Structures of aspartic acid-96 in the L and N intermediates of bacteriorhodopsin: analysis by Fourier transform infrared spectroscopy.
The light-induced difference Fourier transform infrared spectrum between the L or N intermediate minus light-adapted bacteriorhodopsin (BR) was measured in order to examine the protonated states and the changes in the interactions of carboxylic acids of Asp-96 and Asp-115 in these intermediates. Vibrational bands due to the protonated and unprotonated carboxylic acid were identified by isotope shift and band depletion upon substitution of Asp-96 or -115 by asparagine. While the signal due to the deprotonation of Asp-96 was clearly observed in the N intermediate, this residue remained protonated in L. Asp-115 was partially deprotonated in L. The C = O stretching vibration of protonated Asp-96 of L showed almost no shift upon 2H2O substitution, in contrast to the corresponding band of Asp-96 or Asp-115 of BR, which shifted by 9-12 cm-1 under the same conditions. In the model system of acetic acid in organic solvents, such an absence of the shift of the C = O stretching vibration of the protonated carboxylic acid upon 2H2O substitution was seen only when the O-H of acetic acid is hydrogen-bonded. The non-hydrogen-bonded monomer showed the 2H2O-dependent shift. Thus, the O-H bond of Asp-96 enters into hydrogen bonding upon conversion of BR to L. Its increased hydrogen bonding in L is consistent with the observed downshift of the O-H stretching vibration of the carboxylic acid of Asp-96.
Maeda
A
A
Department of Biophysics, Faculty of Science, Kyoto University, Japan.
Sasaki
J
J
Shichida
Y
Y
Yoshizawa
T
T
Chang
M
M
Ni
B
B
Needleman
R
R
Lanyi
J K
JK
D000085:Acetates / Q000737:chemistry; D019342:Acetic Acid; D000595:Amino Acid Sequence; D001224:Aspartic Acid / Q000737:chemistry*; D001436:Bacteriorhodopsins / Q000737:chemistry*; D002247:Carbon Isotopes; D003903:Deuterium; D005583:Fourier Analysis; D006860:Hydrogen Bonding; D009154:Mutation; D011522:Protons; D012545:Schiff Bases; D013055:Spectrophotometry, Infrared; D013329:Structure-Activity Relationship
D000085:Acetates; D002247:Carbon Isotopes; D011522:Protons; D012545:Schiff Bases; D001224:Aspartic Acid; D001436:Bacteriorhodopsins; D003903:Deuterium; D019342:Acetic Acid