Induction changes in photosystems I and II in plant leaves upon modulation of membrane ion transport

The steady-state regime of linear photosynthetic electron transport implies concerted operation of photosystems I and II (PSI and PSII) in plant leaves. Acidification of the thylakoid lumen is known to cause down-regulation of PSII photochemical activity but it is not yet clear how the proton accumulation in the lumen affects the PSI activity and coordinated operation of the two photosystems in intact leaves. Chlorophyll fluorescence and absorbance of oxidized chlorophyll P700 in the near-infrared region ΔA _810–870 (ΔA ₈₁₀) are convenient noninvasive indicators of the redox state of PSII and PSI components, respectively. Simultaneous measurements of chlorophyll fluorescence and ΔA ₈₁₀ in pea leaves revealed that some kinetic stages in the induction curves occur synchronously both in dark-adapted and preilluminated leaves. After the treatment of leaves with ionophores promoting or inhibiting the light-induced thylakoid pH gradient (valinomycin, nigericin, monensin), the induction curves of ΔA ₈₁₀ and chlorophyll fluorescence were consistently modified. The results suggest that characteristic stages of ΔA ₈₁₀ induction curve, representing the second and the third waves of P700 photooxidation, are closely related to ΔpH generation, although the bases of ΔpH dependence differ for these two stages. The second wave of ΔA ₈₁₀ depends presumably on stroma alkalinization as a precondition for photoactivation of electron flow from PSI to terminal acceptors. The third wave of ΔA ₈₁₀ is apparently due to retardation of electron flow between PSII and PSI upon acidification of the lumen.     

Electrochemistry of the CuA domain of Thermus thermophilus cytochrome ba 3

 The electrochemistry of a water-soluble fragment from the Cu_A domain of Thermus thermophilus cytochrome ba ₃ has been investigated. At 25  °C, Cu_A exhibits a reversible reduction at a pyridine-4-aldehydesemicarbazone-modified gold electrode (0.1 M Tris, pH 8) with E° = 0.24 V vs NHE. Thermodynamic parameters for the [Cu(Cys)₂Cu]^+/0 electrode reaction were determined by variable-temperature electrochemistry (ΔS°_rc = –5.4(12) eu, ΔS° = –21.0(12) eu, ΔH° = –11.9(4) kcal/mol;ΔG° = –5.6 (11) kcal/mol). The relatively small reaction entropy is consistent with a low reorganization energy for [Cu(Cys)₂Cu]^+/0 electron transfer. An irreversible oxidation of [Cu(Cys)₂Cu]⁺ at 1 V vs NHE confirms that the Cu^II:Cu^II state of Cu_A is significantly destabilized relative to the Cu^II state of analogous blue-copper proteins. Received: 3 June 1996 / Accepted: 26 August 1996     

Pharmacological properties of the potassium-activated inward current in pyramidal hippocampal neurons

Elevation of the external potassium concentration induced a two-phase inward current in freshly isolated pyramidal hippocampal neurons. This current was voltage-dependent and demonstrated strong inward rectification. The current consisted of a leakage current and a time-dependent current (τ=40–50 msec at 21°C); the latter was designated asI _ΔK. As was shown earlier, K⁺ is a major charge carrier in the development of slow potassium-activated current. The pharmacological properties ofI _ΔK were studied using a patch-clamp technique.I _ΔK was completely blocked by external 10 mM TEA or 5 mM Ba²⁺ (IC₅₀=480±90mM) and exhibited low sensitivity to extracellular Cs⁺ (2 mM). This current was not affected by 1 mM 4-aminopyridine and was insensitive to a muscarinic agonist, carbachol (50 μM), and to 1 mM extracellular Cd²⁺. Elevation of external Ca²⁺ from 2.5 mM to 10 mM did not changeI _ΔK. Our data indicate that the pharmacological properties ofI _ΔK differ from those of other voltage-gated potassium currents, but more specific blockers must be used to make this evidence conclusive.     

A model of photosystem II for the analysis of fast fluorescence rise in plant leaves

The polyphasic patterns of fluorescence induction rise in pea leaves in vivo and after the treatment with ionophores have been studied using a Plant Efficiency Analyzer. To analyze in detail photosystem II (PS II) electron transfer processes, an extended PS II model was applied, which included the sums of exponential functions to specify explicitly the light-driven formation of the transmembrane electric potential (ΔΨ(t)) as well as pH in the lumen (pH_L(t)) and stroma (pH_S(t)). PS II model parameters and numerical coefficients in ΔΨ(t), pH_L(t), and pH_S(t) were evaluated to fit fluorescence induction data for different experimental conditions: leaf in vivo or after ionophore treatment at low or high light intensity. The model imitated changes in the pattern of fluorescence induction rise due to the elimination of transmembrane potential in the presence of ionophores, when ΔΨ = 0 and pH_L(t), pH_S(t) changed to small extent relative to control values in vivo, with maximum ΔΨ(t) ∼ 90 mV and ΔΨ(t) ∼ 40 mV for the stationary state at ΔpH ≅ 1.8. As the light intensity was increased from 300 to 1200 μmol m⁻² s⁻¹, the heat dissipation rate constants increased threefold for nonradiative recombination of P680⁺Phe⁻ and by ∼30% for P680⁺Q_A⁻. The parameters ΔΨ, pH_S and pH_L were analyzed as factors of PS II redox state populations and fluorescence yield. The kinetic mechanism of fluorescence quenching is discussed, which is related with light-induced lumen acidification, when ⁺Q_A⁻ and P680⁺ recombination probability increases to regulate the Q_A reduction.     

Dependence of chlorophyll P700 redox transients during the induction period on the transmembrane distribution of protons in chloroplasts of pea leaves

Differential absorbance measurements and fluorometry were applied to examine the impact of dicyclohexylcarbodiimide (DCCD, an inhibitor of H⁺ conductance in thylakoid membranes) and nigericin (a K⁺/H⁺ antiporter) on photoinduced redox state transients of chlorophyll P700 and the induction curves of chlorophyll fluorescence in pea (Pisum sativum L., cv. Premium) leaves. The treatment of leaves with DCCD strongly modified the kinetics of P700⁺ absorbance changes (ΔA ₈₁₀) by promoting rapid photooxidation of P700. These characteristic changes in ΔA ₈₁₀ induction kinetics and P700⁺ accumulation did not appear when the leaves were treated with DCCD in the presence of nigericin. In addition to opposite modifications of ΔA ₈₁₀ kinetics evoked by permeability-modifying agents, the fluorescence induction curves differed conspicuously depending on leaf incubation in DCCD solutions with or without nigericin. The observed modifications of fluorescence induction curves and ΔA ₈₁₀ indicate that DCCD suppresses electron transport from photosystem II (PSII) to P700, whereas this inhibition is removed by nigericin. The results suggest that slowing down of the electron transport rate in the presence of DCCD was caused by elevation of ΔpH in thylakoids. The prevention of pH gradient formation in the presence of protonophore lowered also the steady-state P700⁺ level in far-red irradiated leaves and accelerated the subsequent dark reduction of P700. These findings indicate that PSI-driven cyclic electron flow is accelerated after the removal of the pH gradient.     

Effects of the antidepressant pirlindole and its dehydro-derivative on the activity of monoamine oxidase-a and on GABAA receptors

The effects of pirlindole and dehydro-pirlindole on GABA_A receptors and MAO-A activity were investigated in vitro. Pirlindole was inactive as a GABA antagonist. Dehydro-pirlindole exhibited partial and selective blockade of a subset of GABA_A receptors with an EC₅₀ of 12μM and maximum reversal (ΔB_opt) of 42%. Inhibition of rat brain and human placenta MAO-A by both compounds was much more potent (with IC₅₀ range 0.3–0.005 μM). Their effects on MAO-A activity were partially reversible in vitro. In contrast to pirlindole, dehydro-pirlindole may act not only as MAO-A inhibitor but also as a clozapine-like selective GABA_A receptor blocker, preferentially blocking a subset of GABA_A receptors that are not sensitive to DMCM or Ro 5-4864. Pirlindole is the generic name of the drug pyrazidol.     

Kinetic and thermodynamic analysis of dimerization inhibitors binding to HIV protease monomers by surface plasmon resonance

The analysis of kinetic and thermodynamic parameters of binding of peptide and nonpeptide dimerization inhibitors of HIV protease (HIVp) to the enzyme monomers immobilized on an optical chip has been studied by surface plasmon resonance. The molecular interactions were investigated at different inhibitor concentrations (0–80 μM) and temperatures (15–35°C). Determination of kinetic (k _on, k _off), equilibrium (K _d), and thermodynamic (ΔG, ΔH, and -TΔS) has shown that both inhibitors are characterized by similar interaction parameters and the entropic term (-TΔS) of about −20 kcal/mol is the main driving force for the HIVp complex formation with the inhibitors, while the positive value (14 kcal/mol) of the enthalpic term (ΔH) counteracted the complex formation.     

Carbon isotopes and water use efficiency: sense and sensitivity

We revisit the relationship between plant water use efficiency and carbon isotope signatures (δ¹³C) of plant material. Based on the definitions of intrinsic, instantaneous and integrated water use efficiency, we discuss the implications for interpreting δ¹³C data from leaf to landscape levels, and across diurnal to decadal timescales. Previous studies have often applied a simplified, linear relationship between δ¹³C, ratios of intercellular to ambient CO₂ mole fraction (C _i/C _a), and water use efficiency. In contrast, photosynthetic ¹³C discrimination (Δ) is sensitive to the ratio of the chloroplast to ambient CO₂ mole fraction, C _c/C _a (rather than C _i/C _a) and, consequently, to mesophyll conductance. Because mesophyll conductance may differ between species and over time, it is not possible to determine C _c/C _a from the same gas exchange measurements as C _i/C _a. On the other hand, water use efficiency at the leaf level depends on evaporative demand, which does not directly affect Δ. Water use efficiency and Δ can thus vary independently, making it difficult to obtain trends in water use efficiency from δ¹³C data. As an alternative approach, we offer a model available at to explore how water use efficiency and ¹³C discrimination are related across leaf and canopy scales. The model provides a tool to investigate whether trends in Δ indicate changes in leaf functional traits and/or environmental conditions during leaf growth, and how they are associated with trends in plant water use efficiency. The model can be used, for example, to examine whether trends in δ¹³C signatures obtained from tree rings imply changes in tree water use efficiency in response to atmospheric CO₂ increase. This is crucial for predicting how plants may respond to future climate change. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Variation in leaf physiology of <Emphasis Type="Italic">Salix arctica</Emphasis> within and across ecosystems in the High Arctic: test of a dual isotope (Δ<Superscript>13</Superscript>C and Δ<Superscript>18</Superscript>O) conceptual model

Leaf carbon isotope discrimination (Δ¹³C) varies with the balance between net photosynthesis (A) and stomatal conductance (g _s ). Inferences that can be made with Δ¹³C are limited, as changes could reflect variation in A and/or g _s . Investigators have suggested that leaf δ¹⁸O enrichment above source water (Δ¹⁸O) may enable differentiation between sources of variation in Δ¹³C, as leaf Δ¹⁸O varies with transpiration rate (E), which is closely correlated with g _s when leaves experience similar leaf to air vapor pressure differences. We examined leaf gas exchange of Salix arctica at eight sites with similar air temperatures and relative humidities but divergent soil temperatures and soil water contents near Pituffik, Greenland (76°N, 38°W). We found negative correlations at the site level between g _s and Δ¹⁸O in bulk leaf tissue (r ² = 0.62, slope = −17.9‰/mol H₂O m⁻² s⁻¹, P = 0.02) and leaf α-cellulose (r ² = 0.83, slope = −11.5‰ mol H₂O m⁻² s⁻¹, P < 0.01), consistent with the notion that leaf water enrichment declines with increasing E. We also found negative correlations at the site-level between intrinsic water-use efficiency (iWUE) and Δ¹³C in bulk leaf tissue (r ² = 0.65, slope = −0.08‰/μmol CO₂ /mol H₂O, P = 0.02) and leaf α-cellulose (r ² = 0.50, slope = −0.05 ‰/[μmol CO₂ /mol H₂O], P = 0.05). When increasing Δ¹³C was driven by increasing g _s alone, we found negative slopes between Δ¹³C and Δ¹⁸O for bulk leaf tissue (−0.664) and leaf α-cellulose (−1.135). When both g _s and A _max increased, we found steeper negative slopes between Δ¹³C and Δ¹⁸O for bulk leaf tissue (−2.307) and leaf α-cellulose (−1.296). Our results suggest that the dual isotope approach is capable of revealing the qualitative contributions of g _s and A _max to Δ¹³C at the site level. In our study, bulk leaf tissue was a better medium than leaf α-cellulose for application of the dual isotope approach.     

Biodegradation of dioxins by recombinant Escherichia coli expressing rat CYP1A1 or its mutant

Among polychlorinated dibenzo-p-dioxins (PCDDs), 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TetraCDD) is the most toxic one. Recently, we reported that rat CYP1A1 mutant, F240A, expressed in yeast showed metabolic activity toward 2,3,7,8-TetraCDD. In this study, we successfully expressed N-terminal truncated P450s (Δ1A1 and ΔF240A) in Escherichia coli cells. Kinetic analysis using membrane fractions prepared from the recombinant E. coli cells revealed that ΔF240A has enzymatic properties similar to F240A expressed in yeast. The metabolism of PCDDs by recombinant E. coli cells expressing both ΔF240A and human NADPH-P450 reductase was also examined. When 2,3,7-TriCDD was added to the E. coli cell culture at a final concentration of 10 μM, approximately 90% of the 2,3,7-TriCDD was converted into multiple metabolites within 8 h. These results indicate the possible application of prokaryotic cells expressing ΔF240A to the bioremediation of PCDD-contaminated soil.     

Enhanced Solubility,Stability, and Transcorneal Permeability of Delta-8-Tetrahydrocannabinol in the Presence of Cyclodextrins

The purpose of this study was to investigate the effect of cyclodextrins (CDs) on aqueous solubility, stability, and in vitro corneal permeability of delta-8-tetrahydrocannabinol (Δ⁸-THC). Phase solubility of Δ⁸-THC was studied in the presence of 2-hydroxypropyl-β-cyclodextrin (HPβCD), randomly methylated-β-cyclodextrin (RMβCD) and sulfobutyl ether-β-cyclodextrin sodium salt (SβCD). Stability of Δ⁸-THC in 5% w/v aqueous CD solutions, as a function of pH, was studied following standard protocols. In vitro corneal permeation of Δ⁸-THC (with and without CDs) across excised rabbit cornea was also determined. Phase-solubility profile of Δ⁸-THC in the presence of both HPβCD and RMβCD was of the A_P type, whereas, with SβCD an A_L type was apparent. Aqueous solubility of Δ⁸-THC increased to 1.65, 2.4, and 0.64 mg/mL in the presence of 25% w/v HPβCD, RMβCD, and SβCD, respectively. Significant degradation of Δ⁸-THC was not observed within the study period at the pH values studied, except for at pH 1.2. Transcorneal permeation of Δ⁸-THC was dramatically improved in the presence of CDs. The results demonstrate that CDs significantly increase aqueous solubility, stability, and transcorneal permeation of Δ⁸-THC. Thus, topical ophthalmic formulations containing Δ⁸-THC and modified beta CDs may show markedly improved ocular bioavailability.     

Inorganic carbon uptake by an Antarctic sea-ice diatom,Nitzschia frigida

There have been no studies to date on the mechanisms of inorganic carbon acquisition by Antarctic microalgae. Consequently, we have examined inorganic carbon (DIC) use inNitzschia frigida, a diatom typical of the Antarctic bottom-ice community. The K_0.5(CO₂) of photosynthesis in this organism was estimated to be 1.09 μM at pH 7.5. The internal concentration of DIC was approximately 4050 μM at an external [DIC] of 45 μM. At air-equilibration levels of inorganic carbon this would be sufficient for a ten-fold accumulation ratio of CO₂. Cells ofN. frigida are capable of carbon-dependent photosynthesis at rates that exceed that expected from uncatalysed CO₂ supply to the cell. About 25% of the total carbonic anhydrase activity appears to be associated with the cell surface inN. frigida. These results support the hypothesis thatN. frigida, like many microalgae from temperate waters, has an active carbon-concentrating mechanism, associated with the ability to utilize external HCO ₃ ⁻ for photosynthesis.     

Phosphorylation of the pheromone-responsive Gβ protein of Saccharomyces cerevisiae does not affect its mating-specific signaling function

The pheromone-responsive G_β subunit of Saccharomyces cerevisiae (encoded by STE4) is rapidly phosphorylated at multiple sites when yeast cells are exposed to mating pheromone. It has been shown that a mutant form of Ste4 lacking residues 310–346, ste4Δ310–346, cannot be phosphorylated, and that its expression leads to defects in recovery from pheromone stimulation. Based on these observations, it was proposed that phosphorylation of Ste4 is associated with an adaptive response to mating pheromone. In this study we used site-directed mutagenesis to create two phosphorylation null (Pho⁻) alleles of STE4: ste4-T320 A/S335A and ste4-T322 A/S335A and ste4-T322A/S335A. When expressed in yeast, these mutant forms of Ste4 remained unphosphorylated upon pheromone stimulation. The elimination of Ste4 phosphorylation has no discernible effect on either signaling or adaptation. In addition, disruption of the FUS3 gene, which encodes a pheromone-specific MAP kinase, leads to partial loss of pheromone-induced Ste4 phosphorylation. Two-hybrid analysis suggests that the ste4Δ310–346 deletion mutant is impaired in its interaction with Gpa1, the pheromone-responsive G_α of yeast, whereas the Ste4-T320A/S335A mutant has normal affinity for Gpa1. Taken together, these results indicate that pheromone-induced phosphorylation of Ste4 is not an adaptive mechanism, and that the adaptive defect exhibited by the 310–346 deletion mutant is likely to be due to disruption of the interaction between Ste4 and Gpa1. Received: 14 February 1998 / Accepted: 28 February 1998     

Voltage changes involving photosystem II quinone–iron complex turnover

An electrometrical technique was used to investigate proton-coupled electron transfer between the primary plastoquinone acceptor Q_A⁻ and the oxidized non-heme iron Fe³⁺ on the acceptor side of photosystem II core particles incorporated into phospholipid vesicles. The sign of the transmembrane electric potential difference Δψ (negative charging of the proteoliposome interior) indicates that the iron–quinone complex faces the interior surface of the proteoliposome membrane. Preoxidation of the non-heme iron was achieved by addition of potassium ferricyanide entrapped into proteoliposomes. Besides the fast unresolvable kinetic phase (τ ∼ 0.1 μs) of Δψ generation related to electron transfer between the redox-active tyrosine Y_Z and Q_A, an additional phase in the submillisecond time domain (τ ∼ 0.1 ms at 23°C, pH 7.0) and relative amplitude ∼ 20% of the amplitude of the fast phase was observed under exposure to the first flash. This phase was absent under the second laser flash, as well as upon the first flash in the presence of DCMU, an inhibitor of electron transfer between Q_A and the secondary quinone Q_B. The rate of the additional electrogenic phase is decreased by about one-half in the presence of D₂O and is reduced with the temperature decrease. On the basis of the above observations we suggest that the submillisecond electrogenic reaction induced by the first flash is due to the vectorial transfer of a proton from external aqueous phase to an amino acid residue(s) in the vicinity of the non-heme iron. The possible role of the non-heme iron in cyclic electron transfer in photosystem II complex is discussed.     

Genetic determination of the effect of post-translational modification on the innate immune response to the 19 kDa lipoprotein of Mycobacterium tuberculosis

Background  

The 19 kDa lipoprotein of Mycobacterium tuberculosis (MTB) is an important target of the innate immune response. To investigate the effect of post-translation modification of this protein on innate recognition in the context of the whole bacillus, we derived a recombinant M. tuberculosis H37Rv that lacked the 19 kDa gene (Δ19) and complemented this strain by reintroduction of the 19 kDa gene into the chromosome as a single copy to produce Δ19::19. We also reintroduced the 19 kDa gene in two modified forms that lacked motifs for acylation (Δ19::19NA) and O-glycosylation (Δ19::19NOG).     

Remodeling of the glycosylation pathway in the methylotrophic yeast Hansenula polymorpha to produce human hybrid-type N-glycans

As a step forward to achieve the generation of human complex-type N-glycans in the methylotrophic yeast Hansenula polymorpha, we here report the modification of the yeast glycosylation pathway by heterologous expression of the human gene encoding β-1,2-N-acetylglucosaminyltransferase I (GnTI). For the optimal expression of human GnTI in the yeast Golgi compartment, the catalytic domain of the GnTI was fused to various N-terminal leader sequences derived from the yeast type II membrane proteins. The vectors containing GnTI fusion constructs were introduced into the H. polymorpha och1Δ single and och1Δalg3Δ double mutant strains expressing the ER-targeted Aspergillus saitoi α-1,2 mannosidase, respectively. Both of the glycoengineered Hpoch1Δ and Hpoch1ΔHpalg3Δ strains were shown to produce successfully the hybrid-type glycans with a monoantennary N-acetylglucosamine (GlcNAc₁Man₅GlcNAc₂ and GlcNAc₁Man₃GlcNAc₂, respectively) by N-glycan profile analysis of cell wall proteins. Furthermore, by comparative analysis of byproduct formation and the glycosylation site occupancy, we propose that the Hpoch1Δ strain would be more suitable than the Hpoch1ΔHpalg3Δ strain as a host for the production of recombinant proteins with humanized glycans.     

Membrane responses to changes in the extracellular potassium concentration in isolated hippocampal pyramidal neurons

The responses of freshly isolated hippocampal pyramidal neurons to rapid, elevations of the external potassium concentration ([K⁺] _out ) were investigated using the whole-cell variation of a patch-clamp technique. An elevation of [K⁺] _out induced a two-phase inward current at the membrane potentials more negative than the reversal potential for K ions. This current consisted of a leakage, current and a time-dependent current (τ=40–50 msec at 21°C), the latter designated below asI _ΔK. It displayed first-order activation kinetics that showed neither voltage, nor concentration dependence. The amplitude of this current was determined by the external K⁺ concentration and increased with hyperpolarization. Voltage dependence ofI _ΔK measured within the range from −20 to −120 mV was similar to that for inward rectifier. Activation ofI _ΔK was utterly dependent on Na⁺; substitution of extracellular Na⁺ with choline chloride almost completely depressedI _ΔK.I _ΔK was absent in the cells freshly dissociated from the nodosal and dorsal root ganglia. This suggests that this earlier unrecognized current is instrumental in preserving densely packed hippocampal pyramidal neurons from sudden increases in [K⁺] _out and following spontaneous over-excitation. It prevents the neurons from responding to K⁺-induced depolarizations by slowing down potassium influx.     

Femtosecond stage of electron transfer in reaction centers of the triple mutant SL178K/GM203D/LM214H of <Emphasis Type="Italic">Rhodobacter sphaeroides</Emphasis>

Coherent processes in an initial phase of charge transfer in reaction centers (RCs) of the triple mutant S(L178)K/G(M203)D/L(M214)H of Rhodobacter sphaeroides were investigated by difference (light — dark) absorption spectroscopy with 18 fsec time resolution. Electron transfer in the B cofactor branch is activated in this mutant, while the A-branch electron transfer is slowed in comparison with native RCs of Rba. sphaeroides. A bulk of absorption difference spectra was analyzed in the 940–1060 nm range (stimulated emission of excited bacteriochlorophyll dimer P* and absorption of bacteriochlorophyll anions B_A⁻ and β⁻, where β is a bacteriochlorophyll substituting the native bacteriopheophytin H_A) and in the 735–775 nm range (bleaching of the absorption band of the bacteriopheophytin H_B in the B-branch) in the −0.1 to 4 psec range of delays with respect to the moment of photoexcitation of P at 870 nm. Spectra were measured at 293 and 90 K. The kinetics of P* stimulated emission at 940 nm shows its decay with a time constant of ∼14 psec at 90 K and ∼18 psec at 293 K, which is accompanied by oscillations with a frequency of ∼150 cm⁻¹. A weak absorption band is found at 1018 nm that is formed ∼100 fsec after excitation of P and reflects the electron transfer from P* to β and/or B_A with accumulation of the P⁺β⁻ and/or P⁺B_A⁻ states. The kinetics of ΔA at 1018 nm contains the oscillations at ∼150 cm⁻¹ and distinct low-frequency oscillations at 20–100 cm⁻¹; also, the amplitude of the oscillations at 150 cm⁻¹ is much smaller at 293 than at 90 K. The oscillations in the kinetics of the 1018 nm band do not contain a 32 cm⁻¹ mode that is characteristic for native Rba. sphaeroides RCs having water molecule HOH55 in their structure. The ΔA kinetics at 751 nm reflects the electron transfer to HB with formation of the P⁺H_B⁻ state. The oscillatory part of this kinetics has the form of a single peak with a maximum at ∼50 fsec completely decaying at ∼200 fsec, which might reflect a reversible electron transfer to the B-branch. The results are analyzed in terms of coherent nuclear wave packet motion induced in the P* excited state by femtosecond light pulses, of an influence of the incorporated mutations on the mutual position of the energy levels of charge separated states, and of the role of water HOH55 in the dynamics of the initial electron transfer.     

Variation in carbon isotope discrimination within and among Sphagnum species in a temperate wetland

Field samples of bryophytes are highly variable in carbon isotope discrimination values (Δ, a measure of ¹³CO₂ uptake relative to ¹²CO₂), but it is unknown what affects Δ under field conditions, or how variation in Δ relates to bryophyte performance. This study employed field and greenhouse common garden studies to evaluate the influence of microsite, seasonal, and genetic variation on Δ in peatmosses. Three species of Sphagnum that occupy hollow (S. recurvum), carpet (S. palustre), and hummock (S. tenerum) habitats were sampled for relative growth rates (RGR), C:N ratio, and Δ throughout a growing season. Values of Δ ranged from 19.0 to 27.1‰. This variation was unrelated to species (P=0.61). However, Δ varied seasonally (P<0.001), with lower discrimination in the spring (mean 22.5‰), followed by summer (23.8‰) and winter (24.7‰). There was also significant microsite variation (P=0.015) which disappeared when plants were grown in a common garden. In both spring and summer, microsite variation in Δ was inversely related to RGR (P<0.001), but unrelated to C:N ratios (P>0.08). These results suggest that environmental, not genetic, variation at microsites affects Δ in non-vascular plants. However, environmental control of Δ is unlike that in vascular plants where water limitation lowers chloroplastic demand and increases resistance to carbon uptake. In non-vascular plants, water limitation lowers chloroplastic demand and decreases resistance to carbon uptake. These processes have additive effects and generate high spatial and seasonal variability in Δ. Received: 29 April 1999 / Accepted: 8 November 1999     

Sequence variants of chemokine receptor genes and susceptibility to HIV-1 infection

Genetic susceptibility to HIV infection was previously proven to be influenced by some chemokine receptor polymorphisms clustering on chromosome 3p21. Here the influence of 5 genetic variants was studied: Δ32CCR5, G(-2459)ACCR5, G190ACCR2, G744ACX3CR1 and C838TCX3CR1. They were screened in a cohort of 168 HIV-1 positive adults [HIV(+) group] and 151 newborns [control group] from northwestern Poland. PCR-RFLP was performed to screen for the variants (except for A32CCR5 polymorphism, where PCR fragment size was sufficient to identify the alleles) and then electrophoresed on agarose gel to determine fragment size. Distribution of genotypes and alleles was not significantly different between the groups except for theCCR5 polymorphisms, with the A32 allele and the (-2459)ACCR5 allele more frequent among neonates than in the HIV(+) group. No Δ32/Δ32 homozygotes were found in the HIV(+) group, but 16.1% were Δ32/wt heterozygotes. In the control group, 1.3% were Δ32/Δ32 homozygotes and 26.0% were Δ32/wt heterozygotes. Linkage between the chemokine polymorphisms was calculated using the most informative loci for haplotype reconstruction. Haplotypes containing Δ32 CCR5,190GCCR2 and 744ACX3CR1 were found to be significantly more common in the control group. This suggests an association between these haplotypes and resistance to HIV-1 infection.