Studying Lipid–Protein Interactions with Electron Paramagnetic Resonance Spectroscopy of Spin-Labeled Lipids

Páli, Tibor; Kóta, Zoltán

doi:10.1007/978-1-4939-9512-7_22

Cited by 6 publications

(5 citation statements)

References 137 publications

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“…In contrast to 4-oxo-TEMP, the EPR spectrum of the spin label 5-SASL evidenced its incorporation into the lipid phase (Kóta et al, 2002;Páli and Kóta, 2019) which is the site where ROS are formed (we observe negligible aqueous 5-SASL signal, with sharp lines). In principle, the loss of 5-SASL EPR intensity over time should reflect the ROS produced during irradiation of the samples -0.06 and 0.11 mol/mol Chl for LHCII in detergent and membranes, respectively, after 30 min.…”

Section: Photobleaching Is Caused By Singlet Oxygen Produced By Chl Tripletsmentioning

confidence: 57%

“…Stearic acid spin labels, such as 5-SASL, partition between the membrane and the aqueous buffer with very high preference toward membranes. The spectra of thylakoid lipid vesicles, LHCII proteoliposomes as well as of detergent-solubilized LHCII all showed features typical for 5-SASL in membrane phase (see, e.g., Kóta et al, 2002;Páli and Kóta, 2019), whereas the aqueous-phase EPR signature was less than 1% and could be neglected. 5-SASL can react with various radicals thereby losing its EPR signal, via either one electron oxidation or reduction.…”

Section: Electron Paramagnetic Resonancementioning

confidence: 99%

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Photobleaching of Chlorophyll in Light-Harvesting Complex II Increases in Lipid Environment

et al. 2020

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Excess light causes damage to the photosynthetic apparatus of plants and algae primarily via reactive oxygen species. Singlet oxygen can be formed by interaction of chlorophyll (Chl) triplet states, especially in the Photosystem II reaction center, with oxygen. Whether Chls in the light-harvesting antenna complexes play direct role in oxidative photodamage is less clear. In this work, light-induced photobleaching of Chls in the major trimeric light-harvesting complex II (LHCII) is investigated in different molecular environments – protein aggregates, embedded in detergent micelles or in reconstituted membranes (proteoliposomes). The effects of intense light treatment were analyzed by absorption and circular dichroism spectroscopy, steady-state and time-resolved fluorescence and EPR spectroscopy. The rate and quantum yield of photobleaching was estimated from the light-induced Chl absorption changes. Photobleaching occurred mainly in Chl a and was accompanied by strong fluorescence quenching of the remaining unbleached Chls. The rate of photobleaching increased by 140% when LHCII was embedded in lipid membranes, compared to detergent-solubilized LHCII. Removing oxygen from the medium or adding antioxidants largely suppressed the bleaching, confirming its oxidative mechanism. Singlet oxygen formation was monitored by EPR spectroscopy using spin traps and spin labels to detect singlet oxygen directly and indirectly, respectively. The quantum yield of Chl a photobleaching in membranes and detergent was found to be 3.4 × 10 –5 and 1.4 × 10 –5 , respectively. These values compare well with the yields of ROS production estimated from spin-trap EPR spectroscopy (around 4 × 10 –5 and 2 × 10 –5 ). A kinetic model is proposed, quantifying the generation of Chl and carotenoid triplet states and singlet oxygen. The high quantum yield of photobleaching, especially in the lipid membrane, suggest that direct photodamage of the antenna occurs with rates relevant to photoinhibition in vivo . The results represent further evidence that the molecular environment of LHCII has profound impact on its functional characteristics, including, among others, the susceptibility to photodamage.

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Section: Photobleaching Is Caused By Singlet Oxygen Produced By Chl Tripletsmentioning

confidence: 57%

Section: Electron Paramagnetic Resonancementioning

confidence: 99%

Photobleaching of Chlorophyll in Light-Harvesting Complex II Increases in Lipid Environment

et al. 2020

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“…Therefore, by using diluted samples to change the contribution of the five-peak component (not shown), the three components could be separated: We first obtained the pure five-peak component from the series of spectra of MTLS at different label concentrations. Then, the five-peak component was subtracted from the composite spectra, leaving the normal mobile plus immobile components, which were then treated with the routine technique for the two components [59]. Although we see the immobile component as observed earlier [31], it is almost undetectable; hence, the outer splittings could not be determined.…”

Section: Spin Labelling With Mtslmentioning

confidence: 77%

“…The protein-containing samples are qualitatively similar to those in previous EPR reports on spin-labelled fatty acids in the presence of serum albumins [20,27,28,30,33], showing composite spectra with different contributions from a mobile and an immobile component. Since the mobile component was identical with that of 5-SASL in water, optimised subtraction allowed us to determine the shape and relative contribution of the spectrum from 5-SASL bound to BSA, and also the mobile component (see [59] for a detailed description of the technique). The spectra of BSA and BSA+TiO 2 have almost only the immobile component, with only a few percentages of the mobile component.…”

Section: Spin Labelling With 5-saslmentioning

confidence: 99%

Spin-Label Electron Paramagnetic Resonance Spectroscopy Reveals Effects of Wastewater Filter Membrane Coated with Titanium Dioxide Nanoparticles on Bovine Serum Albumin

Sebők-Nagy,

Kóta,

Kincses

et al. 2023

Molecules

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The accumulation of proteins in filter membranes limits the efficiency of filtering technologies for cleaning wastewater. Efforts are ongoing to coat commercial filters with different materials (such as titanium dioxide, TiO2) to reduce the fouling of the membrane. Beyond monitoring the desired effect of the retention of biomolecules, it is necessary to understand what the biophysical changes are in water-soluble proteins caused by their interaction with the new coated filter membranes, an aspect that has received little attention so far. Using spin-label electron paramagnetic resonance (EPR), aided with native fluorescence spectroscopy and dynamic light scattering (DLS), here, we report the changes in the structure and dynamics of bovine serum albumin (BSA) exposed to TiO2 (P25) nanoparticles or passing through commercial polyvinylidene fluoride (PVDF) membranes coated with the same nanoparticles. We have found that the filtering process and prolonged exposure to TiO2 nanoparticles had significant effects on different regions of BSA, and denaturation of the protein was not observed, neither with the TiO2 nanoparticles nor when passing through the TiO2-coated filter membranes.

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“…3D ) 83 and endogenous cyclic hydroxylamines (CHAs). Usefully, as the SASL probe is lipophilic, 84 it can easily attach to the lipid phase of membrane vesicles, which is the major site of free radical production in cells. This has been successfully exploited to track total radical production in reconstituted membranes of light harvesting complex II from pea, showing spin decay in the lipid phase following irradiation, but not in the aqueous phase.…”

Section: Direct Measurement Of Ros Species Using Chemical Probes Biosensors and Electron Paramagnetic Resonancementioning

confidence: 99%

Measuring ROS and redox markers in plant cells

et al. 2021

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Studying Lipid–Protein Interactions with Electron Paramagnetic Resonance Spectroscopy of Spin-Labeled Lipids

Cited by 6 publications

References 137 publications

Photobleaching of Chlorophyll in Light-Harvesting Complex II Increases in Lipid Environment

Photobleaching of Chlorophyll in Light-Harvesting Complex II Increases in Lipid Environment

Spin-Label Electron Paramagnetic Resonance Spectroscopy Reveals Effects of Wastewater Filter Membrane Coated with Titanium Dioxide Nanoparticles on Bovine Serum Albumin

Measuring ROS and redox markers in plant cells

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