Energy Transduction and Sensory Signalling in Haloarchaea Structural Characterisation of the Photocycles of Bacteriorhodopsin and Sensory Rhodopsin II
Archaeal rhodopsins belong a subfamily of heptahelical transmembrane proteins. All contain a buried retinal chromophore covalently bound to a conserved lysine residue in helix G via a protonated Schiff base. Upon light-activation, retinal is isomerised from an all-trans to a 13-cis configuration, which initiates a sequence of specific structural rearrangements. For bacteriorhodopsin, this results in the vectorial transport of a proton across the cell membrane. For sensory rhodopsin II this initiates a photosignal, which is communicated to a tightly bound transducer molecule. The transducer, in turn, triggers a phosphorylation cascade inducing a negative phototaxis response in the host archaea.
The kinetic crystallography studies presented in this thesis reveal the early structural rearrangements of bacteriorhodopsin's photocycle, leading up to the primary proton transfer between the protonated Schiff base and the negatively charged Asp85. Upon photoactivation, retinal isomerisation induces a structural rearrangement in the attached lysine residue which disorders a key water molecule. In the resting state, this water molecule is hydrogen bonded to both the primary proton donor and acceptor. Its removal initiates a sequence of events, reversing the relative proton affinities of the primary proton transfer groups. Moreover, a local bend in helix C, partially driven by the mutual electrostatic attraction between the Schiff base and Asp85, facilitates the creation of a low barrier pathway for the proton transfer. From the early rearrangements observed here and the structural results from later intermediates, a model for vectorial proton pumping is proposed.
The mechanism of proton pumping by bacteriorhodopsin is contrasted against the early structural rearrangements observed in the photocycle of sensory rhodopsin II. As indicated by their strong structural relationship, the mechanism for the primary proton transfer event appears to be conserved. Moreover, the putative binding site for the cognate transducer suggests a common structural theme connecting the latter half of the bacteriorhodopsin photocycle and the mechanism of signal relay in sensory rhodopsin II. However, slight modifications of the retinal binding pocket lead to subtle differences in the early relaxation of photoisomerised retinal. These results illustrate how the two systems have been optimised for the different functions of energy transduction and sensory signalling.