Pheophytin-mediated energy storage of photosystem II particles detected by photoacoustic spectroscopy

The photoacoustic (PA) characteristics (energy storage and heat dissipation) of photosystem II (PSII) core-enriched particles from barley were studied (i) in conditions where there was electron flow, i.e., in the presence of a combination of the electron acceptor K₃ Fe (CN)₆, referred to as FeCN, and the electron donor diphenylcarbazide (DPC), and (ii) in conditions where electron flow was suppressed, i.e., in the absence of FeCN and DPC. The experimental data show that a decrease of heat dissipation with a minimum at 540 nm can be interpreted as energy storage resulting from the presence of pheophytin (Pheo) in the PSII particles. On account of the capability of the PA method to measure the energy absorbed by the chromophores which is converted to heat, it is suggested that the PA detection of Pheo present in the PSII complex will permit to clarify the function of processes involving non-radiative relaxation of excited states in P680-Pheo-Q_A interactions.Abbreviations -Car -Carotene - Chl Chlorophyll - DPC Diphenylcarbazide - EPR Electron Paramagnetic Resonance - FeCN potassium ferricyanide - HEPES N-2-hydroxyethylenepiperazine-N-2-ethanesulfonate - P680 reaction center of PSII - PA Photoacoustic - Pheo pheophytin - PSI photosystem I - PSII photosystem II - Q_A primary electron acceptor of PSII     

Improvement of four sigma analysis for the investigation of oxygen evolution by Photosystem II

We present here an improvement to the analysis of oxygen evolution with four sigma coefficients (4-S) by computing z, the sum of the S-state probabilities, which was introduced earlier (Delrieu and Rosengard 1987, Biochim Biophys Acta 892: 163–171). We demonstrate that z is equal to the ratio of two consecutive Mean Y (the estimation of the steady state oxygen production based on local properties) found by three sigma analysis. The quantity z is useful for computing double-hits, and for showing the inactivation/activation processes of PS II complexes. Three sigma analysis assumes z=1 exactly; since this is not verified, it is argued that four sigma analysis is closer to the real workings of the water oxidizing complex. Oxygen evolution can then be interpreted in the frame of a modified Kok's model where the sum of the probabilities equals z. We therefore suggest that the closer fitting of four sigma analysis to oxygen production data is not simply due to an extra, unnecessary variable, but to the fact that PS II complexes can be inactivated and reactivated under flashing light. Finally, in order to facilitate the use of four sigma analysis, a computer program is made available upon request.     

The refined 2.15 A X-ray crystal structure of human liver cathepsin B: the structural basis for its specificity.

From the lysosomal cysteine proteinase cathepsin B, isolated from human liver in its two-chain form, monoclinic crystals were obtained which contain two molecules per asymmetric unit. The molecular structure was solved by a combination of Patterson search and heavy atom replacement methods (simultaneously with rat cathepsin B) and refined to a crystallographic R value of 0.164 using X-ray data to 2.15 A resolution. The overall folding pattern of cathepsin B and the arrangement of the active site residues are similar to the related cysteine proteinases papain, actinidin and calotropin DI. 166 alpha-carbon atoms out of 248 defined cathepsin B residues are topologically equivalent (with an r.m.s. deviation of 1.04 A) with alpha-carbon atoms of papain. However, several large insertion loops are accommodated on the molecular surface and modify its properties. The disulphide connectivities recently determined for bovine cathepsin B by chemical means were shown to be correct. Some of the primed subsites are occluded by a novel insertion loop, which seems to favour binding of peptide substrates with two residues carboxy-terminal to the scissile peptide bond; two histidine residues (His110 and His111) in this "occluding loop' provide positively charged anchors for the C-terminal carboxylate group of such polypeptide substrates. These structural features explain the well-known dipeptidyl carboxypeptidase activity of cathepsin B. The other subsites adjacent to the reactive site Cys29 are relatively similar to papain; Glu245 in the S2 subsite favours basic P2-side chains. The above mentioned histidine residues, but also the buried Glu171 might represent the group with a pKa of approximately 5.5 near the active site, which governs endo- and exopeptidase activity. The "occluding loop' does not allow cystatin-like protein inhibitors to bind to cathepsin B as they do to papain, consistent with the reduced affinity of these protein inhibitors for cathepsin B compared with the related plant enzymes.     

Electron Transport-Dependent Chlorophyll-a Fluorescence Quenching by O(2) in Various Algae and Higher Plants

A comparison of chlorophyll-a fluorescence in brown algae (Macrocystis integrifolia, Fucus vesiculosis), green algae (Scenedesmus obliquus, Ulva sp.) and higher plants (bean, corn) show differences in the relative fluorescence intensities and induction time courses which characterize each type of plant. These differences are not reflected in either the maximum fluorescence emission in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (F_max) or the nonvariable fluorescence (F_o). Constancy of F_o and F_max suggests functional similarities of photosystem II and associated antennae pigments in the various classes of plants. The time course differences are observed only in the absence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea and appear, therefore, to be electron transport dependent. During induction, the peak in fluorescence (F_p) is much lower in all of the algae studied than in the higher plants. Exogenous O₂ strongly quenches F_p in all plants studied and our data indicate that the low F_p in the algae can be partially accounted for by endogenous O₂ quenching.     

Differential activation of the mouse beta-globin promoter by enhancers.

A series of plasmids was constructed to study the effect of two enhancers, the simian virus 40 72-base-pair repeat and the Harvey sarcoma virus 73-base-pair repeat, on the mouse beta maj-globin promoter. These plasmids contain the mouse beta maj-globin promoter linked to the Escherichia coli galK gene, thus allowing galactokinase enzyme activity to be used as a measure of promoter function. In CV-1 (primate) cells, it was found that an enhancer is required for optimal promoter activity and that the simian virus 40 (primate) enhancer increases galactokinase fourfold more than the Harvey sarcoma virus (mouse) enhancer. In L (mouse) cells, however, the Harvey sarcoma virus enhancer is 1.3-fold stronger than the simian virus 40 enhancer. These data support the hypothesis that enhancer activity can be species specific. Furthermore, when both enhancers are present on the same plasmid, their effect is additive on the beta-globin promoter whether the plasmid is in CV-1 cells or L cells.     

HIV-1 and its envelope glycoprotein down-regulate chemotactic ligand receptors and chemotactic function of peripheral blood monocytes

Peripheral blood monocytes from AIDS patients exhibit defective migratory responses to chemotactic stimuli in vitro and to inflammatory sites in vivo. In studies presented here, normal monocytes were infected with the HIV-1/HTLV-IIIBa-L isolate in vitro and evaluated for chemotactic responsiveness. Within 2 days after viral exposure, but before evidence of virus production in the monocytes, chemotactic activity was significantly impaired. Decreased chemotactic activity was associated with modulation of receptors for the chemotactic ligands, C5a and FMLP, on the monocyte cell surface. In addition to HIV-1, monocytes treated with purified HIV-1 envelope glycoprotein gp120 demonstrated a comparable modulation of chemotactic ligand receptors and migratory function. In addition, the HIV-1 or HIV-1 gp120-treated monocytes were induced to undergo differentiation as monitored by HLA-DR expression. Immunoprecipitation of the gp120 with a specific antibody reversed its effects on monocyte chemotaxis and HLA-DR expression. Taken together, these data indicate that the initial interaction of HIV-1 with the monocyte is not passive, but that the binding of HIV-1 and/or HIV-1 gp120 to the CD4R on monocytes transduces a signal leading to transient monocyte activation.     

Individual risk prediction: Comparing random forests with Cox proportional-hazards model by a simulation study

With big data becoming widely available in healthcare, machine learning algorithms such as random forest (RF) that ignores time-to-event information and random survival forest (RSF) that handles right-censored data are used for individual risk prediction alternatively to the Cox proportional hazards (Cox-PH) model. We aimed to systematically compare RF and RSF with Cox-PH. RSF with three split criteria [log-rank (RSF-LR), log-rank score (RSF-LRS), maximally selected rank statistics (RSF-MSR)]; RF, Cox-PH, and Cox-PH with splines (Cox-S) were evaluated through a simulation study based on real data. One hundred eighty scenarios were investigated assuming different associations between the predictors and the outcome (linear/linear and interactions/nonlinear/nonlinear and interactions), training sample sizes (500/1000/5000), censoring rates (50%/75%/93%), hazard functions (increasing/decreasing/constant), and number of predictors (seven, 15 including noise variables). Methods' performance was evaluated with time-dependent area under curve and integrated Brier score. In all scenarios, RF had the worst performance. In scenarios with a low number of events (⩽70), Cox-PH was at least noninferior to RSF, whereas under linearity assumption it outperformed RSF. Under the presence of interactions, RSF performed better than Cox-PH as the number of events increased whereas Cox-S reached at least similar performance with RSF under nonlinear effects. RSF-LRS performed slightly worse than RSF-LR and RSF-MSR when including noise variables and interaction effects. When applied to real data, models incorporating survival time performed better. Although RSF algorithms are a promising alternative to conventional Cox-PH as data complexity increases, they require a higher number of events for training. In time-to-event analysis, algorithms that consider survival time should be used.     

EFFECTS OF LONG TERM EXPOSURE TO LOW TEMPERATURE ON THE PHOTOSYNTHETIC APPARATUS OF DUNALIELLA TERTIOLECTA (CHLOROPHYCEAE)1

The effects of long term exposure to suboptimal growth temperature on the photosynthetic apparatus of Dunaliella tertiolecta Butcher were investigated using carbon fixation rate versus irradiance curves and the variable fluorescence induction method. Carbon fixation rates per unite chlorophyll a at saturating (p^B_m) and subsaturating (α^B) irradiances were 55% and 39% lower, respectively, at 12° C than at 20° C. Chlorophyll a quotas and the spectrally averaged in vivo absorption cross section normalized to chlorophyll a (a^*) were not significantly different at these two temperatures. Analysis of the fluorescence kinetics revealed 1) no significant variations of the amount of PSII photoactive reaction centers per unit chlorophyll a, 2) a 14% decrease of the PSII quantum yield(+) and 3) a 29% decrease of the energy transfer efficiency between the light harvesting chlorophyll a pigment bed and the PSII reaction centers. The decrease in energy transfer efficiency between the antennae and the PSII reaction centers at 12° C was interpreted as a mechanism to avoid photoinhibition.