Search Results (16)

Photosynthetic reaction centers (RCs) are membrane proteins converting photonic excitations into electric gradients. The heliobacterial RCs (HbRCs) are assumed to be the precursors of all known RCs, making them a compelling subject for investigating structural and functional relationships. A comprehensive picture of the electronic structure of the HbRCs is still missing. In this work, the combination of selective isotope labelling of ¹³C and ¹⁵N nuclei and the utilization of photo-CIDNP MAS NMR (photochemically induced dynamic nuclear polarization magic-angle spinning nuclear magnetic resonance) allows for highly enhanced signals from the radical-pair-forming cofactors. The remarkable magnetic-field dependence of the solid-state photo-CIDNP effect allows for observation of positive signals of the electron donor cofactor at 4.7 T, which is interpreted in terms of a dominant contribution of the differential relaxation (DR) mechanism. Conversely, at 9.4 T, the emissive signals mainly originate from the electron acceptor, due to the strong activation of the three-spin mixing (TSM) mechanism. Consequently, we have utilized two-dimensional homonuclear photo-CIDNP MAS NMR at both 4.7 T and 9.4 T. These findings from experimental investigations are corroborated by calculations based on density functional theory (DFT). This allows us to present a comprehensive investigation of the electronic structure of the cofactors involved in electron transfer (ET). Full article

5-Deazaflavins are analogs of naturally occurring flavin cofactors. They serve as substitutes for natural flavin cofactors to investigate and modify the reaction pathways of flavoproteins. Demethylated 5-deazaflavins are potential candidates for artificial cofactors, allowing us to fine-tune the reaction kinetics and absorption characteristics of flavoproteins. In this contribution, demethylated 5-deazariboflavin radicals are investigated (1) to assess the influence of the methyl groups on the electronic structure of the 5-deazaflavin radical and (2) to explore their photophysical properties with regard to their potential as artificial cofactors. We determined the proton hyperfine structure of demethylated 5-deazariboflavins using photochemically induced dynamic nuclear polarization (photo-CIDNP) spectroscopy, as well as density functional theory (DFT). To provide context, we compare our findings to a study of flavin mononucleotide (FMN) derivatives. We found a significant influence of the methylation pattern on the absorption properties, as well as on the proton hyperfine coupling ratios of the xylene moiety, which appears to be solvent-dependent. This effect is enhanced by the replacement of N5 by C5-H in 5-deazaflavin derivatives compared to their respective flavin counterparts. Full article

The time-resolved CIDNP method can provide information about degenerate exchange reactions (DEEs) involving short-lived radicals. In the temperature range from 8 to 65 °C, the DEE reactions of the guanosine-5′-monophosphate anion GMP(-H)⁻ with the neutral radical GMP(-H)^•, of the N-acetyl tyrosine anion N-AcTyrO⁻ with a neutral radical N-AcTyrO^•, and of the tyrosine anion TyrO⁻ with a neutral radical TyrO^• were studied. In all the studied cases, the radicals were formed in the reaction of quenching triplet 2,2′-dipyridyl. The reorganization energies were obtained from Arrhenius plots. The rate constant of the reductive electron transfer reaction in the pair GMP(-H)^•/TyrO⁻ was determined at T = 25 °C. Rate constants of the GMP(-H)^• radical reduction reactions with TyrO⁻ and N-AcTyrO⁻ anions calculated by the Marcus cross-relation differ from the experimental ones by two orders of magnitude. The rate constants of several other electron transfer reactions involving GMP(-H)⁻/GMP(-H)^•, N-AcTyrO⁻/N-AcTyrO^•, and TyrO⁻/TyrO^• pairs calculated by cross-relation agree well with the experimental values. The rate of nuclear paramagnetic relaxation was found for the 3,5 and β-protons of TyrO^• and N-AcTyrO^•, the 8-proton of GMP(-H)^•, and the 3,4-protons of DPH^• at each temperature. In all cases, the dependences of the rate of nuclear paramagnetic relaxation on temperature are described by the Arrhenius dependence. Full article

Schiff bases and similar molecules forming metal complexes may cause redox effects, which may also be influenced by light. Anthraquinones such as doxorubicin and idarubicin are widely used antitumor agents, which can generate reactive oxygen species (ROS), stimulated by both the presence of iron and copper ions and also by light. The generated ROS can cause DNA scission, cell membrane oxidation, and many other toxic effects. The redox activity of the quinone-quinoline chelator 2-phenyl-4-(butylamino)naphtho [2,3-h]quinoline-7,12-dione (Q1) was investigated in the presence of iron, copper, and zinc. The influence of light in these interactions was also examined. The chemically induced dynamic nuclear polarization (CIDNP), nuclear magnetic resonance (NMR), and electron paramagnetic resonance (EPR) methods were used to elucidate the molecular changes and ROS generation effects of the Q1 metal interactions. A model electron transfer reaction system between 1,4-dihydropyridine and Q1 was utilized to demonstrate that the chelate complexes of Q1 with both Fe(III) and Cu(II) ions were more redox active than Q1 itself. Similarly, CIDNP and NMR data showed that the concentration dependence of the free radicals yield is much higher in the presence of Fe(III) and Cu(II) ions, in comparison to Zn(II), and also that it increased in the presence of light. These findings underline the role of transition metal ions and Q1 in cyclic redox chain reactions and increase the prospect of the development of copper- and iron-based chelating agents, including Q1 and its derivatives, for anticancer therapy. Furthermore, these findings also signify the effect of light on enhancing ROS formation by Q1 and the prospect of utilizing such information for designing target specific anticancer drugs for photodynamic therapy. Full article

It is well-known that non-covalent interactions play an essential role in the functioning of biomolecules in living organisms. The significant attention of researchers is focused on the mechanisms of associates formation and the role of the chiral configuration of proteins, peptides, and amino acids in the association. We have recently demonstrated the unique sensitivity of chemically induced dynamic nuclear polarization (CIDNP) formed in photoinduced electron transfer (PET) in chiral donor–acceptor dyads to non-covalent interactions of its diastereomers in solutions. The present study further develops the approach for quantitatively analyzing the factors that determine the association by examples of dimerization of the diastereomers with the RS, SR, and SS optical configurations. It has been shown that, under the UV irradiation of dyads, CIDNP is formed in associates, namely, homodimers (SS-SS), (SR-SR), and heterodimers (SS-SR) of diastereomers. In particular, the efficiency of PET in homo-, heterodimers, and monomers of dyads completely determines the forms of dependences of the CIDNP enhancement coefficient ratio of SS and RS, SR configurations on the ratio of diastereomer concentrations. We expect that the use of such a correlation can be useful in identifying small-sized associates in peptides, which is still a problem. Full article

This short review reports the surprising phenomenon of nuclear hyperpolarization occurring in chemical reactions, which is called CIDNP (chemically induced dynamic nuclear polarization) or photo-CIDNP if the chemical reaction is light-driven. The phenomenon occurs in both liquid and solid-state, and electron transfer systems, often carrying flavins as electron acceptors, are involved. Here, we explain the physical and chemical properties of flavins, their occurrence in spin-correlated radical pairs (SCRP) and the possible involvement of flavin-carrying SCRPs in animal magneto-reception at earth’s magnetic field. Full article

Optical isomers of short peptide Lysine-Tryptophan-Lysine (Lys-{L/D-Trp}-Lys) and Lys-Trp-Lys with an acetate counter-ion were used to study photoinduced intramolecular and intermolecular processes of interest in photobiology. A comparison of L- and D-amino acid reactivity is also the focus of scientists’ attention in various specialties because today, the presence of amyloid proteins with D-amino acids in the human brain is considered one of the leading causes of Alzheimer’s disease. Since aggregated amyloids, mainly Aβ42, are highly disordered peptides that cannot be studied with traditional NMR and X-ray techniques, it is trending to explore the reasons for differences between L- and D-amino acids using short peptides, as in our article. Using NMR, chemically induced dynamic nuclear polarization (CIDNP) and fluorescence techniques allowed us to detect the influence of tryptophan (Trp) optical configuration on the peptides fluorescence quantum yields, bimolecular quenching rates of Trp excited state, and the photocleavage products formation. Thus, compared with the D-analog, the L-isomer shows a greater Trp excited state quenching efficiency with the electron transfer (ET) mechanism. There are experimental confirmations of the hypothesis about photoinduced ET between Trp and the CONH peptide bond, as well as between Trp and another amide group. Full article

Electron transfer plays a crucial role in ROS generation in living systems. Molecular oxygen acts as the terminal electron acceptor in the respiratory chains of aerobic organisms. Two main mechanisms of antioxidant defense by exogenous antioxidants are usually considered. The first is the inhibition of ROS generation, and the second is the trapping of free radicals. In the present study, we have elucidated both these mechanisms of antioxidant activity of glycyrrhizin (GL), the main active component of licorice root, using the chemically induced dynamic nuclear polarization (CIDNP) technique. First, it was shown that GL is capable of capturing a solvated electron, thereby preventing its capture by molecular oxygen. Second, we studied the effect of glycyrrhizin on the behavior of free radicals generated by UV irradiation of xenobiotic, NSAID—naproxen in solution. The structure of the glycyrrhizin paramagnetic intermediates formed after the capture of a solvated electron was established from a photo-CIDNP study of the model system—the dianion of 5-sulfosalicylic acid and DFT calculations. Full article

The mechanisms of stereoselectivity of the interaction of chiral drugs with active sites of enzymes and cell receptors attract significant attention. The first reason is the difference in therapeutic activity of the enantiomers of the common drugs. Another reason is the interest in the role of chiral inversion of amino acids involved in various peptides in the development of many diseases including Alzheimer’s, Parkinson’s, type II diabetes, and a number of other pathological conditions. In our study we use elementary chemical process—electron transfer (ET) to simulate individual stages of ligand–receptor and enzyme–substrate interactions. In particular, previous studies of photoinduced ET in chiral donor-acceptor dyads consisting of the nonsteroidal anti-inflammatory drug (R/S)-ketoprofen and (L)-tryptophan show the stereo and spin selectivity of ET in diastereomers. The present study is devoted to the interaction of (S)-ketoprofen with L- and D-enantiomers of tryptophan in homogeneous aqueous solution and in phospholipid membranes. The study was done using the NMR technique and molecular modeling. These approaches confirm efficient penetration of ketoprofen into the lipid bilayer and binding with tryptophan molecule. The short-lived paramagnetic intermediates formed during the photoinduced ET from electron donor tryptophan to ketoprofen have been detected using the chemically induced dynamic nuclear polarization (CIDNP) technique. It was found that S-ketoprofen interacts stereoselectively with tryptophan enantiomers in the lipid membrane. The formation of the ketyl radical of ketoprofen under irradiation leads to the oxidation of membrane lipids and may be the cause of ketoprofen phototoxicity. However, in contrast to a homogeneous solution in phosphate buffer saline, where the amino acid tryptophan accelerates the photodecomposition of KP due to intramolecular hydrogen transfer, tryptophan in a lipid membrane significantly reduces the rate of photodegradation due to a reversible electron (or hydrogen) transfer reaction. The stereoselectivity in the rate of KP and lipids decomposition under UV irradiation of S-ketoprofen in the presence of tryptophan enantiomers in lipid bilayer has been detected. Full article

The damage of cell membranes induced by photosensitive drugs has attracted the significant attention of researchers in various fields of medicine. Ketoprofen (KP) is known to be the most photosensitive among the nonsteroidal anti-inflammatory drugs. The phototoxic side effects of KP and other non-steroidal anti-inflammatory drugs are associated with the action of free radicals, but there is insufficient information about the nature of these radicals. In the present study, free radicals formed upon KP irradiation within lipid membranes were studied using nuclear magnetic resonance (NMR) and chemically induced dynamic nuclear polarization (CIDNP) methods, as well as a molecular dynamics simulation. Our study confirmed the effective penetration of KP into the lipid bilayer and showed a significant effect of the nature of the medium on the photolysis mechanism. While, in a homogeneous solution, the main channel of KP photolysis is free radical-mediated monomolecular decomposition with formation of radical pairs of benzyl and CO₂H^● radicals, then, in the lipid membrane, the reaction route shifts towards the bimolecular reaction of KP photoreduction. In addition, the effect of the presence an electron donor (the amino acid tryptophan) on lipid oxidation has been studied. It was found that photoreaction of KP with tryptophan proceeds more efficiently than with lipid molecules. Full article

Oxidation of methionine (Met) is an important reaction that plays a key role in protein modifications during oxidative stress and aging. The first steps of Met oxidation involve the creation of very reactive and short-lived transients. Application of complementary time-resolved radiation and photochemical techniques (pulse radiolysis and laser flash photolysis together with time-resolved CIDNP and ESR techniques) allowed comparing in detail the one-electron oxidation mechanisms initiated either by ^●OH radicals and other one-electron oxidants or the excited triplet state of the sensitizers e.g., 4-,3-carboxybenzophenones. The main purpose of this review is to present various factors that influence the character of the forming intermediates. They are divided into two parts: those inextricably related to the structures of molecules containing Met and those related to external factors. The former include (i) the protection of terminal amine and carboxyl groups, (ii) the location of Met in the peptide molecule, (iii) the character of neighboring amino acid other than Met, (iv) the character of the peptide chain (open vs cyclic), (v) the number of Met residues in peptide and protein, and (vi) the optical isomerism of Met residues. External factors include the type of the oxidant, pH, and concentration of Met-containing compounds in the reaction environment. Particular attention is given to the neighboring group participation, which is an essential parameter controlling one-electron oxidation of Met. Mechanistic aspects of oxidation processes by various one-electron oxidants in various structural and pH environments are summarized and discussed. The importance of these studies for understanding oxidation of Met in real biological systems is also addressed. Full article

The peculiarities of spin effects in photoinduced electron transfer (ET) in diastereomers of donor-acceptor dyads are considered in order to study the influence of chirality on reactivity. Thus, the spin selectivity—the difference between the enhancement coefficients of chemically induced dynamic nuclear polarization (CIDNP)—of the dyad’s diastereomers reflects the difference in the spin density distribution in its paramagnetic precursors that appears upon UV irradiation. In addition, the CIDNP coefficient itself has demonstrated a high sensitivity to the change of chiral centers: when one center is changed, the hyperpolarization of all polarized nuclei of the molecule is affected. The article analyzes the experimental values of spin selectivity based on CIDNP calculations and molecular dynamic modeling data in order to reveal the effect of optical configuration on the structure and reactivity of diastereomers. In this way, we succeeded in tracing the differences in dyads with L- and D-tryptophan as an electron donor. Since the replacement of L-amino acid with D-analog in specific proteins is believed to be the cause of Alzheimer’s and Parkinson’s diseases, spin effects and molecular dynamic simulation in model dyads can be a useful tool for investigating the nature of this phenomenon. Full article

A choline-binding module from pneumococcal LytA autolysin, LytA_239–252, was reported to have a highly stable nativelike β-hairpin in aqueous solution, which turns into a stable amphipathic α-helix in the presence of micelles. Here, we aim to obtain insights into this DPC-micelle triggered β-hairpin-to-α-helix conformational transition using photo-CIDNP NMR experiments. Our results illustrate the dependency between photo-CIDNP phenomena and the light intensity in the sample volume, showing that the use of smaller-diameter (2.5 mm) NMR tubes instead of the conventional 5 mm ones enables more efficient illumination for our laser-diode light setup. Photo-CIDNP experiments reveal different solvent accessibility for the two tyrosine residues, Y249 and Y250, the latter being less accessible to the solvent. The cross-polarization effects of these two tyrosine residues of LytA_239–252 allow for deeper insights and evidence their different behavior, showing that the Y250 aromatic side chain is involved in a stronger interaction with DPC micelles than Y249 is. These results can be interpreted in terms of the DPC micelle disrupting the aromatic stacking between W241 and Y250 present in the nativelike β-hairpin, hence initiating conversion towards the α-helix structure. Our photo-CIDNP methodology represents a powerful tool for observing residue-level information in switch peptides that is difficult to obtain by other spectroscopic techniques. Full article

The study of the L- and D-amino acid properties in proteins and peptides has attracted considerable attention in recent years, as the replacement of even one L-amino acid by its D-analogue due to aging of the body is resulted in a number of pathological conditions, including Alzheimer’s and Parkinson’s diseases. A recent trend is using short model systems to study the peculiarities of proteins with D-amino acids. In this report, the comparison of the excited states quenching of L- and D-tryptophan (Trp) in a model donor–acceptor dyad with (R)- and (S)-ketoprofen (KP-Trp) was carried out by photochemically induced dynamic nuclear polarization (CIDNP) and fluorescence spectroscopy. Quenching of the Trp excited states, which occurs via two mechanisms: prevailing resonance energy transfer (RET) and electron transfer (ET), indeed demonstrates some peculiarities for all three studied configurations of the dyad: (R,S)-, (S,R)-, and (S,S)-. Thus, the ET efficiency is identical for (S,R)- and (R,S)-enantiomers, while RET differs by 1.6 times. For (S,S)-, the CIDNP coefficient is almost an order of magnitude greater than for (R,S)- and (S,R)-. To understand the source of this difference, hyperpolarization of (S,S)-and (R,S)- has been calculated using theory involving the electron dipole–dipole interaction in the secular equation. Full article

Photoinduced elementary processes in chiral linked systems, consisting of drugs and tryptophan (Trp) residues, attract considerable attention due to several aspects. First of all, these are models that allow one to trace the full and partial charge transfer underlying the binding of drugs to enzymes and receptors. On the other hand, Trp fluorescence is widely used to establish the structure and conformational mobility of proteins due to its high sensitivity to the microenvironment. Therefore, the study of mechanisms of Trp fluorescence quenching in various systems has both fundamental and practical interest. An analysis of the photo-chemically induced dynamic nuclear polarization (CIDNP) and Trp fluorescence quenching in (R/S)-ketoprofen-(S)-tryptophan ((S/R)-KP-(S)-Trp) dyad carried out in this work allowed us to trace the intramolecular reversible electron transfer (ET) and obtain evidence in favor of the resonance energy transfer (RET). The fraction of dyad’s singlet excited state, quenched via ET, was shown to be 7.5 times greater for the (S,S)-diastereomer than for the (R,S) analog. At the same time, the ratio of the fluorescence quantum yields shows that quenching effectiveness of (S,S)-diastereomer to be 5.4 times lower than for the (R,S) analog. It means that the main mechanism of Trp fluorescence quenching in (S/R)-KP-(S)-Trp dyad is RET. Full article