Boichenko V A, Hou J M, Mauzerall D
The Rockefeller University, 1230 York Avenue, New York, New York 10021, USA.
Biochemistry. 2001 Jun 19;40(24):7126-32. doi: 10.1021/bi010374k.
The volume and enthalpy changes for charge transfer in the 0.1-10 micros time window in photosynthetic reaction centers of the intact cells of Synechocystis PCC 6803 were determined using pulsed, time-resolved photoacoustics. This required invention of a method to correct for the cell artifact at the temperature of maximum density of water caused by the heterogeneous system. Cells grown under either white or red light had different PS I/PS II molar ratios, approximately 3 and approximately 1.7, respectively, but invariable action spectra and effective antenna sizes of the photosystems. In both cultures, the photoacoustic measurements revealed that their thermodynamic parameters differed strongly in the spectral regions of predominant excitation of PS I (680 nm) and PS II (625 nm). On correcting for contribution of the two photosystems at these wavelengths, the volume change was determined to be -27 +/- 3 and -2 +/- 3 A3 for PS I and PS II, respectively. The energy storage on the approximately 1 micros time scale was estimated to be 80 +/- 15% and 45 +/- 10% per trap in PS I and PS II, respectively. These correspond to enthalpies of -0.33 +/- 0.2 and -1 +/- 0.2 eV for the assumed formation of ion radical pairs P700+F(AB-) and Y(Z*)P680Q(A-), respectively. Taking the free energy of the above reactions as the differences of their redox potentials in situ, apparent entropy changes were estimated to be +0.4 +/- 0.2 and -0.2 +/- 0.2 eV for PS I and PS II, respectively. These values are similar to that obtained in vitro for the purified reaction center complexes on the microsecond time scale [Hou et al. (2001) Biochemistry 40, 7109-7116, 7117-7125]. The constancy of these thermodynamic values over a 2-fold change of the ratio of PS I/PS II is support for this method of in vivo analysis. Our pulsed PA method can correct the "cell" or heterogeneous artifact and thus opens a new route for studying the thermodynamics of electron transfer in vivo.
利用脉冲时间分辨光声技术测定了集胞藻PCC 6803完整细胞光合反应中心在0.1 - 10微秒时间窗口内电荷转移的体积和焓变。这需要发明一种方法来校正由异质系统在水的最大密度温度下引起的细胞假象。在白光或红光下生长的细胞具有不同的PSI/PSII摩尔比，分别约为3和约1.7，但光系统的作用光谱和有效天线大小不变。在两种培养物中，光声测量表明，它们的热力学参数在PSI（680nm）和PSII（625nm）的主要激发光谱区域有很大差异。在校正这两个波长下两个光系统的贡献后，确定PSI和PSII的体积变化分别为-27±3和-2±3 ų。在约1微秒时间尺度上，PSI和PSII中每个陷阱的能量存储估计分别为80±15%和45±10%。这些分别对应于假设形成离子自由基对P700⁺F(AB⁻)和Y(Z*)P680Q(A⁻)时的焓变-0.33±0.2和-1±0.2 eV。将上述反应的自由能作为其原位氧化还原电位的差值，估计PSI和PSII的表观熵变分别为+0.4±0.2和-0.2±0.2 eV。这些值与在微秒时间尺度上对纯化反应中心复合物进行体外测量得到的值相似[Hou等人（2001年）《生物化学》40, 7109 - 7116, 7117 - 7125]。这些热力学值在PSI/PSII比值2倍变化范围内的恒定性支持了这种体内分析方法。我们的脉冲光声方法可以校正“细胞”或异质假象，从而为研究体内电子转移的热力学开辟了一条新途径。
