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PubMed1993

Secondary structure of M13 coat protein in phospholipids studied by circular dichroism, Raman, and Fourier transform infrared spectroscopy.

There is considerable uncertainty about the precise secondary structure adopted by the M13 coat protein when embedded in a phospholipid bilayer. Circular dichroism (CD) spectroscopy suggests that a major change in the structure of the coat protein occurs upon membrane insertion. It is reported that the structure of the protein in the membrane has only about 50% alpha-helix, the rest being mainly in a beta-sheet conformation, whereas the protein is almost completely alpha-helical when intact in the phage. In this study we have undertaken a spectroscopic analysis using Fourier transform infrared, Raman, and CD spectroscopy to characterize the secondary structure of M13 coat protein when present in membranes consisting of dioleoylphosphatidylglycerol and dimyristoylphosphatidylglycerol. In sharp contrast to earlier CD studies, our results indicate that the coat protein in its membrane-embedded state has a very high alpha-helical content with virtually no beta-sheet structures present. This result indicates that the structures of the coat protein when intact in the phage or when embedded in the membrane are similar. Although our results differ from earlier CD studies, they are consistent with a recent NMR study, which showed that the M13 coat protein in sodium dodecyl sulfate micelles is primarily alpha-helical with no evidence for beta-sheet structure [Henry, G. D., & Sykes, B.D. (1992) Biochemistry 31, 5284-5297]. These results lead to the conclusion that the M13 coat protein can insert from the membrane-bound state into a virus particle with a similar secondary structure, without large energy implications.(ABSTRACT TRUNCATED AT 250 WORDS)

Bacteriophage M13↗

PubMed1989

Correlation of structure and function in the Ca2+-ATPase of sarcoplasmic reticulum: a Fourier transform infrared spectroscopy (FTIR) study on the effects of dimethyl sulfoxide and urea.

The effect of dimethyl sulfoxide (DMSO) on the structure of sarcoplasmic reticulum was analyzed by Fourier transform infrared (FTIR) and fluorescence spectroscopy. Exposure of sarcoplasmic reticulum vesicles to 35% DMSO (v/v) at 2 degrees C for several hours in a D2O medium produced no significant change in the phospholipid and protein Amide I regions of the FTIR spectra, but the intensity of the Amide II band decreased, presumably due to proton/deuterium exchange. At 40% to 60% DMSO concentration a shoulder appeared in the FTIR spectra at 1630 cm-1, that is attributed to the formation of new beta or random coil structures; irreversible loss of ATPase activity accompanied this change. At 70% DMSO concentration the intensity of the main Amide I band at 1639 cm-1 decreased and a new band appeared at 1622 cm-1, together with a shoulder at 1682 cm-1. These changes indicate an abrupt shift in the conformational equilibrium of Ca2+-ATPase from alpha to beta structure or to a new structure characterized by weaker hydrogen bonding. Decrease of ionization of aspartate and glutamate carboxyl groups in the presence of DMSO may also contribute to the change in intensity at 1622 cm-1. The changes were partially reversed upon removal of DMSO. Exposure of sarcoplasmic reticulum vesicles to 1.5 kbar pressure for 1 h at 2 degrees C in an EGTA-containing (low Ca2+) medium causes irreversible loss of ATPase activity, with the appearance of new beta structure, and abolition of the Ca2+-induced fluorescence response of FITC covalently bound to the Ca2+-ATPase; DMSO (35%) stabilized the Ca2+-ATPase against pressure-induced changes in structure and enzymatic activity, while urea (0.8 M) had the opposite effect.

Animals↗

PubMed2005

Characterization of historic silk by polarized attenuated total reflectance Fourier transform infrared spectroscopy for informed conservation.

When assessing historic textiles and considering appropriate conservation, display, and storage strategies, characterizing the physical condition of the textiles is essential. Our work has concentrated on developing nondestructive or micro-destructive methodologies that will permit this. Previously, we have demonstrated a correlation between the physical deterioration of unweighted and "pink" tin (IV) chloride weighted silk and certain measurable spectroscopic and chromatographic signatures, derived from polarized Fourier transform infrared attenuated total reflectance (FTIR-ATR) spectroscopy (Pol-ATR) and high-performance liquid chromatography (HPLC) microsampling analyses. The application of the Pol-ATR technique to aged silk characterization has now been extended to include a more comprehensive range of weighting methods and aging regimes. This was intended to replicate the full spectrum of states of deterioration observed in silk textiles, from pristine to heavily degraded. Breaking strength was employed as a measure of the physical integrity of the fibers, and, as expected, decreased with aging. An orientational crystallinity parameter, reflecting the microstructural ordering of the fibroin polymer within the fibers, was derived from the Pol-ATR spectra. A good correlation was observed between the breaking strength of the variety of fibers and this parameter. This suggests that the physical state of historic silk fabrics might be adequately characterized for conservation purposes by such indirect micromethodology.

Animals↗

PubMed1994

The secondary structure of the von Willebrand factor type A domain in factor B of human complement by Fourier transform infrared spectroscopy. Its occurrence in collagen types VI, VII, XII and XIV, the integrins and other proteins by averaged structure predictions.

The type A domain of the von Willebrand Factor is found also in the complement proteins factor B (FB), C2, CR3 and CR4, the integrins, collagen types VI, VII, XII and XIV, and other proteins. FB is a component of the alternative pathway of the complement system of immune defence, and is cleaved into the fragments Bb and Ba during complement activation. Bb contains a von Willebrand Factor type A (vWF) domain of unknown secondary structure and a serine proteinase (SP) domain, whereas Ba contains three short consensus repeat/complement control protein (SCR/CCP) domains. Fourier transform infrared (FT-IR) spectroscopy on a recombinant vWF domain and on FB and its Bb and Ba fragments shows a broad amide I band. In H2O buffer, second derivative spectra of the amide I band show subcomponents at 1654 to 1657 cm-1, which is typical of alpha-helix, and at 1676 to 1685 cm-1 and 1636 to 1637 cm-1, which are typical of beta-strand. alpha-Helix was detected in the vWF domain, the Bb fragment and FB, and the proportion of alpha-helix present decreased in that order. This shows that the vWF domain contains appreciable amounts of alpha-helix, while the SP and SCR/CCP domains are almost entirely beta-sheet in their secondary structures. Quantitative integration of the vWF FT-IR spectrum showed that this contained 31% alpha-helix and 36% beta-sheet. In 2H2O buffer, the alpha-helix content in the vWF domain is sensitive to the solvent, while the beta-sheet content is less so. An alignment of 75 vWF type A sequences from 25 proteins was used for averaged secondary structure predictions of the total length of 206 residues by the Robson and Chou-Fasman methods. In support of the FT-IR analysis, a total of at least five well-predicted alpha-helices (35% of residues) and at least five well-predicted beta-strands (21% of residues) were identified by both predictive methods, all of which were interspersed by regions of coil or turn conformations. Eight of the ten predicted alpha-helices and beta-strands form an alternating arrangement with each other. Since the predicted alpha-helices are mostly amphipathic, and since the alpha-helix FT-IR band is sensitive to solvent, the alpha-helices are inferred to be on the protein surface.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗

PubMed2003

Studies on the interaction of total saponins of panax notoginseng and human serum albumin by Fourier transform infrared spectroscopy.

Total saponins of panax notoginseng (TPNS), isolated from the roots of panax notoginseng (Burk) F.H. Chen, have been considered as the main active components of San-Chi and have various therapeutical actions. Their interactions with human serum albumin have been investigated by Fourier transformed infrared spectrometry and fluorescence methods. The results showed that TPNS combined with HSA through C=O and C-N groups of polypeptide chain. The drug-protein combination caused the significant loss of alpha-helix structure and the microenvironment changes of the tyrosine residues in protein at higher drug concentration. Combining the curve-fitting results of amide I and amide III bands, the alterations of protein secondary structure after drug complexation were quantitatively determined. The alpha-helix structure has a decrease of approximately 6%, from 55 to 49% and the beta-sheet increased approximately 3%, from 23 to 26% at high drug concentration. However, no major alterations were observed for the beta-turn and random coil structures up on drug-protein binding.

Chromatography, High Pressure Liquid↗

PubMed2004

Probing the Q-proton pathway of ba3-cytochrome c oxidase by time-resolved Fourier transform infrared spectroscopy.

In cytochrome c oxidase, the terminal respiratory enzyme, electron transfers are strongly coupled to proton movements within the enzyme. Two proton pathways (K and D) containing water molecules and hydrophobic amino acids have been identified and suggested to be involved in the proton translocation from the mitochondrial matrix or the bacterial cytoplasm into the active site. In addition to the K and D proton pathways, a third proton pathway (Q) has been identified only in ba3-cytochrome c oxidase from Thermus thermophilus, and consists of residues that are highly conserved in all structurally known heme-copper oxidases. The Q pathway starts from the cytoplasmic side of the membrane and leads through the axial heme a3 ligand His-384 to the propionate of the heme a3 pyrrol ring A, and then via Asn-366 and Asp-372 to the water pool. We have applied FTIR and time-resolved step-scan Fourier transform infrared (TRS2-FTIR) spectroscopies to investigate the protonation/deprotonation events in the Q-proton pathway at ambient temperature. The photolysis of CO from heme a3 and its transient binding to CuB is dynamically linked to structural changes that can be tentatively attributed to ring A propionate of heme a3 (1695/1708 cm(-1)) and to deprotonation of Asp-372 (1726 cm(-1)). The implications of these results with respect to the role of the ring A propionate of heme a3-Asp372-H2O site as a proton carrier to the exit/output proton channel (H2O pool) that is conserved among all structurally known heme-copper oxidases, and is part of the Q-proton pathway in ba3-cytochrome c oxidase, are discussed.

Cell Membrane↗

PubMed2003

Molecular recognition of 7-(2-octadecyloxycarbonylethyl)guanine to cytidine at the air/water interface and LB film studied by Fourier transform infrared spectroscopy.

Monolayer behavior of a nucleolipid amphiphile, 7-(2-octadecyloxycarbonylethyl)guanine (ODCG), on aqueous cytidine solution was investigated by means of surface-molecular area (pi-A) isotherms. It indicates that molecular recognition by hydrogen bonding is present between ODCG monolayer and the cytidine in subphase. The Fourier transform infrared (FTIR) transmission spectroscopic result indicates that the cytidine molecules in the subphase can be transferred onto solid substrates by Langmuir-Blodgett (LB) technique as a result of the formation of Watson-Crick base-pairing at the air/water interface. Investigation by rotating polarized FTIR transmission also suggests that the headgroup recognition of this amphiphile to the dissolved cytidine influence the orientation of the tailchains.

Cytidine↗

PubMed2001

Studies on charge transfer properties from mixture of Schiff base and zinc complex in Langmuir-Blodgett film by UV-vis absorption and Fourier transform infrared spectroscopy.

A novel mixed Langmuir Blodgett (LB) film based on a 1:1 (molar ratio) mixture of a non-amphiphile complex (Et4N)2[Zn(dmit)2] (H2dmit = 4,5-dimercapto-1,3-dithiole-2-thione) and Schiff base amphiphile 2,4-dihydroxy-N-octadecylbenzylideneamine (SBC18) was constructed and characterized by Fourier transform infrared (FTIR) spectra and UV-vis electronic absorption spectra. After iodine was doped in situ, magnitude of charge transfer increases, which was demonstrated by FTIR and UV-vis absorption spectral analysis.

Benzylidene Compounds↗

PubMed1996

Characterization of dipalmitoylphosphatidylcholine liposomes containing a soybean-derived sterylglucoside mixture by differential scanning calorimetry, Fourier transform infrared spectroscopy, and enzymatic assay.

Previously, we reported that dipalmitoylphosphatidylcholine liposomes (DPPC-liposomes) containing soybean-derived steryl glucoside mixtures (SG) (DPPC/SG-liposomes) accumulated in the liver, especially in parenchymal cells. DPPC/SG-liposomes and a mixture of DPPC and SG (DPPC/SG mixture) were compared by means of differential scanning calorimetry, Fourier transform infrared (FT-IR), and enzymatic assays. The results suggested that the maximum molar mixing ratios of DPPC and SG in a powder was DPPC:SG = 7:2.2. Enzymatic assays indicated that the glucose group of SG projected outward from the liposomal surface and that the amount of SG on the liposomal surface was limited, the maximum mole fraction of SG in DPPC/SG-liposomes being 0.27 (DPPC: SG = 7:2.6). FT-IR spectra indicated that the glucose group of SG interacts with the phosphate group of DPPC on the surface of liposomes, since the phosphate symmetric and asymmetric stretching bands of DPPC/ SG-liposomes were shifted to lower frequencies with increasing SG. These results suggested that the glucose group of SG projecting outward from the liposomal membrane contributes to the hepatic cellular distribution of DPPC/SG-liposomes.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2004

Characterization of commercial carrageenans by Fourier transform infrared spectroscopy using single-reflection attenuated total reflection.

The purity and composition of commercial carrageenans vary widely and, therefore, have to be checked prior to their use in the food industry. Infrared spectroscopy is an alternative method to the expensive and time-consuming wet chemical and NMR methods to characterize carrageenan samples. The use of an attenuated total reflection accessory coupled to a Fourier transform infrared spectrophotometer allows a direct analysis of the sample without any preparation step, which is an additional benefit for the rapid identification check of raw material at reception in an industrial environment. Using a set of calibration samples, three multivariate calibrations were developed to predict the total carrageenan content as well as the molar ratio of kappa- and iota-carrageenans. A validation with an independent set of samples confirmed the robustness of the calibrations and the accuracy of the predictions. The accuracies of the calibrations given by their respective standard errors of prediction are 5.6 g/100 g, and 6.1 mol %, and 6.6 mol %, respectively, for the total carrageenan content and the molar ratios of kappa- and iota-carrageenans. The total preparation and analysis time is <5 min per sample.

Calibration↗

PubMed1993

Fourier transform infrared spectroscopy characterization of the lamellar and nonlamellar structures of free lipid A and Re lipopolysaccharides from Salmonella minnesota and Escherichia coli.

The structural polymorphism of free lipid A and deep rough mutant lipopolysaccharide (LPS Re) from Salmonella minnesota strain R595 and Escherichia coli strain F515 was characterized by Fourier transform infrared (IR) spectroscopy. For this, the beta <--> alpha phase states and the three-dimensional supramolecular structures, the latter deduced from small-angle synchrotron radiation x-ray diffraction, were investigated at different water contents, Mg2+ concentrations, and temperatures. The analysis of the IR data for vibrations originating from the hydrophobic moiety shows that the beta <--> alpha acyl chain melting is strongly expressed only for the stretching and scissoring modes of the methylene groups. Vibrational groups originating from the interface region sense the acyl chain melting well (ester carbonyl bands) or only weakly (amide bands), and those resulting from the pure polar moiety not at all. From the x-ray data, the existence of lamellar (L), different cubic, and, for lipid A and LPS R595, also inverted hexagonal (HII) structures could be proven in the temperature range 20-80 degrees C with cubic <--> cubic and cubic <--> HII transitions for the Mg(2+)-free and L <--> HII transitions for the Mg(2+)-containing samples. These structural transitions can be characterized most readily by specific changes of the vibrational bands resulting from the interface region: the ester carbonyl and the amide bands. The magnitude of the changes corresponds to that of the structural rearrangement, i.e., is highest for the L <--> HII, lower for the cubic <--> HII, and lowest for the cubic <--> cubic transitions. The structural transitions are only marginally expressed for vibrational bands of the hydrophobic moiety. Similarly, the band contours of vibrations from the hydrophilic region are no indicators of the structural reorientations except for the carboxylate bands of LPS Re. Particularly the stretching vibrations of the phosphate groups are nearly completely invariant; the absolute values of their half bandwidths, however, differ significantly for lipid A and LPS Re, which seems to be of biological relevance. The ability of IR spectroscopy to detect supramolecular changes also beyond the measurability by x-ray diffraction, i.e., at water contents > 95 to 99.5%, is demonstrated.

Biophysical Phenomena↗

PubMed1997

The tertiary structural changes in bacteriorhodopsin occur between M states: X-ray diffraction and Fourier transform infrared spectroscopy.

The tertiary structural changes occurring during the photocycle of bacteriorhodopsin (BR) are assigned by X-ray diffraction to distinct M states, M1 and M2. Purple membranes (PM) of the mutant Asp96Asn at 15, 57, 75 and 100% relative humidity (r.h.) were studied in a parallel X-ray diffraction and Fourier transform infrared (FTIR) spectroscopic investigation. Light-dependent conformational changes of BR-Asp96Asn are observed at high hydration levels (100 and 75% r.h.) but not in partially dehydrated samples (57 and 15% r.h.). The FTIR spectra of continuously illuminated samples at low and high hydration, despite some differences, are characteristic of the M intermediate. The changes in diffraction patterns of samples in the M2 state are of the same magnitude as those of wild-type samples trapped with GuaHCl in the M(G) state. Additional large changes in the amide bands of the FTIR spectra occur between M2 and M(G). This suggests, that the tertiary structural changes between M1 and M2 are responsible for the switch opening the cytoplasmic half-channel of BR for reprotonation to complete the catalytic cycle. These tertiary structural changes seem to be triggered by a charge redistribution which might be a common feature of retinal proteins also in signal transduction.

Amino Acid Sequence↗

PubMed1988

Characterization of complex gramicidin monolayers by light reflection and Fourier transform infrared spectroscopy.

The reflectivities of monomolecular films of water-insoluble fatty alcohols and fatty acid methyl esters are measured at the air/water interface. A correlation between chain length and reflectivity of the monofilm is established which agrees with calculated values derived from a theoretical model. The correlation is used to estimate thickness of a monolecular film of gramicidin A. Fourier transform infrared (FTIR) spectroscopy is applied to transferred mixed layers of ion-channel-forming gramicidin A and dioctadecyldimethylammonium bromide in order to evaluate the structure of gramicidin. Transfer conditions for these monofilms are elaborated. Results of the reflection method and FTIR spectroscopy demonstrate that gramicidin exists as double-stranded beta-helix inside the monolayer at a lateral pressure similar to that found in biomembranes.

Fourier Analysis↗

PubMed2025

Fourier transform infrared spectroscopy enables rapid species discrimination across Malassezia and strain-level typing in M. pachydermatis.

Malassezia pachydermatis is a zoophilic yeast found on the skin and in the outer ear canal of many mammals. It normally maintains a commensal lifestyle but can cause dermatitis and otitis in predisposed hosts, particularly in atopic dogs. M. pachydermatis is genetically diverse, with strains clustering into at least three phylogroups based on molecular typing, a pattern we now confirm through whole-genome sequencing (WGS). Accurate species and strain-level identification is essential for understanding its epidemiology, pathogenic potential, and response to treatment. In this study, we established Fourier Transform Infrared (FTIR) spectroscopy as a rapid, cost-effective method for distinguishing M. pachydermatis from other Malassezia species, including M. globosa, M. furfur, M. restricta, and M. sympodialis. Within M. pachydermatis, FTIR spectroscopy resolved even closely related strains with high accuracy producing clusters congruent with WGS-based phylogeny. The incorporation of an Artificial Neural Network classifier further enhanced the discriminatory power, enabling robust and automated strain assignment. These findings demonstrate the potential of FTIR spectroscopy as a practical tool for large-scale epidemiological surveillance of M. pachydermatis and for clinical and veterinary applications where strain-level identification could inform treatment and management of Malassezia-associated diseases.

Fourier Transform Infrared (FTIR) spectroscopy↗

PubMed1975

Sulfhydryl groups in hemoglobin. A new molecular probe at the alpha1 beta 1 interface studied by Fourier transform infrared spectroscopy.

Infrared absorption bands due to sulfhydryl groups (VSH) of alpha-104(G11) and beta-112 (G14) cysteine residues of human carboxyhemoglobin (HbCO) have been observed near 2560 cm- minus 1 by use of Fourier transform infrared (FTIR) spectroscopy. The beta-93 cysteine SH groups absorb infrared radiation so weakly that they are not distinguished from background. Only single SH absorption bands due to the alpha-104 cysteines of pig and horse hemoglobin are observed. The SH absorption bands from human HbCO disappear in alkali, are broadened by detergent or guanidinium chloride, and show a complex titration curve, and an isotopic frequency shift (VSD/VSH equals to 0.7267) virtually identical with that reported for methanethiol. The integrated absorption coefficient (epsilonmM(area)) for 0.1 M ethanetiol increased with H-bond acceptor solvents in the order: CCl4 (0.07), water (0.21), acetone (0.43), and N,N-dimethyl-acetamide (1.35 mM- minus 1 cm- minus 2). Comparison of the integrated absorption coefficients for the alpha-104 cysteine SH (2.43), and the beta-112 SH (0.80), of human HbCO with those of ethanetiol solutions suggested specifically H-bonded structures with peptide carbonyl groups 4 (or 3) residues back in the G helices. This was found to agree with a molecular model of the alpha-chain G helix of horse HbO2 built to coordinates from M. F. Perutz.

Animals↗

PubMed2005

Scanning tunneling microscopy, Fourier transform infrared spectroscopy, and electrochemical characterization of 2-naphthalenethiol self-assembled monolayers on the Au surface: a study of bridge-mediated electron transfer in Ru(NH3)6(2+)/Ru(NH3)6(3+) redox reactions.

We have studied the structure, adsorption kinetics, and barrier properties of self-assembled monolayers of 2-naphthalenethiol on Au using electrochemical techniques, grazing-angle Fourier transform infrared (FTIR) spectroscopy, and scanning tunneling microscopy (STM). The results of cyclic voltammetric and impedance measurements using redox probes show that 2-naphthalenethiol on Au forms a stable and reproducible, but moderately blocking, monolayer. Annealing of the self-assembled monolayer (SAM)-modified surface at 72 +/- 2 degrees C remarkably improves the blocking property of the monolayer of 2-naphthalenethiol on Au. From the study of kinetics of SAM formation, we find that the self-assembly follows Langmuir adsorption isotherm. Our STM and FTIR results show that the molecules are adsorbed with the naphthalene ring tilted from the surface normal by forming a square root 3 x 3 R30 degrees overlayer structure. From our studies, we conclude that the electron-transfer reaction of ferro/ferricyanide in the freshly formed monolayer occurs predominantly through the pinholes and defects present in the monolayer. However, in the case of thermally annealed specimen, although the ferro/ferricyanide reaction is almost completely blocked, the electron-transfer reaction of hexaammineruthenium(III) chloride is not significantly inhibited. It is proposed that the electron-transfer reaction in the case of the ruthenium redox couple takes place by a tunneling mechanism through the high-electron-density aromatic naphthalene ring acting as a bridge between the monolayer-modified electrode and the ruthenium complex.

Journal Article↗

PubMed1995

Experimental evidence for hydrogen-bonded network proton transfer in bacteriorhodopsin shown by Fourier-transform infrared spectroscopy using azide as catalyst.

Experimental evidence for proton transfer via a hydrogen-bonded network in a membrane protein is presented. Bacteriorhodopsin's proton transfer mechanism on the proton uptake pathway between Asp-96 and the Schiff base in the M-to-N transition was determined. The slowdown of this transfer by removal of the proton donor in the Asp-96-->Asn mutant can be accelerated again by addition of small weak acid anions such as azide. Fourier-transform infrared experiments show in the Asp-96-->Asn mutant a transient protonation of azide bound to the protein in the M-to-N transition and, due to the addition of azide, restoration of the IR continuum band changes as seen in wild-type bR during proton pumping. The continuum band changes indicate fast proton transfer on the uptake pathway in a hydrogen-bonded network for wild-type bR and the Asp-96-->Asn mutant with azide. Since azide is able to catalyze proton transfer steps also in several kinetically defective bR mutants and in other membrane proteins, our finding might point to a general element of proton transfer mechanisms in proteins.

Amino Acid Sequence↗

PubMed1999

Low-frequency fourier transform infrared spectroscopy of the oxygen-evolving and quinone acceptor complexes in photosystem II.

The low-frequency (<1000 cm-1) region of the IR spectrum has the potential to provide detailed structural and mechanistic insight into the photosystem II/oxygen evolving complex (PSII/OEC). A cluster of four manganese ions forms the core of the OEC and diagnostic manganese-ligand and manganese-substrate modes are expected to occur in the 200-900 cm-1 range. However, water also absorbs IR strongly in this region, which has limited previous Fourier transform infrared (FTIR) spectroscopic studies of the OEC to higher frequencies (>1000 cm-1). We have overcome the technical obstacles that have blocked FTIR access to low-frequency substrate, cofactor, and protein vibrational modes by using partially dehydrated samples, appropriate window materials, a wide-range MCT detector, a novel band-pass filter, and a closely regulated temperature control system. With this design, we studied PSII/OEC samples that were prepared by brief illumination of O2 evolving and Tris-washed preparations at 200 K or by a single saturating laser flash applied to O2 evolving and inhibited samples at 250 K. These protocols allowed us to isolate low-frequency modes that are specific to the QA-/QA and S2/S1 states. The high-frequency FTIR spectra recorded for these samples and parallel EPR experiments confirmed the states accessed by the trapping procedures we used. In the S2/S1 spectrum, we detect positive bands at 631 and 602 cm-1 and negative bands at 850, 679, 664, and 650 cm-1 that are specifically associated with these two S states. The possible origins of these IR bands are discussed. For the low-frequency QA-/QA difference spectrum, several modes can be assigned to ring stretching and bending modes from the neutral and anion radical states of the quinone acceptor. These results provide insight into the PSII/OEC and demonstrate the utility of FTIR techniques in accessing low-frequency modes in proteins.

Electron Spin Resonance Spectroscopy↗