Coordinating the structural rearrangements associated with unidirectional proton transfer in the bacteriorhodopsin photocycle induced by deprotonation of the proton-release group: a time-resolved difference FTIR spectroscopic study.
In the photocycle of bacteriorhodopsin at pH 7, proton release from the proton releasing group (PRG) to the extracellular medium occurs during formation of the M intermediate. This proton release is inhibited at acidic pH, below the pK(a) of the PRG, approximately 6 in M, and instead occurs later in the cycle as the initial state is restored from the O intermediate. Here, structural changes related to deprotonation of the PRG have been investigated by time-resolved FTIR spectroscopy at 25 degrees C. The vibrational features at 2100-1790, 1730-1685, 1661, and 1130-1045 cm(-1) have greater negative intensity in the pure M-minus-BR spectrum and even in the M-minus-BR spectrum, that is present earlier together with the L-minus-BR spectrum, at pH 7, than in the corresponding M-minus-BR spectra at pH 5 or 4. The D212N mutation abolishes the decreases in the intensities of the broad feature between 1730 and 1685 cm(-1) and the band at 1661 cm(-1). The 1730-1685 cm(-1) feature may arise from transition dipole coupling of the backbone carbonyl groups of Glu204, Phe208, Asp212, and Lys216 interacting with Tyr57 and C(15)-H of the chromophore. The 1661 cm(-1) band, which is insensitive to D(2)O substitution, may arise by interaction of the backbone carbonyl of Asp212 with C(15)-H. The 2100-1790 cm(-1) feature with a trough at 1885 cm(-1) could be due to a water cluster. Depletion of these bands upon deprotonation of the PRG is attributable to disruption of a coordinated structure, held in place by interactions of Asp212. Deprotonation of the PRG is also accompanied by disruption of the interaction of the water molecule near Arg82. The liberated Asp212 may stabilize the protonated state of Asp85 and thus confer unidirectionality to the transport.
Morgan
Joel E
JE
Department of Biology, Center for Biotechnology and Interdisciplinary Studies, Room 2137, Rensselaer Polytechnic Institute, 110 Eighth Street, Troy, New York 12180, USA.
Vakkasoglu
Ahmet S
AS
Lanyi
Janos K
JK
Gennis
Robert B
RB
Maeda
Akio
A
D001436:Bacteriorhodopsins / Q000737:chemistry* / Q000378:metabolism* / Q000528:radiation effects; D019617:Halobacterium salinarum / Q000378:metabolism / Q000528:radiation effects; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D010777:Photochemistry; D013053:Spectrophotometry; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods; D013472:Sunlight; D014732:Vibration; D014867:Water / Q000032:analysis
D014867:Water; D001436:Bacteriorhodopsins