in H2O and D2O during dark adaptation was investigated at 30—55°C at neutral pH. In this temperature range, phase transition of purple membrane and destruction of the tertiary structure of bacteriorhodopsin did not take place. We found that the solvent isotope effect is inverted below about 45°C; i.e., kf (D2O) / kf (H2O) > 1. Applying the transition state theory, the changes in enthalpy from the initial state to the transition state along the thermal trans-to-cis forward reaction coordinate, ΔH*f, were determined to be 24.7 ± 1.2 and 20.1 ± 0.4 kcal mol−1 in H2O and D2O, respectively. The relative entropic change of the transition state in H2O and D2O, ΔΔS*f = ΔS*f (D2O) − ΔS*f (H2O), was −14.4 ± 3.9 cal mol−1 K−1. In addition, the Gibbs free energy of trans-to-cis thermal isomerization reaction in D2O is 0.4—0.7 kcal mol−1 lower than that in H2O. It is the first time the entropy and enthalpy of the transition state have been quantified to elucidate the solvent isotope effect in the retinal thermal isomerization of bacteriorhodopsin during dark adaptation. The solvent isotope effect on the thermodynamics properties and kinetics implied that the hydrogen bonding in the transition state during the dark adaptation of bR is stronger than that in the initial state.