Multiple retinal isomerizations during the early phase of the bestrhodopsin photoreaction.

Abstract

Bestrhodopsins constitute a class of light-regulated pentameric ion channels that consist of one or two rhodopsins in tandem fused with bestrophin ion channel domains. Here, we report on the isomerization dynamics in the rhodopsin tandem domains of <i>Phaeocystis antarctica</i> bestrhodopsin, which binds all-trans retinal Schiff-base (RSB) absorbing at 661 nm and, upon illumination, converts to the meta-stable P540 state with an unusual 11-<i>cis</i> RSB. The primary photoproduct P682 corresponds to a mixture of highly distorted 11<i>-cis</i> and 13-<i>cis</i> RSB directly formed from the excited state in 1.4 ps. P673 evolves from P682 in 500 ps and contains highly distorted 13-<i>cis</i> RSB, indicating that the 11-<i>cis</i> fraction in P682 converts to 13-<i>cis</i>. Next, P673 establishes an equilibrium with P595 in 1.2 µs, during which RSB converts to 11-<i>cis</i> and then further proceeds to P560 in 48 µs and P540 in 1.0 ms while remaining 11-<i>cis</i>. Hence, extensive isomeric switching occurs on the early ground state potential energy surface (PES) on the hundreds of ps to µs timescale before finally settling on a metastable 11-<i>cis</i> photoproduct. We propose that P682 and P673 are trapped high up on the ground-state PES after passing through either of two closely located conical intersections that result in 11-<i>cis</i> and 13-<i>cis</i> RSB. Co-rotation of C11=C12 and C13=C14 bonds results in a constricted conformational landscape that allows thermal switching between 11<i>-cis</i> and 13-<i>cis</i> species of highly strained RSB chromophores. Protein relaxation may release RSB strain, allowing it to evolve to a stable 11-<i>cis</i> isomeric configuration in microseconds.