Distal Oxygen Research Articles

Reaction Mechanism of the Terminal Plastoquinone QB in Photosystem II as Revealed by Time-Resolved Infrared Spectroscopy.

The secondary plastoquinone (PQ) electron acceptor QB in photosystem II (PSII) undergoes a two-step photoreaction through electron transfer from the primary PQ electron acceptor QA, converting into plastoquinol (PQH2). However, the detailed mechanism of the QB reactions remains elusive. Here, we investigated the reaction mechanism of QB in cyanobacterial PSII core complexes using two time-revolved infrared (TRIR) methods: dispersive-type TRIR spectroscopy and rapid-scan Fourier transform infrared spectroscopy. Upon the first flash, the ∼140 μs phase is attributed to electron transfer from QA•- to QB, while the ∼2.2 and ∼440 ms phases are assigned to the binding of an internal PQ in a nearby cavity to the vacant QB site and an external PQ traveling to the QB site through channels, respectively, followed by immediate electron transfer. The resultant QB•- is suggested to be in equilibrium with QBH•, which is protonated at the distal oxygen. Upon the second flash, the ∼130 μs and ∼3.3 ms phases are attributed to electron transfer to QBH• and the protonation of QB•- followed by electron transfer, respectively, forming QBH-, which then immediately accepts a proton from D1-H215 at the proximal oxygen to become QBH2. The resultant D1-H215 anion is reprotonated in ∼22 ms via a pathway involving the bicarbonate ligand. The final ∼490 ms phase may reflect the release of PQH2 and its replacement with PQ. The present results highlight the importance of time-resolved infrared spectroscopy in elucidating the mechanism of QB reactions in PSII.

Phe265 of the D1 protein is required to stabilize plastoquinone binding in the QB-binding site of photosystem II in Synechocystis sp. PCC 6803

In Photosystem II electrons from water splitting pass through a primary quinone electron acceptor (QA) to the secondary plastoquinone (QB). The D2 protein forms the QA-binding site and the D1 protein forms the QB-binding site. A non-heme iron sits between QA and QB resulting in a quinone-Fe-acceptor complex that must be activated before assembly of the oxygen-evolving complex can occur. An extended loop (residues 223–266) between the fourth (helix D) and fifth (helix E) helices of the D1 protein activates forward electron transfer via a conformational change that stabilizes a bidentate bicarbonate ligand to the non-heme iron while simultaneously stabilizing the binding of QB. We show that positioning of D1:Phe265 to provide a hydrogen bond to the distal oxygen of QB is required for forward electron transfer. In addition, mutations targeting D1:Phe265, resulted in a 50 mV decrease in the QB/QB– midpoint potential.

DFT mechanistic study of biomimetic diiron complex catalyzed dehydrogenation: Unexpected Fe(III)Fe(III)-1,1-μ-hydroperoxy active species for hydride abstraction

The diiron active site is pivotal in catalyzing transformations in both biological and chemical systems. Recently, a range of biomimetic diiron catalysts have been synthesized, drawing inspiration from the active architecture of soluble methane monooxygenase (sMMO). These catalysts have been successfully deployed for the dehydrogenation of indolines, marking a significant advancement in the field. Using density functional theory (DFT) calculations, we have identified a novel mechanistic pathway that governs the dehydrogenation of indolines catalyzed by a biomimetic diiron complex. Specifically, this reaction is facilitated by the transfer of a hybrid atom from the C1 position of the substrate to the distal oxygen atom of the Fe(III)Fe(III)-1,1-μ-hydroperoxy active species. This transfer serves as the rate-limiting step for the heterolytic cleavage of the OO bond, ultimately generating the substrate cation. The mechanism we propose aligns well with mechanistic investigations incorporating both kinetic isotope effect (KIE) measurements and evaluations of stereochemical selectivity. This research contributes to the broader scientific understanding of catalysis involving biomimetic diiron complexes and offers valuable insights into the catalytic behaviors of non-heme diiron metalloenzymes.

The efficacy of apneic oxygenation during intubation using a prototype of an oxygenation laryngoscope - a technical simulation

BackgroundRecently, a non-commercial oxygenation laryngoscope was able to maintain apneic oxygenation during simulated intubation efforts. Since that prototype was 3 mm wider than a standard Macintosh laryngoscope blade, the intubation performance of this device may differ from standard blades. A new prototype of an oxygenation laryngoscope was developed, consisting of a standard-size Macintosh blade and a fixed oxygen supply line to the side. Actually, it is unclear at which point of this blade the oxygen supply line should end to facilitate the best possible oxygen supply for apneic oxygenation.MethodsIn this simulation study using a standardized human airway manikin, the efficacy of apneic oxygenation by oxygen insufflation using standard and modified Macintosh blades was compared: a standard Macintosh blade without oxygen supply line as control, one with an additional oxygen supply line ending proximal near the handle, one with the line ending at the middle of the blade, and one with the line ending near the tip. A preoxygenated test lung was connected to an oximeter with a flow rate of 200ml/min, simulating oxygen consumption of a male adult, and to the trachea of an anatomically correctly shaped airway manikin. Apneic oxygenation was performed and oxygen content was measured over a 20-minutes observation period. Experiments were repeated five times for each laryngoscope blade.ResultsOxygen percentage in the test lung dropped from 100 ± 0% at the start of the experiment to 53 ± 1.5% in the room air control group (p < 0.001 compared to all other groups), and to 74 ± 2.5% in the proximal oxygen line group, whereas oxygen percentage remained at 100% in both the medium and distal oxygen line groups (p = 1 between these groups; p < 0.001 between all other groups).ConclusionsIn this simulation study with a preoxygenated airway manikin, the use of a modified Macintosh laryngoscope blade with oxygen line attached at the tip or at the middle were able to maintain apneic oxygenation without measurable drop of oxygen content over 20 min. Proximal placement of the oxygen supply line still showed an advantage against room air, however it did not completely prevent room air from entering the airway.Trial registrationNot applicable.

Pyocyanin-dependent electrochemical inhibition of Pseudomonas aeruginosa biofilms is synergistic with antibiotic treatment.

Pseudomonas aeruginosa biofilms are common in chronic wound infections and recalcitrant to treatment. Survival of cells within oxygen-limited regions in these biofilms is enabled by extracellular electron transfer (EET), whereby small redox active molecules act as electron shuttles to access distal oxidants. Here, we report that electrochemically controlling the redox state of these electron shuttles, specifically pyocyanin (PYO), can impact cell survival within anaerobic P. aeruginosa biofilms and can act synergistically with antibiotic treatment. Prior results demonstrated that under anoxic conditions, an electrode poised at sufficiently oxidizing potential (+100 mV vs Ag/AgCl) promotes EET within P. aeruginosa biofilms by re-oxidizing PYO for reuse by the cells. Here, when a reducing potential (-400 mV vs Ag/AgCl) was used to disrupt PYO redox cycling by maintaining PYO in the reduced state, we observed a 100-fold decrease in colony forming units within these biofilms compared with those exposed to electrodes poised at +100 mV vs Ag/AgCl. Phenazine-deficient Δphz* biofilms were unaffected by the potential applied to the electrode but were re-sensitized by adding PYO. The effect at -400 mV was exacerbated when biofilms were treated with sub-MICs of a range of antibiotics. Most notably, addition of the aminoglycoside gentamicin in a reductive environment almost completely eradicated wild-type biofilms but had no effect on the survival of Δphz* biofilms in the absence of phenazines. These data suggest that antibiotic treatment combined with the electrochemical disruption of PYO redox cycling, either through the toxicity of accumulated reduced PYO or the disruption of EET, or both, can lead to extensive killing. IMPORTANCE Biofilms provide a protective environment but also present challenges to the cells living within them, such as overcoming nutrient and oxygen diffusion limitations. Pseudomonas aeruginosa overcomes oxygen limitation by secreting soluble redox active phenazines, which act as electron shuttles to distal oxygen. Here, we show that electrochemically blocking the re-oxidation of one of these electron shuttles, pyocyanin, decreases cell survival within biofilms and acts synergistically with gentamicin to kill cells. Our results highlight the importance of the role that the redox cycling of electron shuttles fulfills within P. aeruginosa biofilms.

Do Patients With Arterial Occlusive Disease of Different Etiologies Benefit Equally From Cilostazol?

Cilostazol is a guideline-recommended drug that improves intermittent claudication and quality of life in patients with chronic atherosclerotic peripheral arterial disease. The drug is used for most etiologies of arterial occlusive diseases in clinical practice. This study aimed to evaluate whether patients benefit equally from cilostazol regardless of etiology. Patients on cilostazol were divided into 4 groups according to arterial occlusive disease etiology: (1) atherosclerosis, (2) diabetic angiopathy, (3) embolism/thrombosis, and (4) Buerger disease. Patients' maximum walking distance, ankle-brachial index score and distal tissue oxygen saturation (Sto2), clinical improvement onset time, ability to reach maximum benefit time, vascular surgeries, and wounds were compared before they started cilostazol and after 12 months. Results were evaluated at a statistical significance of P < .05. In 194 patients, 307 target extremities were evaluated in the 4 disease groups. After cilostazol use, maximum walking distance, ankle-brachial index score, and distal Sto2 increased significantly in all groups (P < .001), but distal Sto2 in the diabetic angiopathy and Buerger disease groups was significantly lower than in the atherosclerosis group (P < .001). Ankle-brachial index and distal Sto2 differences in the Buerger disease group were significantly lower (both P < .001). The vascular surgery counts decreased significantly in the atherosclerosis and embolism/thrombosis groups (P = .019 and P = .004, respectively). Patients with nonatherosclerotic arterial occlusive disease also benefit from cilostazol, but patients with Buerger disease or diabetic angiopathy seem to benefit less. Combining cilostazol with anticoagulant or antiaggregant agents and closer monitoring of these patients may produce better results.

Evaluation of distal skin temperature and tissue oxygen saturation determined by near-infrared spectroscopy for predicting ultrasound-guided lateral infraclavicular block success.

Changes in tissue oxygen saturation determined by near-infrared spectroscopy (NIRS) may help predict and determine the success of a lateral infraclavicular (LIC) block.We investigated whether evaluation of tissue oxygen saturation determined by NIRS couldbe an indicator of LIC block success. Forty patients scheduled for hand or forearm surgery under LIC block were studied. NIRS sensors were placed on the ventral aspect of both mid-forearms, and the contralateral hand was used as the control group. NIRS values were recorded before the block andat regular intervals during the following 30 min. NIRS values were significantly higher in the successfully blocked patients whencompared to the complete failure, partial failure, and contralateral hand groups at the 10thmin. In the successfully blocked patients, NIRS values (mean ± SD [change in %]) increasedby 11.09 ± 4.86 (16.03%), 15.00 ± 4.53 (21.76%), 16.35 ± 5.14 (23.77%), 16.38 ± 4.88(23.85%), 16.67 ± 5.04 (24.29%), and 16.96 ± 5.71 (24.78%), respectively, from baselineto 5, 10, 15, 20, 25, and 30 min. ΔTs values were significantly higher in the successfullyblocked patients than in the complete failure patients and contralateral hand at the 30thmin. However, there was no statistically significant difference when comparing ΔTs values ofsuccessful block and partial failure block patients at the 30th min. We conclude that measurement of tissue oxygen saturation by NIRS withinthe scope of evaluation of the lateral infraclavicular block is a rapid, effective, and applicabletechnique.

Supracondylar fracture humerus with pink pulseless hand and angiogram confirmed vascular injury – Case report

Supracondylar fracture of the humerus is a common injury in paediatric population. Neurovascular damage is a well-known serious complication of this injury. This should be managed urgently by the treating clinician in order to prevent severe disability to the child. In this case report, we illustrate the presentation, management and follow up of a 7-year-old boy presented with a supracondylar humerus fracture, highlighting the diagnostic and therapeutic measures taken in sequential manner.&#x0D; The child presented following a domestic fall and having pain, swelling and acute deformity of the left elbow. Clinically, no distal radial pulse or oxygen saturation using pulse oximetry detected on admission, even though the left hand was warm and pink with normal capillary refill time. X-ray confirmed the clinical diagnosis of supracondylar fracture humerus, and it was classified as Gartland type III.&#x0D; Manipulation and casting done without internal fixation on the next available casualty list and even after 72 hours of the procedure no return of distal pulse despite warm, pink hand. After vascular opinion, CT Angiogram revealed no flow segment of 2.2cm of brachial artery at the trauma site and minimum demonstrable flow distally. However, watchful waiting was selected as the management option by both vascular and orthopaedic teams as this child continues to have warm, pink hand with normal capillary refilling and minimal pain.&#x0D; In 6 months follow up, the child has gained both radial and ulnar pulses, full range of elbow motion, complete neurological recovery and good hand function.

Formation and stabilization of C4a-hydroperoxy-FAD by the Arg/Asn pair in HadA monooxygenase.

HadA monooxygenase catalyses the detoxification of halogenated phenols and nitrophenols via dehalogenation and denitration respectively. C4a-hydroperoxy-FAD is a key reactive intermediate wherein its formation, protonation and stabilization reflect enzyme efficiency. Herein, transient kinetics, site-directed mutagenesis and pH-dependent behaviours of HadA reaction were employed to identify key features stabilizing C4a-adducts in HadA. The formation of C4a-hydroperoxy-FAD is pH independent, whereas its decay and protonation of distal oxygen are associated with pKa values of 8.5 and 8.4 respectively. These values are correlated with product formation within a pH range of 7.6-9.1, indicating the importance of adduct stabilization to enzymatic efficiency. We identified Arg101 as a key residue for reduced FAD (FADH- ) binding and C4a-hydroperoxy-FAD formation due to the loss of these abilities as well as enzyme activity in HadAR101A and HadAR101Q . Mutations of the neighbouring Asn447 do not affect the rate of C4a-hydroperoxy-FAD formation; however, they impair FADH- binding. The disruption of Arg101/Asn447 hydrogen bond networking in HadAN447A increases the pKa value of C4a-hydroperoxy-FAD decay to 9.5; however, this pKa was not altered in HadAN447D (pKa of 8.5). Thus, Arg101/Asn447 pair should provide important interactions for FADH- binding and maintain the pKa associated with H2 O2 elimination from C4a-hydroperoxy-FAD in HadA. In the presence of substrate, the formation of C4a-hydroxy-FAD at the hydroxylation step is pH insensitive, and it dehydrates to form the oxidized FAD with pKa of 7.9. This structural feature might help elucidate how the reactive intermediate was stabilized in other flavin-dependent monooxygenases.

Computational Study of Aromatic Hydroxylation Catalyzed by the Iron-Dependent Hydroxylase PqqB Involved in the Biosynthesis of Redox Cofactor Pyrroloquinoline Quinone.

PqqB from Methylobacterium extorquens is a unique nonheme iron-dependent hydroxylase involved in the biosynthesis of redox cofactor pyrroloquinoline quinone (PQQ). A series of recent experiments have demonstrated that PqqB catalyzes the stepwise insertions of two oxygen atoms into the tyrosine ring of the diamino acid substrate, generating the quinone moiety of PQQ; however, the reaction details have not been elucidated yet. In this paper, on the basis of the crystal structures, the enzyme-substrate complex models were constructed, and the catalytic mechanism of PqqB was explored by performing a series of combined QM/MM calculations. Our results confirmed that the first hydroxylation is performed by the highly reactive FeIV-oxo species and follows the typical H-abstraction/hydroxyl rebound mechanism, which is similar to the common aliphatic hydroxylation catalyzed by the α-KG enzymes. Nevertheless, the second hydroxylation is achieved by the Fe-O2 species, and the reactant complex can be described as an intermediate radical-FeII-superoxide, that is, the dioxygen is activated by accepting an electron from the bidentate coordination intermediate. Since both the dioxygen and intermediate are activated by electron transfer, the distal oxygen of superoxide can directly attack the carbonyl carbon of substrate to form an alkylperoxo intermediate, then the O-O heterolysis occurs to afford the epoxide intermediate, which finally evolves into the product by rearrangement. It is the bidentate coordination of catechol moiety to iron that leads to the one-electron oxidation of the substrate by the dioxygen, which significantly activates the substrate and promotes the superoxide radical attack. During the catalysis, Asp90 and His240 in the second sphere play an important role by acting as acid-base catalysts to mediate the proton transfer and manipulate the suitable orientation of superoxide. These findings may provide useful information for understanding the unique reaction mechanism of PqqB that employs both the FeIV-oxo and FeII-superoxide to carry out the aromatic hydroxylation.

Updating the Paradigm: Redox Partner Binding and Conformational Dynamics in Cytochromes P450.

This Account summarizes recent findings centered on the role that redox partner binding, allostery, and conformational dynamics plays in cytochrome P450 proton coupled electron transfer. P450s are one of Nature's largest enzyme families and it is not uncommon to find a P450 wherever substrate oxidation is required in the formation of essential molecules critical to the life of the organism or in xenobiotic detoxification. P450s can operate on a remarkably large range of substrates from the very small to the very large, yet the overall P450 three-dimensional structure is conserved. Given this conservation of structure, it is generally assumed that the basic catalytic mechanism is conserved. In nearly all P450s, the O2 O-O bond must be cleaved heterolytically enabling one oxygen atom, the distal oxygen, to depart as water and leave behind a heme iron-linked O atom as the powerful oxidant that is used to activate the nearby substrate. For this process to proceed efficiently, externally supplied electrons and protons are required. Two protons must be added to the departing O atom while an electron is transferred from a redox partner that typically contains either a Fe2S2 or FMN redox center. The paradigm P450 used to unravel the details of these mechanisms has been the bacterial CYP101A1 or P450cam. P450cam is specific for its own Fe2S2 redox partner, putidaredoxin or Pdx, and it has long been postulated that Pdx plays an effector/allosteric role by possibly switching P450cam to an active conformation. Crystal structures, spectroscopic data, and direct binding experiments of the P450cam-Pdx complex provide some answers. Pdx shifts the conformation of P450cam to a more open state, a transition that is postulated to trigger the proton relay network required for O2 activation. An essential part of this proton relay network is a highly conserved Asp (sometimes Glu) that is known to be critical for activity in a number of P450s. How this Asp and proton delivery networks are connected to redox partner binding is quite simple. In the closed state, this Asp is tied down by salt bridges, but these salt bridges are ruptured when Pdx binds, leaving the Asp free to serve its role in proton transfer. An alternative hypothesis suggests that a specific proton relay network is not really necessary. In this scenario, the Asp plays a structural role in the open/close transition and merely opening the active site access channel is sufficient to enable solvent protons in for O2 protonation. Experiments designed to test these various hypotheses have revealed some surprises in both P450cam and other bacterial P450s. Molecular dynamics and crystallography show that P450cam can undergo rather significant conformational gymnastics that result in a large restructuring of the active site requiring multiple cis/trans proline isomerizations. It also has been found that X-ray driven substrate hydroxylation is a useful tool for better understanding the role that the essential Asp and surrounding residues play in catalysis. Here we summarize these recent results which provide a much more dynamic picture of P450 catalysis.

Theoretical (DFT) study on the hydroxylation mechanism of Sn(IV)porphyrin: How does Sn(IV)porphyrin inhibit heme oxygenase catalysis function

We have performed quantum mechanics (QM) calculation on the mechanism of the tin porphyrin hydroxylation, analogous process to the first step of heme degradation. We investigated a concerted mechanism, whereby the distal oxygen of hydroperoxo ligand (O2, Scheme 2a) of [SnIVporphyrin(OOH)]+ complex attacks directly to porphyrin α‑meso carbon, followed by the OOH bond cleavage and the formation of a hydroxyled porphyrin macrocycle. The first and rate-determining step of the process involves the distortion of macrocycle. We have also investigated an alternative stepwise mechanism involving hydrogen bonding interaction between hyrdoperoxo group and one water molecule, of water cluster; in which dioxygen bond of heydroperoxo breaks homolyticly. The concerted, direct attack pathway, is computed to have a higher energy barrier compared with OO homolytic cleavage mechanism. Furthermore, calculations reveal that in comparison to heme degradation process, hydroxylation of porphyrin α‑meso carbon in SnIVporphyrin complexes is a much higher-energy pathway. Therefore, according to this study, it could be concluded that in the enzymatic environment Sn complexes bounded to HO enzyme remain un-degraded.

Computational Survey of Recent Experimental Developments in the Hydroxylation Mechanism of Kynurenine 3-Monooxygenase.

Recently, two new mechanistic proposals for the kynurenine 3-monooxygenase (KMO) catalyzed hydroxylation reaction of l-Kynurenine (l-Kyn) have been proposed. According to the first proposal, instead of the distal oxygen, the proximal oxygen of the hydroperoxide intermediate of flavin adenine dinucleotide (FAD) is transferred to the substrate ring. The second study proposes that l-Kyn participates in its base form in the reaction. To address these proposals, the reaction was reconsidered with a 386 atom quantum cluster model that is based on a recent X-ray structure (PDB id: 6FOX). The computations were carried out at the UB3LYP/6-311+G(2d,2p)//UB3LYP/6-31G(d,p) level with solvation (polarizable continuum model) and dispersion (DFT-D3(BJ)) corrections. To supplement the results of the density functional theory (DFT) calculations, molecular dynamics (MD) simulations of the protein-substrate complex were employed. The comparison of a proximal oxygen transfer mechanism to the distal oxygen transfer mechanism revealed that the former requires too high of a barrier energy while the latter validated our previous results. According to the MD simulations, the hydroperoxy moiety does not favor an alignment that might promote the proximal oxygen transfer mechanism. In the second part of the study, hydroxylation reaction with the base form of l-Kyn was sought. Although DFT calculations confirmed a much more facile reaction with the base form of l-Kyn, a mechanism which would allow the deprotonation of the l-Kyn before the oxygen transfer could not be determined with the X-ray-based positions. A concerted mechanism with both the oxygen transfer and the deprotonation required a high barrier energy. A stepwise mechanism involving the deprotonation of l-Kyn was found, starting from an MD frame. The overall barrier of the oxygen transfer step of this model was found to be in the range of that of with neutral l-Kyn. MD simulations supported the idea of ineffectiveness of the nearby shell surrounding the utilized active site core on the deprotonation of l-Kyn.

UVPD Spectroscopy of Differential Mobility-Selected Prototropic Isomers of Rivaroxaban.

Two ion populations of protonated Rivaroxaban, [C19H18ClN3O5S + H]+, are separated under pure N2 conditions using differential mobility spectrometry prior to characterization in a hybrid triple quadrupole linear ion trap mass spectrometer. These populations are attributed to bare protonated Rivaroxaban and to a proton-bound Rivaroxaban-ammonia complex, which dissociates prior to mass-selecting the parent ion. Ultraviolet photodissociation (UVPD) and collision-induced dissociation (CID) studies indicate that both protonated Rivaroxaban ion populations are comprised of the computed global minimum prototropic isomer. Two ion populations are also observed when the collision environment is modified with 1.5% (v/v) acetonitrile. In this case, the protonated Rivaroxaban ion populations are produced by the dissociation of the ammonium complex and by the dissociation of a proton-bound Rivaroxaban-acetonitrile complex prior to mass selection. Again, both populations exhibit a similar CID behavior; however, UVPD spectra indicate that the two ion populations are associated with different prototropic isomers. The experimentally acquired spectra are compared with computed spectra and are assigned to two prototropic isomers that exhibit proton sharing between distal oxygen centers.

Mechanistic Insights into the Oxygen Atom Transfer Reactions by Nonheme Manganese Complex: A Computational Case Study on the Comparative Oxidative Ability of Manganese-Hydroperoxo vs High-Valent MnIV═O and MnIV–OH Intermediates

Understanding the comparative oxidative abilities of high-valent metal-oxo/hydroxo/hydroperoxo species holds the key to robust biomimic catalysts that perform desired organic transformations with very high selectivity and efficiency. The comparative oxidative abilities of popular high-valent iron-oxo and manganese-oxo species are often counterintuitive, for example, oxygen atom transfer (OAT) reaction by [(Me2EBC)MnIV-OOH]3+, [(Me2EBC)MnIV-OH]3+, and [(Me2EBC)MnIV═O]2+ (Me2EBC = 4,11-dimethyl-1,4,8,11-tetraazabicyclo[6.6.2]hexadecane) shows extremely high reactivity for MnIV-OOH species and no reactivity for MnIV-OH and MnIV═O species toward alkyl/aromatic sulfides. Using a combination of density functional theory (DFT) and ab initio domain-based local pair natural orbital coupled-cluster with single, double, and perturbative triples excitation (DLPNO-CCSD(T)) and complete-active space self-consistent field/N-electron valence perturbation theory second order (CASSCF/NEVPT2) calculations, here, we have explored the electronic structures and sulfoxidation mechanism of these species. Our calculations unveil that MnIV-OOH reacts through distal oxygen atom with the substrate via electron transfer (ET) mechanism with a very small kinetic barrier (16.5 kJ/mol), placing this species at the top among the best-known catalysts for such transformations. The MnIV-OH and MnIV═O species have a much larger barrier. The mechanism has also been found to switch from ET in the former to concerted in the latter, rendering both unreactive under the tested experimental conditions. Intrinsic differences in the electronic structures, such as the presence and absence of the multiconfigurational character coupled with the steric effects, are responsible for such variations observed. This comparative oxidative ability that runs contrary to the popular iron-oxo/hydroperoxo reactivity will have larger mechanistic implications in understanding the reactivity of biomimic catalysts and the underlying mechanisms in PSII.

Iron‐Catalyzed N‐Alkylation of Secondary Amines with Alcohols Using Borrowing Hydrogen Strategy

AbstractIron‐based catalysts have been widely used in N‐alkylation of amines with alcohol using borrowing hydrogen strategy, due to low toxicity and cost. Herein, based on Saito and co‐workers’ experiment, we discuss a possible mechanism for N‐alkylation of secondary amines with alcohol under an iron‐based catalyst using DFT calculations. Contrary to N‐alkylation of primary amines, only one hydrogen on alcohol benzyl group is taken by the iron‐catalyst during radical transfer process. Thus, instead of forming an imine, a structure with a radical is formed. Overall, there are three parts to this mechanism: 1) hydrogen borrowing by the iron‐based catalyst; 2) radical addition; 3) hydrogen reduction. There are two possible pathways of hydrogen borrowing: either by a distal oxygen or by the oxygen directly coordinating with iron. According to the computed free energy barrier, the former one is more favorable. We also found four possible pathways of radical addition. Our calculations suggest a direct dehydration step with short steps and lower energy barrier was more favorable.

Unraveling the mechanism of latex clearing protein Lcp&lt;sub&gt;K30&lt;/sub&gt; catalysed oxidative cleavage of poly(&lt;italic&gt;cis&lt;/italic&gt;-1,4-isoprene)

<p indent="0mm">Natural rubber or synthetic rubber materials have been extensively used in our daily life and industrial production for more than <sc>100 years</sc> due to their unique physical properties. However, a amount of rubber waste has been released into the environment, posing great challenges to the disposal of these polymers. In contrast to the disposal strategies such as combustion or stockpiling, enzyme catalyzed rubber degradation constitutes an environmentally friendly strategy. Latex clearing protein (Lcp) is an iron-heme containing endotype dioxygenase that catalyses the oxidative cleavage of the chemically inert poly(<italic>cis</italic>-1,4-isoprene) which is the main component of rubber, affording the products of aldehyde and ketone. It was found that active site E148 residue is key to the reactivity of Lcp based on the mutation experiments. However, the role of E148 and the overall mechanism are still controversial. It is generally accepted that both of dioxygen atoms are incorporated into the substrate. In previous studies, two possible mechanisms of Lcp catalyzed cleavage of poly(<italic>cis</italic>-1,4-isoprene) are proposed. In the first one, the distal oxygen atom of Fe(III)−O₂⁻ complex attacks onto the C=C double bond, leading to a three-membered epoxide intermediate. This is followed by the attack of Fe=O complex on the epoxide intermediate and subsequent C−C bond cleavage, affording the final ketone and aldehyde products. While in the second one, the proximal oxygen atom of Fe(III)−O₂⁻ complex attacks onto the C=C double bond of the substrate, which is followed by the attack of the distal oxygen atom onto the substrate radical to generate a four-membered cyclic dioxetane intermediate. The final O−O as and C−C bond cleavage in the dioxetane intermediate would afford the final products. In this study, based on the crystal structure of Lcp_K30, the combined molecular dynamic (MD) simulations and quantum mechanics/molecular mechanics (QM/MM) calculations are performed to elucidate the catalytic mechanism of Lcp catalysed cleavage of central double bond of poly(<italic>cis</italic>-1,4-isoprene). Our study shows E148 is protonated during the reaction and forms stable H bond interactions with Fe(III)−O₂⁻ species. Thus, the plausible role of E148 is related to the positioning of both Fe(III)−O₂⁻ species and the substrate. Moreover, it can be ruled out that the E148 acts as a base to abstract the proton from the substrate. The proposed mechanism of Lcp works as follows. Firstly, the distal oxygen of Fe(III)−O₂⁻ attacks onto the carbon−carbon double bond to generate the peroxide alky radical intermediate. This is followed by the rebound of the proximal oxygen to the carbon radical to generate the dioxetane intermediate, with Fe(III) being reduced to Fe(II). The following O−O bond cleavage occurs via the Fe(II)-mediated reductive cleavage of O−O bond of the dioxetane intermediate, leading to the vicinal diol radical anion intermediate. Finally, a facile C−C cleavage is undergone to yield the ketone and aldehyde products. In particular, our calculations show that the direct O−O cleavage of the dioxetane intermediate in the absence of Fe(II) catalysis requires a remarkably high barrier of <sc>160 kJ/mol,</sc> indicating the Fe(II) catalysis is vital for the further transformation of the dioxetane intermediate. For comparison, we also investigated the previously proposed mechanism involving the three-membered epoxide intermediate. However, the attack of the epoxide by Fe^IV=O complex is found to be highly unfavourable thermodynamically and thus can be ruled out. As such, this study may not only expand our understanding of the catalytic mechanism of Lcp_K30, but also provide useful information for rational design or engineering of Lcp enzyme toward the degradation of synthetic rubber materials.

Sural Nerve Perfusion in Mice.

Peripheral nerve function is metabolically demanding and nerve energy failure has been implicated in the onset and development of diabetic peripheral neuropathy and neuropathic pain conditions. Distal peripheral nerve oxygen supply relies on the distribution of red blood cells (RBCs) in just a few, nearby capillary-sized vessels and is therefore technically challenging to characterize. We developed an approach to characterize distal sural nerve hemodynamics in anesthetized, adult male mice using in vivo two-photon laser scanning microscopy. Our results show that RBC velocities in mouse sural nerve vessels are higher than those previously measured in mouse brain, and are sensitive to hindlimb temperatures. Nerve blood flow, measured as RBC flux, however, was similar to that of mouse brain and unaffected by local temperature. Power spectral density analysis of fluctuations in RBC velocities over short time intervals suggest that the technique is sufficiently sensitive and robust to detect subtle flow oscillations over time scales from 0.1 to tens of seconds. We conclude that in vivo two-photon laser scanning microscopy provides a suitable approach to study peripheral nerve hemodynamics in mice, and that local temperature control is important during such measurements.

Non-radical reactions in persulfate-based homogeneous degradation processes: A review

Recently there is vigorous growth of reports on persulfate (PS)-based advanced oxidation processes (AOPs), inclining to development of newfangled catalysts and treatment of a wide range of organic pollutants. PS-related non-radical oxidation pathways are lately proposed, whereas there still exist debatable observations and explanation on these issues. While non-radical reactions in heterogeneous systems have been well commented, this review focuses on the latest discovery of non-radical oxidation in homogeneous persulfate (especially peroxymonosulfate (PMS)) systems. The application of direct oxidation process is firstly summarized and the roles of PMS distal oxygen in attack on organic pollutants like antibiotics are highlighted. Besides, the effects of exogenic and endogenic halogen on the reactivity and mechanisms of HOX in PS activation are discussed. Then the formation and reaction pathways of singlet oxygen ( 1 O 2 ) triggered by phenol/carbonyl compounds and nucleophiles are elaborated. Recent discovery on formation of high valent metal species by PS activation and their reactivity is reviewed lastly. By sorting out a large number of cutting-edge research reports, this review conducts a detailed and critical analysis and overview on the homogeneous PS non-radical mechanisms, and points out the controversies and challenges so as to better understand the reaction mechanism and further improve their treatment performance. • Direct oxidation and halides induced non-radical processes are summarized. • 1 O 2 and high-valent metal species oxidation mechanisms are discussed. • Debatable role of 1 O 2 in non-radical oxidation is emphasized. • Prospective and implications of PS non-radical reaction are proposed.

Mechanistic Insights into the Oxidative Rearrangement Catalyzed by the Unprecedented Dioxygenase ChaP Involved in Chartreusin Biosynthesis.

ChaP is a non-heme iron-dependent dioxygenase belonging to the vicinal oxygen chelate (VOC) enzyme superfamily that catalyzes the final α-pyrone ring formation in the biosynthesis of chartreusin. In contrast to other common dioxygenases, for example, 2,3-catechol dioxygenase which uses the dioxygen molecule as the oxidant, ChaP requires the flavin-activated oxygen (O22-) as the equivalent. Previous experiments showed that the ChaP-catalyzed ring rearrangement contains two successive C-C bond cleavages and one lactonization; however, the detailed reaction mechanism is unknown. In this work, on the basis of the recently obtained crystal structure of ChaP, the computational model was constructed and the catalytic mechanism of ChaP was explored by performing quantum mechanical/molecular mechanical (QM/MM) calculations. Our calculation results reveal that ChaP uses the proximal oxygen in iron-coordinated HOO- to attack the carbonyl carbon of the substrate, whereas the previous proposal that Asp49 acts as a base to deprotonate the iron-coordinated HOO- to generate O22- is unlikely. In the first stage reaction, owing to the coordination of the substrate with iron, the substrate is activated by accepting an electron from iron and the resulting oxy-radical intermediate formed by O-O cleavage can easily trigger the ring rearrangement. In the final decarboxylation, the phenolic anion of the substrate cooperatively accepts the proton of iron-coordinated HOO- to facilitate the attack of the distal oxygen, and the proton-coupled electron transfer (PCET) from the substrate to the FeIV═O plays a key role for the decarboxylation. These findings may provide useful information for understanding the ChaP-catalyzed oxidative rearrangement and other flavin-dependent non-heme dioxygenases.