Long-wavelength chlorophylls in photosystem I of cyanobacteria: origin, localization, and functions.

The structural organization of photosystem I (PSI) complexes in cyanobacteria and the origin of the PSI antenna long-wavelength chlorophylls and their role in energy migration, charge separation, and dissipation of excess absorbed energy are discussed. The PSI complex in cyanobacterial membranes is organized preferentially as a trimer with the core antenna enriched with long-wavelength chlorophylls. The contents of long-wavelength chlorophylls and their spectral characteristics in PSI trimers and monomers are species-specific. Chlorophyll aggregates in PSI antenna are potential candidates for the role of the long-wavelength chlorophylls. The red-most chlorophylls in PSI trimers of the cyanobacteria Arthrospira platensis and Thermosynechococcus elongatus can be formed as a result of interaction of pigments peripherally localized on different monomeric complexes within the PSI trimers. Long-wavelength chlorophylls affect weakly energy equilibration within the heterogeneous PSI antenna, but they significantly delay energy trapping by P700. When the reaction center is open, energy absorbed by long-wavelength chlorophylls migrates to P700 at physiological temperatures, causing its oxidation. When the PSI reaction center is closed, the P700 cation radical or P700 triplet state (depending on the P700 redox state and the PSI acceptor side cofactors) efficiently quench the fluorescence of the long-wavelength chlorophylls of PSI and thus protect the complex against photodestruction.

[Variability of light-induced circular dichroism spectra of photosystem I complexes of cyanobacteria].

Circular dichroism (CD) spectra of photosystem I (PSI) complexes of the cyanobacteria Thermosynechococcus elongatus, Arthrospira platensis and Synechocystis sp. PCC 6803 were studied. CD spectra of dark-adapted PSI trimers and monomers, measured at 77 K, show common bands at 669-670(+), 673(+), 680(-), 683-685(-), 696-697(-), 702(-) and 711(-) nm. The intensities of these bands are species specific. In addition, bands at 683-685(-) and 673(+) nm differ in intensity for trimeric and monomeric PSI complexes. CD difference spectra (P700(+)-P700) of PSI complexes at 283 K exhibit conservative bands at 701(-) and 691(+) nm due to changes in resonance interaction of chlorophylls in the reaction center upon oxidation of P700. Additional bands are observed at 671(-), 678(+), 685(-), 693(-) nm and in the region 720-725 nm those intensities correlate with intensities of analogous bands of antenna chlorophylls in dark-adapted CD spectra. It is suggested that the variability of CD difference spectra of PSI complexes is determined by changes in resonance interaction of reaction center chlorophylls with closely located antenna chlorophylls.

[Classification of lectins as universal regulator molecules of biological systems].

Modern concepts of lectin classification reflecting tendencies in the development of functional classification of lectins (Ln) and Ln-like proteins are presented. Classification of bacterial Ln is proposed. Classification of Ln and Ln-like proteins of plants, animals, and bacteria is based on 8 major elements of protein secondary structure that allows for the evaluation of topographic diversity of their surfaces, potential compatibility of bacterial, vegetable, and mammalian Ln molecules and possible variants of their co-functioning.

Isolation from Spirulina membranes of two photosystem I-type complexes, one of which contains chlorophyll responsible for the 77 K fluorescence band at 760 nm.

Two types of chlorophyll-protein complexes of photosystem I (PSIa, PSIc) have been isolated from the membranes of Spirulina platensis using a Triton X-100 treatment and chromatography on DEAE-Toyopearl. The complexes are equally enriched with P700 (Chl: P700 = 100-110) but show different electrophoretic molecular masses--140 (PSIa) and 320 kDa (PSIc)--and differ in the content of long-wavelength absorbing Chl. PSIa has a typical PSI fluorescence band at 730 nm (F730) as the main band at 77 K, whereas PSIc is responsible for F760, the intensity of which depends on the redox state of P700. PSIc only shows 77 K light-induced variable fluorescence at 760 typical of Spirulina membranes and cells.

Trimeric forms of the photosystem I reaction center complex pre-exist in the membranes of the cyanobacterium Spirulina platensis.

Oligomeric and monomeric forms of chlorophyll-protein complexes of photosystem I (PSI) have been isolated from the mesophilic cyanobacterium Spirulina [(1992) FEBS Lett. 309, 340-342]. Electron microscopic analysis of the complexes showed that the oligomeric form is a trimer of the shape and dimensions similar to those of the trimer from thermophilic cyanobacteria. The chlorophyl ratio in the isolated trimer and monomer was found to be 7:3. The trimeric form of PSI complex in contrast to the monomeric one contains the chlorophyll emitting at 760 nm (77K), which is also found in Spirulina membranes and therefore could be used as an intrinsic probe for the trimeric complex. The 77K circular dichroism spectrum of the trimeric form is much more similar to that of Spirulina membranes than the spectrum of the monomer. Thus, the trimeric PSI complexes exist and dominate in the Spirulina membranes.

[Determination of secondary structure of globular proteins using circular dichroism spectra]. / Opredelenie vtorichnoi struktury globuliarnykh belkov po spektram krugovogo dikhroizma.

A ridge regression method is presented for prediction of the secondary structure of proteins by the circular dichroism spectra (CD) from 190 to 236 nm. Eight types of the secondary structure were calculated on a microcalculator. The method is based on the X-ray data of Kabsh and Sander. The teaching rule is constructed on CD spectra of 30 proteins of all structural classes of the globular proteins (alpha, alpha/beta, alpha + beta and beta-proteins). The errors of the methods are analysed by removing each protein from the reference set and analyzing its structure in terms of the remaining proteins. Correlation coefficients and root-mean square deviations between CD and X-ray data were: 0.99 and 0.03 for alpha-helix, 0.86 and 0.02 for 3(10)-helix, 0.92 and 0.06 for antiparallel beta-sheet, 0.86 and 0.03 for parallel beta-sheet, 0.94 and 0.01 for T3 beta-turn, 0.85 and 0.02 for other beta-turn, 0.84 and 0.03 for S-bends, 0.83 and 0.04 for "random" structure.

[Interaction of the photosystem 1 reaction center with antenna chlorophyll in a pigment-protein complex isolated from pea chloroplasts]. / Vzaimodeistvie reaktsionnogo tsentra fotosistemy 1 s antennym khlorofillom v pigment-belkovom komplekse, vydelennom iz khloroplastov gorokha.

Using a specially developed phosporoscopic attachment to spectropolarimeter, light induced spectra of circular dichroism (CD) in region 600-750 nm were measured for a pigment protein complex of photosystem 1 (PC-1) isolated from pea chloroplast (chlorophyll : P700 = 40). Minor components at 672 and 678 nm are observed in light induced spectra besides the components of dimer splitting of P700 Qy transition at 691 and 698 nm. Haussian deconvolution of light induced CD spectra of P700 and low temperature CD spectrum of PC-1 indicates that minor components are due to forms of antenna chlorophylls Chl672 and Chl678, rotational strength of that is changed by 2-4% as a result of P700 oxidation. Long term incubation of PC-1 with Triton X-100 inhibits P700 and destroys longwave optically active chlorophyll forms. A strong relation between dichroic density of 693 nm band in CD spectrum of PC-1 and the value of light induced absorption change at 698 nm could be used to determine P700 concentration on the basis of CD spectrum of PC-1. Such a relation shows that Chl693 is an important component of photo-system 1 reaction center. It is suggested that P700 is not an isolated dimer but it is included in the local complex from 8-10 chlorophyll molecules (Chl672, Chl678, Chl686, Chl693).

[Comparative study of the circular dichroism of rabbit liver and muscle glycogen phosphorylases a and b]. / Sravnitel'noe issledovanie krugovogo dikhroizma glikogenfosforilaz a i b iz myshts i pecheni krolika.

Circular dichroism (CD) spectra of glycogen phosphorylase a and b from rabbit liver have been measured in the presence of various ligands in the near- and far-ultraviolet regions. Positive circular dichroism was detected in the absorption band of protein-bound pyridoxal phosphate (333 nm). The mean residue ellipticity of this dichroic band (35 deg cm2dmol-1) is of the same order for muscle and liver phosphorylase a and b and does not change upon binding of glucose-1-phosphate and AMP. Only glucose induces small changes in the ellipticity in this region. The CD spectra of muscle and liver phosphorylase a and b in the 250-300 nm region have at least five positive dichroic bands namely at 259, 264, 273, 281 and 288 nm and have strong resemblances for all these forms of the enzyme in spite of the fundamental differences in their properties. The binding of AMP and glucose to phosphorylase from both sources induces distinct perturbations in CD spectra; the changes are much larger for muscle and liver phosphorylase a than for phosphorylase b which indicates that conformational perturbations induced by binding of activator and inhibitor to the inactive form of phosphorylase are probably more local than for the active form. The CD spectra in far-ultraviolet region are similar for all forms of phosphorylase. The percent of alpha-helices calculated according to Chen is about 50; this value coincides very well with the value 51% received for muscle phosphorylase a by X-ray crystallographic analysis at 2.5 A resolution.

[Spectral properties and structure of bacteriopheophytin a dimer]. / Spektral'nye svoistva i struktura dimera bakteriofeofitina a.

In alcohol-glycerol solutions of bacteriopheophytin alpha takes place dimer formation with a long wave absorption band at 857 nm, the dimerization constant enhances with glycerol content increasing in solution. The measurement of absorption, fluorescence and CD spectra allowed to characterize the main electronic transitions (Qy, Qx, Bxy) of the dimer. For the first electronic transition (Qy) of the dimer as compared to the monomer the same oscillator strength, spectral bands narrowing of splitting components and significant increase of rotation strength in CD spectrum were found. The structural model of bacteriopheophytin a dimer is suggested on the basis of the data obtained. Coincidence of the values of the long wave shift and exitonic splitting of Qy transition for the dimer investigated and antennae bacteriochlorophylls shows that the appearance of these properties does not require pigment protein interaction. Significant (20--30 times) exceeding of the rotation strength of Qy transition of the bacteriopheophytin dimer as compared ato the antennae complex pigments can indicate the absence of this dimer in intact structures.

[Comparative study of bacteriopheophytin a and b aggregation]. / Sravnitel'noe izuchenie agregatsii bakteriofeofitinov a i b.

Aggregation of bacteriopheophytins a and b (BPh) in ethanol--glycerol solution and a comparison of spectral properties of obtained aggregated pigment forms were studied. The dependence of aggregation process on percentage of glycerol and pigment concentration on alcohol solution was studied. The aggregation of BPh was followed by very strong deformation of electron absorption and circular dichroism (CD) spectra. Some changes in the first and second electron bands and in Soret region were found. An analysis of BPh a and b aggregates CD spectra shows its general similarity in all spectral regions. It may be connected with structural identity of the obtained aggregated forms. The molar coefficient extinction of BPh aggregates was determined.

[Study of energy migration between native forms of chlorophyll at -196 degrees C by the method of sensitized fluorescence]. / Issledovanie migratsii énergii mezhdu nativnymi formami khlorofilla pri -196 gradusakh C metodom sensibilizirovannoi fluorestsentsii

Mathematical decomposition of fluorescence spectra of chloroplasts and cells measured under monochromatic excitation in the regions of absorption maxima of the main native forms allowed the authors to calculate the efficiency of energy migration between forms. It is shown that 80--100% of absorbed quanta flow to the Chl 738/708 form (bean) in photosystem 1, whereas in photosystem 2, the Chl 695/690 and Chl 703/698 forms are terminal energy acceptors. Efficient backward energy transfer between the Chl 680/676 and Chl 686/682 forms and the existence of energy migration pathways passing one or two steps of the energy stairway formed by the energy levels of the chlorophyll forms are found. It is shown that chlorphyll b is not an intermediate link in the process of energy transfer from carotenoids to chlorophyll a. The intrinsic fluoresence quantum yields of the native forms are calculated.

[Variable fluorescence of native forms of chlorophyll at -196 degrees C]. / Peremennaia fluorestsentsiia nativnykh form khlorofilla pri -196 gradusakh C

The value (f = Ioo/Io) of variable fluorescence (delta I) of individual forms of chlorophyll a are calculated. Absence of delta I in the forms phiCl indicates that there is no energy transfer from phiC2 to phiCL.

[Study of different types of chlorophyll a aggregates in solutions and films by absorption and luminescence derivative spectroscopy]. / Issledovaine razlichnykh tipov agregatov khlorofilla a v rastvorah i plenkakh metodom absorbtsinnoi i liuminestsentnoi proizvodnoi spektroskopii

The second derivative of absorption, fluorescence and fluorescence excitation spectra of chlorophyll a in concentrated solutions and films was investigated. More than 14 forms of pigment aggregates, which can be divided into two types--with narrow 8-10nm) and wide (25-40nm) low temperature (-196 degrees C) spectra bands, were found. For the most part of the aggregated forms, the position and half width of the bands, as well as the Stokes shift and relative quantum yield were determined. The comparison of the spectral characteristics points to the indentity of the aggregates and corresponding native forms of Chl. a. It is shown that the universal relationship between absorption and fluorescence bands in applicable to the aggregates of the two types and the energy of resonance interaction between monomers in the aggregates is evaluated.

[Activation and inhibition of photoinduced proton absorption in Rhodospirillum rubrum chromatophores by detergents and solvents]. / Aktivatsiia i ingibirovanie fotoindutsirovannogo pogloshcheniia protonov v khromatoforakh Rhodospirillum rubrum detergentami i rastvoriteliami.

The effects of detergents (Triton X-100) and solvents (diethyl ether, metanol) on the reversible light-induced proton uptake, photophosphorylation and band shift of the carotenoid in chromatophores from R. rubrum are described. All these compounds were found to stimulate the extent of light-induced proton uptake with subsequent inhibition when the concentrations were increased. Stimulation of proton uptake is accompanied by inhibition of both phosphorylation and carotenoid absorbance shift.

[Nature of the electron excited state in pigment redox reactions. II. Analysis of the scheme of primary processes in the photooxidation reaction of chlorophylls a and b and pheophytin a ]. / Priroda élektronno-vozbuzhdennogo sostoianiia v okislitel'no-vosstanovitel'nykh reaktsiiakh pigmentov. II. Analiz skhemy pervichnykh protsessov v reaktsii fotookisleniia khlorofillov a i b i feofitina a.

A scheme of primary reactions in photooxidation of pigments was considered assuming that electron transfer processes can occur via singlet excited as well as triplet states. The results of analysis are compared with the experimental data on relative yield values of chlorophylls a, b, and pheophytin a cation-radicals, as well as with the data on fluorescence quenching. A conclusion has been drawn that photooxidation of pigments proceeds exclusively via the triplet state. The dependence of rate constant quenching values of chlorophyll a triplet state by certain electron acceptors on values of half cell potentials was given.

[Nature of the electron-excited state in redox reactions of pigments. I. Photooxidation of chlorophyll a by n-benzoquine]. / Priroda élektronno-vozbuzhdennogo sostoianiia v okislitel'no-vosstanovitel'nykh reaktsiiakh pigmentov. I. Fotookislenie khlorofilla a n-benzokhimonom.

A method for studying the nature of electronic excited state under photooxidation is proposed. It is shown by an example of the oxidation of chlorophyll a with p-benzoquinone that the formation of cation-radicals of pigments proceeds only through the triplet state. On the basis of experimental data the values of the rate constants of formation of chlorophyll cation-radicals through the singlet-excited state (K2(1)--10(7) M-1 s-1) and triplet state (Kr=10(9) M-1 s-1) of the pigment are evaluated.

[Effect of secondary structure on the rate dark reduction of P700+ with ascorbate of the photosystem I complex]. / Vliianie vtorichnoi struktury na skorost' temnovogo vosstanovleniia P700+ askorbatom kompleksa fotosistemy I.

Relationship was studied between the spectra of KD, absorption and fluorescence and kinetics of P700+ dark reduction with exogenous donor and sodium dodecyl concentration inducing a conformation transition of the protein globule. It has been shown that with a decrease of the detergent concentration from 0.01 to 0.06% an increase of the portion of the protein globule-helical regions (from 10 to 30%) and an essential decrease of the time of P700+ dark reduction with ascorbate take place. The changes of KD, absorption and fluorescence spectra of chlorophyll a observed in the same region of detergent concentration do not correlate with the changes of the secondary structure of the protein part of the pigment-protein complex. It is suggested that accelerated reduction of P700+ with ascorbate is conditioned by an increase of accessibility of exogenous donor to the reaction centre resulting from conformation changes of the complex protein part.

Structural changes in R-phycoerythrin upon CdS quantum dot synthesis in tunnel cavities of protein molecules.

Structural changes in R-phycoerythrin used as a matrix for the synthesis of CdS quantum dots have been analyzed by circular dichroism spectrometry. In deionized water, quantum dot synthesis in the tunnel cavity of the R-phycoerythrin molecule proved to be accompanied by uncoiling of α-helices and changes in the conformation of its chromophore groups, with consequent decay of protein fluorescence. Since R-phycoerythrin fluorescence is important for practical applications, conditions for quantum dot synthesis have been optimized by replacing deionized water with 0.01 M MES buffer, pH 5.7. Under such conditions, the size of the CdS quantum dots (determined from atomic force microscopy images) remains the same as in deionized water, but quantum dots cause only minor structural changes in protein molecules, as follows from circular dichroism and absorption spectra. The thermostability of R-phycoerythrin is enhanced, as indicated by an increase in the experimental activation energy for denaturation (from 140.8 to 149.9 kJ/mol) and the intensity of R-phycoerythrin fluorescence is also enhanced approximately twofold.

Opioid peptides derived from food proteins suppress aggregation and promote reactivation of partly unfolded stressed proteins.

A new view of the opioid peptides is presented. The potential of small peptides derived from precursor food proteins, to bind to partly unfolded stressed proteins to prevent their irreversible aggregation and inactivation has been demonstrated in various in vitro test systems: dithiothreitol-induced aggregation of alpha-lactalbumin (LA), heat-induced aggregation of alcohol dehydrogenase (ADH), and aggregation and inactivation of bovine erythrocyte carbonic anhydrase (CA) in the process of its refolding after removal of stress conditions. Using dynamic light scattering (DLS), turbidimetry, fluorescence, and circular dichroism measurements protective effects of the synthetic opioid peptides: exorphin C from wheat gluten (Tyr-Pro-Ile-Ser-Leu), rubiscolin-5 from spinach ribulose-bisphosphate-carboxylase/oxygenase (Rubisco) (Tyr-Pro-Leu-Asp-Leu), and hemorphin-6 from bovine hemoglobin (Tyr-Pro-Trp-Thr-Gln-Arg) have been revealed. We have demonstrated the concentration-dependent suppression of light scattering intensity of aggregates of LA and ADH in the presence of the peptides, the population of nanoparticles with higher hydrodynamic radii being shifted to the lower ones, accompanied by an increase in the lag period of aggregation. The presence of the peptides in the refolding solution was shown to assist reactivation of CA and enhance the yield of the CA soluble protein. The results suggest that bioactive food protein fragments may be regarded as exogenous supplements to the endogenous defense mechanisms of the human organism under stress conditions.

Molecular arrangement of pigment-protein complex of photosystem 1.

The circular dichroism (CD) method was applied to study the molecular organization of P700, antenna chlorophyll and protein of photosystem 1 complexes (CP1), isolated from chloroplasts under mild treatment with Triton X-100. Analysis of CD spectra and protein: chlorophyll: P700 ratios for CP1 complexes that were different in their chlorophyll content indicate that CP1 preparations can be considered as a mixture of CP1-RC, containing P700 (10-20%), and CP1-LH without P700 (80-90%). Both types of complexes contain approximately 25 chlorophyll molecules, and the destruction of their spatial organization with detergents represents a cooperative transition. The rate of chlorophyll destruction in CP1-LH is much higher than that in CP1-RC. In both complexes a 65 kDa polypeptide predominates, whose secondary structure (typical for α/ß proteins) is stable to Triton X-100 and does not depends on the chlorophyll content. Chlorophyll seems to be grouped in clusters (5-7 molecules) in the hydrophobic cores of 2-3 parallel α/ß domains of the 65 kDa protein. Only one of the clusters in CP1-RC includes P700; on P700 photooxidation the change of its interaction with the nearest pigment environment results in a complicated shape of the light-induced CD spectra.