Measurements Of Fluorescence Induction Kinetics Research Articles

Ca2+ effects on Fe(II) interactions with Mn-binding sites in Mn-depleted oxygen-evolving complexes of photosystem II and on Fe replacement of Mn in Mn-containing, Ca-depleted complexes.

Fe(II) cations bind with high efficiency and specificity at the high-affinity (HA), Mn-binding site (termed the "blocking effect" since Fe blocks further electron donation to the site) of the oxygen-evolving complex (OEC) in Mn-depleted, photosystem II (PSII) membrane fragments (Semin et al. in Biochemistry 41:5854, 2002). Furthermore, Fe(II) cations can substitute for 1 or 2Mn cations (pH dependent) in Ca-depleted PSII membranes (Semin et al. in Journal of Bioenergetics and Biomembranes 48:227, 2016; Semin et al. in Journal of Photochemistry and Photobiology B 178:192, 2018). In the current study, we examined the effect of Ca2+ cations on the interaction of Fe(II) ions with Mn-depleted [PSII(-Mn)] and Ca-depleted [PSII(-Ca)] photosystem II membranes. We found that Ca2+ cations (about 50mM) inhibit the light-dependent oxidation of Fe(II) (5µM) by about 25% in PSII(-Mn) membranes, whereas inhibition of the blocking process is greater at about 40%. Blocking of the HA site by Fe cations also decreases the rate of charge recombination between QA- and YZ•+ from t1/2 = 30 ms to 46ms. However, Ca2+ does not affect the rate during the blocking process. An Fe(II) cation (20µM) replaces 1Mn cation in the Mn4CaO5 catalytic cluster of PSII(-Ca) membranes at pH 5.7 but 2 Mn cations at pH 6.5. In the presence of Ca2+ (10mM) during the substitution process, Fe(II) is not able to extract Mn at pH 5.7 and extracts only 1Mn at pH 6.5 (instead of two without Ca2+). Measurements of fluorescence induction kinetics support these observations. Inhibition of Mn substitution with Fe(II) cations in the OEC only occurs with Ca2+ and Sr2+ cations, which are also able to restore oxygen evolution in PSII(-Ca) samples. Nonactive cations like La3+, Ni2+, Cd2+, and Mg2+ have no influence on the replacement of Mn with Fe. These results show that the location and/or ligand composition of one Mn cation in the Mn4CaO5 cluster is strongly affected by calcium depletion or rebinding and that bound calcium affects the redox potential of the extractable Mn4 cation in the OEC, making it resistant to reduction.

Photosystem II Functionality in Barley Responds Dynamically to Changes in Leaf Manganese Status.

A catalytic manganese (Mn) cluster is required for the oxidation of water in the oxygen-evolving complex (OEC) of photosystem II (PSII) in plants. Despite this essential role of Mn in generating the electrons driving photosynthesis, limited information is available on how Mn deficiency affects PSII functionality. We have here used parameters derived from measurements of fluorescence induction kinetics (OJIP transients), non-photochemical quenching (NPQ) and PSII subunit composition to investigate how latent Mn deficiency changes the photochemistry in two barley genotypes differing in Mn efficiency. Mn deficiency caused dramatic reductions in the quantum yield of PSII and led to the appearance of two new inflection points, the K step and the D dip, in the OJIP fluorescence transients, indicating severe damage to the OEC. In addition, Mn deficiency decreased the ability to induce NPQ in the light, rendering the plants incapable of dissipating excess energy in a controlled way. Thus, the Mn deficient plants became severely affected in their ability to recover from high light-induced photoinhibition, especially under strong Mn deficiency. Interestingly, the Mn-efficient genotype was able to maintain a higher NPQ than the Mn-inefficient genotype when exposed to mild Mn deficiency. However, during severe Mn deficiency, there were no differences between the two genotypes, suggesting a general loss of the ability to disassemble and repair PSII. The pronounced defects of PSII activity were supported by a dramatic decrease in the abundance of the OEC protein subunits, PsbP and PsbQ in response to Mn deficiency for both genotypes. We conclude that regulation of photosynthetic performance by means of maintaining and inducing NPQ mechanisms contribute to genotypic differences in the Mn efficiency of barley genotypes growing under conditions with mild Mn deficiency.

Laser-Induced Chlorophyll Fluorescence Measurement System to Assess Photosynthetic Status within Leaves

Chlorophyll (Chl) fluorescence measured on a leaf surface only provides the photosynthetic status of chloroplasts near the surface due to self-shading effect of Chl. Here, we report a laser-induced Chl fluorescence measurement system, which enables measurement of fluorescence induction kinetics at different tissues within a leaf as well as on both leaf surfaces, to assess photosynthetic status within leaves. The logarithmic time-scaled Chl fluorescence induction kinetics obtained from Chenopodium album leaves showed polyphasic transients in which four inflection points designated as O, J, I and P, were observed. Adaxial surface and palisade mesophyll showed significantly higher fluorescence intensities at O than abaxial surface and spongy mesophyll, respectively. In contrast, fluorescence intensities at J, I and P were significantly higher at abaxial surface and spongy mesophyll. Using these fluorescence intensities, the JIP test was performed. The results of JIP test indicated that adaxial surface and palisade mesophyll are characterized by lower maximum quantum yield of photosystem II (PSII) and net rate of PSII closure, but higher rate of energy transfer into electron transport chain than abaxial surface and spongy mesophyll. Considering these dorsiventral and intra-leaf variations in Chl fluorescence parameters, this study suggested the necessity of Chl fluorescence measurements on tissues as well as both surfaces to evaluate photosynthetic status of a whole leaf.

Elucidating the site of action of oxalate in photosynthetic electron transport chain in spinach thylakoid membranes

The effects of oxalate on PS II and PS I photochemistry were studied. The results suggested that in chloride-deficient thylakoid membranes, oxalate inhibited activity of PS II as well as PS I. To our knowledge, this is the only anion so far known which inhibits both the photosystems. Measurements of fluorescence induction kinetics, YZ* decay, and S2 state multiline EPR signal suggested that oxalate inhibited PS II at the donor side most likely on the oxygen evolving complex. Measurements of re-reduction of P700+ signal in isolated PS I particles in oxalate-treated samples suggested a binding site of oxalate on the donor, as well as the acceptor side of PS I.

Cl- channel inhibitors of the arylaminobenzoate type act as photosystem II herbicides: a functional and structural study.

The Cl- channel blocker NPPB (5-nitro-2-(3-phenylpropylamino) benzoic acid) inhibited photosynthetic oxygen evolution of isolated thylakoid membranes in a pH-dependent manner with a K(i) of about 2 microM at pH 6. Applying different electron acceptors, taking electrons either directly from photosystem II (PS II) or photosystem I (PS I), the site of inhibition was localized within PS II. Measurements of fluorescence induction kinetics and thermoluminescence suggest that the binding of NPPB to the QB binding site of PS II is similar to the herbicide DCMU (3-(3,4-dichlorophenyl)-1,1-dimethylurea). The effects of different arylaminobenzoate derivatives and other Cl- channel inhibitors on photosynthetic electron transport were investigated. The structure--activity relationship of the inhibitory effect on PS II shows interesting parallels to the one observed for the arylaminobenzoate block of mammalian Cl- channels. A molecular modeling approach was used to fit NPPB into the QB binding site and to identify possible molecular interactions between NPPB and the amino acid residues of the binding site in PS II. Taken together, these data give a detailed molecular picture of the mechanism of NPPB binding.

Effects of dehydration on the electron transport of Chlorella. An in vivo fluorescence study

Chlorella was used to study the effects of dehydration on photosynthetic activities. The use of unicellular green algae assured that the extent of dehydration was uniform throughout the whole cell population during the course of desiccation. Changes in the activities of the cells were monitored by measurements of fluorescence induction kinetics. It was found that inhibition of most of the photosynthetic activities started at a similar level of cellular water content. They included CO2 fixation, photochemical activity of Photosystem II and electron transport through Photosystem I. The blockage of electron flow through Photosystem I was complete and the whole transition occurred within a relative short time of dehydration. On the other hand, the suppression of Photosystem II activity was incomplete and the transition took a longer time of dehydration. Upon rehydration, the inhibition of Photosystem II activity was fully reversible when samples were in the middle of the transition, but was not thereafter. The electron transport through Photosystem I was also reversible during the transition, but was only partially afterward.

Chlorophyll Fluorescence as a Probe of the Photosynthetic Competence of Leaves in the Field: A Review of Current Instrumentation

Chlorophyll fluorescence has been widely used in laboratory studies in understanding both the mechanism of photosynthesis itself and the mechanisms by which a range of environmental factors alter photosynthetic capacity. The measurement of chlorophyll fluorescence is both non-destructive and non-invasive, and thus has considerable potential for use in the field situation. Applications range simply from a means of rapidly identifying injury to leaves in the absence of visible symptoms to a detailed analysis of causes of change in photosynthetic capacity. This paper introduces the topic of chlorophyll fluorescence, its interpretation and its application in field studies, giving particular attention to interpretation and measurement of fluorescence induction kinetics and to the application of recently developed modulated light fluorimeters. Three commercial fluorimeters designed for field applications were compared: (1) Plant Stress Meter, BioMonitor AB, Sweden; (2) MFMS, Hansatech Ltd, UK; (3) PAM 101, H. Walz, Federal Republic of Germany. The structure and potential of each are briefly reviewed. It was beyond the scope of this comparison to examine the full potential of each instrument but measurements of the widely used parameter Fv/Fmax were made with each on: (1) Triticum aestivum L. leaves treated with the herbicide Atrazine; and (2) Picea abies (L.) Karst. samples collected from sites which were known to receive different levels of ambient air pollution. In both experiments, the results obtained from the three fluorimeters showed good agreement. The relative merits of each instrument to field applications are discussed. Key-words: Chlorophyll fluorescence, stress detection, fluorimeters, Atrazine treatment, photosynthesis, air pollutants, photosystem II