Solid state NMR study of [epsilon-13C]Lys-bacteriorhodopsin: Schiff base photoisomerization.
Previous solid state 13C-NMR studies of bacteriorhodopsin (bR) have inferred the C = N configuration of the retinal-lysine Schiff base linkage from the [14-13C]retinal chemical shift (1-3). Here we verify the interpretation of the [14-13C]-retinal data using the [epsilon-13C]lysine 216 resonance. The epsilon-Lys-216 chemical shifts in bR555 (48 ppm) and bR568 (53 ppm) are consistent with a C = N isomerization from syn in bR555 to anti in bR568. The M photointermediate was trapped at pH 10.0 and low temperatures by illumination of samples containing either 0.5 M guanidine-HCl or 0.1 M NaCl. In both preparations, the [epsilon-13C]Lys-216 resonance of M is 6 ppm downfield from that of bR568. This shift is attributed to deprotonation of the Schiff base nitrogen and is consistent with the idea that the M intermediate contains a C = N anti chromophore. M is the only intermediate trapped in the presence of 0.5 M guanidine-HCl, whereas a second species, X, is trapped in the presence of 0.1 M NaCl. The [epsilon-13C]Lys-216 resonance of X is coincident with the signal for bR568, indicating that X is either C = N anti and protonated or C = N syn and deprotonated.
Farrar
M R
MR
Department of Chemistry, Brandeis University, Waltham, Massachusetts 02254-9110.
Lakshmi
K V
KV
Smith
S O
SO
Brown
R S
RS
Raap
J
J
Lugtenburg
J
J
Griffin
R G
RG
Herzfeld
J
J
D001436:Bacteriorhodopsins / Q000737:chemistry* / Q000528:radiation effects; D055592:Biophysical Phenomena; D001703:Biophysics; D002247:Carbon Isotopes; D019617:Halobacterium salinarum / Q000737:chemistry / Q000528:radiation effects; D008239:Lysine / Q000737:chemistry / Q000528:radiation effects; D009682:Magnetic Resonance Spectroscopy; D010777:Photochemistry; D012545:Schiff Bases / Q000737:chemistry / Q000528:radiation effects; D013696:Temperature
D002247:Carbon Isotopes; D012545:Schiff Bases; D001436:Bacteriorhodopsins; D008239:Lysine