The cysteine-proximal aspartates in the Fx-binding niche of photosystem I. Effect of alanine and lysine replacements on photoautotrophic growth, electron transfer rates, single-turnover flash efficiency, and EPR spectral properties.

The FX electron acceptor in Photosystem I (PS I) is a highly electronegative (Em = -705 mV) interpolypeptide [4Fe-4S] cluster ligated by cysteines 556 and 565 on PsaB and cysteines 574 and 583 on PsaA in Synechocystis sp. PCC 6803. An aspartic acid is adjacent to each of these cysteines on PsaB and adjacent to the proline-proximal cysteine on PsaA. We investigated the effect of D566PsaB and D557PsaB on electron transfer through FX by changing each aspartate to the neutral alanine or to the positively charged lysine either singly (D566APsaB, D557APsaB, D566KPsaB, and D557KPsaB) or in pairs (D557APsaB/D566APsaB and D557KPsaB/D566APsaB). All mutants except for D557KPsaB/D566APsaB grew photoautotrophically, but the growth of D557KPsaB and D557APsaB/D566APsaB was impaired under low light. The doubling time was increased, and the chlorophyll content per cell was lower in D557KPsaB and D557APsaB/D566APsaB relative to the wild type and the other mutants. Nevertheless, the rates of NADP+ photoreduction in PS I complexes from all mutants were no less than 75% of that of the wild type. The kinetics of back-reaction of the electron acceptors on a single-turnover flash showed efficient electron transfer to the terminal acceptors FA and FB in PS I complexes from all mutants. The EPR spectrum of FX was identical to that in the wild type in all but the single and double D566APsaB mutants, where the high-field resonance was shifted downfield. We conclude that the impaired growth of some of the mutants is related to a reduced accumulation of PS I rather than to photosynthetic efficiency. The chemical nature and the charge of the amino acids adjacent to the cysteine ligands on PsaB do not appear to be significant factors in the efficiency of electron transfer through FX.

Dynamics of the excited state of the primary electron donor in reaction centers of Rhodopseudomonas viridis as revealed by hole burning at 1.7K. Different conformational states.

The spectra of absorbance changes (delta A) due to the formation of P+Q- (P, primary electron donor, Q, primary quinone acceptor) at 1.7K in Rhodopseudomonas viridis reaction centers (RCs) excited at 1014 nm has been shown to include, besides a progression of broad (170-190 cm-1) Gaussian vibronic bands separated by 150 cm-1, a 'narrow' structure near 1014 nm which can be simulated by a Lorentian zero-phonon hole (ZPH) and Lorentian one-mode (26.8 cm-1) phonon wings. The widths of ZPH of approximately 17 cm-1 for delta A reflecting the formation of P+Q- decaying in the ms time domain and of 6.8 +/- 0.4 cm-1 for P+Q- decaying in the min time domain at 1.7K, seems to correspond to different conformations of RCs with a relaxation time of P* of approximately 0.6 ps (in agreement with measurements in this time domain) and 1.6 +/- 0.1 ps, respectively. The comparison of the spectra of delta A in the region of the BL band for slow (min) and fast (ms) decaying components suggests a different mutual arrangement of P and BL for different conformations of RCs. It is assumed that the broad and narrow structures of the P band reflect the transitions to two configurations with different P-protein interactions. 'Narrow' structure of delta A spectrum with essentially the same phonon wings and ZPH (width of 3.8 +/- 0.4 cm-1) was observed within the P band when HL was photoreduced at 1.7K.

Sub-picosecond dynamics of excited state of primary electron donor in reaction centers of Rhodopseudomonas viridis as revealed by hole burning at 1.7K broad and narrow holes.

Within the QF band of the primary electron donor (P), the spectra of absorbance changes due to the formation of a state of P+QA- (QA is the primary quinone) at 1.7K in Rhodopseudomonas viridis reaction centers excited at 1014 nm have been shown to involve two spectral features characterized by: (i) a progression of broad (170-190 cm-1) Gaussian vibronic bands (S-factor = 1.4) separated by 150 cm-1 and (ii) a 'narrow' structure near 1014 nm, characterized by 0-0 transition at 1014 nm with a width of approximately 50 cm-1 and 0-1 transition at 1000 nm with the width of approximately 100 cm-1, and S-factor = 0.9. The width of 50 cm-1 can be related to either zero-phonon hole (ZPH) width or the structure involving phonon wings and ZPH being unresolved. Since dichroic value (approximately 0.37) is unvarying over the P band, the vibrations involved are totally symmetric. The ZPH (width of approximately 3 cm-1) and phonon wings (frequency of approximately 30 cm-1) are resolved within the P band near 1014 nm when the spectrum of delta A due to the formation of bacteriopheophytinL- is measured at 1.7K.

Quantitative method for studying orientation of transition dipoles in membrane vesicles of spherical symmetry.

Pigmented vesicular membranes embedded in polyacrylamide gel exhibit linear dichroism when the gel sample is squeezed [Abdourakhmanov, I.A., Ganago, A.O., Erokhin, Yu.E., Solov'ev, A.A. and Chugunov, V.A. (1979) Biochim. Biophys. Acta 546, 183-186]. The orientation technique of gel-squeezing was modified to enhance polarization effects in membrane vesicles of spherical symmetry. Model calculations were carried out to provide a tool for the quantitative evaluation of the dichroism of squeezed gel samples. The orientation angles of the dipoles can be calculated with reasonable precision by measuring two quantities: (i) the macroscopic deformation parameter of the gel sample, and (ii) a parameter (e.g. the polarization ratio of the fluorescence emission) characterizing the orientation of the transition dipoles in the membranes embedded in the squeezed gel. The validity of the model was confirmed through a series of polarization measurements relating to the fluorescence of chlorophyll a in membranes of osmotically shocked chloroplasts, 'blebs'.

[Polarization fluorescence of chloroplasts oriented in polyacrylamide gel. Experiment and theoretical analysis]. / Poliarizovannaia fluorestsentsiia khloroplastov, orientirovannykh v poliakrilamidnom gele. Eksperiment i teoreticheskii analiz.

Maize mesophyll chloroplasts were oriented in polyacrylamide gel upon squeezing of gel samples. Fluorescence polarization was measured as a function of the gel deformation parameter. Linearly polarized fluorescence spectra were recorded at -140 degrees. The results show that chloroplasts were oriented in polyacrylamide gel as effectively as in magnetic field. Theoretical models are analyzed for orientation of disc-shaped and rod-shaped chloroplasts. Of these two, only the model for disc-shaped chloroplasts can account for the experimental results. Based on experimental data, the approaches are proposed to estimate the order of alignment of thylakoid membranes within chloroplasts, and to calculate the angles made by transition dipoles with the thylakoid membrane plane. Internal structure of chloroplasts (alignment of thylakoid membranes) is shown to be unchanged upon squeezing of the gel samples.

[Selective action of ruby laser radiation on the pigment apparatus of higher plants]. / Selektivnoe deistvie izlucheniia rubinovogo lazera na pigmentnyi apparat vysshikh rastenii.

Low-temperature (77 K) luminescence of pea chloroplasts and sub-chloroplast particles enriched with different components of photosynthetic pigment apparatus, and changes of luminescence induced by ruby laser pulse irradiation (694.3 nm) at various intensities were studied. The measured spectra showed selective laser - induced damage revealed by a predominant decrease of longer - wavelength luminescence band (near 730 nm). The effect is associated with light subchloroplast particles. Damage in PSI reaction centres occurred at power densities higher than a threshold value (5-8) X 10(10) W . m-2. No significant selective changes in luminescence spectra of PS2 pigment forms were observed at power densities up to 10(12) W . m-2. PSI - associated pigment forms that luminescence at 750-760 nm appear to be the most sensitive to laser irradiation. These forms are presumably the "sinks" for extra excitation energy in PSI.

[Analysis of the linear dichroism of reaction centers oriented in polyacrylamide gel]. / Analiz lineinogo dikhroizma reaktsionnykh tsentrov, orientirovannykh v poliakrilamidnom gele.

A model for orientation of rigid disc-shaped and rod-shaped macromolecules in polymer is analysed. Analytical expressions are given for the dependence of linear dichroism value on the angle between the transition dipole vector and the axis of the macromolecules, for certain deformation of the sample. The reaction centers from Rhodopseudomonas sphaeroides R-26 are oriented as rod-shaped particles. The angles between the long axis of the reaction center and transition dipoles for bacteriochlorophyll 31.8 +/- 2,4 degrees (870 nm), 42,3 +/- 0.9 degrees (796 nm), 68.0 +/- 0.6 degrees (600 nm) and for bacteriopheophytin 76.5 +/- 1.9 degrees (760 nm), 39.8 +/- 1.5 degrees (540 nm) are determined from absorption spectra.

Orientation and linear dichroism of the reaction centers from Rhodopseudomonas sphaeroides R-26.

Linear dicroism of chromatophores and isolated reaction centers from the photosynthetic bacterium Rhodopseudomonas sphaeroides strain R-26 was studied using a novel technique of orientation. The results are discussed in view of the reaction center structure and its position in the membrane. The advantages of the new orientation technique are also outlined.