TY - JOUR
T1 - Detection of threonine structural changes upon formation of the M-intermediate of bacteriorhodopsin
T2 - Evidence for assignment to Thr-89
AU - Liu, Xiaomei
AU - Lee, Min Joo
AU - Coleman, Matthew
AU - Rath, Parshuram
AU - Nilsson, Anders
AU - Fischer, Wolfgang B.
AU - Bizounok, Marina
AU - Herzfeld, Judith
AU - Jan Karstens, Willem F.
AU - Raap, Jan
AU - Lugtenburg, Johan
AU - Rothschild, Kenneth J.
N1 - Funding Information:
We wish to thank Drs. J. Olejnik, S. Williams and L. Smilowitz for helpful discussions. This work was supported by a grant from the NSF (MCB9419059) to K.J.R. and from the NIH (GM36810) to J.H. A.N. is supported by a postdoctoral fellowship from the Wenner-Gren Center Foundation (Sweden). M.C. is supported by an NIH Molecular Biophysics Training Grant (GM08291-06).
PY - 1998/7/20
Y1 - 1998/7/20
N2 - The behavior of threonine residues in the bacteriorhodopsin (bR) photocycle has been investigated by Fourier transform infrared difference spectroscopy. l-Threonine labeled at the hydroxyl group with 18O (l-[3-18O]threonine) was incorporated into bR and the bR→M FTIR difference spectra measured. Bands are assigned to threonine vibrational modes on the basis of 18O induced isotope frequency shifts and normal mode calculations. In the 3500 cm-1 region, a negative band is assigned to the OH stretch of threonine. In the 1125 cm-1 region, a negative band is assigned to a mixed CH3 rock/CO stretch mode. The frequency of both these bands indicates the presence of at least one hydrogen bonded threonine hydroxyl group in light adapted bR which undergoes a change in structure by formation of the M intermediate. Spectral changes induced by the substitution Thr-89→Asn but not Thr-46→Asn or Asp-96→Asn are consistent with the assignment of these bands to Thr-89. These results along with another related study on the mutant Thr-89→Asn indicate that the active site of bR includes Thr-89 and that its interaction with the retinylidene Schiff base and Asp-85 may play an important role in regulating the color of bacteriorhodopsin and the transfer of a proton to the Schiff base. Copyright (C) 1998 Elsevier Science B.V.
AB - The behavior of threonine residues in the bacteriorhodopsin (bR) photocycle has been investigated by Fourier transform infrared difference spectroscopy. l-Threonine labeled at the hydroxyl group with 18O (l-[3-18O]threonine) was incorporated into bR and the bR→M FTIR difference spectra measured. Bands are assigned to threonine vibrational modes on the basis of 18O induced isotope frequency shifts and normal mode calculations. In the 3500 cm-1 region, a negative band is assigned to the OH stretch of threonine. In the 1125 cm-1 region, a negative band is assigned to a mixed CH3 rock/CO stretch mode. The frequency of both these bands indicates the presence of at least one hydrogen bonded threonine hydroxyl group in light adapted bR which undergoes a change in structure by formation of the M intermediate. Spectral changes induced by the substitution Thr-89→Asn but not Thr-46→Asn or Asp-96→Asn are consistent with the assignment of these bands to Thr-89. These results along with another related study on the mutant Thr-89→Asn indicate that the active site of bR includes Thr-89 and that its interaction with the retinylidene Schiff base and Asp-85 may play an important role in regulating the color of bacteriorhodopsin and the transfer of a proton to the Schiff base. Copyright (C) 1998 Elsevier Science B.V.
KW - Bacteriorhodopsin
KW - Fourier transform infrared
KW - Isotope labeling
KW - Proton transport
KW - Site directed mutagenesis
KW - Threonine
UR - http://www.scopus.com/inward/record.url?scp=0032551761&partnerID=8YFLogxK
U2 - 10.1016/S0005-2728(98)00088-7
DO - 10.1016/S0005-2728(98)00088-7
M3 - Article
C2 - 9711293
AN - SCOPUS:0032551761
SN - 0005-2728
VL - 1365
SP - 363
EP - 372
JO - Biochimica et Biophysica Acta - Bioenergetics
JF - Biochimica et Biophysica Acta - Bioenergetics
IS - 3
ER -