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Chromophore Distortions in Photointermediates of Proteorhodopsin Visualized by Dynamic Nuclear Polarization-Enhanced Solid-State NMR

Mehler, Michaela, Eckert, Carl Elias, Leeder, Alexander J., Kaur, Jagdeep, Fischer, Tobias, Kubatova, Nina, Brown, Lynda J., Brown, Richard C. D., Becker-Baldus, Johanna, Wachtveitl, Josef, Glaubitz, Clemens
Journal of the American Chemical Society 2017 v.139 no.45 pp. 16143-16153
carbon, models, mutation, nitrogen, nuclear magnetic resonance spectroscopy, proton pump, rhodopsin, schiff bases, stable isotopes
Proteorhodopsin (PR) is the most abundant retinal protein on earth and functions as a light-driven proton pump. Despite extensive efforts, structural data for PR photointermediate states have not been obtained. On the basis of dynamic nuclear polarization (DNP)-enhanced solid-state NMR, we were able to analyze the retinal polyene chain between positions C10 and C15 as well as the Schiff base nitrogen in the ground state in comparison to light-induced, cryotrapped K- and M-states. A high M-state population could be achieved by preventing reprotonation of the Schiff base through a mutation of the primary proton donor (E108Q). Our data reveal unexpected large and alternating ¹³C chemical shift changes in the K-state propagating away from the Schiff base along the polyene chain. Furthermore, two different M-states have been observed reflecting the Schiff base reorientation after the deprotonation step. Our study provides novel insight into the photocycle of PR and also demonstrates the power of DNP-enhanced solid-state NMR to bridge the gap between functional and structural data and models.