Citation
Diller, A. (2007, September 18). Photo-CIDNP MAS NMR Studies on photosynthetic reaction centers. Retrieved from https://hdl.handle.net/1887/12365
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Summary
Photosynthesis is the process through which plants and photosynthetic organisms convert solar energy into chemical energy. The fundamen- tal primary reaction of photosynthesis is charge separation which takes place in the photosynthetic RCs. Among all photosynthetic RCs, only PS II provides a redox force that is sufficiently strong for water oxida- tion. Based on this capability, the production of atmospheric oxygen is possible. Despite decades of research, the origin of this oxidizing power is not yet understood, however, there is converging evidence that it is not related to a single evolutionary event, such as a single mutation. Rather it arose from multiple stepwise processes.
Photo-CIDNP proves to be a valuable technique to gain informa- tion on the primary step of photosynthesis. The solid-state photo- CIDNP effect is caused by photochemical reactions which shuffle the nuclear spin system out of its Boltzmann equilibrium. The effect can be observed by solid-state NMR methods as a strong modification of NMR signal intensities. Three mechanism are known to build up net nuclear polarization in the solid-state: TSM, DD and DR.
This thesis contains the results of photo-CIDNP studies on various photosynthetic organisms and discusses perspectives of the application of photo-CIDNP. InChapter 2, experiments have been performed on bacterial RCs of Rb. sphaeroides WT and its carotenoid-less mutant R26. It has been observed that the recovery of the NMR signal after a solid-state photo-CIDNP experiment is up to a factor of 30 faster than expected from the longitudinal relaxation time. Theoretical anal- ysis shows that the faster recovery is a consequence of additional decay channels for the nuclear polarization during the photocycle of quinone- blocked photosynthetic RCs. These channels are opened up by the same processes that create net polarization in photo-CIDNP. Coherent mixing between electron and nuclear spin states due to pseudosecular hf coupling within the radical pair state as it occurs in the TSM and DD mechanisms provides such a coherent loss channel for nuclear po- larization. The experiments on R26 RCs, where the occurrence of the additional DR leads to further production of net nuclear polarization, shows that an additional relaxation channel allows for accelerated sig-
In Chapter 3,13C-photo-CIDNP data on unlabeled PS I and PS II from plants are reported in order to explore the electron donors.
In a previous study, as the origin of the observed asymmetry, a local electrostatic field was proposed, pulling the electron charge towards the C-131 carbonyl, stabilizing the frontier orbitals and increasing the redox force. The data presented here allow a more detailed view which corroborates the earlier interpretation. The maximum of electron spin density is found on rings III and V. In addition, electron spin density is observed on aromatic amino acid side chains or carotenoid molecules.
The comparison between PS I and PS II reveals that the electronic structure of the P680 radical cation is a Chl a cofactor with strong matrix interaction, while the radical cation of P700, the donor of PS I, appears to be a Chl a which is essentially undisturbed.
Chapter 4 presents photo-CIDNP studies of PS I and PS II on uniformly 15N-labeled spinach. The interpretation of 15N photo- CIDNP MAS NMR of isotope labeled RCs is more straightforward, compared with 13C signal patterns of unlabeled RCs as presented in Chapter 3, as each 15N signal can be assigned easily to one of the four pyrrole subunits. The results show that the electron spin density distribution in PS I, apart from its known delocalization over 2 Chl molecules, reveals no marked disturbance, whereas the pattern of the electron-spin density distribution in PS II shows a strong asymmetry of electron spin density towards the pyrrole ring IV in the oxidized radical state. This is in-line with the strong asymmetry of electron spin density detected by 13C photo-CIDNP MAS NMR in Chapter 3 demonstrating maximum electron spin density at the neighboring C-15 methine carbon. Based on the chemical shifts and an analysis of the X-ray structure of PS II, the electron spin density observed on aromatic side chains in Chapter 3, can be assigned here most likely to the axial His of the PS II donor. Based on these observations, a ’hinge model’ for the donor of PS II is proposed, explaining the inversion of electron spin density based on a tilt of the axial His towards pyrrole ring IV causing π-π overlap of both aromatic systems.
Chapter 5 presents photo-CIDNP experiments on selectively iso- tope labeled purified RCs as well as on membrane-bound RCs from Rb. sphaeroides WT. Isotope labeling provides an additional gain
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in NMR sensitivity and selectivity. While photo-CIDNP experiments on selectively isotope labeled purified RCs show no influence on the sign of the photo-CIDNP effect, the sign of the photo-CIDNP effect of membrane bound RCs shows an inversion and appears as absorp- tive (positive) signals of the donor upon isotope labeling. The data presented here ruled out that the sign-change effect observed in iso- tope labeled chromatophores is caused by the centrifugal orientation or magnetic field-orientation. The effect appears to depend on the sur- rounding of the RCs in terms of proteins and membranes. Since the observed sign-change effect is limited to the donor signals, the anal- ogy to the DR mechanism observed in RCs of Rb. sphaeroides R26, is discussed. The spectroscopic distinction between donor signals and acceptor signals may allow to use photo-CIDNP as a valuable tool for spectral editing.
The bottom line of the work described in this thesis and prospects for further research on the donor of PS II and applications of photo- CIDNP are discussed in Chapter 6.
