1-X-3 motif in inter-protein recognition: structures, widespreading and possible practical application.

Similarities in the discrete mode and size of contact areas of a wide range of protein complexes allows us to suggest the existence of a limited number of types of inter-protein interactions. Comparison of structures of bound determinants indicates that the double-module, 1-X-3 type of motif is widespread in recognition processes. Thus, in many cases, the sites of ligand recognition are formed by two significant amino acids and separated by insignificant ones. Typical examples of such motifs are the RGD sequence of some adhesive and haemostatic proteins, the primary sites for plasminogen sorption on the fibrin network, the reactive sites of protein inhibitors of serine proteinases, and the sites for activation of the hydrolysis of protein pro-forms and receptors. It is assumed that there is widespread double-module determinants in many inter-protein interactions.     

Regulation of chloroplast metabolism in leaves: Evidence that NADP-dependent glyceraldehydephosphate dehydrogenase,but not ferredoxin-NADP reductase,controls electron flow to phosphoglycerate in the dark-light transition

P₇₀₀ is rapidly, but only transiently photooxidized upon illuminating dark-adapted leaves. Initial oxidation is followed by a reductive phase even under far-red illumination which excites predominantly photosystem (PS) I. In this phase, oxidized P₇₀₀ is reduced by electrons coming from PSII. Charge separation in the reaction center of PSI is prevented by the unavailability of electron acceptors on the reducing side of PSI. It is subsequently made possible by the opening of an electron gate which is situated between PSI and the electron acceptor phosphoglycerate. Electron acceptors immediately available for reduction while the gate is closed corresponded to 10 nmol · (mg chlorophyll)^–1 electrons in geranium leaves, 16 nmol · (mg chlorophyll)^–1 in sunflower and 22 nmol · (mg chlorophyll)^–1 in oleander. Reduction of NADP during the initial phase of P₇₀₀ oxidation showed that the electron gate was not represented by ferredoxin-NADP reductase. Availability of ATP indicated that electron flow was not hindered by deactivation of the thylakoid ATP synthetase. It is concluded that NADP-dependent glyceraldehydephosphate dehydrogenase is completely deactivated in the dark and activated in the light. The rate of activation depends on the length of the preceding dark period. As chloroplasts contain both NAD- and NADP-dependent glyceraldehydephosphate dehydrogenases, deactivation of the NADP-dependent enzyme disconnects chloroplast NAD and NADP systems and prevents phosphoglycerate reduction in the dark at the expense of NADPH and ATP which are generated by glucose-6-phosphate oxidation and glycolytic starch breakdown, respectively.Abbreviations Chl chlorophyll - P₇₀₀ electron donor pigment in the reaction center of photosystem I Cooperation of the Institute of Botany of the University of Würzburg with the Institute of Astrophysics and Atmospheric Physics of the Estonian Academy of Sciences in Tartu was supported by the Deutsche Forschungsgemeinschaft and the Estonian Academy of Sciences. This work was performed within the Sonderforschungsbereich 251 of the University of Würzburg.     

Acclimation of barley to changes in light intensity: photosynthetic electron transport activity and components

Barley seedlings (Hordeum vulgare L. Boone) were grown at 20°C with 16 h/8 h light/dark cycle of either high (H) intensity (500 mole m^-2 s^-1) or low (L) intensity (55 mole m^-2 s^-1) white light. Plants were transferred from high to low (H L) and low to high (L H) light intensity at various times from 4 to 8 d after leaf emergence from the soil. Primary leaves were harvested at the beginning of the photoperiod. Thylakoid membranes were isolated from 3 cm apical segments and assayed for photosynthetic electron transport, Photosystem II (PS II) atrazine-binding sites (Q_B), cytochrome f(Cytf) and the P-700 reaction center of Photosystem I (PS I). Whole chain, PS I and PS II electron transport activities were higher in H than in L controls. Q_B and Cytf were elevated in H plants compared with L plants. The acclimation of H L plants to low light occurred slowly over a period of 7 days and resulted in decreased whole chain and PS II electron transport with variable effects on PS I activity. The decrease in electron transport of H L plants was associated with a decrease in both Q_B and Cytf. In L H plants, acclimation to high light occurred slowly over a period of 7 days with increased whole chain, PS I and PS II activities. The increase in L H electron transport was associated with increased levels of Q_B and Cytf. In contrast to the light intensity effects on Q_B levels, the P-700 content was similar in both control and transferred plants. Therefore, PS II/PS I ratios were dependent on light environment.Abbreviations Asc ascorbate - BQ 2,5-dimethyl-p-benzoquinone - DBMIB 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone - DCIP 2,6-dichlorophenolindophenol - H control plants grown under high light intensity - H L plants transferred from high to low light intensity - L low control plants grown under low light intensity - L H plants transferred from low to high light intensity - MV methyl viologen - P-700 photoreaction center of Photosystem I - Q_B atrazine binding site - TMPD N,N,N,N-tetramethyl-p-phenylenediamine Cooperative investigations of the United States Department of Agriculture, Agricultural Research Service, and the North Carolina Agricultural Research Service, Raleigh, NC. Paper No. 11990 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh, NC 27695-7643, USA.     

Inhibitor binding to isolated chloroplast cytochrome bf complex

Effects of three inhibitors of quinol oxidation in the chloroplast cytochrome bf complex (stigmatellin, tridecylstigmatellin and dibromothymoquinone) were studied in an isolated system comprising Photosystem I (PS I) particles, plastocyanin (PC) and cytochrome bf complex, in the absence of quinol or quinone. Addition of these inhibitors increased the extent of cytochrome f oxidation after a laser flash created oxidised PS I reaction centre (P700) and PC, and decreased somewhat the extent of PC oxidation. The re-reduction of oxidised P700 was more complete than when inhibitor was absent. The data were simulated with reactions which included the putative reduction of cytochrome f by the Rieske centre (FeS) and different rate-coefficients according as to whether inhibitor was bound to the bf complex or not. It was concluded that under the conditions studied the Rieske centre donated electrons to oxidised cytochrome f and plastocyanin with an average rate coefficient of 35 s^–1. This electron transfer was prevented by any of the three inhibitors, which also increased the equilibrium coefficient for the cytochrome f/PC reaction by a maximum factor of two. This increase corresponded to a decrease in the back reaction coefficient and an increase in the forward rate. The equilibrium coefficient for the reduction of oxidised P700 by PC was about 2 in the absence of inhibitor but increased to about 20 in their presence, but only if cytochrome bf complex was additionally present. This was attributed to the transient formation of complexes between P700 with bound plastocyanin, and bf complex. The operative mid-point potential of FeS, if that of cytochrome f is 370 mV, was 390 mV. Deviations in midpoint potentials (P700/plastocyanin) from solution values were attributed to the bound state of the reactants. Estimates were made of the binding coefficient of each of the three inhibitors to p-sites in the cytochrome bf complex in the absence of competing quinol. A stoichiometry of two inhibitors per bf dimer was necessary to cause the above changes in reduction potential of cyt f and PC. A result of one inhibitor per dimer was statistically unlikely, particularly in the case of tridecylstigmatellin.Abbreviations Cyt- cytochrome - DBMIB(H₂)- 2,5-dibromo-3--ethyl-6-isopropyl-p-benzoquinone (reduced) - E _m- midpoint reduction potential of a couple relative to the standard hydrogen electrode - e-t- electron transfer - FeS (or R)- Rieske iron-sulphur centre - HEPES- N-2-hydroxyethylpiperazine-N-2-ethanesulphonic acid - Mega-9- nonoyl-N-methylglucamide - n-site (Q_r-site)- quinone reduction site in cytochrome bf complex - PC- plastocyanin - p-site (Q_o-site)- quinol oxidation site in cytochrome bf complex - PQ- plastoquinone - PSI- Photosystem I - P700- reaction centre in Photosystem I - TDS- tridecyl stigmatellin     

Vertical canopy structure of Dutch chalk grasslands in relation to their management

Analysis of leaf canopy overtopping relationships was carried out using a non-destructive point quadrat method followed by a destructive stratified harvest of the above-ground phytomass in Dutch chalk grasslands with different management: summer sheep grazing and annual autumn mowing. The two methods of analysis are compared: e.g. relative leaf overtopping can be recorded by the point quadrat method but it is obscured in vertical vegetation profiles based on stratified phytomass distribution. However the stratified harvest method describes the relationship between canopy phytomass and light microclimate, recorded by measuring Photosynthetically Active Radiation (PAR) at different heights in the vegetation. Plant growth form during peak standing crop is of greater significance than Raunkiaerian life form in determining structure of chalk grassland vegetation. In annually mown grasslands, the tall graminoid growth form, shown by Brachypodium pinnatum, reduces PAR in the lower canopy and suppresses most other growth forms except those which can reach the higher canopy levels, e.g. clumped herbs such as Origanum vulgare. In contrast, grazing can result in a reduction of dominance from the tall graminoids and reduction of the abundance of taller, grazing-sensitive herbs, e.g. clumped and climbing herbs such as Origanum and Lathyrus pratensis, and an increase in grazing-tolerant species, e.g. smaller rosette herb growth forms, e.g. Leontodon hispidus and shorter rhizomatous or stoloniferous graminoids, e.g. Carex flacca and Briza media. Changes in both the overtopping hierarchy at the peak of the growing season and the intensity of overtopping in the course of a growing season are of conclusive importance in determining the relative abundance of species in the vegetation.     

Range of photosynthetic control of postillumination P700+ reduction rate in sunflower leaves

The kinetics of the postillumination reduction of P700⁺ which reflects the rate constant for plastoquinol (PQH₂) oxidation was recorded in sunflower leaves at different photon absorption densities (PAD), CO₂ and O₂ concentrations. The P700 oxidation state was calculated from the leaf transmittance at 830 nm logged at 50 s intervals. The P700⁺ dark reduction kinetics were fitted with two exponents with time constants of 6.5 and about 45 ms at atmospheric CO₂ and O₂ concentrations. The time constant of the fast component, which is the major contributor to the linear electron transport rate (ETR), did not change over the range of PADs of 14.5 to 134 nmol cm^-2 s^-1 in 21% O₂, but it increased up to 40 ms under severe limitation of ETR at low O₂ and CO₂. The acceptor side of Photosystem I (PS I) became reduced in correlation with the downregulation of the PQH₂ oxidation rate constant. It is concluded that thylakoid pH-related downregulation of the PQH₂ oxidation rate constant (photosynthetic control) is not present under normal atmospheric conditions but appears under severe limitation of the availability of electron acceptors. The measured range of photosynthetic control fits with the maximum variation of ETR under natural stress in C₃ plants. Increasing the carboxylase/oxygenase specificity would lead to higher reduction of the PS I acceptor side under stress.Abbreviations Cyt b ₆ f cytochrome b ₆ f complex - C_w cell-wall CO₂ concentration, M - ETR electron transport rate - Fd ferredoxin - FNR ferredoxin-NADP reductase - FRL far-red light - PC plastocyanin - PAD photon absorption density nmol cm^-2 s^-1 - PFD photon flux density nmol cm^-2 s^-1 - PS I Photosystem I complex - PQ plastoquinon - PQH₂ plastoquinol - PS II Photosystem II complex - P700 Photosystem I donor pigment, reduced - S830 830 nm signal (D830, difference of S830 from the dark level) - WL white light - Y_l maximum quantum yield of PS I electron transport, rel. un     

Effects of pH on the kinetics of redox reactions in and around the cytochromebf complex in an isolated system

Rate-coefficients describing the electron transfer reactions between P700 and plastocyanin, between cytochromef in cytochromebf complexes and plastocyanin, and between decyl plastoquinol and cytochromebf complexes were determined as a function of pH in the range 4–10 from flash-induced absorbancy changes at four wavelengths. The reactions between P700 and plastocyanin, and between cytochromef and plastocyanin were optimised when there was electrostatic interaction between ionised acidic groups in plastocyanin with a pK_a of 4.3–4.7 and ionised basic constituents in P700 (assumed to be in the PSI-F subunit) and in cytochromef, with a pK_b of 8.9–9.4. The basic groups are thought to be lysine rather than arginine. This mechanism agrees with that inferred from effects of ionic strength changes on rate-coefficients. The relation between the second-order rate-coefficient for decyl plastoquinol oxidation by thebf complex and pH was characterised by a pK_a of 6.1. This is interpreted as showing that the anion radical form of that quinol, which has a pK_a of 6, and which becomes progressively protonated when pH is changed from 7 to 5, is essential to reduce cytochromeb-563 (low potential) during quinol oxidation. Above pH 9, permanent effects were observed on this rate-coefficient, which were absent in the reactions between P700, plastocyanin and cytochromef.     

Resource availability at three presumed stages of a heathland succession on the Lüneburger Heide,Germany

Abstract. The availability of maj or plant resources was investigated in three vegetation types that were assumed to represent different stages of a secondary succession on heathland on the Lüneburger Heide, northwestern Germany. Canopy transmission and absorption of photosynthetically active radiation (PAR), soil-water availability, and nutrient (Ca, K, Mg, N, P) availability were monitored with high spatial and temporal resolution in (1) a Calluna vulgaris heathland, (2) a pioneer birch-pine forest and (3) a late-successional oak-beech forest, situated close to each other on comparable geological substrate (diluvial). Mean fractional transmission of PAR during summer decreased from 0.48 in the heathland to 0.04 in the oak-beech forest while the fractional canopy absorption increased from 0.49 to 0.92. Soil-water availability as indicated by the soil-water potential, was significantly influenced by differential canopy interception loss and characteristic rooting patterns in the three vegetation types. Annual mean nutrient concentrations in the equilibrium soil solution were similar or, for some elements, increased from the heathland to the birch-pine and the oak-beech forest despite a growing demand. A marked increase was found for the total nutrient pools in the soil-organic layer between early and late successional vegetation types. On the poor glacial parent material, nutrient pools seem to be strongly dependent on stand productivity and litter production which increased with succession. Thus, for nutrients, facilitation seems to be important in this type of succession.     

Measurement of the relative electron-transport capacity of Photosystem I and Photosystem II in spinach chloroplasts

The ratio of Photosystem (PS) II to PS I electron-transport capacity in spinach chloroplasts was compared from reaction-center and steady-state rate measurements. The reaction-center electron-transport capacity was based upon both the relative concentrations of the PS II_α, PS II_β and PS I centers, and the number of chlorophyll molecules associated with each type of center. The reaction-center ratio of total PS II to PS I electron-transport capacity was about 1.8:1. Steady-state electron-transport capacity data were obtained from the rate of light-induced absorbance-change measurements in the presence of ferredoxin-NADP⁺, potassium ferricyanide and 2,5-dimethylbenzoquinone (DMQ). A new method was developed for determining the partition of reduced DMQ between the thylakoid membrane and the surrounding aqueous phase. The ratio of membrane-bound to aqueous DMQH₂ was experimentally determined to be 1.3:1. When used at low concentrations (200 μM), potassium ferricyanide is shown to be strictly a PS I electron acceptor. At concentrations higher than 200 μM, ferricyanide intercepted electrons from the reducing side of PS II as well. The experimental rates of electron flow through PS II and PS I defined a PS II/PS I electron-transport capacity ratio of 1.6:1.     

Surface potential and reaction of the membrane-bound electron transfer components. II. Integrity of the chloroplast membrane and reaction of P-700

Electrostatic characteristics of the membrane surface in the vicinity of P-700 were estimated by analyzing the salt and detergent effects on its reaction rate with ionic reagents using the Gouy-Chapman diffuse double layer theory in various preparations of chloroplasts.

Upon disruption of thylakoid membranes by sonic treatment or by treatment with digitonin, the reaction rate markedly increased, while the estimated surface charge density became smaller.

It was concluded that the membrane surface which determines the reaction rate between P-700 and the ionic reagents changed as the disruption of thylakoid structure. The outer thylakoid surface had more negative charges than the inner one.

Changes in the electrical potential profile across the thylakoid membrane during the illumination were also discussed from these results.