Localized Electronic Structure of Nitrogenase FeMoco Revealed by Selenium K-edge High Resolution X-ray Absorption Spectroscopy

The size and complexity of Mo-dependent nitrogenase, a multicomponent enzyme capable of reducing dinitrogen to ammonia, have made a detailed understanding of the FeMo cofactor (FeMoco) active site electronic structure an ongoing challenge. Selective substitution of sulfur by selenium in FeMoco affords a unique probe wherein local Fe–Se interactions can be directly interrogated via high-energy resolution fluorescence detected X-ray absorption spectroscopic (HERFD XAS) and extended X-ray absorption fine structure (EXAFS) studies. These studies reveal a significant asymmetry in the electronic distribution of the FeMoco, suggesting a more localized electronic structure picture than is typically assumed for iron–sulfur clusters. Supported by experimental small molecule model data in combination with time dependent density functional theory (TDDFT) calculations, the HERFD XAS data is consistent with an assignment of Fe2/Fe6 as an antiferromagnetically coupled diferric pair. HERFD XAS and EXAFS have also been applied to Se-substituted CO-inhibited MoFe protein, demonstrating the ability of these methods to reveal electronic and structural changes that occur upon substrate binding. These results emphasize the utility of Se HERFD XAS and EXAFS for selectively probing the local electronic and geometric structure of FeMoco

Evaluation of two sets of immunohistochemical and Western blot confirmatory methods in the detection of typical and atypical BSE cases

<p>Abstract</p> <p>Background</p> <p>Three distinct forms of bovine spongiform encephalopathy (BSE), defined as classical (C-), low (L-) or high (H-) type, have been detected through ongoing active and passive surveillance systems for the disease.</p> <p>The aim of the present study was to compare the ability of two sets of immunohistochemical (IHC) and Western blot (WB) BSE confirmatory protocols to detect C- and atypical (L- and H-type) BSE forms.</p> <p>Obex samples from cases of United States and Italian C-type BSE, a U.S. H-type and an Italian L-type BSE case were tested in parallel using the two IHC sets and WB methods.</p> <p>Results</p> <p>The two IHC techniques proved equivalent in identifying and differentiating between C-type, L-type and H-type BSE. The IHC protocols appeared consistent in the identification of PrP^Scdistribution and deposition patterns in relation to the BSE type examined. Both IHC methods evidenced three distinct PrP^Scphenotypes for each type of BSE: prevailing granular and linear tracts pattern in the C-type; intraglial and intraneuronal deposits in the H-type; plaques in the L-type.</p> <p>Also, the two techniques gave comparable results for PrP^Scstaining intensity on the C- and L-type BSE samples, whereas a higher amount of intraglial and intraneuronal PrP^Scdeposition on the H-type BSE case was revealed by the method based on a stronger demasking step.</p> <p>Both WB methods were consistent in identifying classical and atypical BSE forms and in differentiating the specific PrP^Scmolecular weight and glycoform ratios of each form.</p> <p>Conclusions</p> <p>The study showed that the IHC and WB BSE confirmatory methods were equally able to recognize C-, L- and H-type BSE forms and to discriminate between their different immunohistochemical and molecular phenotypes. Of note is that for the first time one of the two sets of BSE confirmatory protocols proved effective in identifying the L-type BSE form. This finding helps to validate the suitability of the BSE confirmatory tests for BSE surveillance currently in place.</p

Aberrant Mitochondrial Homeostasis in the Skeletal Muscle of Sedentary Older Adults

The role of mitochondrial dysfunction and oxidative stress has been extensively characterized in the aetiology of sarcopenia (aging-associated loss of muscle mass) and muscle wasting as a result of muscle disuse. What remains less clear is whether the decline in skeletal muscle mitochondrial oxidative capacity is purely a function of the aging process or if the sedentary lifestyle of older adult subjects has confounded previous reports. The objective of the present study was to investigate if a recreationally active lifestyle in older adults can conserve skeletal muscle strength and functionality, chronic systemic inflammation, mitochondrial biogenesis and oxidative capacity, and cellular antioxidant capacity. To that end, muscle biopsies were taken from the vastus lateralis of young and age-matched recreationally active older and sedentary older men and women (N = 10/group; ♀  =  ♂). We show that a physically active lifestyle is associated with the partial compensatory preservation of mitochondrial biogenesis, and cellular oxidative and antioxidant capacity in skeletal muscle of older adults. Conversely a sedentary lifestyle, associated with osteoarthritis-mediated physical inactivity, is associated with reduced mitochondrial function, dysregulation of cellular redox status and chronic systemic inflammation that renders the skeletal muscle intracellular environment prone to reactive oxygen species-mediated toxicity. We propose that an active lifestyle is an important determinant of quality of life and molecular progression of aging in skeletal muscle of the elderly, and is a viable therapy for attenuating and/or reversing skeletal muscle strength declines and mitochondrial abnormalities associated with aging

Selenium Valence-to-Core X-ray Emission Spectroscopy and Kβ HERFD X-ray Absorption Spectroscopy as Complementary Probes of Chemical and Electronic Structure

Selenium X-ray absorption spectroscopy (XAS) has found widespread use in investigations of Se-containing materials, geochemical processes, and biological active sites. In contrast to sulfur Kβ X-ray emission spectroscopy (XES), which has been found to contain electronic and structural information complementary to S XAS, Se Kβ XES remains comparatively under-explored. Herein, we present the first Se Valence-to-Core (VtC) XES studies of reduced Se-containing compounds and FeSe dimers. Se VtC XES is found to be sensitive to changes in covalent Se bonding interactions (Se–Se/Se–C/Se–H bonding) while relatively insensitive to changes in Fe oxidation states as selenide bridges in FeSe dimers ([Fe2Se2]2+ vs [Fe2Se2]+). Contrastingly, Se Kβ HERFD XAS is demonstrated to be quite sensitive to changes in Fe-oxidation state, with Se Kβ HERFD XAS demonstrating experimental resolution equivalent to K HERFD XAS. Additionally, computational studies reveal both Se VtC XES and XAS to be sensitive to selenium protonation in FeSe complexes

Do patients perceive a link between a fragility fracture and osteoporosis?

Abstract Background To evaluate factors associated with whether patients associate their fracture with future fracture risk. Methods Fragility fracture patients participated in a telephone interview. Unadjusted odds ratios (OR, [95% CI]) were calculated to identify factors associated with whether patients associate their fracture with increased fracture risk or osteoporosis. Predictors identified in univariate analysis were entered into multivariable logistic regression models. Results 127 fragility fracture patients (82% female) participated in the study, mean (SD) age 67.5 (12.7) years. An osteoporosis diagnosis was reported in 56 (44%) participants, but only 17% thought their fracture was related to osteoporosis. Less than 50% perceived themselves at increased risk of fracture. The odds of an individual perceiving themselves at increased risk for fracture were higher for those that reported a diagnosis of osteoporosis (OR 22.91 [95%CI 7.45;70.44], p Conclusion Many fragility fracture patients do not associate their fracture with osteoporosis. It is crucial for physicians to communicate to patients that an osteoporosis diagnosis, increasing age or a fragility fracture increases the risk for future fracture.</p

Highly Activated Terminal Carbon Monoxide Ligand in an Iron–Sulfur Cluster Model of FeMco with Intermediate Local Spin State at Fe

Nitrogenases, the enzymes that convert N2 to NH3, also catalyze the reductive coupling of CO to yield hydrocarbons. CO-coordinated species of nitrogenase clusters have been isolated and used to infer mechanistic information. However, synthetic FeS clusters displaying CO ligands remain rare, which limits benchmarking. Starting from a synthetic cluster that models a cubane portion of the FeMo cofactor (FeMoco), including a bridging carbyne ligand, we report a heterometallic tungsten–iron–sulfur cluster with a single terminal CO coordination in two oxidation states with a high level of CO activation (νCO = 1851 and 1751 cm–1). The local Fe coordination environment (2S, 1C, 1CO) is identical to that in the protein making this system a suitable benchmark. Computational studies find an unusual intermediate spin electronic configuration at the Fe sites promoted by the presence the carbyne ligand. This electronic feature is partly responsible for the high degree of CO activation in the reduced cluster

A sedentary lifestyle exacerbates functional decline and systemic inflammation in the elderly.

<p>(A) Maximal isometric torque (N.m) in young, AO, and SO subjects. Each point represents an individual who underwent strength testing as described in the methods. (B) Walk test and timed stair-climb test in AO and SO subjects as described in the methods. (C) CRP levels (ng.mg of protein⁻¹) in the <i>vastus lateralis</i> of young, AO, and SO subjects (N = 10/group; ♀  =  ♂). Asterisk denotes significant changes <i>vs.</i> young, and dagger denotes significant changes <i>vs.</i> AO (P≤0.05).</p