Energy Transduction on the Nanosecond Time Scale: Early Structural Events in a Xanthopsin Photocycle

Abstract

Photoactive yellow protein (PYP) is a member of the xanthopsin family of eubacterial blue-light photoreceptors. On absorption of light, PYP enters a photocycle that ultimately transduces the energy contained in a light signal into an altered biological response. Nanosecond time-resolved x-ray crystallography was used to determine the structure of the short-lived, red-shifted, intermediate state denoted [pR], which develops within 1 nanosecond after photoelectronic excitation of the chromophore of PYP by absorption of light. The resulting structural model demonstrates that the [pR] state possesses the cis conformation of the 4-hydroxyl cinnamic thioester chromophore, and that the process of trans to cis isomerization is accompanied by the specific formation of new hydrogen bonds that replace those broken upon excitation of the chromophore. Regions of flexibility that compose the chromophore-binding pocket serve to lower the activation energy barrier between the dark state, denoted pG, and [pR], and help initiate entrance into the photocycle. Direct structural evidence is provided for the initial processes of transduction of light energy, which ultimately translate into a physiological signal.

Watching a signaling protein function in real time via 100-ps time-resolved Laue crystallography

“…4; Movies S3 and S4). Structures similar to pR 1 , pR 2 , and pB 0 have been reported previously (21,22,24,25), but pR 0 is unique. The simplest possible kinetic model connecting these intermediates is sequential with a reversible pR 0 -to-pR 1 transition, and with a pR 2 -to-pB 0 transition that short-circuits to the ground pG state approximately half the time (Fig.…”

“…3, curved arrows). This unusual structure has not been trapped in cryotrapping experiments (18)(19)(20), and its lifetime is too short to be captured in prior nanosecond time-resolved Laue crystallography experiments (21)(22)(23). The driving force for generating this intermediate is the trans-to-cis photoisomerization of pCA, during which the distance between the Cys69 sulfur and the terminal phenolate oxygen atom (Fig.…”

On the absorbance changes in the photocycle of the photoactive yellow protein: A quantum-chemical analysis

“…A structure determined by nanosecond time-resolved x-ray crystallography (10) has been assigned to the spectroscopically observed intermediate PYP L , which develops in solution within 1 ns after electronic excitation (5-7). It should be noted, however, that the assignment performed by Perman et al (10) is not confirmed by the present contribution.…”

“…The crystal structure obtained in the absence of light, which corresponds to PYP G (9)(10)(11), has the chromophore buried in a hydrophobic pocket with a trans conformation. The negative charge of the dark form is stabilized by a network of hydrogen bonds.…”

An efficient Monte Carlo method for calculating ab initio transition state theory reaction rates in solution

“…Recent advances in ultrafast lasers, 1, 2 X-ray 3 and other spectroscopies as well as in computational chemistry 4,5 have made possible the study of most gas-phase and some condensed-phase reactions in molecular detail. However, most experimental investigations of complex reaction mechanisms taking place in liquid environments are still inferred from isotope and solvent (medium) effects on the reaction rate.…”