Exploring the Electron Transfer Pathways in Photosystem I by High-Time-Resolution Electron Paramagnetic Resonance: Observation of the B-Side Radical Pair P700 +A1B – in Whole Cells of the Deuterated Green Alga Chlamydomonas reinhardtii at Cryogenic Temperatures

• Published in: Journal of the American Chemical Society Vol. 134; no. 12; pp. 5563 - 5576
• Main Authors: Berthold, Thomas, Donner von Gromoff, Erika, Santabarbara, Stefano, Stehle, Patricia, Link, Gerhard, Poluektov, Oleg G, Heathcote, Peter, Beck, Christoph F, Thurnauer, Marion C, Kothe, Gerd
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 28-03-2012
• Subjects: Chlamydomonas reinhardtii - chemistry; Chlamydomonas reinhardtii - cytology; Cold Temperature; Deuterium - chemistry; Electron Spin Resonance Spectroscopy; Electron Transport; Models, Molecular; Photosystem I Protein Complex - chemistry; Plant Proteins - chemistry
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/ja208806g

Crystallographic models of photosystem I (PS I) highlight a symmetrical arrangement of the electron transfer cofactors which are organized in two parallel branches (A, B) relative to a pseudo-C 2 symmetry axis that is perpendicular to the membrane plane. Here, we explore the electron transfer pathways of PS I in whole cells of the deuterated green alga Chlamydomonas reinhardtii using high-time-resolution electron paramagnetic resonance (EPR) at cryogenic temperatures. Particular emphasis is given to quantum oscillations detectable in the tertiary radical pairs P700 +A1A – and P700 +A1B – of the electron transfer chain. Results are presented first for the deuterated site-directed mutant PsaA-M684H in which electron transfer beyond the primary electron acceptor A0A on the PsaA branch of electron transfer is impaired. Analysis of the quantum oscillations, observed in a two-dimensional Q-band (34 GHz) EPR experiment, provides the geometry of the B-side radical pair. The orientation of the g tensor of P700 + in an external reference system is adapted from a time-resolved multifrequency EPR study of deuterated and 15N-substituted cyanobacteria (Link, G.; Berthold, T.; Bechtold, M.; Weidner, J.-U.; Ohmes, E.; Tang, J.; Poluektov, O.; Utschig, L.; Schlesselman, S. L.; Thurnauer, M. C.; Kothe, G. J. Am. Chem. Soc. 2001, 123, 4211–4222). Thus, we obtain the three-dimensional structure of the B-side radical pair following photoexcitation of PS I in its native membrane. The new structure describes the position and orientation of the reduced B-side quinone A1B – on a nanosecond time scale after light-induced charge separation. Furthermore, we present results for deuterated wild-type cells of C. reinhardtii demonstrating that both radical pairs P700 +A1A – and P700 +A1B – participate in the electron transfer process according to a mole ratio of 0.71/0.29 in favor of P700 +A1A –. A detailed comparison reveals different orientations of A1A – and A1B – in their respective binding sites such that formation of a strong hydrogen bond from A1 – to the protein backbone is possible only in the case of A1A –. We suggest that this is relevant to the rates of forward electron transfer from A1A – or A1B – to the iron–sulfur center FX, which differ by a factor of 10. Thus, the present study sheds new light on the orientation of the phylloquinone acceptors in their binding pockets in PS I and the effect this has on function.