ABSTRACT
The spectral-kinetic characteristics of the fluorescence of the tryptophan molecule in an aqueous solution and in the composition of a protein (albumin) were studied in the temperature range from -170 to 25°C. To explain the observed changes in the spectra and the tryptophan fluorescence lifetime with temperature, a model of transitions between the excited and ground states involving a charge-transfer state was used, which takes into account the nonlinear nature of the dynamics of these transitions. In these processes, an important role is played by the interaction of tryptophan molecules with its microenvironment, as well as rearrangements in the system of hydrogen bonds of the water-protein matrix surrounding the tryptophan molecule.
Subject(s)
Serum Albumin, Bovine/chemistry , Tryptophan/chemistry , Water/chemistry , Animals , Cattle , Fluorescence , Hydrogen Bonding , Kinetics , Serum Albumin, Bovine/metabolism , Spectrometry, Fluorescence , Temperature , Tryptophan/metabolism , Water/metabolismABSTRACT
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 50 mM) inhibit the light-dependent oxidation of Fe(II) (5 µM) 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 YZâ¢+ from t1/2 = 30 ms to 46 ms. However, Ca2+ does not affect the rate during the blocking process. An Fe(II) cation (20 µM) 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+ (10 mM) 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.
Subject(s)
Calcium/metabolism , Ferrous Compounds/metabolism , Manganese/metabolism , Oxygen/metabolism , Photosystem II Protein Complex/metabolism , Spinacia oleracea/physiology , Binding Sites , Cations/metabolism , Fluorescence , Kinetics , Oxidation-Reduction , PhotochemistryABSTRACT
Enhancement of optical properties of photosensitizers by additional light-harvesting antennas is promising for the improvement of the photodynamic therapy. However, large number of parameters determine interactions of nanoparticles and photosensitizers in complex and, thus the photodynamic efficacy of the hybrid structure. In order to achieve high efficiency of energetic coupling and photodynamic activity of such complexes it is important to know the location of the photosensitizer molecule on the nanoparticle, because it affects the spectral properties of the photosensitizer and the stability of the hybrid complex in vitro/in vivo. In this work complexes of polycationic aluminum phthalocyanines and CdSe/ZnS quantum dots were obtained. We used quantum dots which outer shell consists of polymer with carboxyl groups and provides water solubility and the negative charge of the nanoparticle. We found that phthalocyanine molecules could penetrate deeply into the polymer shell of quantum dot, leading thereby to significant changes in the spectral and photodynamic properties of phthalocyanines. We also showed that noncovalent interactions between phthalocyanine and quantum dot provide possibility for a release of the phthalocyanine from the hybrid complex and its binding to both Gram-positive and Gram-negative bacterial cells. Also, detailed characterization of the nanoparticle core and shell sizes was carried out.
Subject(s)
Drug Carriers/chemistry , Indoles/chemistry , Organometallic Compounds/chemistry , Quantum Dots/chemistry , Cadmium Compounds/chemistry , Fluorescence Resonance Energy Transfer , Gram-Negative Bacteria/drug effects , Gram-Positive Bacteria/drug effects , Indoles/pharmacology , Microscopy, Confocal , Microscopy, Electron, Transmission , Organometallic Compounds/pharmacology , Selenium Compounds/chemistry , Singlet Oxygen/chemistry , Singlet Oxygen/metabolism , Sulfides/chemistry , Zinc Compounds/chemistryABSTRACT
The temperature dependences of tryptophan fluorescence decay kinetics in aqueous glycerol and 1 M trehalose solutions were examined. The fluorescence decay kinetics were recorded in the spectral region of 292.5-417.5 nm with nanosecond time resolution. The kinetics curves were approximated by the sum of three exponential terms, and the spectral distribution (DAS) of these components was determined. An antisymbatic course of fluorescence decay times of two (fast and medium) components in the temperature range from -60 to +10°C was observed. The third (slow) component showed only slight temperature dependence. The antisymbatic behavior of fluorescence lifetimes of the fast and medium components was explained on the assumption that some of the excited tryptophan molecules are transferred from a short-wavelength B-form with short fluorescence lifetime to a long-wavelength R-form with an intermediate fluorescence lifetime. This transfer occurred in the indicated temperature range.
Subject(s)
Fluorescence , Temperature , Tryptophan/chemistry , Glycerol/chemistry , Half-Life , Kinetics , Solutions , Spectrometry, Fluorescence , Trehalose/chemistry , WaterABSTRACT
The technique for recording of chlorophyll a fluorescence induction kinetics has been used for assessment of the physiological state of leaves of tillet (Tilia cordata), pendent white birch (Betula pendula), American arborvitae (Thuja occidentalis) in urban environments. Different sensitivity of plants to adverse growing conditions was observed. The most sensitive JIP test parameters such as PI(ABS), F(V)/F(0), F(V)/F(M), R(fd) are determined to be used as indicators of the physiological state of urban phytocenosis. Recommendations for the application of this technique in monitoring studies are given.
Subject(s)
Betula/physiology , Chlorophyll/chemistry , Photosynthesis , Plant Physiological Phenomena , Chlorophyll A , Environmental Monitoring , Fluorescence , Humans , Plant Leaves/physiology , Spectrometry, FluorescenceABSTRACT
Topographic studies of crystalline fractions from different morphological layers of the young adult bovine lens were conducted. Crystallin profiles were obtained for each lens layer, using thin-layer isoelectric focusing in polyacrylamide gel (IEF). Water soluble (WS) crystallins from the lens equator revealed a separation into HM (high molecular weight) alpha(L)-, beta(H)-, beta(L)-, beta(S)-, and gamma-crystallins. The nature of the water insoluble (WI) protein fraction in the separated lens layers reflected the aggregated state of alpha(L)-, beta(L)-, beta(S)-, and gamma-crystallins in different regions of the lens, concealed in the central cavity of the alpha-crystallin chaperone model. The IEF data demonstrate a possible chaperone-like function for alpha-crystallin in the nucleus and inner cortex of the lens, but not in the outer cortex. The water binding properties of bovine lens alpha-crystallin, calf skin collagen, and bovine serum albumin (BSA) were investigated with various techniques. The water adsorptive capacity was obtained in high vacuum desorption experiments volumetrically, and also gravimetrically in controlled atmosphere experiments. The NMR spin-echo technique was used to study the hydration of protein samples and to determine the spin-spin relaxation times (T(2)) from the protons of water adsorbed on the proteins. Isolated bovine lenses were sectioned into 11-12 morphological layers (from anterior cortex through nucleus to posterior cortex). The water content in relation to dry weight of proteins was measured in individual morphological lens layers. During water vapor uptake at relative humidity P/P(0) = 0.75, alpha-crystallin did not adsorb water suggesting that hydrophobic regions of the protein are exposed to the aqueous solvent. At relative humidity P/P(0) = 1.0, the adsorption of water by alpha-crystallin was 17% with a single component decay character of spin echo (T(2) = 3 msec). Addition of water to alpha-crystallin to about 50% of its weight/weight in the protein sample showed T(2) = 8 msec with only one single component decay of the spin-echo signal. The single component decay character of the spin echo indicates water tightly bound by alpha-crystallin. Under a relative humidity P/P(0) = 1.0, collagen and BSA adsorbed, correspondingly, 19.3 and 28% of water and showed a two-component decay curve with T(2) about 5 and 40 msec. The findings demonstrate the presence of two water fractions in collagen and BSA which are separated in space. The IEF data suggest a tight binding of water with alpha-crystallin with similar distribution patterns in the lens layers. To conclude, it was found that alpha-crystallin can immobilize water to a greater extent than other proteins such as collagen and BSA. These results shed new light on structural properties of alpha-crystallin and its superhydration properties and have important implications for understanding the mechanism of the chaperone-like action of this protein in the lens and non-ocular tissues.