Stable Radical Isoporphyrin Copolymer Prepared with Di(phenylphosphane).

Two diphosphanes with variable-length ligands tested as nucleophiles to prepare isoporphyrin copolymers in the presence of ditolylporphyrin of zinc (ZnT2P) prevented the oxidation of the diphosphine ligand. This paper demonstrates the power of this approach and describes the photoelectrocatalytic properties. The obtained copolymers were characterized by UV-vis spectroscopy, X-ray photoelectron spectroscopy, atomic force micrograph (AFM), EQCM (Electrochemical Quartz Cristal Microbalance) and electrochemistry. Their impedance properties (EIS) were studied and their photovoltaic performances were also investigated by photocurrent transient measurements under visible light irradiation.

Site-Selective Radical Aromatic C-H Functionalization of Alloxazine and Flavin through Ground-State Single Electron Transfer.

Flavins and their alloxazine isomers are key chemical scaffolds for bioinspired electron transfer strategies. Their properties can be fine-tuned by functional groups, which must be introduced at an early stage of the synthesis as their aromatic ring is inert towards post-functionalization. We show that the introduction of a remote metal-binding redox site on alloxazine and flavin activates their aromatic ring towards direct C-H functionalization. Mechanistic studies are consistent with a synthetic sequence involving ground-state single electron transfer (SET) with an electrophilic source followed by radical-radical coupling. This unprecedented reactivity opens new opportunities in molecular editing of flavins by direct aromatic post-functionalization and the utility of the method is demonstrated with the site-selective C6 functionalization of alloxazine and flavin with a CF3 group, Br or Cl, that can be further elaborated into OH and aryl for chemical diversification.

Alloxazine-Based Ligands Appended with Coordinating Groups: Synthesis, Electrochemical Studies, and Formation of Coordination Polymers.

Three new ligands based on the alloxazine core appended with pyridyl coordinating groups have been designed, synthesized, and characterized. The ligands are revealed to be redox-active in DMF solution, as attested to by CV and combined CV/EPR studies. The spin of the reduced species appears to be delocalized on the alloxazine core, as attested to by DFT calculations. The coordination abilities of one of the ligands toward Cu2+ or Ni2+ 3d cations revealed the formation of the first alloxazine-based 3D coordination polymers, presenting strong π-π stacking and substantial cavities. Preliminarily charge/discharge experiments in Li batteries evidence Li+ insertion in such systems.

Assessing the Extent of Potential Inversion by Cyclic Voltammetry: Theory, Pitfalls, and Application to a Nickel Complex with Redox-Active Iminosemiquinone Ligands.

Potential inversion refers to the situation where a protein cofactor or a synthetic molecule can be oxidized or reduced twice in a cooperative manner; that is, the second electron transfer is easier than the first. This property is very important regarding the catalytic mechanism of enzymes that bifurcate electrons and the properties of bidirectional redox molecular catalysts that function in either direction of the reaction with no overpotential. Cyclic voltammetry is the most common technique for characterizing the thermodynamics and kinetics of electron transfer to or from these molecules. However, a gap in the literature is the absence of analytical predictions to help interpret the values of the voltammetric peak potentials when potential inversion occurs; the cyclic voltammograms are therefore often analyzed by simulating the data, with no discussion of the possibility of overfitting and often no estimation of the error on the determined parameters. Here we formulate the theory for the voltammetry of freely diffusing or surface-confined two-electron redox species in the experimentally relevant irreversible limit where the peak separation depends on the scan rate. We explain why the model is intrinsically underdetermined, and we illustrate this conclusion by analysis of the voltammetry of a nickel complex with redox-active iminosemiquinone ligands. Being able to characterize the thermodynamics of two-electron electron-transfer reactions will be crucial for designing more efficient catalysts.

Mechanisms of Radical Formation on Chemically Modified Graphene Oxide under Near Infrared Irradiation.

In this work, the mechanisms of radical generation on different functionalized graphene oxide (GO) conjugates under near-infrared (NIR) light irradiation are investigated. The GO conjugates are designed to understand how chemical functionalization can influence the generation of radicals. Both pristine and functionalized GO are irradiated by a NIR laser, and the production of different reactive oxygen species (ROS) is investigated using fluorimetry and electron paramagnetic resonance to describe the type of radicals present on the surface of GO. The mechanism of ROS formation involves a charge transfer from the material to the oxygen present in the media, via the production of superoxide and singlet oxygen. Cytotoxicity and effects of ROS generation are then evaluated using breast cancer cells, evidencing a concentration dependent cell death associated to the heat and ROS. The study provides new hints to understand the photogeneration of radicals on the surface of GO upon near infrared irradiation, as well as, to assess the impact on these radicals in the context of a combined drug delivery system and phototherapeutic approach. These discoveries open the way for a better control of phototherapy-based treatments employing graphene-based materials.

Dual-Readout of the Mechanical Response of a Bis-acridinium [2]Rotaxane.

A [2]rotaxane built around a multi-responsive bis-acridinium macrocycle has been synthesized. Structural investigation has confirmed the interlocked nature of the molecule, and MD simulations illuminated its conformational dynamics with atomic resolution. Both halochromic and redox-switching properties were explored to shed light on the mechanical response and electronic changes that occur in the bis-acridinium [2]rotaxane. The topology of the rotaxane led to different mechanical behaviors upon addition of hydroxide ions or reduction that were easily detected by UV/Vis spectroscopy and electrochemistry.

A Single Bioinspired Hexameric Nickel Catechol-Alloxazine Catalyst Combines Metal and Radical Mechanisms for Alkene Hydrosilylation.

Mechanisms combining organic radicals and metallic intermediates hold strong potential in homogeneous catalysis. Such activation modes require careful optimization of two interconnected processes: one for the generation of radicals and one for their productive integration towards the final product. We report that a bioinspired polymetallic nickel complex can combine ligand- and metal-centered reactivities to perform fast hydrosilylation of alkenes under mild conditions through an unusual dual radical- and metal-based mechanism. This earth-abundant polymetallic complex incorporating a catechol-alloxazine motif as redox-active ligand operates at low catalyst loading (0.25 mol%) and generates silyl radicals and a nickel-hydride intermediate through a hydrogen atom transfer (HAT) step. Evidence of an isomerization sequence enabling terminal hydrosilylation of internal alkenes points towards the involvement of the nickel-hydride species in chain walking. This single catalyst promotes a hybrid pathway by combining synergistically ligand and metal participation in both inner- and outer- sphere processes.

Photoinduced Arylation of Acridinium Salts: Tunable Photoredox Catalysts for C-O Bond Cleavage.

A photoinduced arylation of N-substituted acridinium salts has been developed and has exhibited a high functional group tolerance (e.g., halogen, nitrile, ketone, ester, and nitro). A broad range of well-decorated C9-arylated acridinium-based catalysts with fine-tuned photophysical and photochemical properties, namely, excited-state lifetimes and redox potentials have been synthetized in a one-step procedure. These functionalized acridinium salts were later evaluated in the photoredox-catalyzed fragmentation of 1,2-diol derivatives (lignin models). Among them, 2-bromophenyl substituted N-methyl acridinium has outperformed all photoredox catalysts, including commercial Fukuzumi's catalyst, for the selective CßO-Ar bond cleavage of diol monoarylethers to afford 1,2-diols in good yields.

Design of a Targeting and Oxygen-Independent Platform to Improve Photodynamic Therapy: A Proof of Concept.

Photodynamic therapy (PDT) is a promising technique to treat different kinds of disease especially cancer. PDT requires three elements: molecular oxygen, a photoactivatable molecule called the photosensitizer (PS), and appropriate light. Under illumination, the PSs generate, in the presence of oxygen, the formation of reactive oxygen species including singlet oxygen, toxic, which then destroys the surrounding tissues. Even if PDT is used with success to treat actinic keratosis or prostate cancer for example, PDT suffers from two major drawbacks: the lack of selectivity of most of the PSs currently used clinically as well as the need for oxygen to be effective. To remedy the lack of selectivity, targeting the tumor neovessels is a promising approach to destroy the vascularization and cause asphyxia of the tumor. KDKPPR peptide affinity for the neuropilin-1 (NRP-1) receptor overexpressed on endothelial cells has already been proven. To compensate for the lack of oxygen, we focused on photoactivatable alkoxyamines (Alks), molecules capable of generating toxic radicals by light activation. In this article, we describe the synthesis of a multifunctional platform combining three units: a PS for an oxygen-dependent PDT, a peptide to target tumor neovessels, and an Alk for an oxygen-independent activity. The synthesis of the compound was successfully carried out, and the study of its photophysical properties showed that the PS retained its capacity to form singlet oxygen and the affinity tests confirmed the affinity of the compound for NRP-1. Thanks to the electron paramagnetic resonance spectroscopy, a technique of choice for radical investigation, the radicals generated by the illumination of the Alk could be detected. The proof of concept was thus successfully established.

Interdigitated conducting tetrathiafulvalene-based coordination networks.

Assembly of a novel ethylenedithio-tetrathiafulvalene (EDT-TTF) derivative bearing two adjacent 4-thiopyridyl groups with M(NCS)2 nodes (M = Fe, Co) leads to two isostructural 1D coordination polymers showing an enhancement of their electronic conductivity by six orders of magnitude (10-6vs. 10-12 S cm-1), upon surface oxidation by iodine and subsequent generation of EDT-TTF-based radicals.

A Highly Stable Organic Radical Cation.

Functionalization of a methylviologen with four methyl ester substituents significantly facilitates the first two reduction steps. The easily generated radical cation shows markedly improved air stability compared to the parent methylviologen, making this derivative of interest in organic electronic applications.

Using Hyperfine Electron Paramagnetic Resonance Spectroscopy to Define the Proton-Coupled Electron Transfer Reaction at Fe-S Cluster N2 in Respiratory Complex I.

Energy-transducing respiratory complex I (NADH:ubiquinone oxidoreductase) is one of the largest and most complicated enzymes in mammalian cells. Here, we used hyperfine electron paramagnetic resonance (EPR) spectroscopic methods, combined with site-directed mutagenesis, to determine the mechanism of a single proton-coupled electron transfer reaction at one of eight iron-sulfur clusters in complex I, [4Fe-4S] cluster N2. N2 is the terminal cluster of the enzyme's intramolecular electron-transfer chain and the electron donor to ubiquinone. Because of its position and pH-dependent reduction potential, N2 has long been considered a candidate for the elusive "energy-coupling" site in complex I at which energy generated by the redox reaction is used to initiate proton translocation. Here, we used hyperfine sublevel correlation (HYSCORE) spectroscopy, including relaxation-filtered hyperfine and single-matched resonance transfer (SMART) HYSCORE, to detect two weakly coupled exchangeable protons near N2. We assign the larger coupling with A(1H) = [-3.0, -3.0, 8.7] MHz to the exchangeable proton of a conserved histidine and conclude that the histidine is hydrogen-bonded to N2, tuning its reduction potential. The histidine protonation state responds to the cluster oxidation state, but the two are not coupled sufficiently strongly to catalyze a stoichiometric and efficient energy transduction reaction. We thus exclude cluster N2, despite its proton-coupled electron transfer chemistry, as the energy-coupling site in complex I. Our work demonstrates the capability of pulse EPR methods for providing detailed information on the properties of individual protons in even the most challenging of energy-converting enzymes.

Dimerization interface and dynamic properties of yeast IF1 revealed by Site-Directed Spin Labeling EPR spectroscopy.

The mitochondrial ATPase inhibitor, IF1, regulates the activity of the mitochondrial ATP synthase. The oligomeric state of IF1 related to pH is crucial for its inhibitory activity. Although extensive structural studies have been performed to characterize the oligomeric states of bovine IF1, only little is known concerning those of yeast IF1. While bovine IF1 can be found as an inhibitory dimer at low pH and a non-inhibitory tetramer at high pH, a monomer/dimer equilibrium has been described for yeast IF1, high pH values favoring the monomeric state. Combining different strategies involving the grafting of nitroxide spin labels combined with Electron Paramagnetic Resonance (EPR) spectroscopy, the present study brings the first structural characterization, at the residue level, of yeast IF1 in its dimeric form. The results show that the dimerization interface involves the central region of the peptide revealing that the dimer corresponds to a non-inhibitory state. Moreover, we demonstrate that the C-terminal region of the peptide is highly dynamic and that this segment is probably folded back onto the central region. Finally, the pH-dependence of the inter-label distance distribution has been observed indicating a conformational change between two structural states in the dimer.

Exploring intrinsically disordered proteins using site-directed spin labeling electron paramagnetic resonance spectroscopy.

Proteins are highly variable biological systems, not only in their structures but also in their dynamics. The most extreme example of dynamics is encountered within the family of Intrinsically Disordered Proteins (IDPs), which are proteins lacking a well-defined 3D structure under physiological conditions. Among the biophysical techniques well-suited to study such highly flexible proteins, Site-Directed Spin Labeling combined with EPR spectroscopy (SDSL-EPR) is one of the most powerful, being able to reveal, at the residue level, structural transitions such as folding events. SDSL-EPR is based on selective grafting of a paramagnetic label on the protein under study and is limited neither by the size nor by the complexity of the system. The objective of this mini-review is to describe the basic strategy of SDSL-EPR and to illustrate how it can be successfully applied to characterize the structural behavior of IDPs. Recent developments aimed at enlarging the panoply of SDSL-EPR approaches are presented in particular newly synthesized spin labels that allow the limitations of the classical ones to be overcome. The potentialities of these new spin labels will be demonstrated on different examples of IDPs.

Diversification of EPR signatures in Site Directed Spin Labeling using a ß-phosphorylated nitroxide.

Site Directed Spin Labeling (SDSL) combined with EPR spectroscopy is a very powerful approach to investigate structural transitions in proteins in particular flexible or even disordered ones. Conventional spin labels are based on nitroxide derivatives leading to classical 3-line spectra whose spectral shapes are indicative of the environment of the labels and thus constitute good reporters of structural modifications. However, the similarity of these spectral shapes precludes probing two regions of a protein or two partner proteins simultaneously. To overcome the limitation due to the weak diversity of nitroxide label EPR spectral shapes, we designed a new spin label based on a ß-phosphorylated nitroxide giving 6-line spectra. This paper describes the synthesis of this new spin label, its grafting at four different positions of a model disordered protein able to undergo an induced α-helical folding and its characterization by EPR spectroscopy. For comparative purposes, a classical nitroxide has been grafted at the same positions of the model protein. The ability of the new label to report on structural transitions was evaluated by analyzing the spectral shape modifications induced either by the presence of a secondary structure stabilizer (trifluoroethanol) or by the presence of a partner protein. Taken together the results demonstrate that the new phosphorylated label gives a very distinguishable signature which is able to report from subtle to larger structural transitions, as efficiently as the classical spin label. As a complementary approach, molecular dynamics (MD) calculations were performed to gain further insights into the binding process between the labeled NTAIL and PXD. MD calculations revealed that the new label does not disturb the interaction between the two partner proteins and reinforced the conclusion on its ability to probe different local environments in a protein. Taken together this study represents an important step forward in the extension of the panoply of SDSL-EPR approaches.

Dynamics of the intrinsically disordered protein CP12 in its association with GAPDH in the green alga Chlamydomonas reinhardtii: a fuzzy complex.

CP12 is a widespread regulatory protein of oxygenic photosynthetic organisms that contributes to the regulation of the Calvin cycle by forming a supra-molecular complex with at least two enzymes: glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoribulokinase (PRK). CP12 shares some similarities with intrinsically disordered proteins (IDPs) depending on its redox state. In this study, site-directed spin labeling (SDSL) combined with EPR spectroscopy was used to probe the dynamic behavior of CP12 from Chlamydomonas reinhardtii upon binding to GAPDH, the first step towards ternary complex formation. The two N-terminal cysteine residues were labeled using the classical approach while the tyrosine located at the C-terminal end of CP12 was modified following an original procedure. The results show that the label grafted at the C-terminal extremity is in the vicinity of the interaction site whereas the N-terminal region remains fully disordered upon binding to GAPDH. In conclusion, GAPDH-CP12 is a fuzzy complex, in which the N-terminal region of CP12 keeps a conformational freedom in the bound form. This fuzziness could be one of the keys to facilitate binding of PRK to CP12-GAPDH and to form the ternary supra-molecular complex.