Molecular dynamics and quantum chemistry studies of the protein bacteriorhodopsin.

摘要

Molecular dynamics (MD) simulations and Quantum Chemistry (QC) calculations are employed to study the structure and function of the protein bacteriorhodopsin (bR), a 26 kD protein which residues in the purple membrane of the bacterium Halobacterium halobium. Bacteriorhodopsin undergoes a light-driven cyclic process, which pumps protons across the membrane, in order to maintain a proton gradient necessary for ATP synthesis. The cycle is initiated through a trans → cis isomerization of the chromophore retinal, which is bound to a lysine residue via a protonated Schiff base linkage. Initially, MD simulations are used to develop a refined three-dimensional structure of the protein, using the experimentally determined electron-microscopy structure of bR as a basis, and to determine equilibrium positions for several water molecules within the protein interior. MD simulations are then used to model the early isomerization reaction events in the bR trans and 13-cis photocycles. The simulations yield proposed structures consistent with the J, K, and L intermediates observed for these photocycles and offers a suggestion as to why an unprotonated retinal Schiff base intermediate, i.e., an M state, is not formed in the 13-cis photocycle. The simulations suggest that leakage from the 13-cis to the trans cycle arises due to an initial 13-cis,15- syn → all-trans,15-anti "bicycle peddle" photoisomerization. Electronic excitations and conformational potential surfaces of retinal in vacuo and in bacteriorhodopsin are determined by means of the ab initio CASSCF method. The calculations account for the protein environment by explicitly including all protein partial atomic charges into the electronic Hamiltonian. The calculations are applied to an ensemble of bacteriorhodopsins generated by molecular dynamics simulations using a CHARMM force field with special parameters for retinal torsions. Spectral calculations successfully reproduce a shift in absorption maxima between native bacteriorhodopsin and its D85N mutant and demonstrate that the broad absorption spectrum of bacteriorhodopsin is mainly caused by the fluctuations of retinal's internal degrees of freedom. Calculations are also carried out to describe the potential surface governing the dark adaptation of an ensemble of bacteriorhodopsins. The resulting rate of dark adaptation at room temperature k∼10-5s-1 and its 1000-fold enhancement through Asp-85 protonation are in agreement with experiment. The primary all-trans → 13- cis photoisomerization of retinal in bacteriorhodopsin has been investigated by means of quantum chemical and combined classical/quantum mechanical simulations employing the density matrix evolution method. Ab initio calculations on an analogue of a protonated Schiff base of retinal in vacuo reveal two closely lying excited states S1 and S2, the potential surfaces of which intersect along the reaction coordinate through an avoided crossing, and then exhibit a second weakly avoided crossing or a conical intersection with the ground state surface.