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[FeFe]-hydrogenases catalyze the reversible two-electron reduction of two protons to molecular hydrogen. Although these enzymes are among the most efficient H2-converting biocatalysts in nature, their catalytic cofactor (termed H-cluster) is irreversibly destroyed upon contact with dioxygen. The [FeFe]-hydrogenase CbA5H from Clostridium beijerinckii has a unique mechanism to protect the H-cluster from oxygen-induced degradation. The protective strategy of CbA5H was proposed based on a partial protein structure of CbA5H's oxygen-shielded form. Here, we present a cryo-EM structure of 2.2 Å resolution from the entire enzyme in its dimeric and active state and elucidate the structural parameters of the reversible cofactor protection mechanism. We found that both subunits of the homodimeric structure of CbA5H have a Zn2+-binding four-helix domain, which does not play a role in electron transport as described for other complex protein structures. Biochemical data instead confirm that two [4Fe-4S] clusters are responsible for electron transfer in CbA5H, while the identified zinc atom is critical for oligomerization and protein stability. [ABSTRACT FROM AUTHOR]

Abstract Background Aim of the present study is the evaluation of ultrasound as a physical method for virus inactivation in human plasma products prior to transfusion. Our study is focused on achieving a high level of virus inactivation simultaneously leaving blood products unaltered, measured by the level of degradation of coagulation factors, especially in third world countries where virus contamination of blood products poses a major problem. Virus inactivation plays an important role, especially in the light of newly discovered or unknown viruses, which cannot be safely excluded via prior testing. Methods Taking into account the necessary protection of the relevant coagulation activity for plasma, the basis for a sterile virus inactivation under shielding gas insufflation was developed for future practical use. Influence of frequency and power density in the range of soft and hard cavitation on the inactivation of transfusion-relevant model viruses for Hepatitis-(BVDV = bovine diarrhea virus), for Herpes-(SFV = Semliki Forest virus, PRV = pseudorabies virus) and Parvovirus B19 (PPV = porcine parvovirus) were examined. Coagulation activity was examined via standard time parameters to minimize reduction of functionality of coagulation proteins. A fragmentation of coagulation proteins via ultrasound was ruled out via gel electrophoresis. The resulting virus titer was examined using end point titration. Results Through CO2 shielding gas insufflation—to avoid radical emergence effects—the coagulation activity was less affected and the time window for virus inactivation substantially widened. In case of the non-lipidated model virus (AdV-luc = luciferase expressing adenoviral vector), the complete destruction of the virus capsid through hard cavitation was proven via scanning electron microscopy (SEM). This can be traced back to microjets and shockwaves occurring in hard cavitation. The degree of inactivation seems to depend on size and compactness of the type of viruses. Using our pre-tested and subsequently chosen process parameters with the exception of the small PPV, all model viruses were successfully inactivated and reduced by up to log 3 factor. For a broad clinical usage, protection of the coagulation activities may require further optimization. Conclusions Building upon the information gained, an optimum inactivation can be reached via raising of power density up to 1200 W and simultaneous lowering of frequency down to 27 kHz. In addition, the combination of the two physical methods UV treatment and ultrasound may yield optimum results without the need of substance removal after the procedure.

Abstract Background Intensive care unit (ICU) capacity is a scant and precious resource in hospitals. Therefore, an optimal occupancy rate as well as detailed occupation planning is of great importance. Most literature deals with admission to the ICU, while only few discuss discharge from the ICU. Specifically, a delay of transfer from the ICU can cause a shortness of beds, jeopardize urgent patient treatment and lead to a decrease in treatment quality as well as economic downsides. This study examined the incidence, costs and reasons for delayed discharge from the ICU and analyzed the influence of the department the patient was admitted to. Methods Over the course of 12 months, the discharges of all 1643 patients of two surgical intensive care units of a large academic medical center were analyzed. Delay in minutes and reasons were recorded and translated into financial figures. A univariate logistic regression model was developed to evaluate the impact of length of stay at the ICU, age, gender, subspecialty and specific ICU on the delay of transfer. In a next step, significant factors of the univariate logistic regression were incorporated into a multivariate regression model. Results In 326 out of 1312 patients ready for discharge (24.8%), the transfer to the floor was delayed. Time of delay for all patients added up to a total of 265,691 min in 1 year. The application of the internal cost allocation, in which 1 min corresponds to 0.75 Euro cents, led to costs of 199,268 Euros (~ $240,000) for the study period. In 91.7% of the cases, the reason for the delay was the lack of an available or appropriate bed on the regular ward. Multivariate regression analysis revealed that the type of department the patient is admitted to poses a significantly influencing factor for delayed discharge from the ICU. Conclusion Delay in discharge from the ICU is a common problem of economic relevance. The main reason is a lack of appropriate floor beds. Patients from certain specific departments are at a higher risk to be discharged with delay. A solution to this problem lies in the focus on the downstream units. A proper use of the scarce resources is to be pursued because of ethical as well as economic reasons in an increasingly aging population.

Friedreich’s ataxia (FRDA) is the most important recessive ataxia in the Caucasian population. It is caused by a deficit of the mitochondrial protein frataxin. Despite its pivotal effect on biosynthesis of iron-sulfur clusters and mitochondrial energy production, little is known about the influence of frataxin depletion on homeostasis of the cellular mitochondrial network. We have carried out a forward genetic screen to analyze genetic interactions between genes controlling mitochondrial homeostasis and Drosophila frataxin. Our screen has identified silencing of Drosophila mitofusin (Marf) as a suppressor of FRDA phenotypes in glia. Drosophila Marf is known to play crucial roles in mitochondrial fusion, mitochondrial degradation and in the interface between mitochondria and endoplasmic reticulum (ER). Thus, we have analyzed the effects of frataxin knockdown on mitochondrial morphology, mitophagy and ER function in our fly FRDA model using different histological and molecular markers such as tetramethylrhodamine, ethyl ester (TMRE), mitochondria-targeted GFP (mitoGFP), p62, ATG8a, LAMP1, Xbp1 and BiP/GRP78. Furthermore, we have generated the first Drosophila transgenic line containing the mtRosella construct under the UAS control to study the progression of the mitophagy process in vivo. Our results indicated that frataxin-deficiency had a small impact on mitochondrial morphology but enhanced mitochondrial clearance and altered the ER stress response in Drosophila. Remarkably, we demonstrate that downregulation of Marf suppresses ER stress in frataxin-deficient cells and this is sufficient to improve locomotor dysfunction, brain degeneration and lipid dyshomeostasis in our FRDA model. In agreement, chemical reduction of ER stress by means of two different compounds was sufficient to ameliorate the effects of frataxin deficiency in three different fly FRDA models. Altogether, our results strongly suggest that the protection mediated by Marf knockdown in glia is mainly linked to its role in the mitochondrial-ER tethering and not to mitochondrial dynamics or mitochondrial degradation and that ER stress is a novel and pivotal player in the progression and etiology of FRDA. This work might define a new pathological mechanism in FRDA, linking mitochondrial dysfunction due to frataxin deficiency and mitofusin-mediated ER stress, which might be responsible for characteristic cellular features of the disease and also suggests ER stress as a therapeutic target.

Optimal material resource planning is crucial to run safe and cost-efficient hospital services. In this article, we investigate a real problem in hospitals, motivated by an environmental and economically inefficient use of disposable, single-use, endoscopes. We develop a mathematical model and create a decision support tool to determine when reusable, multi-use, bronchoscopes should be sent for inspection including information to what extent single-use bronchoscopes can cover the remaining demand. The results show that the proposed approach can contain operational costs which consist of costs for buying single-use devices, inspection costs and reprocessing costs, i.e., sterilization of reusable devices. Our tool can assist hospitals to predict when reusable bronchoscopes should undergo inspection and whether the current inventory of reusable devices is sufficient to cover the demand. Finally, we evaluate the impact of variation in demand on total costs. [ABSTRACT FROM AUTHOR]