Chromophore distortions in photointermediates of proteorhodopsin visualized by Dynamic Nuclear Polarization-enhanced solid-state NMR

Abstract

Proteorhodopsin (PR) is the most abundant retinal protein on earth and functions as a light-driven proton pump. Despite of extensive efforts, structural data for PR photointermediate states have not been obtained. Based on 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 de-protonation 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.

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ORCID for Alexander J. Leeder: orcid.org/0000-0001-9271-6038

ORCID for Lynda J. Brown: orcid.org/0000-0002-5678-0814

ORCID for Richard C.D. Brown: orcid.org/0000-0003-0156-7087

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