Ultrafast Two-Photon Spectroscopy of Carotenoids involved in the Regulation of Photosynthesis

吸收光能的利用和耗散之间的快速平衡对于天然植物和基因修饰植物的存活是必不可少的。[1-3]然而，尽管进行了深入的研究（见最近在《自然》和《科学》上的出版物）[1，4-6]，这些调节过程的生物物理机制仍然知之甚少。分子齿轮移位模型提出，叶黄素类胡萝卜素的化学结构的可逆的和逐步的酶促修饰使得能够在类胡萝卜素至叶绿素的光捕获和叶绿素至类胡萝卜素的淬灭之间切换。[7]不幸的是，所涉及的类胡萝卜素的第一激发单重态，Car S1，是光学禁止的[8]，这使得不可能通过常规吸收或荧光光谱法直接研究其作用。在过去，我们已经表明，双光子激发（TPE）允许一个选择性的人口的禁态，因为光学选择规则是不同的这个非线性过程。[9-15]初步的双光子敏化荧光实验[9]表明，所提出的类胡萝卜素-叶绿素能量流的变化确实发生在植物中。我们现在计划研究分离的色素蛋白复合物，以及类囊体和叶绿体制剂含有明确定义的量的高或低光叶黄素。这些双光子实验的目的是为所提出的机制的主要贡献提供一个明确的证明或反驳，并阐明哪些色素-蛋白质复合物参与了这种调节。

A quick balancing between utilization and dissipation of absorbed light energy is essential for the survival of natural and gene modified plants.[1-3] However, despite intensive investigations (see recent publications in Nature and Science)[1, 4-6] the biophysical mechanisms of these regulation processes are still poorly understood. The molecular gear shift model proposes that a reversible and step-wise enzymatic modification of the chemical structure of xanthophyll carotenoids enables a switching between carotenoid-to-chlorophyll lightharvesting and chlorophyll-to-carotenoid quenching.[7] Unfortunately, the involved firstexcited singlet state of the carotenoids, Car S1, is optically forbidden[8] which renders impossible a direct investigation of its role by conventional absorption or fluorescence spectroscopy. In the past we have shown that two-photon excitation (TPE) allows a selective population of the forbidden state because the optical selection rules are different for this nonlinear process.[9-15] Preliminary two-photon sensitized fluorescence experiments[9] indicated that the proposed change in carotenoid-chlorophyll energy flow indeed takes place in plants. We now plan to investigate isolated pigment-protein complexes as well as thylakoid and chloroplast preparations containing well defined amounts of high- or low-light xanthophylls. The aim of these two-photon experiments is to provide a clear prove or disprove for a major contribution of the proposed mechanism and to elucidate which of the pigment-protein complexes are involved in such a regulation.