Radical-Induced Changes in Transition Metal Interfacial Magnetic Properties: A Blatter Derivative on Polycrystalline Cobalt.

In this work, we study the interface obtained by depositing a monolayer of a Blatter radical derivative on polycrystalline cobalt. By examining the occupied and unoccupied states at the interface, using soft X-ray techniques, combined with electronic structure calculations, we could simultaneously determine the electronic structure of both the molecular and ferromagnetic sides of the interface, thus obtaining a full understanding of the interfacial magnetic properties. We found that the molecule is strongly hybridized with the surface. Changes in the core level spectra reflect the modification of the molecule and the cobalt electronic structures inducing a decrease in the magnetic moment of the cobalt atoms bonded to the molecules which, in turn, lose their radical character. Our method allowed us to screen, beforehand, organic/ferromagnetic interfaces given their potential applications in spintronics.

Chiral π-Conjugated Double Helical Aminyl Diradical with the Triplet Ground State.

We report a neutral high-spin diradical of chiral C2-symmetric bis[5]diazahelicene with ΔEST ≈ 0.4 kcal mol-1, as determined by EPR spectroscopy/SQUID magnetometry. The diradical is the most persistent among all high-spin aminyl radicals reported to date by a factor of 20, with a half-life of up to 6 days in 2-MeTHF at room temperature. Its triplet ground state and excellent persistence may be associated with the unique spin density distribution within the dihydrophenazine moiety, which characterizes two effective 3-electron C-N bonds analogous to the N-O bond of a nitroxide radical. The enantiomerically enriched (ee ≥ 94%) (MM)- and (PP)-enantiomers of the precursors to the diradicals are obtained by either preparative chiral supercritical fluid chromatography or resolution via functionalization with the chiral auxiliary of the C2-symmetric racemic tetraamine. The barrier for the racemization of the solid tetraamine is ΔG‡ = 43 ± 0.01 kcal mol-1 in the 483-523 K range. The experimentally estimated lower limit of the barrier for the racemization of a diradical, ΔG‡ ≥ 26 kcal mol-1 in 2-MeTHF at 293 K, is comparable to the DFT-determined barrier of ΔG‡ = 31 kcal mol-1 in the gas phase at 298 K. While the enantiomerically pure tetraamine displays strong chiroptical properties, with anisotropy factor |g| = |Δε|/ε = 0.036 at 376 nm, |g| ≈ 0.005 at 548 nm of the high-spin diradical is comparable to that recently reported triplet ground-state diradical dication. Notably, the radical anion intermediate in the generation of diradical exhibits a large SOMO-HOMO inversion, SHI = 35 kcal mol-1.

Cucurbit[7]uril Enhances Distance Measurements of Spin-Labeled Proteins.

We report complex formation between the chloroacetamide 2,6-diazaadamantane nitroxide radical (ClA-DZD) and cucurbit[7]uril (CB-7), for which the association constant in water, Ka = 1.9 × 106 M-1, is at least 1 order of magnitude higher than the previously studied organic radicals. The radical is highly immobilized by CB-7, as indicated by the increase in the rotational correlation time, τrot, by a factor of 36, relative to that in the buffer solution. The X-ray structure of ClA-DZD@CB-7 shows the encapsulated DZD guest inside the undistorted CB-7 host, with the pendant group protruding outside. Upon addition of CB-7 to T4 Lysozyme (T4L) doubly spin-labeled with the iodoacetamide derivative of DZD, we observe the increase in τrot and electron spin coherence time, Tm, along with the narrowing of interspin distance distributions. Sensitivity of the DEER measurements at 83 K increases by a factor 4-9, compared to the common spin label such as MTSL, which is not affected by CB-7. Interspin distances of 3 nm could be reliably measured in water/glycerol up to temperatures near the glass transition/melting temperature of the matrix at 200 K, thus bringing us closer to the goal of supramolecular recognition-enabled long-distance DEER measurements at near physiological temperatures. The X-ray structure of DZD-T4L 65 at 1.12 Å resolution allows for unambiguous modeling of the DZD label (0.88 occupancy), indicating an undisturbed structure and conformation of the protein.

From Stable Radicals to Thermally Robust High-Spin Diradicals and Triradicals.

Stable radicals and thermally robust high-spin di- and triradicals have emerged as important organic materials due to their promising applications in diverse fields. New fundamental properties, such as SOMO/HOMO inversion of orbital energies, are explored for the design of new stable radicals, including highly luminescent ones with good photostability. A relation with the singlet-triplet energy gap in the corresponding diradicals is proposed. Thermally robust high-spin di- and triradicals, with energy gaps that are comparable to or greater than a thermal energy at room temperature, are more challenging to synthesize but more rewarding. We summarize a number of high-spin di- and triradicals, based on nitronyl nitroxides that provide a relation between the experimental pairwise exchange coupling constant J/k in the high-spin species vs experimental hyperfine coupling constants in the corresponding monoradicals. This relation allows us to identify outliers, which may correspond to radicals where J/k is not measured with sufficient accuracy. Double helical high-spin diradicals, in which spin density is delocalized over the chiral π-system, have been barely explored, with the sole example of such high-spin diradical possessing alternant π-system with Kekulé resonance form. Finally, we discuss a high-spin diradical with electrical conductivity and derivatives of triangulene diradicals.

Cucurbit[7]uril Enhances Distance Measurements of Spin-Labeled Proteins.

We report complex formation between the chloroacetamide 2,6-diazaadamantane nitroxide radical (ClA-DZD) and cucurbit[7]uril (CB-7), for which the association constant in water, Ka = 1.9 × 106 M-1, is at least one order of magnitude higher than the previously studied organic radicals. The radical is highly immobilized by CB-7, as indicated by the increase of the rotational correlation time, τrot, by a factor of 36, relative to that in the buffer solution. The X-ray structure of ClA-DZD@CB-7 shows the encapsulated DZD guest inside the undistorted CB-7 host, with the pendant group protruding outside. Upon addition of CB-7 to T4 Lysozyme (T4L) doubly spin-labeled with the iodoacetamide derivative of DZD, we observe the increase in τrot and electron spin coherence time, Tm, along with the narrowing of inter-spin distance distributions. Sensitivity of the DEER measurements at 83 K increases by a factor 4 - 9, compared to the common spin label such as MTSL, which is not affected by CB-7. Inter-spin distances of 3-nm could be reliably measured in water/glycerol up to temperatures near the glass transition/melting temperature of the matrix at 200 K, thus bringing us closer to the goal of supramolecular recognition-enabled long-distance DEER measurements at near physiological temperatures. The X-ray structure of DZD-T4L 65 at 1.12 Å resolution allows for unambiguous modeling of the DZD label (0.88 occupancy), indicating undisturbed structure and conformation of the protein.

Long-Term Degradation Mechanisms in Application-Implemented Radical Thin Films.

Blatter radical derivatives are very attractive due to their potential applications, ranging from batteries to quantum technologies. In this work, we focus on the latest insights regarding the fundamental mechanisms of radical thin film (long-term) degradation, by comparing two Blatter radical derivatives. We find that the interaction with different contaminants (such as atomic H, Ar, N, and O and molecular H2, N2, O2, H2O, and NH2) affects the chemical and magnetic properties of the thin films upon air exposure. Also, the radical-specific site, where the contaminant interaction takes place, plays a role. Atomic H and NH2 are detrimental to the magnetic properties of Blatter radicals, while the presence of molecular water influences more specifically the magnetic properties of the diradical thin films, and it is believed to be the major cause of the shorter diradical thin film lifetime in air.

Thermally Ultrarobust S = 1/2 Tetrazolinyl Radicals: Synthesis, Electronic Structure, Magnetism, and Nanoneedle Assemblies on Silicon Surface.

Open-shell organic molecules, including S = 1/2 radicals, may provide enhanced properties for several emerging technologies; however, relatively few synthesized to date possess robust thermal stability and processability. We report the synthesis of S = 1/2 biphenylene-fused tetrazolinyl radicals 1 and 2. Both radicals possess near-perfect planar structures based on their X-ray structures and density-functional theory (DFT) computations. Radical 1 possesses outstanding thermal stability as indicated by the onset of decomposition at 269 °C, based on thermogravimetric analysis (TGA) data. Both radicals possess very low oxidation potentials <0 V (vs. SCE) and their electrochemical energy gaps, Ecell ≈ 0.9 eV, are rather low. Magnetic properties of polycrystalline 1 are characterized by superconducting quantum interference device (SQUID) magnetometry revealing a one-dimensional S = 1/2 antiferromagnetic Heisenberg chain with exchange coupling constant J'/k ≈ -22.0 K. Radical 1 in toluene glass possesses a long electron spin coherence time, Tm ≈ 7 µs in the 40-80 K temperature range, a property advantageous for potential applications as a molecular spin qubit. Radical 1 is evaporated under ultrahigh vacuum (UHV) forming assemblies of intact radicals on a silicon substrate, as confirmed by high-resolution X-ray photoelectron spectroscopy (XPS). Scanning electron microscope (SEM) images indicate that the radical molecules form nanoneedles on the substrate. The nanoneedles are stable for at least 64 hours under air as monitored by using X-ray photoelectron spectroscopy. Electron paramagnetic resonance (EPR) studies of the thicker assemblies, prepared by UHV evaporation, indicate radical decay according to first-order kinetics with a long half-life of 50 ± 4 days at ambient conditions.

Ultrafast Bioorthogonal Spin-Labeling and Distance Measurements in Mammalian Cells Using Small, Genetically Encoded Tetrazine Amino Acids.

Site-directed spin-labeling (SDSL)─in combination with double electron-electron resonance (DEER) spectroscopy─has emerged as a powerful technique for determining both the structural states and the conformational equilibria of biomacromolecules. DEER combined with in situ SDSL in live cells is challenging since current bioorthogonal labeling approaches are too slow to allow for complete labeling with low concentrations of spin label prior to loss of signal from cellular reduction. Here, we overcome this limitation by genetically encoding a novel family of small, tetrazine-bearing noncanonical amino acids (Tet-v4.0) at multiple sites in proteins expressed in Escherichia coli and in human HEK293T cells. We achieved specific and quantitative spin-labeling of Tet-v4.0-containing proteins by developing a series of strained trans-cyclooctene (sTCO)-functionalized nitroxides─including a gem-diethyl-substituted nitroxide with enhanced stability in cells─with rate constants that can exceed 106 M-1 s-1. The remarkable speed of the Tet-v4.0/sTCO reaction allowed efficient spin-labeling of proteins in live cells within minutes, requiring only sub-micromolar concentrations of sTCO-nitroxide. DEER recorded from intact cells revealed distance distributions in good agreement with those measured from proteins purified and labeled in vitro. Furthermore, DEER was able to resolve the maltose-dependent conformational change of Tet-v4.0-incorporated and spin-labeled MBP in vitro and support assignment of the conformational state of an MBP mutant within HEK293T cells. We anticipate the exceptional reaction rates of this system, combined with the relatively short and rigid side chains of the resulting spin labels, will enable structure/function studies of proteins directly in cells, without any requirements for protein purification.

Aromatic C-C Bond Cleavage in a Curved π-System of Aminocorannulene.

We report a metal- and oxidant-free aromatic C-C bond cleavage in the curved corannulene skeleton. Reaction of 1-aminocorannulene with hydrazonyl chloride generates an amidrazone intermediate that undergoes facile intramolecular proton migrations and ring annulation to give a 1,2,4-triazole derivative of planar benzo[ghi]fluoranthene, in which the release of strain associated with the curved π-surface and the formation of an aromatic triazole moiety are the driving forces. This report provides new insights into the aromatic C-C bond cleavage.

Ultra-Fast Bioorthogonal Spin-Labeling and Distance Measurements in Mammalian Cells Using Small, Genetically Encoded Tetrazine Amino Acids.

Studying protein structures and dynamics directly in the cellular environments in which they function is essential to fully understand the molecular mechanisms underlying cellular processes. Site-directed spin-labeling (SDSL)-in combination with double electron-electron resonance (DEER) spectroscopy-has emerged as a powerful technique for determining both the structural states and the conformational equilibria of biomacromolecules. In-cell DEER spectroscopy on proteins in mammalian cells has thus far not been possible due to the notable challenges of spin-labeling in live cells. In-cell SDSL requires exquisite biorthogonality, high labeling reaction rates and low background signal from unreacted residual spin label. While the bioorthogonal reaction must be highly specific and proceed under physiological conditions, many spin labels display time-dependent instability in the reducing cellular environment. Additionally, high concentrations of spin label can be toxic. Thus, an exceptionally fast bioorthogonal reaction is required that can allow for complete labeling with low concentrations of spin-label prior to loss of signal. Here we utilized genetic code expansion to site-specifically encode a novel family of small, tetrazine-bearing non-canonical amino acids (Tet-v4.0) at multiple sites in green fluorescent protein (GFP) and maltose binding protein (MBP) expressed both in E. coli and in human HEK293T cells. We achieved specific and quantitative spin-labeling of Tet-v4.0-containing proteins by developing a series of strained trans -cyclooctene (sTCO)-functionalized nitroxides-including a gem -diethyl-substituted nitroxide with enhanced stability in cells-with rate constants that can exceed 10 6 M -1 s -1 . The remarkable speed of the Tet-v4.0/sTCO reaction allowed efficient spin-labeling of proteins in live HEK293T cells within minutes, requiring only sub-micromolar concentrations of sTCO-nitroxide added directly to the culture medium. DEER recorded from intact cells revealed distance distributions in good agreement with those measured from proteins purified and labeled in vitro . Furthermore, DEER was able to resolve the maltose-dependent conformational change of Tet-v4.0-incorporated and spin-labeled MBP in vitro and successfully discerned the conformational state of MBP within HEK293T cells. We anticipate the exceptional reaction rates of this system, combined with the relatively short and rigid side chains of the resulting spin labels, will enable structure/function studies of proteins directly in cells, without any requirements for protein purification.