Chloroplast import motor subunits FtsHi1 and FtsHi2 are located on opposite sides of the inner envelope membrane.

Protein import into chloroplasts is powered by ATP hydrolysis in the stroma. Establishing the identity and functional mechanism of the stromal ATPase motor that drives import is critical for understanding chloroplast biogenesis. Recently, a complex consisting of Ycf2, FtsHi1, FtsHi2, FtsHi4, FtsHi5, FtsH12, and malate dehydrogenase was shown to be important for chloroplast protein import, and it has been proposed to act as the motor driving protein translocation across the chloroplast envelope into the stroma. To gain further mechanistic understanding of how the motor functions, we performed membrane association and topology analyses on two of its subunits, FtsHi1 and FtsHi2. We isolated cDNA clones encoding FtsHi1 and FtsHi2 preproteins to perform in vitro import experiments in order to determine the exact size of each mature protein. We also generated antibodies against the C-termini of the proteins, i.e., where their ATPase domains reside. Protease treatments and alkaline and high-salt extractions of chloroplasts with imported and endogenous proteins revealed that FtsHi1 is an integral membrane protein with its C-terminal portion located in the intermembrane space of the envelope, not the stroma, whereas FtsHi2 is a soluble protein in the stroma. We further complemented an FtsHi1-knockout mutant with a C-terminally tagged FtsHi1 and obtained identical results for topological analyses. Our data indicate that the model of a single membrane-anchored pulling motor at the stromal side of the inner membrane needs to be revised and suggest that the Ycf2-FtsHi complex may have additional functions.

Hemorrhage risk prediction after computed tomography-guided lung biopsy: Clinical parameters and quantitative pulmonary vascular analysis.

BACKGROUND/PURPOSE: We evaluated the utility of combining quantitative pulmonary vasculature measures with clinical factors for predicting pulmonary hemorrhage after computed tomography (CT)-guided lung biopsy. METHODS: Patients who underwent CT-guided lung biopsy were retrospectively included in this study. Clinical and radiographic vasculature variables were evaluated as predictors of pulmonary hemorrhage. The radiographic pulmonary vascular analysis included vessel count, density, diameter, and area, and also blood volume in small vessels with a cross-sectional area ≤5 mm2 (BV5) and total blood vessel volume (TBV) in the lungs. Univariate and multivariate logistic regressions were used to identify the independent risk factors of higher-grade pulmonary hemorrhage and establish the prediction model presented as a nomogram. RESULTS: The study included 126 patients; discovery cohort n = 103, and validation cohort n = 23. All pulmonary hemorrhage, higher-grade (grade ≥2) pulmonary hemorrhage, and hemoptysis occurred in 42.9%, 15.9%, and 3.2% of patients who underwent CT-guided lung biopsies. In the discovery cohort, patients with larger lesion depth (p = 0.013), higher vessel density (p = 0.033), and higher BV5 (p = 0.039) were more likely to experience higher-grade hemorrhage. The nomogram prediction model for higher-grade hemorrhage built by the discovery cohort showed similar performance in the validation cohort. CONCLUSIONS: Higher-grade pulmonary hemorrhage may occur after CT-guided lung biopsy. Lesion depth, vessel density, and BV5 are independent risk factors for higher-grade pulmonary hemorrhage. Nomograms integrating clinical parameters and radiographic pulmonary vasculature measures offer enhanced capability for assessing hemorrhage risk following CT-guided lung biopsy, thereby facilitating improved patient clinical care.

New Generation of Benzidine Wide-Gap Conjugated Materials: Solution Processing a Classic Evaporated Scaffold.

Carbazole- and fluorene-substituted benzidine blocks have been functionalized with two different solubilizing pendant groups, in order to enhance the material's solubility in greener solvents. Preserving the optical and electrochemical properties, the aromatic function and substitution showed an important influence on the solvent affinity, achieving concentrations up to 150 mg/mL in o-xylenes for the glycol-containing materials and decent solubility in alcohols for the compounds functionalized with ionic chains. The latter solution proved to be ideal for the preparation of luminescence slot-die coating film on top of flexible-substrates up to 33 cm×2 cm. As a proof of concept, the materials have been implemented in different organic electronic devices, highlighting the low turn-on voltage (4 V) presented by organic light-emitting diodes (OLEDs), which is comparable with vacuum-processed devices. A structure-solubility relationship and a synthetic strategy are disentangled in this manuscript to tailor organic semiconductors and adapt their solubility towards the desired solvent and application.

First insights on value-based healthcare of elders using ICHOM older person standard set reporting.

BACKGROUND: Clinical guidelines for specific conditions fragment care provision for elders. The International Consortium for Health Outcomes Measurement (ICHOM) has developed a global standard set of outcome measures for comprehensive assessment of older persons. The goal of this study was to report value-based health metrics in Taiwan using this ICHOM toolset. METHODS: The cross-sectional study of baseline data excerpted from a prospective longitudinal cohort, which recruited people ≥65 years old with ≥3 chronic medical conditions between July and December 2018. All participants received measurements of physical performance, anthropometric characteristics, health-related behaviors, Charlson Comorbidity Index, and Montreal Cognitive Assessment. The ICHOM toolset comprises three tiers: 1 includes frailty and having chosen a preferred place of death; 2 includes polypharmacy, falls, and participation in decision-making; and 3 includes loneliness, activities of daily living, pain, depression, and walking speed. These items were converted into a 0-10 point value-based healthcare score, with high value-based health status defined as ≥8/10 points. RESULTS: Frequencies of individual ICHOM indicators were: frail 11.7%, chose preferred place of death 14.4%, polypharmacy 31.5%, fell 17.1%, participated in decision-making 81.6%, loneliness 26.8%, limited activities of daily living 22.4%, pain 10.4%, depressed mood 13.0%, and slowness 38.5%. People with high disease burden (OR 0.40, 95% CI 0.21-0.76, p = 0.005) or cognitive impairment (OR 0.49, 95%CI 0.27-0.87, p = 0.014) were less likely to have high value-based healthcare status. CONCLUSIONS: The ICHOM Standard Set Older Person health outcome measures provide an opportunity to shift from a disease-centric medical paradigm to whole person-focused goals. This study identified advanced age, chronic disease burden and cognitive impairment as important barriers to achieving high value-based healthcare status.

Protocadherin 10 suppresses tumorigenesis and metastasis in colorectal cancer and its genetic loss predicts adverse prognosis.

Protocadherin 10 (PCDH10), a novel tumor suppressor gene in human cancers, is located in a common deleted region at chromosome 4q28 in colorectal cancer (CRC). This study aimed to ascertain the genetic loss of PCDH10 and its clinical relevance in CRC and to explore the tumor suppressor function of PCDH10. The genetic deletion of PCDH10 was determined in 171 pairs of primary tumors and corresponding normal mucosae by loss of heterozygosity study. In total, 53 carcinomas were positive for allelic loss of PCDH10. The genetic aberration was significantly associated with tumor progression and distant metastasis (p = 0.021 and p = 0.018, respectively) and was an independent predictor of poor survival for CRC patients (p = 0.005). Expression of PCDH10 gene was silenced or markedly down-regulated in all of 12 CRC cell lines tested and in 41 of 53 colorectal carcinomas compared with their matched normal mucosae. Ectopic expression of PCDH10 suppressed cancer cell proliferation, anchorage-independent growth, migration and invasion in vitro. Subcutaneous injection of PCDH10-expressing CRC cells into SCID mice revealed the reduction of tumor growth compared with that observed in mock-inoculated mice. Furthermore, through intrasplenic implantation, the re-expression of PCDH10 in silenced cells restrained liver metastasis and improved survival in SCID mice. In conclusion, PCDH10 is a pivotal tumor suppressor in CRC, and the loss of its function promotes not only tumor progression but also liver metastasis. In addition, the genetic deletion of PCDH10 represents an adverse prognostic marker for the survival of patients with CRC.

Direct C-H electrophilic borylation with (C6F5)2B-NTf2 to generate B-N dibenzo[a,h]pyrenes.

The borylation of aryl substituted pyridines is an effective way of preparing B-N doped conjugated organic frameworks. Trihaloborane Lewis acids are often employed for this protocol, and may require further functionalization to replace the remaining halides on boron. We report a new, fully characterized, electrophilic borylating agent, (C6F5)2B(κ2-NTf2), that smoothly incorporates a -B(C6F5)2 unit into the model substrate 2-phenylpyridine. To demonstrate its utility in preparing more complex B-N doped structures, we use it to prepare seven examples of the 6a,13a-diaza-7,14-dibora-dibenzo[a,h]pyrene framework, with substituents of varying donor properties. The structural, redox, and photophysical properties of this new family of B-N doped polycyclic hydrocarbon compounds were probed experimentally and computationally.

Ruthenium polyhydrides supported by rigid PCP pincer ligands: dynamic behaviour and reactions with CO2.

Two rigid ß-elimination immune PCcarbeneP pincer ligands, differing in their electron donor properties by variation of the substitution pattern on the aromatic linker arms, were complexed to ruthenium to form the dichlorides LRRuCl2 (R = H or NMe2). These compounds were converted to hydrido chlorides by treatment with dihydrogen (H2) and a base. By converting to tert-butoxide derivatives in situ under an atmosphere of H2, the poly hydride PCalkylP complexes LHRRu(H)3 compounds were generated. In these complexes, H2 has added across the RuC bond in the PCcarbeneP starting materials. The polyhydrides are dynamic in solution and extensive NMR studies helped to elucidate the speciation and fluxional processes operative in this dynamic system. The polyhydride complexes react rapidly with CO2 to give the PCcarbeneP formato hydride complexes LRRu(H)-κ2-O2CH. For R = H, the 1,2-hydride shift from the anchoring alkyl of the PCalkylP carbon to the metal is reversible, but for R = NMe2 it is irreversible. The CO2 incorporated into the formato ligand of these compounds exchanges with free CO2via a bimolecular mechanism that is more rapid for R = NMe2 than for R = H; plausible explanations for this observation are proffered. Experiments designed to evaluate the efficacy of the R = NMe2 formato hydride complex as a catalyst precursor for CO2 hydrogenation to formate salts reveal poor performance in comparison to state-of-the-art ruthenium-based catalysts. This is due primarily to the precipitation of a dimeric µ-κ2-κ1-CO3 carbonate complex that is not an active catalyst for the reaction.

Oversampling to overcome overfitting: exploring the relationship between data set composition, molecular descriptors, and predictive modeling methods.

The traditional biological assay is very time-consuming, and thus the ability to quickly screen large numbers of compounds against a specific biological target is appealing. To speed up the biological evaluation of compounds, high-throughput screening is widely used in the fields of biomedical, biological information, and drug discovery. The research presented in this study focuses on the use of support vector machines, a machine learning method, various classes of molecular descriptors, and different sampling techniques to overcome overfitting to classify compounds for cytotoxicity with respect to the Jurkat cell line. The cell cytotoxicity data set is imbalanced (a few active compounds and very many inactive compounds), and the ability of the predictive modeling methods is adversely affected in these situations. Commonly imbalanced data sets are overfit with respect to the dominant classified end point; in this study the models routinely overfit toward inactive (noncytotoxic) compounds when the imbalance was substantial. Support vector machine (SVM) models were used to probe the proficiency of different classes of molecular descriptors and oversampling ratios. The SVM models were constructed from 4D-FPs, MOE (1D, 2D, and 21/2D), noNP+MOE, and CATS2D trial descriptors pools and compared to the predictive abilities of CATS2D-based random forest models. Compared to previous results in the literature, the SVM models built from oversampled data sets exhibited better predictive abilities for the training and external test sets.

Template-based de novo design for type II kinase inhibitors and its extented application to acetylcholinesterase inhibitors.

There is a compelling need to discover type II inhibitors targeting the unique DFG-out inactive kinase conformation since they are likely to possess greater potency and selectivity relative to traditional type I inhibitors. Using a known inhibitor, such as a currently available and approved drug or inhibitor, as a template to design new drugs via computational de novo design is helpful when working with known ligand-receptor interactions. This study proposes a new template-based de novo design protocol to discover new inhibitors that preserve and also optimize the binding interactions of the type II kinase template. First, sorafenib (Nexavar) and nilotinib (Tasigna), two type II inhibitors with different ligand-receptor interactions, were selected as the template compounds. The five-step protocol can reassemble each drug from a large fragment library. Our procedure demonstrates that the selected template compounds can be successfully reassembled while the key ligand-receptor interactions are preserved. Furthermore, to demonstrate that the algorithm is able to construct more potent compounds, we considered kinase inhibitors and other protein dataset, acetylcholinesterase (AChE) inhibitors. The de novo optimization was initiated using a template compound possessing a less than optimal activity from a series of aminoisoquinoline and TAK-285 inhibiting type II kinases, and E2020 derivatives inhibiting AChE respectively. Three compounds with greater potency than the template compound were discovered that were also included in the original congeneric series. This template-based lead optimization protocol with the fragment library can help to design compounds with preferred binding interactions of known inhibitors automatically and further optimize the compounds in the binding pockets.

Electrocatalyst decomposition pathways: torsional strain in a second sphere proton relay shuts off CO2RR in a Re(2,2'-bipyridyl)(CO)3X type electrocatalyst.

Group 7 tris(carbonyl) bipyridine complexes have been well explored as important CO2 reduction reaction (CO2RR) electrocatalysts and now represent an excellent platform for catalyst design. Recent synthetic focus has been on the installation of proton sources/relays within the primary/secondary coordination sphere. These proton sources have been implicated in directly assisting catalysis by acting as shuttles for proton transfer or through the stabilization of transition states through hydrogen bonding. Herein, we report a new ligand system for CO2RR electrocatalysts, which features an aryl amine appended to a quinoline-bipyridine core. While the geometrical arrangement of the aryl amine seems amenable to assisting CO2RR electrocatalysis, we find, through spectroelectrochemical and chemical reduction studies, the torsional strain imposed on the ligand induces a structural reorganization through loss of a hydrogen atom radical. This new complex, which utilizes the anionic nitrogen as a donor atom, and other Re complexes with the same coordination motif, have been found to be entirely inactive for CO2RR. Subsequent reduction yields hydrogenation of the complex through dearomatization of the quinoline backbone concomitant with decomposition products. While the electrocatalytic capability of the reported complexes is moderate, the study represents an important investigation into the deactivation of CO2RR electrocatalysts as a consequence of typical proton shuttle moieties and guides future ligand design by highlighting an oft overlooked structural parameter.

Effects of incorporating multidomain interventions into integrated primary care on quality of life: a randomised controlled trial.

BACKGROUND: Integrating primary prevention into care pathways for older adults is a core strategy of healthy ageing, but evidence remains limited. We aimed to determine whether incorporating a multidomain intervention into primary health care could improve standard value-based health outcomes and quality of life. METHODS: For this Taiwan Integrated Geriatric Care (TIGER) study, a pragmatic randomised controlled trial, we recruited community-dwelling outpatients aged 65 years or older with at least three chronic medical conditions. We excluded people with malignancies undergoing chemotherapy, people with a life expectancy of less than 12 months, or people who were insufficiently able to communicate with study staff. Participants were randomly assigned (1:1) to usual care or to the integrated multidomain intervention using block randomisation. The integrated multidomain intervention entailed 16 2-h sessions per year, comprising communal physical exercise, cognitive training, nutrition and disease education, plus individualised treatment by specialists in integrated geriatric care. The primary outcome was changes from baseline quality of life, based on 36-item Short Form Health Survey (SF-36) scores, at 3, 6, 9, and 12 months. Intervention effects were analysed per protocol using a generalised linear mixed model. This trial is registered with ClinicalTrials.gov, NCT03528005. FINDINGS: Between June 25, 2018, and Feb 15, 2019, 628 participants were screened, of whom 398 were assigned to the integrated multidomain intervention (n=199) or to usual care (n=199). 335 (84%) participants completed the 12-month follow-up. Compared with the usual care group, the integrated multidomain intervention group had significantly higher mean SF-36 physical component scores across all timepoints (overall difference 0·8, 95% CI 0·2-1·5; p=0·010), but differences at 3, 6, 9, and 12 months did not reach statistical significance. The SF-36 mental component scores did not differ significantly overall, but were significantly higher in the integrated multidomain intervention group at the 12-month follow-up (55·3 [SD 7·6] vs 57·2 [7·0]; p=0·019). No serious adverse events occurred. INTERPRETATION: Incorporating multidomain interventions into integrated health care improved quality of life. Our standardised protocol is amenable to inclusion in policies to promote value-based care and healthy ageing. FUNDING: National Health Research Institutes, Taiwan, and Ministry of Science and Technology, Taiwan.

Effect of Wind on Horizontal Displacement of Fatal Fall from a Height.

The horizontal displacement of the human body resulting from fatal fall from a height is an important variable commonly used to inversely determine the cause or identify other forensic aspects of the fall. When examining the horizontal displacement, the wind effect is generally ignored. This technical note reports analytical modeling of the falling process, utilizing previous measurements of wind force acting on the human body, for determining the functional relationship between the wind speed and the horizontal displacement in the falling process. The result reveals that it does not take extremely rare wind conditions to cause a considerable shift of the human body, highlighting the importance to consider wind as a factor in investigations of fatal falls.

Forensic Application of Analytical Mechanics in Fatal Fall From a Height.

Fatal fall from a height initiated with a swinging motion on the sagittal plane of the victim's body is examined. Method of analytical mechanics is adopted to establish the functional relations between the swinging angle of the body and the force needed to sustain the body until release, and the amount of the horizontal displacement that would occur after release. The analysis is applied to support an expert opinion presented in a criminal court in which two accused were convicted of murder for dropping a female victim from a bridge. The results indicate that it is unlikely for the body to locate as recorded by the police investigation report in the case. This study demonstrates that a straightforward mechanical analysis can be utilized to examine uncommon falling scenarios.

Design of accurate predictors for DNA-binding sites in proteins using hybrid SVM-PSSM method.

In this paper, we investigate the design of accurate predictors for DNA-binding sites in proteins from amino acid sequences. As a result, we propose a hybrid method using support vector machine (SVM) in conjunction with evolutionary information of amino acid sequences in terms of their position-specific scoring matrices (PSSMs) for prediction of DNA-binding sites. Considering the numbers of binding and non-binding residues in proteins are significantly unequal, two additional weights as well as SVM parameters are analyzed and adopted to maximize net prediction (NP, an average of sensitivity and specificity) accuracy. To evaluate the generalization ability of the proposed method SVM-PSSM, a DNA-binding dataset PDC-59 consisting of 59 protein chains with low sequence identity on each other is additionally established. The SVM-based method using the same six-fold cross-validation procedure and PSSM features has NP=80.15% for the training dataset PDNA-62 and NP=69.54% for the test dataset PDC-59, which are much better than the existing neural network-based method by increasing the NP values for training and test accuracies up to 13.45% and 16.53%, respectively. Simulation results reveal that SVM-PSSM performs well in predicting DNA-binding sites of novel proteins from amino acid sequences.

Microwave Assisted Reduction of Pt-Catalyst by N-Phenyl-p-Phenylenediamine for Proton Exchange Membrane Fuel Cells.

The presence of N-phenyl-p-phenylenediamine (PPDA: a dimer of aniline) during microwave (MW) irradiation can significantly improve Pt-loading on the XC72 carbon matrix as a catalyst support of proton exchange membrane fuel cells (PEMFCs). PPDA is converted to an emeraldine base state during MW-assisted redox reaction, which is characterized by both FTIR and Raman spectra. The increased degree of conjugation from the formation of quinone-state of PPDA is confirmed by UV-VIS spectra. TEM micrographs and residue weights obtained from the TGA thermograms illustrate the particle size and Pt-loading percent of Pt nanoparticles (NPs) after MW irradiation, respectively. X-ray diffraction patterns indicate Pt NPs are successfully loaded on XC72 by MW irradiation corresponding to hydrothermal method. The single cell performance demonstrates an increasing power and maximum current density when Pt-catalyst of membrane exchanged assembly (MEA) is prepared by MW-assisted reduction in the presence of PPDA.

Entropic stabilization of a deubiquitinase provides conformational plasticity and slow unfolding kinetics beneficial for functioning on the proteasome.

Human ubiquitin C-terminal hydrolyase UCH-L5 is a topologically knotted deubiquitinase that is activated upon binding to the proteasome subunit Rpn13. The length of its intrinsically disordered cross-over loop is essential for substrate recognition. Here, we showed that the catalytic domain of UCH-L5 exhibits higher equilibrium folding stability with an unfolding rate on the scale of 10-8 s-1, over four orders of magnitudes slower than its paralogs, namely UCH-L1 and -L3, which have shorter cross-over loops. NMR relaxation dynamics analysis confirmed the intrinsic disorder of the cross-over loop. Hydrogen deuterium exchange analysis further revealed a positive correlation between the length of the cross-over loop and the degree of local fluctuations, despite UCH-L5 being thermodynamically and kinetically more stable than the shorter UCHs. Considering the role of UCH-L5 in removing K48-linked ubiquitin to prevent proteasomal degradation of ubiquitinated substrates, our findings offered mechanistic insights into the evolution of UCH-L5. Compared to its paralogs, it is entropically stabilized to withstand mechanical unfolding by the proteasome while maintaining structural plasticity. It can therefore accommodate a broad range of substrate geometries at the cost of unfavourable entropic loss.

Random-Coil Behavior of Chemically Denatured Topologically Knotted Proteins Revealed by Small-Angle X-ray Scattering.

Recent studies on the mechanisms by which topologically knotted proteins attain their natively knotted structures have intrigued theoretical and experimental biophysicists. Of particular interest is the finding that YibK and YbeA, two small trefoil knotted proteins, remain topologically knotted in their chemically denatured states. Using small-angle X-ray scattering (SAXS), we examine whether these chemically denatured knotted proteins are different from typical random coils. By revisiting the scaling law of radius of gyration (Rg) as a function of polypeptide chain length for chemically denatured proteins and natively folded proteins, we find that the chemically denatured knotted proteins in fact follow the same random coil-like behavior, suggesting that the formation of topological protein knots do not necessarily require global compaction while the loosely knotted polypeptide chains are capable of maintaining the correct chirality without defined secondary or tertiary structures.

Tumor cells require thymidylate kinase to prevent dUTP incorporation during DNA repair.

The synthesis of dTDP is unique because there is a requirement for thymidylate kinase (TMPK). All other dNDPs including dUDP are directly produced by ribonucleotide reductase (RNR). We report the binding of TMPK and RNR at sites of DNA damage. In tumor cells, when TMPK function is blocked, dUTP is incorporated during DNA double-strand break (DSB) repair. Disrupting RNR recruitment to damage sites or reducing the expression of the R2 subunit of RNR prevents the impairment of DNA repair by TMPK intervention, indicating that RNR contributes to dUTP incorporation during DSB repair. We identified a cell-permeable nontoxic inhibitor of TMPK that sensitizes tumor cells to doxorubicin in vitro and in vivo, suggesting its potential as a therapeutic option.