Understanding Covalent versus Spin–Orbit Coupling Contributions to Temperature-Dependent Electron Spin Relaxation in Cupric and Vanadyl Phthalocyanines

Recent interest in transition-metal complexes as potential quantum bits (qubits) has reinvigorated the investigation of fundamental contributions to electron spin relaxation in various ligand scaffolds. From quantum computers to chemical and biological sensors, interest in leveraging the quantum properties of these molecules has opened a discussion of the requirements to maintain coherence over a large temperature range, including near room temperature. Here we compare temperature-, magnetic field position-, and concentration-dependent electron spin relaxation in copper(II) phthalocyanine (CuPc) and vanadyl phthalocyanine (VOPc) doped into diamagnetic hosts. While VOPc demonstrates coherence up to room temperature, CuPc coherence times become rapidly T₁-limited with increasing temperature, despite featuring a more covalent ground-state wave function than VOPc. As rationalized by a ligand field model, this difference is ascribed to different spin–orbit coupling (SOC) constants for Cu(II) versus V(IV). The manifestation of SOC contributions to spin–phonon coupling and electron spin relaxation in different ligand fields is discussed, allowing for a further understanding of the competing roles of SOC and covalency in electron spin relaxation

Understanding Covalent versus Spin–Orbit Coupling Contributions to Temperature-Dependent Electron Spin Relaxation in Cupric and Vanadyl Phthalocyanines

Recent interest in transition-metal complexes as potential quantum bits (qubits) has reinvigorated the investigation of fundamental contributions to electron spin relaxation in various ligand scaffolds. From quantum computers to chemical and biological sensors, interest in leveraging the quantum properties of these molecules has opened a discussion of the requirements to maintain coherence over a large temperature range, including near room temperature. Here we compare temperature-, magnetic field position-, and concentration-dependent electron spin relaxation in copper(II) phthalocyanine (CuPc) and vanadyl phthalocyanine (VOPc) doped into diamagnetic hosts. While VOPc demonstrates coherence up to room temperature, CuPc coherence times become rapidly T₁-limited with increasing temperature, despite featuring a more covalent ground-state wave function than VOPc. As rationalized by a ligand field model, this difference is ascribed to different spin–orbit coupling (SOC) constants for Cu(II) versus V(IV). The manifestation of SOC contributions to spin–phonon coupling and electron spin relaxation in different ligand fields is discussed, allowing for a further understanding of the competing roles of SOC and covalency in electron spin relaxation

On the occurrence of cytochrome P450 in viruses

Author Posting. © The Author(s), 2019. This is the author's version of the work. It is posted here by permission of National Academy of Sciences for personal use, not for redistribution. The definitive version was published in Proceedings of the National Academy of Sciences of the United States of America 116(25), (2019):12343-12352, doi:10.1073/pnas.1901080116.Genes encoding cytochrome P450 (CYP; P450) enzymes occur widely in the Archaea, Bacteria, and Eukarya, where they play important roles in metabolism of endogenous regulatory molecules and exogenous chemicals. We now report that genes for multiple and unique P450s occur commonly in giant viruses in the Mimiviridae, Pandoraviridae, and other families in the proposed order Megavirales. P450 genes were also identified in a herpesvirus (Ranid herpesvirus 3) and a phage (Mycobacterium phage Adler). The Adler phage P450 was classified as CYP102L1, and the crystal structure of the open form was solved at 2.5 Å. Genes encoding known redox partners for P450s (cytochrome P450 reductase, ferredoxin and ferredoxin reductase, and flavodoxin and flavodoxin reductase) were not found in any viral genome so far described, implying that host redox partners may drive viral P450 activities. Giant virus P450 proteins share no more than 25% identity with the P450 gene products we identified in Acanthamoeba castellanii, an amoeba host for many giant viruses. Thus, the origin of the unique P450 genes in giant viruses remains unknown. If giant virus P450 genes were acquired from a host, we suggest it could have been from an as yet unknown and possibly ancient host. These studies expand the horizon in the evolution and diversity of the enormously important P450 superfamily. Determining the origin and function of P450s in giant viruses may help to discern the origin of the giant viruses themselves.We thank Dr. David Nes (Texas Tech University) for providing sterols and Dr. Matthieu Legendre and Dr. Chantal Abergel (CNRS, Marseille) for access to the P. celtis sequences. Drs. Irina Arkhipova, Mark Hahn, Judith Luborsky, and Ann Bucklin commented on the manuscript. The research was supported by a USA-UK Fulbright Scholarship and a Royal Society grant (to D.C.L.), the Boston University Superfund Research Program [NIH Grant 5P42ES007381 (to J.J.S. and J.V.G.) and NIH Grant 5U41HG003345 (to J.V.G.)], the European Regional Development Fund and Welsh Government Project BEACON (S.L.K.), the Woods Hole Center for Oceans and Human Health [NIH Grant P01ES021923 and National Science Foundation Grant OCE-1314642 (to J.J.S.)], and NIH Grant R01GM53753 (to T.L.P.).2019-12-0

Changes in an Enzyme Ensemble During Catalysis Observed by High Resolution XFEL Crystallography

Enzymes populate ensembles of structures with intrinsically different catalytic proficiencies that are difficult to experimentally characterize. We use time-resolved mix-and-inject serial crystallography (MISC) at an X-ray free electron laser (XFEL) to observe catalysis in a designed mutant (G150T) isocyanide hydratase (ICH) enzyme that enhances sampling of important minor conformations. The active site exists in a mixture of conformations and formation of the thioimidate catalytic intermediate selects for catalytically competent substates. A prior proposal for active site cysteine charge-coupled conformational changes in ICH is validated by determining structures of the enzyme over a range of pH values. A combination of large molecular dynamics simulations of the enzyme in crystallo and timeresolved electron density maps shows that ionization of the general acid Asp17 during catalysis causes additional conformational changes that propagate across the dimer interface, connecting the two active sites. These ionization-linked changes in the ICH conformational ensemble permit water to enter the active site in a location that is poised for intermediate hydrolysis. ICH exhibits a tight coupling between ionization of active site residues and catalysis-activated protein motions, exemplifying a mechanism of electrostatic control of enzyme dynamics

μ‑Oxo Dimerization Effects on Ground- and Excited-State Properties of a Water-Soluble Iron Porphyrin CO₂ Reduction Catalyst

Iron 5,10,15,20-tetra(para-N,N,N-trimethylanilinium)porphyrin (Fe-p-TMA) is a water-soluble catalyst capable of electrochemical and photochemical CO2 reduction. Although its catalytic ability has been thoroughly investigated, the mechanism and associated intermediates are largely unknown. Previous studies proposed that Fe-p-TMA enters catalytic cycles as a monomeric species. However, we demonstrate herein that, in aqueous solutions, Fe-p-TMA undergoes formation of a μ-oxo porphyrin dimer that exists in equilibrium with its monomeric form. The propensity for μ-oxo formation is highly dependent on the solution pH and ionic strength. Indeed, the μ-oxo form is stabilized in the presence of electrolytes that are key components of catalytically relevant conditions. By leveraging the ability to chemically control and spectrally address both species, we characterize their ground-state electronic structures and excited-state photodynamics. Global fitting of ultrafast transient absorption data reveals two distinct excited-state relaxation pathways: a three-component sequential model consistent with monomeric relaxation and a two-component sequential model for the μ-oxo species. Relaxation of the monomeric species is best described as a ligand-to-metal charge transfer (τ1 = ∼500 fs), an ionic strength-dependent metal-to-ligand charge transfer (τ2 = 2–4 ps), and finally relaxation of a ligand field excited state to the ground state (τ3 = 5 ps). Conversely, excited-state relaxation of the μ-oxo species proceeds via cleavage of an FeIII–O bond to generate transient FeIVO and FeII porphyrin species (τ1 = 2 ps) that recombine to the ground-state μ-oxo species (τ2 = ∼1 ns). This latter lifetime extends to timescales relevant for chemical reactivity. It is therefore emphasized that further consideration of catalyst speciation and chemical microenvironments is necessary for elucidating the mechanisms of catalytic CO2 reduction reactions

Selective C–H Bond Cleavage with a High-Spin Fe^IV–Oxido Complex

Non-heme Fe monooxygenases activate C–H bonds using intermediates with high-spin FeIV–oxido centers. To mimic these sites, a new tripodal ligand [pop]3− was prepared that contains three phosphoryl amido groups that are capable of stabilizing metal centers in high oxidation states. The ligand was used to generate [FeIVpop(O)]−, a new FeIV–oxido complex with an S = 2 spin ground state. Spectroscopic measurements, which included low-temperature absorption and electron paramagnetic resonance spectroscopy, supported the assignment of a high-spin FeIV center. The complex showed reactivity with benzyl alcohol as the external substrate but not with related compounds (e.g., ethyl benzene and benzyl methyl ether), suggesting the possibility that hydrogen bonding interaction(s) between the substrate and [FeIVpop(O)]− was necessary for reactivity. These results exemplify the potential role of the secondary coordination sphere in metal-mediated processes

Selective C–H Bond Cleavage with a High-Spin FeIV–Oxido Complex

Non-heme Fe monooxygenases activate C-H bonds using intermediates with high-spin FeIV-oxido centers. To mimic these sites, a new tripodal ligand [pop]3- was prepared that contains three phosphoryl amido groups that are capable of stabilizing metal centers in high oxidation states. The ligand was used to generate [FeIVpop(O)]-, a new FeIV-oxido complex with an S = 2 spin ground state. Spectroscopic measurements, which included low-temperature absorption and electron paramagnetic resonance spectroscopy, supported the assignment of a high-spin FeIV center. The complex showed reactivity with benzyl alcohol as the external substrate but not with related compounds (e.g., ethyl benzene and benzyl methyl ether), suggesting the possibility that hydrogen bonding interaction(s) between the substrate and [FeIVpop(O)]- was necessary for reactivity. These results exemplify the potential role of the secondary coordination sphere in metal-mediated processes

An Artificial Peroxidase based on the Biotin-Streptavidin Technology that Rivals the Efficiency of Natural Peroxidases

Horseradish peroxidase (HRP) is an archetypal heme-containing metalloenzyme that uses peroxide to oxidize various substrates. Capitalizing on a well-established catalytic mechanism, diverse peroxidase mimics have been widely investigated and optimized. Herein, we report on the design, assembly, characterization, and genetic engineering of an artificial heme-based peroxidase relying on the biotin-streptavidin technology. The crystal structure of both the wild-type and the best-performing double mutant of artificial peroxidase provided valuable insight regarding the nearby residues that were strategically mutated to optimize the peroxidase activity (i.e. Sav S112E K121H). We hypothesize that these two residues mimic the two key second coordination residues involved in activating the bound peroxide in HRP (i.e. Arg 38 and His 42). The evolved artificial peroxidase exhibited best-in-class activity for oxidizing two standard substrates (TMB and ABTS) in the presence of hydrogen peroxide

Artificial Metalloproteins with Dinuclear Iron–Hydroxido Centers

Dinuclear iron centers with a bridging hydroxido or oxido ligand form active sites within a variety of metalloproteins. A key feature of these sites is the ability of the protein to control the structures around the Fe centers, which leads to entatic states that are essential for function. To simulate this controlled environment, artificial proteins have been engineered using biotin-streptavidin (Sav) technology in which Fe complexes from adjacent subunits can assemble to form [FeIII-(μ-OH)-FeIII] cores. The assembly process is promoted by the site-specific localization of the Fe complexes within a subunit through the designed mutation of a tyrosinate side chain to coordinate the Fe centers. An important outcome is that the Sav host can regulate the Fe···Fe separation, which is known to be important for function in natural metalloproteins. Spectroscopic and structural studies from X-ray diffraction methods revealed uncommonly long Fe···Fe separations that change by less than 0.3 Å upon the binding of additional bridging ligands. The structural constraints imposed by the protein host on the di-Fe cores are unique and create examples of active sites having entatic states within engineered artificial metalloproteins

Artificial Iron Proteins: Modeling the Active Sites in Non-Heme Dioxygenases

An important class of non-heme dioxygenases contains a conserved Fe binding site that consists of a 2-His-1-carboxylate facial triad. Results from structural biology show that, in the resting state, these proteins are six-coordinate with aqua ligands occupying the remaining three coordination sites. We have utilized biotin-streptavidin (Sav) technology to design new artificial Fe proteins (ArMs) that have many of the same structural features found within active sites of these non-heme dioxygenases. An Sav variant was isolated that contains the S; 112; E mutation, which installed a carboxylate side chain in the appropriate position to bind to a synthetic Fe; II; complex confined within Sav. Structural studies using X-ray diffraction (XRD) methods revealed a facial triad binding site that is composed of two N donors from the biotinylated ligand and the monodentate coordination of the carboxylate from S; 112; E. Two aqua ligands complete the primary coordination sphere of the Fe; II; center with both involved in hydrogen bond networks within Sav. The corresponding Fe; III; protein was also prepared and structurally characterized to show a six-coordinate complex with two exogenous acetato ligands. The Fe; III; protein was further shown to bind an exogenous azido ligand through replacement of one acetato ligand. Spectroscopic studies of the ArMs in solution support the results found by XRD