Structural basis of TFIIH activation for nucleotide excision repair.

Nucleotide excision repair (NER) is the major DNA repair pathway that removes UV-induced and bulky DNA lesions. There is currently no structure of NER intermediates, which form around the large multisubunit transcription factor IIH (TFIIH). Here we report the cryo-EM structure of an NER intermediate containing TFIIH and the NER factor XPA. Compared to its transcription conformation, the TFIIH structure is rearranged such that its ATPase subunits XPB and XPD bind double- and single-stranded DNA, consistent with their translocase and helicase activities, respectively. XPA releases the inhibitory kinase module of TFIIH, displaces a 'plug' element from the DNA-binding pore in XPD, and together with the NER factor XPG stimulates XPD activity. Our results explain how TFIIH is switched from a transcription to a repair factor, and provide the basis for a mechanistic analysis of the NER pathway

Investigating the release of coprecipitated uranium from iron oxides

The removal of uranium (VI) from zerovalent iron permeable reactive barriers and wetlands can be explained by its association with iron oxides. The long term stability of immobilized U is yet to be addressed. The present study investigates the remobilization of U(VI) from iron oxides via diverse reaction pathways (acidification, reduction, complex formation). Prior, uranium coprecipitation experiments were conducted under various conditions. The addition of various amounts of a pH-shifting agents (pyrite), an iron complexing agent (EDTA) or iron (III) reduction agent (TiCl3) yielded in uranium remobilization, concentrations above the US EPA allowedmaximum contaminant level(MCL=30 æg/l). This study demonstrates that U(VI) release in nature strongly depends on the conditions and the mechanism of its fixation by geological materials.researc

Structural basis of RNA processing by human mitochondrial RNase P

Human mitochondrial transcripts contain messenger and ribosomal RNAs flanked by transfer RNAs (tRNAs), which are excised by mitochondrial RNase (mtRNase) P and Z to liberate all RNA species. In contrast to nuclear or bacterial RNase P, mtRNase P is not a ribozyme but comprises three protein subunits that carry out RNA cleavage and methylation by unknown mechanisms. Here, we present the cryo-EM structure of human mtRNase P bound to precursor tRNA, which reveals a unique mechanism of substrate recognition and processing. Subunits TRMT10C and SDR5C1 form a subcomplex that binds conserved mitochondrial tRNA elements, including the anticodon loop, and positions the tRNA for methylation. The endonuclease PRORP is recruited and activated through interactions with its PPR and nuclease domains to ensure precise pre-tRNA cleavage. The structure provides the molecular basis for the first step of RNA processing in human mitochondria

Job satisfaction and career development of men in nursing

Background: Men are a growing minority in nursing. A deeper understanding of their motivation, career paths, and job satisfaction is needed to target recruitment and retention efforts. Currently the largest male cohort entering nursing is by second career. This study examines if job satisfaction differs for men who choose nursing as their first choice of career, as a convenience after exploring other options, or as a second career. Methods: This mixed-method study examined the relationship between men entering nursing by choice, convenience or second career and their resultant job satisfaction. A convenience sample of 238 men answered an online survey. Entry path, current practice position, and future career plans in five years was by self report. Job satisfaction was measured using Lester’s instrument. Results: Second career men were significantly more likely to work in specialty practice and more likely to aspire to advanced practice than those entering by choice or convenience early in their careers. No significant differences in job satisfaction were found between entry paths. Conclusions: These data raise provocative issues relating to recruiting men and the impact of men on the profession of nursing. Despite the steady but slow growth in the percent of men in nursing, its image remains overwhelmingly female. This survey does not identify substantial differences in job satisfaction between men who enter through different career paths or substantial differences with other studies of male and female nurses

Molecular interactions of FG nucleoporin repeats at high resolution

Proteins that contain repeat phenylalanine-glycine (FG) residues phase separate into oncogenic transcription factor condensates in malignant leukaemias, form the permeability barrier of the nuclear pore complex and mislocalize in neurodegenerative diseases. Insights into the molecular interactions of FG-repeat nucleoporins have, however, remained largely elusive. Using a combination of NMR spectroscopy and cryoelectron microscopy, we have identified uniformly spaced segments of transient β-structure and a stable preformed α-helix recognized by messenger RNA export factors in the FG-repeat domain of human nucleoporin 98 (Nup98). In addition, we have determined at high resolution the molecular organization of reversible FG–FG interactions in amyloid fibrils formed by a highly aggregation-prone segment in Nup98. We have further demonstrated that amyloid-like aggregates of the FG-repeat domain of Nup98 have low stability and are reversible. Our results provide critical insights into the molecular interactions underlying the self-association and phase separation of FG-repeat nucleoporins in physiological and pathological cell activities

Simultaneous computed tomography-guided biopsy and radiofrequency ablation of solitary pulmonary malignancy in high-risk patients

Background: In recent years experience has been accumulated in percutaneous radiofrequency ablation (RFA) of lung malignancies in nonsurgical patients. Objectives: In this study, we retrospectively evaluated a simultaneous diagnostic and therapeutic approach including CT-guided biopsy followed immediately by RFA of solitary malignant pulmonary lesions. Methods: CT-guided transthoracic core needle biopsy of solitary pulmonary lesions suspicious for malignancy was performed and histology was proven based on immediate frozen sections. RFA probes were placed into the pulmonary tumors under CT guidance and the ablation was performed subsequently. The procedure-related morbidity was analyzed. Follow-up included a CT scan and pulmonary function parameters. Results: A total of 33 CT-guided biopsies and subsequent RFA within a single procedure were performed. Morbidity of CT-guided biopsy included pulmonary hemorrhage (24%) and a mild pneumothorax (12%) without need for further interventions. The RFA procedure was not aggravated by the previous biopsy. The rate of pneumothorax requiring chest tube following RFA was 21%. Local tumor control was achieved in 77% with a median follow-up of 12 months. The morbidity of the CT-guided biopsy had no statistical impact on the local recurrence rate. Conclusions: The simultaneous diagnostic and therapeutic approach including CT-guided biopsy followed immediately by RFA of solitary malignant pulmonary lesions is a safe procedure. The potential of this combined approach is to avoid unnecessary therapies and to perform adequate therapies based on histology. Taking the local control rate into account, this approach should only be performed in those patients who are unable to undergo or who refuse surgery. Copyright (C) 2012 S. Karger AG, Base

The clinical drug candidate anle138b binds in a cavity of lipidic alpha-synuclein fibrils

Aggregation of amyloidogenic proteins is a characteristic of multiple neuro- degenerative diseases. Atomic resolution of small molecule binding to such pathological protein aggregates is of interest for the development of ther- apeutics and diagnostics. Here we investigate the interaction between α- synuclein fibrils and anle138b, a clinical drug candidate for disease modifying therapy in neurodegeneration and a promising scaffold for positron emission tomography tracer design. We used nuclear magnetic resonance spectroscopy and the cryogenic electron microscopy structure of α-synuclein fibrils grown in the presence of lipids to locate anle138b within a cavity formed between two β-strands. We explored and quantified multiple binding modes of the com- pound in detail using molecular dynamics simulations. Our results reveal stable polar interactions between anle138b and backbone moieties inside the tubular cavity of the fibrils. Such cavities are common in other fibril structures as well

Neutralization of SARS-CoV-2 by highly potent, hyperthermostable, and mutation-tolerant nanobodies

Monoclonal anti-SARS-CoV-2 immunoglobulins represent a treatment option for COVID-19. However, their production in mammalian cells is not scalable to meet the global demand. Single-domain (VHH) antibodies (also called nanobodies) provide an alternative suitable for microbial production. Using alpaca immune libraries against the receptor-binding domain (RBD) of the SARS-CoV-2 Spike protein, we isolated 45 infection-blocking VHH antibodies. These include nanobodies that can withstand 95°C. The most effective VHH antibody neutralizes SARS-CoV-2 at 17–50 pM concentration (0.2–0.7 µg per liter), binds the open and closed states of the Spike, and shows a tight RBD interaction in the X-ray and cryo-EM structures. The best VHH trimers neutralize even at 40 ng per liter. We constructed nanobody tandems and identified nanobody monomers that tolerate the K417N/T, E484K, N501Y, and L452R immune-escape mutations found in the Alpha, Beta, Gamma, Epsilon, Iota, and Delta/Kappa lineages. We also demonstrate neutralization of the Beta strain at low-picomolar VHH concentrations. We further discovered VHH antibodies that enforce native folding of the RBD in the E. coli cytosol, where its folding normally fails. Such “fold-promoting” nanobodies may allow for simplified production of vaccines and their adaptation to viral escape-mutations