Low Temperature Absorption Spectra of Chlorophyll a in Polar and Nonpolar Solvents

Absorption spectra of chlorophyll a were measured in polar and non-polar solvents, as a function of temperature from 298 degrees to 77 degrees K. Both dilute and concentrated solutions were examined. In both types of solvents at room temperature, the absorption spectra of concentrated solutions differ from dilute ones in that the half width of the main red absorption band is greater, and all bands are shifted to longer wavelengths. These differences are largely due to the presence of dimers when the pigment concentration is high. In dilute ethanol solutions, where the chlorophyll is unassociated, cooling causes a red shift in all bands which is due to the increased polarity of the solvent at low temperature. On cooling at high concentrations in ethanol and EPA, a new band appears near 700 nm. This band is attributed to dimers present prior to cooling, but absorbing at shorter wavelengths at room temperature. In nonpolar solvents, a band near 700 nm appears at the solvent freezing point. In these solvents, the "700" nm absorption is attributed to dimers, and/or small polymers, partly formed by cooling. A change in aggregate geometry when the solvent becomes viscous or frozen can account for the appearance of this "700" nm absorption band at low temperature, in polar and nonpolar media.     

Detection of polymeric forms of dihydroquercetin by optical absorption and light scattering

Two methods for the detection of long polymers in dihydroquercetin (DHQ) preparations has been developed. The first method is based on UV spectrophotometry. It was shown that the quantity of long polymers in aqueous solutions can be estimated by the ratio of the absorption bands at 328 and 290 nm, since the 328-nm band was attributed to the monomeric form of DHQ, whereas the 290-nm band was attributed to both the monomeric and polymeric forms. The second method is based on the high-sensitive measurement of light-scattering intensity in aqueous solutions of diluted DHQ preparations using a spectrofluorometer with crossed monochromators. It has been shown that the filtration of DHQ solutions through Millipore filters with a pore diameter of 0.05–0.45 microns makes it possible to nearly completely eliminate long polymers and their aggregates. Long polymers at high concentrations can aggregate. The longest polymers and their aggregates may be 0.1 mm in length, which leads to fluctuations in the light-scattering intensity on the second and minute time scale.     

西红柿的光生态研究

采用叶片的反射光谱测量来研究西红柿的光生态。西红柿叶片在４５０ｎｍ处、６８０ｎｍ处各有一条带宽为１２０ｎｍ的吸收带,这说明蓝、红光能够促进西红柿的生长。往聚乙烯树脂中添加荧光助剂、着色剂模拟西红柿作用光谱制成农膜。用这种农膜苫盖温室大棚进行西红柿生产。通过产量调查来进一步认证西红柿的光生态。     

The nature of a chlorophyll ligand in Lhca proteins determines the far red fluorescence emission typical of photosystem I

Photosystem I of higher plants is characterized by a typically long wavelength fluorescence emission associated to its light-harvesting complex I moiety. The origin of these low energy chlorophyll spectral forms was investigated by using site-directed mutagenesis of Lhca1-4 genes and in vitro reconstitution into recombinant pigment-protein complexes. We showed that the red-shifted absorption originates from chlorophyll-chlorophyll (Chl) excitonic interactions involving Chl A5 in each of the four Lhca antenna complexes. An essential requirement for the presence of the red-shifted absorption/fluorescence spectral forms was the presence of asparagine as a ligand for the Chl a chromophore in the binding site A5 of Lhca complexes. In Lhca3 and Lhca4, which exhibit the most red-shifted red forms, its substitution by histidine maintains the pigment binding and, yet, the red spectral forms are abolished. Conversely, in Lhca1, having very low amplitude of red forms, the substitution of Asn for His produces a red shift of the fluorescence emission, thus confirming that the nature of the Chl A5 ligand determines the correct organization of chromophores leading to the excitonic interaction responsible for the red-most forms. The red-shifted fluorescence emission at 730 nm is here proposed to originate from an absorption band at approximately 700 nm, which represents the low energy contribution of an excitonic interaction having the high energy band at 683 nm. Because the mutation does not affect Chl A5 orientation, we suggest that coordination by Asn of Chl A5 holds it at the correct distance with Chl B5.     

Thermal sensitivity of the red absorption tail of the photosystem II reaction center complex.

The red tail of the absorption spectrum of the D1-D2-cytb559 complex, defined as the absorption signal not described by the two Gaussian sub-bands associated with the intense electronic transitions at 680 and 683 nm, exhibits anomalous temperature behavior. This tail was analyzed in the temperature interval between 80 and 300 K in terms of the mean square deviation (sigma2) of the total Qy absorption band and by Gaussian sub-band decomposition. The value of the average optical reorganization energy (Snum) obtained from the temperature dependence of sigma2 for the whole absorption band was 32 cm(-1), and changed to 16-20 cm(-1) after subtraction of the sub-bands describing the red tail. This latter value is in agreement with the hole burning literature data for chlorophyll bound to proteins, and indicates that the rather high value for the apparent optical reorganization energy obtained by analysis of the total Qy band of the D1-D2-cytb559 complex is determined by the temperature sensitivity of the red tail. This suggests that the long wavelength absorption tail might be due to vibrational transitions associated with vibrational modes in the range of 80-150 cm(-1) which are thermally accessible and give rise to an absorption signal on the low-energy side of the (0,0) transition. On the basis of this assumption, the electron-phonon coupling strength (S) for these modes is estimated to be in the range 0.028-0.18. This interpretation furthermore supports the idea that the electronic transition near 683 nm is that of a monomer chlorophyll.     

The low-energy forms of photosystem I light-harvesting complexes: spectroscopic properties and pigment-pigment interaction characteristics

In this work the spectroscopic properties of the special low-energy absorption bands of the outer antenna complexes of higher plant Photosystem I have been investigated by means of low-temperature absorption, fluorescence, and fluorescence line-narrowing experiments. It was found that the red-most absorption bands of Lhca3, Lhca4, and Lhca1-4 peak, respectively, at 704, 708, and 709 nm and are responsible for 725-, 733-, and 732-nm fluorescence emission bands. These bands are more red shifted compared to "normal" chlorophyll a (Chl a) bands present in light-harvesting complexes. The low-energy forms are characterized by a very large bandwidth (400-450 cm(-1)), which is the result of both large homogeneous and inhomogeneous broadening. The observed optical reorganization energy is untypical for Chl a and resembles more that of BChl a antenna systems. The large broadening and the changes in optical reorganization energy are explained by a mixing of an Lhca excitonic state with a charge transfer state. Such a charge transfer state can be stabilized by the polar residues around Chl 1025. It is shown that the optical reorganization energy is changing through the inhomogeneous distribution of the red-most absorption band, with the pigments contributing to the red part of the distribution showing higher values. A second red emission form in Lhca4 was detected at 705 nm and originates from a broad absorption band peaking at 690 nm. This fluorescence emission is present also in the Lhca4-N-47H mutant, which lacks the 733-nm emission band.     

Romanowsky dyes and the Romanowsky-Giemsa effect. 3. Microspectrophotometric studies of Romanowsky-Giemsa staining. Spectroscopic evidence of a DNA-azure B-eosin Y complex producing the Romanowsky-Giemsa effect

The Romanowsky-Giemsa staining (RG staining) has been studied by means of microspectrophotometry using various staining conditions. As cell material we employed in our model experiments mouse fibroblasts, LM cells. They show a distinct Romanowsky-Giemsa staining pattern. The RG staining was performed with the chemical pure dye stuffs azure B and eosin Y. In addition we stained the cells separately with azure B or eosin Y. Staining parameters were pH value, dye concentration, staining time etc. Besides normal LM cells we also studied cells after RNA or DNA digestion. The spectra of the various cell species were measured with a self constructed microspectrophotometer by photon counting technique. The optical ray pass and the diagramm of electronics are briefly discussed. The nucleus of RG stained LM cells, pH congruent to 7, is purple, the cytoplasm blue. After DNA or RNA digestion the purple respectively blue coloration in the nucleus or the cytoplasm completely disappeares. Therefore DNA and RNA are the preferentially stained biological substrates. In the spectrum of RG stained nuclei, pH congruent to 7, three absorption bands are distinguishable: They are A1 (15400 cm-1, 649 nm), A2 (16800 cm-1, 595 nm) the absorption bands of DNA-bound monomers and dimers of azure B and RB (18100 cm-1, 552 nm) the distinct intense Romanowsky band. Our extensive experimental material shows clearly that RB is produced by a complex of DNA, higher polymers of azure B (degree of association p greater than 2) and eosin Y. The complex is primarily held together by electrostatic interaction: inding of polymer azure B cations to the polyanion DNA generates positively charged binding sites in the DNA-azure B complex which are subsequently occupied by eosin Y anions. It can be spectroscopically shown that the electronic states of the azure B polymers and the attached eosin Y interact. By this interaction the absorption of eosin Y is red shifted and of the azure B polymers blue shifted. The absorption bands of both molecular species overlap and generate the Romanowsky band. Its strong maximum at 18100 cm-1 is due to the eosin Y part of the DNA-azure B-eosin Y complex. The discussed red shift of the eosin Y absorption is the main reason for the purple coloration of RG stained nuclei. Using a special technique it was possible to prepare an artificial DNA-azure B-eosin Y complex with calf thymus DNA as a model nucleic acid and the two dye stuffs azure B and eosin Y.(ABSTRACT TRUNCATED AT 400 WORDS)     

Characterization of a protein fraction containing cytochromes b and c1 from mitochondria of Leishmania tarentolae

A soluble red band fraction was obtained from Leishmania tarentolae cells by sucrose gradient sedimentation of a Triton X-100 lysate. Spectral analysis indicated that cytochrome b was present in the red band: the reduced minus oxidized difference spectra revealed absorption maxima at 562,527, and 431 nm at room temperature and 562, 530, and 422 nm at 77K. In addition, a 28-kDa protein was identified in this fraction which retained heme-associated peroxidase activity even after denaturation on SDS-polyacrylamide gels. The amino acid composition of this protein showed a strong similarity to cytochrome c1 of both bovine and yeast.     

Excitation energy transfer in the light-harvesting chlorophyll a/b.protein

The "light-harvesting chlorophyll a/b.protein" described by Thornber has been prepared electrophoretically from spinach chloroplasts. The optical properties relevant to energy transfer have been measured in the red region (i.e. 600-700 nm). Measurements of the absorption spectrum, fluorescence excitation spectrum and excitation dependence of the fluorescence emission spectrum of this protein confirm that energy transfer from chlorophyll b to chlorophyll a is highly efficient, as is the case in concentrated chlorophyll solutions and in vivo. The excitiation dependence of the fluorescence polarization shows a minimum polarization of 1.9% at 650 nm which is the absorption maximum of chlorophyll b in the protein and rises steadily to a maximum value of 13.8% at 695 nm, the red edge of the chlorophyll a absorption band. Analysis of these measurements shows that at least two unresolved components must be responsible for the chlorophyll a absorption maximum. Comparison of polarization measurements with those observed in vivo shows that most of the depolarization observed in vivo can take place within a single protein. Circular dichroism measurements show a double structure in the chlorophyll b absorption band which suggest an exciton splitting not resolved in absorption. Analysis of these data yields information about the relative orientation of the So leads to S1 transition moments of the chlorophyll molecules within the protein.     

High-resolution optical absorption-difference spectra of the triplet state of the primary donor in isolated reaction centers of the photosynthetic bacteria Rhodopseudomonas sphaeroides R-26 and Rhodopseudomonas viridis measured with optically detected magnetic resonance at 1.2 K

We have recorded triplet optical absorption-difference spectra of the reaction center triplet state of isolated reaction centers from Rhodopseudomonas sphaeroides R-26 and Rps. viridis with optical absorption-detected electron spin resonance in zero magnetic field (ADMR) at 1.2 K. This technique is one to two orders of magnitude more sensitive than conventional flash absorption spectroscopy, and consequently allows a much higher spectral resolution. Besides the relatively broad bleachings and appearances found previously (see, e.g., Shuvalov V.A. and Parson W.W. (1981) Biochim. Biophys. Acta 638, 50–59) we have found strong, sharp oscillations in the wavelength regions 790–830 nm (Rps. sphaeroides) and 810–890 nm (Rps. viridis). For Rps. viridis these features are resolved into two band shifts (a blue shift at about 830 nm and a red shift at about 855 nm) and a strong, narrow absorption band at 838 nm. For Rps. sphaeroides R-26 the features are resolved into a red shift at about 810 nm and a strong absorption band at 807 nm. We conclude that the appearance of the absorption bands at 807 and 838 nm, respectively, is due to monomeric bacteriochlorophyll. Apparently, the exciton interaction between the pigments constituting the primary donor is much weaker in the triplet state than in the singlet state, and at low temperature the triplet is localized on one of the bacteriochlorophylls on an optical time scale. The fact that for Rps. sphaeroides the strong band shift and the monomeric band found at 1.2 K are absent at 293 K and very weak at 77 K indicates that these features are strongly temperature dependent. It seems, therefore, premature to ascribe the temperature dependence between 293 and 77 K of the intensity of the triplet absorption-difference spectrum at 810 nm (solely) to a delocalization of the triplet state on one of the accessory bacteriochlorophyll pigments.     

Time-resolved fluorescence spectroscopy of NADPH-cytochrome P-450 reductase: demonstration of energy transfer between the two prosthetic groups

Fluorescence as well as fluorescence anisotropy decay parameters have been obtained from NADPH-cytochrome P-450 reductase by time-resolved fluorescence spectroscopy. The two flavins in the enzyme, FMN and FAD, are slightly fluorescent and exhibit heterogeneous fluorescence lifetimes, as observed with other flavoproteins. The time-dependent anisotropy is also multiexponential and is wavelength-dependent. The anisotropy decay is biexponential with two correlation times when the enzyme is excited at the red edge of the first absorption band (514 nm). When the enzyme is excited in the light absorption maximum (458 nm), an additional shorter correlation time is found, which contains information about the rate of energy transfer between the two flavins present in the enzyme. FMN-depleted NADPH-cytochrome P-450 reductase shows also only two correlation times, as does the enzyme in the "air-stable" semiquinone state when excited at 458 nm. Wavelength-dependent steady-state anisotropy measurements of native and FMN-depleted protein show that the former exhibits lower values than the latter in the region of the first absorption band, but when the red edge of the absorption band is reached, the anisotropy becomes equal in both preparations. A similar situation is encountered in model compounds, monomeric and dimeric flavins, immobilized in poly(methyl methacrylate). Both in the models and in the flavoprotein this can be attributed to failure of energy transfer at the red edge of the absorption band. From the results we were able to derive both geometric parameters and dynamic properties of both flavins in the NADPH-cytochrome P-450 reductase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Redshift of the purple membrane absorption band and the deprotonation of tyrosine residues at high pH: Origin of the parallel photocycles of trans-bacteriorhodopsin

At high pH (> 8) the 570 nm absorption band of all-trans bacteriorhodopsin (bR) in purple membrane undergoes a small (1.5 nm) shift to longer wavelengths, which causes a maximal increase in absorption at 615 nm. The pK of the shift is 9.0 in the presence of 167 mM KCl, and its intrinsic pK is ~8.3. The red shift of the trans-bR absorption spectrum correlates with the appearance of the fast component in the light-induced L to M transition, and absorption increases at 238 and 297 nm which are apparently caused by the deprotonation of a tyrosine residue and red shift of the absorption of tryptophan residues. This suggests that the deprotonation of a tyrosine residue with an exceptionally low pK (pK_a ≈ 8.3) is responsible for the absorption shift of the chromophore band and fast M formation. The pH and salt dependent equilibrium between the two forms of bR, “neutral” and “alkaline,” bR ↔ bR_a, results in two parallel photocycles of trans-bR at high pH, differing in the rate of the L to M transition. In the pH range 10-11.8 deprotonation of two more tyrosine residues is observed with pK's ~ 10.3 and 11.3 (in 167 mM KCL). Two simple models discussing the role of the pH induced tyrosine deprotonation in the photocycle and proton pumping are presented.

It is suggested that the shifts of the absorption bands at high pH are due to the appearance of a negatively charged group inside the protein (tyrosinate) which causes electrochromic shifts of the chromophore and protein absorption bands due to the interaction with the dipole moments in the ground and excited states of bR (Stark effect). This effect gives evidence for a significant change in the dipole moment of the chromophore of bR upon excitation.

Under illumination alkaline bR forms, besides the usual photocycle intermediates, a long-lived species with absorption maximum at 500 nm (P500). P500 slowly converts into bR_a in the dark. Upon illumination P500 is transformed into an intermediate having an absorption maximum at 380 nm (P380). P380 can be reconverted to P500 by blue light illumination or by incubation in the dark.

     

Studies on the absorption spectra of chlorophyll a in aqueous dispersions of lipids from the photosynthetic membranes

The absorption spectra of chlorophyll a were studied in aqueousdispersions of four major lipid components present in the thylakoidmembranes. Chlorophyll a in aqueous dispersions of uncharged galactolipidsrevealed two absorption bands, at 670 and 745 nm, when the molecularratio of chlorophyll to lipid was higher than 0.2. The latterband may be due to the formation of microcrystals of chlorophylla. Chlorophyll a in aqueous dispersions of negatively chargedlipids revealed a single absorption band at 670 nm. However,chlorophyll a was decomposed during measurement in these lipiddispersions. The absorption spectra of chlorophyll a in aqueous dispersionsof mixture of galactolipid and charged lipid were apparentlysimilar to those of chlorophyll a in the charged lipid dispersion.Chlorophyll a, however, was not decomposed in these aqueousdispersions of lipid mixtures. It is concluded that the presence of both galactolipid and chargedlipid are necessary to reconstruct the state of chlorophylla dissolved in the lipid phase in the thylakoid membranes. The red absorption band of chlorophyll a in the reconstructedsystem composed of chlorophyll a, charged and uncharged lipids,appeared at 670 nm with a half bandwidth of 22 nm. Analysisof the absorption spectrum in the fourth derivative and thecurve-fitting methods indicated that the red band was composedmainly of a single band with a peak at 670–671 nm. ¹ Present address: Department of Biology, College of GeneralEducation, University of Tokyo, Komaba, Meguro-ku, Tokyo 153,Japan. (Received October 13, 1977; )     

Effects of acid pH and urea on the spectral properties of the LHII antenna complex from the photosynthetic bacterium Ectothiorhodospira sp.

The aim of this study was to investigate the spectral modifications of the LHII antenna complex from the purple bacterium Ectothiorhodospira sp. upon acid pH titration both in the presence and absence of urea. A blue shift specifically and reversibly affected the B850 band around pH 5.5-6.0 suggesting that a histidine residue most probably participated in the in vivo absorption red shifting mechanism. This transition was observed in the presence and absence of urea. Under strong chaotropic conditions, a second transition occurred around pH 2.0, affecting the B800 band irreversibly and the B850 reversibly. Under these conditions a blue shift from 856 to 842 nm occurred and a new and strong circular dichroism signal from the new 842 nm band was observed. Reverting to the original experimental conditions induced a red shift of the B850 band up to 856 nm but the circular dichroism signal remained mostly unaffected. Under the same experimental conditions, i.e. pH 2.1 in the presence of urea, part of the B800 band was irreversibly destroyed with concomitant appearance of a band around 770 nm due to monomeric bacteriochlorophyll from the disrupted B800. Furthermore, Gaussian deconvolution and second derivative of the reverted spectra at pH 8.0 after strong-acid treatment indicated that the new B850 band was actually composed of two bands centered at 843 and 858 nm. We ascribed the 858 nm band to bacteriochlorophylls that underwent reversible spectral shift and the 843 nm band to oligomeric bacteriopheophytin formed from a part of the B850 bacteriochlorophyll. This new oligomer would be responsible for the observed strong and mostly conservative circular dichroism signal. The presence of bacteriopheophytin in the reverted samples was definitively demonstrated by HPLC pigment analysis. The pheophytinization process progressed as the pH decreased below 2.1, and at a certain point (i.e. pH 1.5) all bacteriochlorophylls, including those from the B800 band, became converted to oligomeric bacteriopheophytin, as shown by the presence of a single absorption band around 843 nm and by the appearance of a single main elution peak in the HPLC chromatogram which corresponded to bacteriopheophytin.     

Protochlorophyll forms with different molecular arrangements

Spectral properties of protochlorophyll (PChl) forms were investigated in solid-film model systems by absorption. fluorescence and circular dichroism (CD) spectroscopy. The solid films were prepared from diethyl ether solution of PChl on a cover glass surface by evaporation of the solvent. After preparation the films usually showed an absorption maximum at 635 nm or in some cases at 640 nm. The PChl form with 635 nm absorption maximum had no CD signal, whilst the films with absorption maximum at 640 nm gave an intense negative CD band at about 640 nm and a positive one at 668 nm. The treatment of the films with ammonia or acetone vapour resulted in a red shift of the absorption maximum from 635 nm or 640 nm to 650 nm. The study of the CD spectra of the films with different PChl forms showed that, depending on the treatment, forms of PChl with similar absorption and fluorescence spectra, but with opposite CD signals, can exist. It is suggested that the differences of the CD spectra are mainly due to different arrangements of the aggregates.     

Hydrogen bonding and proton transfer between nitro-phenols and lecithin in apolar solvents

The interaction of p-nitrophenol (p-NP), 2,4-dinitrophenol (DNP) (II) and 2,4,6-trinitrophenol (TNP)(III) with dipalmitoyllecithin in apolar solvents has been examined by IR and UV spectroscopy. Addition of any nitrophenol to the solution of lecithin in CCl₄ causes disappearance of broad absorption band of water bound with lecithin phosphate grops (3150–3600 cm^?1), which was accompanied by an insignificant increase of absorption near 3040 and 2800 cm^?1. Association of phenolic groups of (I) with the lecithin was observed by disappearance of the free OH absorption band. In UV spectra of (I), complex formation with lecithin results in a 30 nm red shift of phenol long-wave absorption band and in the appearance of an isosbestic point at 303 nm. In the case of III, addition of the lecithin causes a red shift and strong hyperchromic effect, which is accompanied by the appearance of a new absorption band near 420 nm. It was concluded, that nitrophenols displace a part of water from the polar groups of lipids and form hydrogen bonded complexes or ion-pair structures, depending upon acidic properties of the proton donor.     

Isolation of an intrinsic antenna chlorophyll a-protein from the photosystem I reaction center complex of the thermophilic cyanobacterium Synechococcus sp

A chlorophyll-protein was isolated from a Synechococcus P700-chlorophyll a-protein complex free from small subunits (CP1-e) by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis after treatment with 2% 2-mercaptoethanol and 2% SDS. In contrast to CP1-e which, when electrophoresed under denaturating conditions, showed two polypeptide bands of 62 and 60 kDa, the chlorophyll-protein contained only the 60-kDa polypeptide and hence is called CP60. The yield of CP60 was maximal with 1-2% SDS and 2-4% sulfhydryl reagents because the chlorophyll-protein was denatured at higher concentrations of the reagents. The absorption spectrum of CP60, which retained more than half of the chlorophyll alpha molecules originally associated with the 60-kDa subunit of the photosystem I reaction center complex, showed a red band maximum at 672 nm and a small absorption band around 700 nm at liquid nitrogen temperature. CP60 emitted a fluorescence band at 717 to 725 nm at 77 degrees K. The temperature dependence of the far red band of CP60 was essentially the same as that of CP1-e between 77 and 273 degrees K. No photoresponse of P700 was detected in CP60. The results suggest that the two polypeptides resolved by SDS-gel electrophoresis from CP1-e are apoproteins of two distinct chlorophyll-proteins and that CP60 represents a chlorophyll-bearing 60-kDa subunit functioning as an intrinsic antenna protein of the photosystem I reaction center complex. It will also be shown that the temperature dependence of the far red fluorescence band is not related to the photosystem I photochemistry.     

Spectral inhomogeneity of photosystem I and its influence on excitation equilibration and trapping in the cyanobacterium Synechocystis sp. PCC6803 at 77 K.

Ultrafast transient absorption spectroscopy was used to probe excitation energy transfer and trapping at 77 K in the photosystem I (PSI) core antenna from the cyanobacterium Synechocystis sp. PCC 6803. Excitation of the bulk antenna at 670 and 680 nm induces a subpicosecond energy transfer process that populates the Chl a spectral form at 685--687 nm within few transfer steps (300--400 fs). On a picosecond time scale equilibration with the longest-wavelength absorbing pigments occurs within 4-6 ps, slightly slower than at room temperature. At low temperatures in the absence of uphill energy transfer the energy equilibration processes involve low-energy shifted chlorophyll spectral forms of the bulk antenna participating in a 30--50-ps process of photochemical trapping of the excitation by P(700). These spectral forms might originate from clustered pigments in the core antenna and coupled chlorophylls of the reaction center. Part of the excitation is trapped on a pool of the longest-wavelength absorbing pigments serving as deep traps at 77 K. Transient hole burning of the ground-state absorption of the PSI with excitation at 710 and 720 nm indicates heterogeneity of the red pigment absorption band with two broad homogeneous transitions at 708 nm and 714 nm (full-width at half-maximum (fwhm) approximately 200--300 cm(-1)). The origin of these two bands is attributed to the presence of two chlorophyll dimers, while the appearance of the early time bleaching bands at 683 nm and 678 nm under excitation into the red side of the absorption spectrum (>690 nm) can be explained by borrowing of the dipole strength by the ground-state absorption of the chlorophyll a monomers from the excited-state absorption of the dimeric red pigments.     

Circular dichroism of the peripheral chlorophylls in photosystem II reaction centers revealed by electrochemical oxidation

Visible absorption spectra and circular dichroism (CD) of the red absorption band of isolated photosystem II reaction centers were measured at room temperature during progressive bleaching by electrochemical oxidation, in comparison with aerobic photochemical destruction, and with anaerobic photooxidation in the presence of the artificial electron acceptor silicomolybdate. Initially, selective bleaching of peripheral chlorophylls absorbing at 672 nm was obtained by electrochemical oxidation at +0.9 V, whereas little selectivity was observed at higher potentials. Illumination in the presence of silicomolybdate did not cause a bleaching but a spectral broadening of the 672-nm band was observed, apparently in response to the oxidation of carotene. The 672-nm absorption band is shown to exhibit a positive CD, which accounts for the 674-nm shoulder in CD spectra at low temperature. The origin of this CD is discussed in view of the observation that all CD disappears with the 680-nm absorption band during aerobic photodestruction.     

Circular dichroism of the peripheral chlorophylls in photosystem II reaction centers revealed by electrochemical oxidation

Visible absorption spectra and circular dichroism (CD) of the red absorption band of isolated photosystem II reaction centers were measured at room temperature during progressive bleaching by electrochemical oxidation, in comparison with aerobic photochemical destruction, and with anaerobic photooxidation in the presence of the artificial electron acceptor silicomolybdate. Initially, selective bleaching of peripheral chlorophylls absorbing at 672 nm was obtained by electrochemical oxidation at +0.9 V, whereas little selectivity was observed at higher potentials. Illumination in the presence of silicomolybdate did not cause a bleaching but a spectral broadening of the 672-nm band was observed, apparently in response to the oxidation of carotene. The 672-nm absorption band is shown to exhibit a positive CD, which accounts for the 674-nm shoulder in CD spectra at low temperature. The origin of this CD is discussed in view of the observation that all CD disappears with the 680-nm absorption band during aerobic photodestruction.