Changes of Low-Frequency Vibrational Modes Induced by Universal¹⁵N- and¹³C-Isotope Labeling in S₂/S₁FTIR Difference Spectrum of Oxygen-Evolving Complex

Abstract

The effects of universal (15)N- and (13)C-isotope labeling on the low- (650-350 cm(-1)) and mid-frequency (1800-1200 cm(-1)) S(2)/S(1) Fourier transform infrared (FTIR) difference spectrum of the photosynthetic oxygen-evolving complex (OEC) were investigated in histidine-tagged photosystem (PS) II core particles from Synechocystis sp. PCC 6803. In the mid-frequency region, the amide II modes were predominantly affected by (15)N-labeling, whereas, in addition to the amide II, the amide I and carboxylate modes were markedly affected by (13)C-labeling. In the low-frequency region, by comparing a light-induced spectrum in the presence of ferricyanide as the electron acceptor, with the double difference S(2)/S(1) spectrum obtained by subtracting the Q(A)(-)/Q(A) from the S(2)Q(A)(-)/S(1)Q(A) spectrum, considerable numbers of bands found in the light-induced spectrum were assigned to the S(2)/S(1) vibrational modes in the unlabeled PS II core particles. Upon (13)C-labeling, changes were observed for most of the prominent bands in the S(2)/S(1) spectrum. Although (15)N-labeling also induced changes similar to those by (13)C-labeling, the bands at 616(-), 605(+), 561(+), 555(-), and 544(-) cm(-1) were scarcely affected by (15)N-labeling. These results indicated that most of the vibrational modes found in the low-frequency spectrum are derived from the coupling between the Mn-cluster and groups containing nitrogen and/or carbon atom(s) in a direct manner and/or through hydrogen bonding. Interestingly, an intensive band at 577(-) cm(-1) was not affected by (15)N- and (13)C-isotope labeling, indicating that this band arises from the mode that does not include either nitrogen or carbon atoms, such as the skeletal vibration of the Mn-cluster or stretching vibrational modes of the Mn-ligand.

Glutamate-354 of the CP43 polypeptide interacts with the oxygen-evolving Mn4Ca cluster of photosystem II: a preliminary characterization of the Glu354Gln mutant

“…Strickler et al unpublished data) indicate that the 1588(C) cm K1 band and features near 1556(K) and 1530(K) cm K1 correspond to n asym (COO K ) modes. The assignment of the 1588(C) cm K1 feature to a n asym (COO K ) mode is consistent with earlier global 15 N-labelling studies ( Noguchi et al 1995;Noguchi & Sugiura 2003;Kimura et al 2003;Yamanari et al 2004). One possibility is that the 1588(C) and 1531(K) cm K1 features in the double-difference spectrum correspond to the n asym (COO K ) mode of CP43-Glu354 in the S 2 and S 1 states, respectively, and that the 1362(C) and 1378(K) cm K1 features correspond to the n sym (COO K ) mode of CP43-Glu354 in the S 2 and S 1 states, respectively.…”

Protein ligation of the photosynthetic oxygen-evolving center

“…Preliminary global 15 N-labeling experiments conducted with both wild-type and CP43-Glu354Gln PSII particles (M. A. Strickler and R. J. Debus, unpublished) indicate that the positive band at 1588 cm −1 band corresponds to a ν asym (COO − ) mode, in agreement with earlier global 15 N-labeling studies [94][95][96]. However, these same experiments confirm that the mutation alters numerous amide I and amide II modes, raising the possibility that structural perturbations introduced by the mutation and/or the loss of the negative charge on CP43-Glu354 might change the distribution of Mn oxidation states within the Mn 4 Ca cluster and cause a different Mn ion to undergo oxidation during the S 1 → S 2 transition.…”

Structural Changes of D1 C-terminal α-Carboxylate during S-state Cycling in Photosynthetic Oxygen Evolution

“…PCC 6803, which lacks the D1 C-terminal extension and bears a His tag on the C-terminal of CP47 (Ala-344-stop) (17,26). For specific 13 C labeling of the ␣-carboxylate (1-C) or ␣-methyl carbon (3-C) of alanine, cells were grown photoautotrophically in BG11 medium supplemented with 0.5 mM L-[1-…”

“…Total incorporation of 13 C into 1-C or 3-C of Ala was evaluated with liquid chromatography/mass spectrometry analysis and found to be ϳ70% in thylakoid membranes. Thylakoid membranes obtained by disrupting cells using a Bead-Beater (Bio-Spec Products) followed by centrifugation were solubilized with n-dodecyl-␤-D-maltoside, and PS II core particles were chromatographically purified using a nickel column (17,26). The sample medium was replaced with 40 mM sucrose, 5 mM NaCl, 5 mM CaCl 2 , 10 mM Mes/NaOH, pH 6.0 (medium A) by precipitation of PS II core particles using 10% (w/v) polyethylene glycol 6000 and extensive washes with medium A for S 2 /S 1 FTIR difference spectra (17) or repeated ultrafiltration in the presence of 0.06% (w/v) n-dodecyl-␤-Dmaltoside for S-state cycling (25).…”

“…FTIR Measurements-Mid-frequency (1800 -1200 cm Ϫ1 ) and low frequency (670 -350 cm Ϫ1 ) FTIR spectra were recorded on a Bruker IFS66v/S spectrophotometer equipped with a mercury cadmium telluride detector (EG&G Optoelectronics D316/6) and on a Bomen MB102 spectrophotometer equipped with a silicon bolometer (infrared, HDL-5), respectively, at 0°C (Ϯ0.03°C) as described previously (25,26). The PS II core particles (ϳ40 g of chlorophyll) suspended in medium A were mixed with 1 l of sodium ferricyanide solution (100 mM stock) as an electron acceptor.…”