An Iron-Sulfur Cluster with a Highly Pyramidalized Three-Coordinate Iron Center and a Negligible Affinity for Dinitrogen.

Attempts to generate open coordination sites for N2 binding at synthetic Fe-S clusters often instead result in cluster oligomerization. Recently, it was shown for Mo-Fe-S clusters that such oligomerization reactions can be prevented through the use of sterically protective supporting ligands, thereby enabling N2 complex formation. Here, this strategy is extended to Fe-only Fe-S clusters. One-electron reduction of (IMes)3Fe4S4Cl (IMes = 1,3-dimesitylimidazol-2-ylidene) forms the transiently stable edge-bridged double cubane (IMes)6Fe8S8, which loses two IMes ligands to form the face-bridged double-cubane, (IMes)4Fe8S8. The finding that the three supporting IMes ligands do not confer sufficient protection to curtail cluster oligomerization prompted the design of a new N-heterocyclic carbene, SIArMe,iPr (1,3-bis(3,5-diisopropyl-2,6-dimethylphenyl)-2-imidazolidinylidene; abbreviated as SIAr), that features bulky groups strategically placed in remote positions. When the reduction of (SIAr)3Fe4S4Cl or [(SIAr)3Fe4S4(THF)]+ is conducted in the presence of SIAr, the formation of (SIAr)4Fe8S8 is indeed suppressed, permitting characterization of the reduced [Fe4S4]0 product. Surprisingly, rather than being an N2 complex, the product is simply (SIAr)3Fe4S4: a cluster with a three-coordinate Fe site that adopts an unusually pyramidalized geometry. Although (SIAr)3Fe4S4 does not coordinate N2 to any appreciable extent under the surveyed conditions, it does bind CO to form (SIAr)3Fe4S4(CO). This finding demonstates that the binding pocket at the unique Fe is not too small for N2; instead, the exceptionally weak affinity for N2 can be attributed to weak Fe-N2 bonding. The differences in the N2 coordination chemistry between sterically protected Mo-Fe-S clusters and Fe-only Fe-S clusters are discussed.

An Open-Cuboidal [Fe3S4] Cluster Characterized in Both Biologically Relevant Redox States.

Synthetic analogues of the three common types of Fe-S clusters found in biology─diamond-core [Fe2S2] clusters, open-cuboidal [Fe3S4] clusters, and cuboidal [Fe4S4] clusters─have been reported in each biologically relevant redox state with one exception: the open-cuboidal [Fe3S4]+ cluster. Here, we describe the synthesis and characterization of an open-cuboidal [Fe3S4] cluster in both biologically relevant redox states: [Fe3S4]+ and [Fe3S4]0. Like their biological counterparts, the oxidized cluster has a spin-canted, S = 1/2 ground state, and the reduced cluster has an S = 2 ground state. Structural analysis reveals that the [Fe3S4] core undergoes substantial contraction upon oxidation, in contrast to the minimal structural changes observed for the only [Fe3S4] protein for which high-resolution structures are available in both redox states (Azotobacter vinelandii ferredoxin I; Av FdI). This difference between the synthetic models and Av FdI is discussed in the context of electron transfer by [Fe3S4] proteins.

Effect of obesity on airway and systemic inflammation in adults with asthma: a systematic review and meta-analysis.

BACKGROUND: Obesity is associated with more severe asthma, however, the mechanisms responsible are poorly understood. Obesity is also associated with low-grade systemic inflammation; it is possible that this inflammation extends to the airways of adults with asthma, contributing to worse asthma outcomes. Accordingly, the aim of this review was to examine whether obesity is associated with increased airway and systemic inflammation and adipokines, in adults with asthma. METHODS: Medline, Embase, CINAHL, Scopus and Current Contents were searched till 11 August 2021. Studies reporting measures of airway inflammation, systemic inflammation and/or adipokines in obese versus non-obese adults with asthma were assessed. We conducted random effects meta-analyses. We assessed heterogeneity using the I2 statistic and publication bias using funnel plots. RESULTS: We included 40 studies in the meta-analysis. Sputum neutrophils were 5% higher in obese versus non-obese asthmatics (mean difference (MD)=5.0%, 95% CI: 1.2 to 8.9, n=2297, p=0.01, I2=42%). Blood neutrophil count was also higher in obesity. There was no difference in sputum %eosinophils; however, bronchial submucosal eosinophil count (standardised mean difference (SMD)=0.58, 95% CI=0.25 to 0.91, p<0.001, n=181, I2=0%) and sputum interleukin 5 (IL-5) (SMD=0.46, 95% CI=0.17 to 0.75, p<0.002, n=198, I2=0%) were higher in obesity. Conversely, fractional exhaled nitric oxide was 4.5 ppb lower in obesity (MD=-4.5 ppb, 95% CI=-7.1 ppb to -1.8 ppb, p<0.001, n=2601, I2=40%). Blood C reactive protein, IL-6 and leptin were also higher in obesity. CONCLUSIONS: Obese asthmatics have a different pattern of inflammation to non-obese asthmatics. Mechanistic studies examining the pattern of inflammation in obese asthmatics are warranted. Studies should also investigate the clinical relevance of this altered inflammatory response. PROSPERO REGISTERATION NUMBER: CRD42021254525.

Airway and parenchyma transcriptomics in a house dust mite model of experimental asthma.

Lung transcriptomics studies in asthma have provided valuable information in the whole lung context, however, deciphering the individual contributions of the airway and parenchyma in disease pathogenesis may expedite the development of novel targeted treatment strategies. In this study, we performed transcriptomics on the airway and parenchyma using a house dust mite (HDM)-induced model of experimental asthma that replicates key features of the human disease. HDM exposure increased the expression of 3,255 genes, of which 212 were uniquely increased in the airways, 856 uniquely increased in the parenchyma, and 2187 commonly increased in both compartments. Further interrogation of these genes using a combination of network and transcription factor enrichment analyses identified several transcription factors that regulate airway and/or parenchymal gene expression, including transcription factor EC (TFEC), transcription factor PU.1 (SPI1), H2.0-like homeobox (HLX), metal response element binding transcription factor-1 (MTF1) and E74-like factor 4 (ets domain transcription factor, ELF4) involved in controlling innate immune responses. We next assessed the effects of inhibiting lung SPI1 responses using commercially available DB1976 and DB2313 on key disease outcomes. We found that both compounds had no protective effects on airway inflammation, however DB2313 (8 mg/kg) decreased mucus secreting cell number, and both DB2313 (1 mg/kg) and DB1976 (2.5 mg/kg and 1 mg/kg) reduced small airway collagen deposition. Significantly, both compounds decreased airway hyperresponsiveness. This study demonstrates that SPI1 is important in HDM-induced experimental asthma and that its pharmacological inhibition reduces HDM-induced airway collagen deposition and hyperresponsiveness.

Characterization and inhibition of inflammasome responses in severe and non-severe asthma.

BACKGROUND: Increased airway NLRP3 inflammasome-mediated IL-1ß responses may underpin severe neutrophilic asthma. However, whether increased inflammasome activation is unique to severe asthma, is a common feature of immune cells in all inflammatory types of severe asthma, and whether inflammasome activation can be therapeutically targeted in patients, remains unknown. OBJECTIVE: To investigate the activation and inhibition of inflammasome-mediated IL-1ß responses in immune cells from patients with asthma. METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from patients with non-severe (n = 59) and severe (n = 36 stable, n = 17 exacerbating) asthma and healthy subjects (n = 39). PBMCs were stimulated with nigericin or lipopolysaccharide (LPS) alone, or in combination (LPS + nigericin), with or without the NLRP3 inhibitor MCC950, and the effects on IL-1ß release were assessed. RESULTS: PBMCs from patients with non-severe or severe asthma produced more IL-1ß in response to nigericin than those from healthy subjects. PBMCs from patients with severe asthma released more IL-1ß in response to LPS + nigericin than those from non-severe asthma. Inflammasome-induced IL-1ß release from PBMCs from patients with severe asthma was not increased during exacerbation compared to when stable. Inflammasome-induced IL-1ß release was not different between male and female, or obese and non-obese patients and correlated with eosinophil and neutrophil numbers in the airways. MCC950 effectively suppressed LPS-, nigericin-, and LPS + nigericin-induced IL-1ß release from PBMCs from all groups. CONCLUSION: An increased ability for inflammasome priming and/or activation is a common feature of systemic immune cells in both severe and non-severe asthma, highlighting inflammasome inhibition as a universal therapy for different subtypes of disease.

Valence Localization in Alkyne and Alkene Adducts of Synthetic [Fe4S4]+ Clusters.

Reported herein are alkyne and alkene adducts of synthetic [Fe4S4]+ clusters that model intermediates and inhibitor-bound states in enzymes involved in isoprenoid biosynthesis. Treatment of the N-heterocyclic carbene-ligated cluster [(IMes)3Fe4S4(OEt2)][BArF4] (IMes = 1,3-dimesitylimidazol-2-ylidene; [BArF4]- = tetrakis(3,5-bis(trifluoromethyl)phenyl)borate) with phenylacetylene (PhCCH) or cis-cyclooctene (COE) results in displacement of the Et2O ligand to yield the corresponding π complexes, [(IMes)3Fe4S4(PhCCH)][BArF4] and [(IMes)3Fe4S4(COE)][BArF4]. EPR spectroscopic analysis demonstrates that both clusters are doublets with giso > 2 and thus are spectroscopically faithful models of the analogous species characterized in the isoprenoid biosynthetic enzymes IspG and IspH. Structural and Mössbauer spectroscopic analysis reveals that both complexes are best described as [Fe4S4]+ clusters in which the unique Fe site engages in modest back-bonding to the π-acidic ligand. Paramagnetic NMR studies show that, even at room temperature, the alkyne/alkene-bound Fe centers harbor minority spin and therefore adopt an Fe2+ valence. We propose that such valence localization could likewise occur in Fe-S enzymes that interact with π-acidic molecules.

Airway and parenchymal transcriptomics in a novel model of asthma and COPD overlap.

BACKGROUND: Asthma and chronic obstructive pulmonary disease (COPD) are common chronic respiratory diseases, and some patients have overlapping disease features, termed asthma-COPD overlap (ACO). Patients characterized with ACO have increased disease severity; however, the mechanisms driving this have not been widely studied. OBJECTIVES: This study sought to characterize the phenotypic and transcriptomic features of experimental ACO in mice induced by chronic house dust mite antigen and cigarette smoke exposure. METHODS: Female BALB/c mice were chronically exposed to house dust mite antigen for 11 weeks to induce experimental asthma, cigarette smoke for 8 weeks to induce experimental COPD, or both concurrently to induce experimental ACO. Lung inflammation, structural changes, and lung function were assessed. RNA-sequencing was performed on separated airway and parenchyma lung tissues to assess transcriptional changes. Validation of a novel upstream driver SPI1 in experimental ACO was assessed using the pharmacological SPI1 inhibitor, DB2313. RESULTS: Experimental ACO recapitulated features of both asthma and COPD, with mixed pulmonary eosinophilic/neutrophilic inflammation, small airway collagen deposition, and increased airway hyperresponsiveness. Transcriptomic analysis identified common and distinct dysregulated gene clusters in airway and parenchyma samples in experimental asthma, COPD, and ACO. Upstream driver analysis revealed increased expression of the transcription factor Spi1. Pharmacological inhibition of SPI1 using DB2313, reduced airway remodeling and airway hyperresponsiveness in experimental ACO. CONCLUSIONS: A new experimental model of ACO featuring chronic dual exposures to house dust mite and cigarette smoke mimics key disease features observed in patients with ACO and revealed novel disease mechanisms, including upregulation of SPI1, that are amenable to therapy.

Relationship between type 2 cytokine and inflammasome responses in obesity-associated asthma.

BACKGROUND: Obesity is a risk factor for asthma, and obese asthmatic individuals are more likely to have severe, steroid-insensitive disease. How obesity affects the pathogenesis and severity of asthma is poorly understood. Roles for increased inflammasome-mediated neutrophilic responses, type 2 immunity, and eosinophilic inflammation have been described. OBJECTIVE: We investigated how obesity affects the pathogenesis and severity of asthma and identified effective therapies for obesity-associated disease. METHODS: We assessed associations between body mass index and inflammasome responses with type 2 (T2) immune responses in the sputum of 25 subjects with asthma. Functional roles for NLR family, pyrin domain-containing (NLRP) 3 inflammasome and T2 cytokine responses in driving key features of disease were examined in experimental high-fat diet-induced obesity and asthma. RESULTS: Body mass index and inflammasome responses positively correlated with increased IL-5 and IL-13 expression as well as C-C chemokine receptor type 3 expression in the sputum of subjects with asthma. High-fat diet-induced obesity resulted in steroid-insensitive airway hyperresponsiveness in both the presence and absence of experimental asthma. High-fat diet-induced obesity was also associated with increased NLRP3 inflammasome responses and eosinophilic inflammation in airway tissue, but not lumen, in experimental asthma. Inhibition of NLRP3 inflammasome responses reduced steroid-insensitive airway hyperresponsiveness but had no effect on IL-5 or IL-13 responses in experimental asthma. Depletion of IL-5 and IL-13 reduced obesity-induced NLRP3 inflammasome responses and steroid-insensitive airway hyperresponsiveness in experimental asthma. CONCLUSION: We found a relationship between T2 cytokine and NLRP3 inflammasome responses in obesity-associated asthma, highlighting the potential utility of T2 cytokine-targeted biologics and inflammasome inhibitors.

Evidence for Low-Valent Electronic Configurations in Iron-Sulfur Clusters.

Although biological iron-sulfur (Fe-S) clusters perform some of the most difficult redox reactions in nature, they are thought to be composed exclusively of Fe2+ and Fe3+ ions, as well as mixed-valent pairs with average oxidation states of Fe2.5+. We herein show that Fe-S clusters formally composed of these valences can access a wider range of electronic configurations─in particular, those featuring low-valent Fe1+ centers. We demonstrate that CO binding to a synthetic [Fe4S4]0 cluster supported by N-heterocyclic carbene ligands induces the generation of Fe1+ centers via intracluster electron transfer, wherein a neighboring pair of Fe2+ sites reduces the CO-bound site to a low-valent Fe1+ state. Similarly, CO binding to an [Fe4S4]+ cluster induces electron delocalization with a neighboring Fe site to form a mixed-valent Fe1.5+Fe2.5+ pair in which the CO-bound site adopts partial low-valent character. These low-valent configurations engender remarkable C-O bond activation without having to traverse highly negative and physiologically inaccessible [Fe4S4]0/[Fe4S4]- redox couples.

Reversible Alkyl-Group Migration between Iron and Sulfur in [Fe4S4] Clusters.

Synthetic [Fe4S4] clusters with Fe-R groups (R = alkyl/benzyl) are shown to release organic radicals on an [Fe4S4]3+-R/[Fe4S4]2+ redox couple, the same that has been proposed for a radical-generating intermediate in the superfamily of radical S-adenosyl-l-methionine (SAM) enzymes. In attempts to trap the immediate precursor to radical generation, a species in which the alkyl group has migrated from Fe to S is instead isolated. This S-alkylated cluster is a structurally faithful model of intermediates proposed in a variety of functionally diverse S transferase enzymes and features an "[Fe4S4]+-like" core that exists as a physical mixture of S = 1/2 and 7/2 states. The latter corresponds to an unusual, valence-localized electronic structure as indicated by distortions in its geometric structure and supported by computational analysis. Fe-to-S alkyl group migration is (electro)chemically reversible, and the preference for Fe vs S alkylation is dictated by the redox state of the cluster. These findings link the organoiron and organosulfur chemistry of Fe-S clusters and are discussed in the context of metalloenzymes that are proposed to make and break Fe-S and/or C-S bonds during catalysis.

Endoplasmic reticulum-unfolded protein response signalling is altered in severe eosinophilic and neutrophilic asthma.

INTRODUCTION: The significance of endoplasmic reticulum (ER) stress in asthma is unclear. Here, we demonstrate that ER stress and the unfolded protein response (UPR) are related to disease severity and inflammatory phenotype. METHODS: Induced sputum (n=47), bronchial lavage (n=23) and endobronchial biopsies (n=40) were collected from participants with asthma with varying disease severity, inflammatory phenotypes and from healthy controls. Markers for ER stress and UPR were assessed. These markers were also assessed in established eosinophilic and neutrophilic murine models of asthma. RESULTS: Our results demonstrate increased ER stress and UPR pathways in asthma and these are related to clinical severity and inflammatory phenotypes. Genes associated with ER protein chaperone (BiP, CANX, CALR), ER-associated protein degradation (EDEM1, DERL1) and ER stress-induced apoptosis (DDIT3, PPP1R15A) were dysregulated in participants with asthma and are associated with impaired lung function (forced expiratory volume in 1 s) and active eosinophilic and neutrophilic inflammation. ER stress genes also displayed a significant correlation with classic Th2 (interleukin-4, IL-4/13) genes, Th17 (IL-17F/CXCL1) genes, proinflammatory (IL-1b, tumour necrosis factor α, IL-8) genes and inflammasome activation (NLRP3) in sputum from asthmatic participants. Mice with allergic airway disease (AAD) and severe steroid insensitive AAD also showed increased ER stress signalling in their lungs. CONCLUSION: Heightened ER stress is associated with severe eosinophilic and neutrophilic inflammation in asthma and may play a crucial role in the pathogenesis of asthma.

Aim2 suppresses cigarette smoke-induced neutrophil recruitment, neutrophil caspase-1 activation and anti-Ly6G-mediated neutrophil depletion.

Increased inflammasome responses are strongly implicated in inflammatory diseases; however, their specific roles are incompletely understood. Therefore, we sought to examine the roles of nucleotide-binding oligomerization domain-like receptor (NLR) family, pyrin domain-containing 3 (NLRP3) and absent in melanoma-2 (AIM2) inflammasomes in cigarette smoke-induced inflammation in a model of experimental chronic obstructive pulmonary disease (COPD). We targeted NLRP3 with the inhibitor MCC950 given prophylactically or therapeutically and examined Aim2-/- mice in cigarette smoke-induced experimental COPD. MCC950 treatment had minimal effects on disease development and/or progression. Aim2-/- mice had increased airway neutrophils with decreased caspase-1 levels, independent of changes in lung neutrophil chemokines. Suppressing neutrophils with anti-Ly6G in experimental COPD in wild-type mice reduced neutrophils in bone marrow, blood and lung. By contrast, anti-Ly6G treatment in Aim2-/- mice with experimental COPD had no effect on neutrophils in bone marrow, partially reduced neutrophils in the blood and had no effect on neutrophils or neutrophil caspase-1 levels in the lungs. These findings identify that following cigarette smoke exposure, Aim2 is important for anti-Ly6G-mediated depletion of neutrophils, suppression of neutrophil recruitment and mediates activation of caspase-1 in neutrophils.

Pyrogallate-Based Metal-Organic Framework with a Two-Dimensional Secondary Building Unit.

We report a metal-organic framework (MOF) with a rare two-dimensional (2D) secondary building unit (SBU). The SBU comprises mixed-valent Fe2+ and Fe3+ metal ions bridged by oxygen atoms pertaining to the polytopic ligand 3,3',4,4',5,5'-hexahydroxybiphenyl, which also define the iron-oxide 2D layers. Overall, the anionic framework exhibits rare topology and evidences strong electronic communication between the mixed-valence iron sites. These results highlight the importance of dimensionality control of MOF SBUs for discovering new topologies in reticular chemistry, and especially for improving electronic communication within the MOF skeleton.

Human ß-defensin-2 suppresses key features of asthma in murine models of allergic airways disease.

BACKGROUND: Asthma is an airway inflammatory disease and a major health problem worldwide. Anti-inflammatory steroids and bronchodilators are the gold-standard therapy for asthma. However, they do not prevent the development of the disease, and critically, a subset of asthmatics are resistant to steroid therapy. OBJECTIVE: To elucidate the therapeutic potential of human ß-defensins (hBD), such as hBD2 mild to moderate and severe asthma. METHODS: We investigated the role of hBD2 in a steroid-sensitive, house dust mite-induced allergic airways disease (AAD) model and a steroid-insensitive model combining ovalbumin-induced AAD with C muridarum (Cmu) respiratory infection. RESULTS: In both models, we demonstrated that therapeutic intranasal application of hBD2 significantly reduced the influx of inflammatory cells into the bronchoalveolar lavage fluid. Furthermore, key type 2 asthma-related cytokines IL-9 and IL-13, as well as additional immunomodulating cytokines, were significantly decreased after administration of hBD2 in the steroid-sensitive model. The suppression of inflammation was associated with improvements in airway physiology and treatment also suppressed airway hyper-responsiveness (AHR) in terms of airway resistance and compliance to methacholine challenge. CONCLUSIONS AND CLINICAL RELEVANCE: These data indicate that hBD2 reduces the hallmark features and has potential as a new therapeutic agent in allergic and especially steroid-resistant asthma.

Critical role for iron accumulation in the pathogenesis of fibrotic lung disease.

Increased iron levels and dysregulated iron homeostasis, or both, occur in several lung diseases. Here, the effects of iron accumulation on the pathogenesis of pulmonary fibrosis and associated lung function decline was investigated using a combination of murine models of iron overload and bleomycin-induced pulmonary fibrosis, primary human lung fibroblasts treated with iron, and histological samples from patients with or without idiopathic pulmonary fibrosis (IPF). Iron levels are significantly increased in iron overloaded transferrin receptor 2 (Tfr2) mutant mice and homeostatic iron regulator (Hfe) gene-deficient mice and this is associated with increases in airway fibrosis and reduced lung function. Furthermore, fibrosis and lung function decline are associated with pulmonary iron accumulation in bleomycin-induced pulmonary fibrosis. In addition, we show that iron accumulation is increased in lung sections from patients with IPF and that human lung fibroblasts show greater proliferation and cytokine and extracellular matrix responses when exposed to increased iron levels. Significantly, we show that intranasal treatment with the iron chelator, deferoxamine (DFO), from the time when pulmonary iron levels accumulate, prevents airway fibrosis and decline in lung function in experimental pulmonary fibrosis. Pulmonary fibrosis is associated with an increase in Tfr1+ macrophages that display altered phenotype in disease, and DFO treatment modified the abundance of these cells. These experimental and clinical data demonstrate that increased accumulation of pulmonary iron plays a key role in the pathogenesis of pulmonary fibrosis and lung function decline. Furthermore, these data highlight the potential for the therapeutic targeting of increased pulmonary iron in the treatment of fibrotic lung diseases such as IPF. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

A Terminal Imido Complex of an Iron-Sulfur Cluster.

We report the synthesis and characterization of the first terminal imido complex of an Fe-S cluster, (IMes)3 Fe4 S4 =NDipp (2; IMes=1,3-dimesitylimidazol-2-ylidene, Dipp=2,6-diisopropylphenyl), which is generated by oxidative group transfer from DippN3 to the all-ferrous cluster (IMes)3 Fe4 S4 (PPh3 ). This two-electron process is achieved by formal one-electron oxidation of the imido-bound Fe site and one-electron oxidation of two IMes-bound Fe sites. Structural, spectroscopic, and computational studies establish that the Fe-imido site is best described as a high-spin Fe3+ center, which is manifested in its long Fe-N(imido) distance of 1.763(2) Å. Cluster 2 abstracts hydrogen atoms from 1,4-cyclohexadiene to yield the corresponding anilido complex, demonstrating competency for C-H activation.

Reversible Formation of Alkyl Radicals at [Fe4S4] Clusters and Its Implications for Selectivity in Radical SAM Enzymes.

All kingdoms of life use the transient 5'-deoxyadenosyl radical (5'-dAdo•) to initiate a wide range of difficult chemical reactions. Because of its high reactivity, the 5'-dAdo• must be generated in a controlled manner to abstract a specific H atom and avoid unproductive reactions. In radical S-adenosylmethionine (SAM) enzymes, the 5'-dAdo• is formed upon reduction of SAM by an [Fe4S4] cluster. An organometallic precursor featuring an Fe-C bond between the [Fe4S4] cluster and the 5'-dAdo group was recently characterized and shown to be competent for substrate radical generation, presumably via Fe-C bond homolysis. Such reactivity is without precedent for Fe-S clusters. Here, we show that synthetic [Fe4S4]-alkyl clusters undergo Fe-C bond homolysis when the alkylated Fe site has a suitable coordination number, thereby providing support for the intermediacy of organometallic species in radical SAM enzymes. Addition of pyridine donors to [(IMes)3Fe4S4-R]+ clusters (R = alkyl or benzyl; IMes = 1,3-dimesitylimidazol-2-ylidene) generates R•, ultimately forming R-R coupled hydrocarbons. This process is facile at room temperature and allows for the generation of highly reactive radicals including primary carbon radicals. Mechanistic studies, including use of the 5-hexenyl radical clock, demonstrate that Fe-C bond homolysis occurs reversibly. Using these experimental insights and kinetic simulations, we evaluate the circumstances in which an organometallic intermediate can direct the 5'-dAdo• toward productive H-atom abstraction. Our findings demonstrate that reversible homolysis of even weak M-C bonds is a feasible protective mechanism for the 5'-dAdo• that can allow selective X-H bond activation in both radical SAM and adenosylcobalamin enzymes.

Crucial role for lung iron level and regulation in the pathogenesis and severity of asthma.

Accumulating evidence highlights links between iron regulation and respiratory disease. Here, we assessed the relationship between iron levels and regulatory responses in clinical and experimental asthma.We show that cell-free iron levels are reduced in the bronchoalveolar lavage (BAL) supernatant of severe or mild-moderate asthma patients and correlate with lower forced expiratory volume in 1 s (FEV1). Conversely, iron-loaded cell numbers were increased in BAL in these patients and with lower FEV1/forced vital capacity (FVC) ratio. The airway tissue expression of the iron sequestration molecules divalent metal transporter 1 (DMT1) and transferrin receptor 1 (TFR1) are increased in asthma, with TFR1 expression correlating with reduced lung function and increased Type-2 (T2) inflammatory responses in the airways. Furthermore, pulmonary iron levels are increased in a house dust mite (HDM)-induced model of experimental asthma in association with augmented Tfr1 expression in airway tissue, similar to human disease. We show that macrophages are the predominant source of increased Tfr1 and Tfr1+ macrophages have increased Il13 expression. We also show that increased iron levels induce increased pro-inflammatory cytokine and/or extracellular matrix (ECM) responses in human airway smooth muscle (ASM) cells and fibroblasts ex vivo and induce key features of asthma in vivo, including airway hyper-responsiveness (AHR) and fibrosis, and T2 inflammatory responses.Together these complementary clinical and experimental data highlight the importance of altered pulmonary iron levels and regulation in asthma, and the need for a greater focus on the role and potential therapeutic targeting of iron in the pathogenesis and severity of disease.

A Synthetic Model of Enzymatic [Fe4S4]-Alkyl Intermediates.

Although alkyl complexes of [Fe4S4] clusters have been invoked as intermediates in a number of enzymatic reactions, obtaining a detailed understanding of their reactivity patterns and electronic structures has been difficult owing to their transient nature. To address this challenge, we herein report the synthesis and characterization of a 3:1 site-differentiated [Fe4S4]2+-alkyl cluster. Whereas [Fe4S4]2+ clusters typically exhibit pairwise delocalized electronic structures in which each Fe has a formal valence of 2.5+, Mössbauer spectroscopic and computational studies suggest that the highly electron-releasing alkyl group partially localizes the charge distribution within the cubane, an effect that has not been previously observed in tetrahedrally coordinated [Fe4S4] clusters.

Controlling Substrate Binding to Fe4S4 Clusters through Remote Steric Effects.

The extraordinary reactivity exhibited by many Fe-S enzymes is due in large part to the influence of the protein scaffold on substrate binding and activation. In principle, the coordination chemistry of synthetic Fe-S clusters could similarly be controlled through remote steric effects. Toward this end, we report the synthesis of 3:1 site-differentiated [Fe4S4] clusters ligated by N -heterocyclic carbene (NHC) ligands with variable steric profiles: IMes (1,3-dimesitylimidazol-2-ylidene) and I iPrMe (1,3-diisopropyl-4,5-dimethylimidazol-2-ylidene). Treatment of (IMes)3Fe4S4Cl with NaBArF4 in ethereal solvents (Et2O and THF) leads to the formation of an ether adduct, [(IMes)3Fe4S4(solv)][BArF4]; solvent can be displaced by addition of tBuNC to form the unusual monoisocyanide adduct [(IMes)3Fe4S4(CN tBu)][BArF4]. Carrying out the same reactions with the less sterically encumbered cluster (I iPrMe)3Fe4S4Cl results in more typical reactivity: undesired ligand redistribution to form the homoleptic cluster [(I iPrMe)4Fe4S4][BArF4] and generation of the triisocyanide adduct [(I iPrMe)3Fe4S4(CN tBu)3][BArF4]. The increased steric profile of the IMes ligands disfavors ligand redistribution and defines a binding pocket at the apical Fe, thereby enabling the generation of a coordinatively unsaturated and substitutionally labile Fe site. This method of controlling the coordination chemistry at the apical Fe site by modifying the sterics of ligands bound to adjacent Fe sites complements existing strategies for generating site-differentiated Fe-S clusters and provides new opportunities to direct reactivity at cuboidal metalloclusters.