Characterization of heterogeneity in nanodisc samples using Feret signatures.

Nanodiscs have become a popular tool in structure determination of membrane proteins using cryogenic electron microscopy and single particle analysis. However, the structure determination of small membrane proteins remains challenging. When the embedded protein is in the same size range as the nanodisc, the nanodisc can significantly contribute to the alignment and classification during the structure determination process. In those cases, it is crucial to minimize the heterogeneity in the nanodisc preparations to assure maximum accuracy in the classification and alignment steps of single particle analysis. Here, we introduce a new in-silico method for the characterization of nanodisc samples that is based on analyzing the Feret diameter distribution of their particle projection as imaged in the electron microscope. We validated the method with comprehensive simulation studies and show that Feret signatures can detect subtle differences in nanodisc morphologies and composition that might otherwise go unnoticed. We used the method to identify a specific biochemical nanodisc preparation with low size variations, allowing us to obtain a structure of the 23-kDa single-span membrane protein Bcl-xL while embedded in a nanodisc. Feret signature analysis can steer experimental data collection strategies, allowing more efficient use of high-end data collection hardware, as well as image analysis investments in studies where nanodiscs significantly contribute to the total volume of the full molecular species.

Environmental and human-health risks of As in soils with abnormal arsenic levels located in irrigated agricultural areas of Paracatu (MG), Brazil.

The municipality of Paracatu (Brazil) is notorious for its large irrigated agricultural area and by abnormal arsenic (As) levels in selected soils of the region. Concerns regarding As exposure via ingestion of water and food are frequent, yet little is known about the behavior of arsenic in irrigated agricultural soils, as well as on As bioaccessibility/bioavailability in agroecosystems of this region. This work evaluated total and available As in agricultural soils cultivated under irrigation and in soils under native vegetation in Paracatu. We also assessed reactive arsenic fractions and As bioaccessibility in the soil, as well as arsenic levels in plant shoots to estimate As risks in these agroecosystems. Soil (different depths) and plant tissue samples were collected in 6 irrigated agricultural areas (CA1 to CA6) and 4 reference areas (RA1 to RA4). Total soil-As did not differ between soil depths, reinforcing that the source of As in agricultural soils is natural. This was evident when counterpointing arsenic and phosphorus contents at different soil depths, as both accumulate on the surface of oxidic soils when added to agroecosystems by anthropogenic routes (e.g., phosphate fertilization for P and irrigation for As). Available As levels in soils and plants were very small (below detection limit). Furthermore, all soils presented very low oral As bioaccessibility. Our findings revealed that the irrigated soils are not As polluted due to the low enrichment and accumulation of arsenic, as well as the prevalence of low ecological risks. There is no non-carcinogenic risk for the local population, except for children in RA2. The estimated carcinogenic risk for children followed the order RA2 > CA3 > CA4 > RA3 > CA2, and for adults, RA2 > CA3. Ultimately, the strategy of comparing the behavior of P and As in the soils of this study proved to be efficient in showing that there are no major risks to humans and the environment in the investigated area. However, periodic monitoring of As bioavailability in these areas is recommended.

Structural characterization of the RH1-LZI tandem of JIP3/4 highlights RH1 domains as a cytoskeletal motor-binding motif.

JIP3 and JIP4 (JNK-interacting proteins 3 and 4) are adaptors for cargo recruitment by dynein/dynactin and kinesin1 motors. Both are dimers that are stabilised by two sections of leucine zipper coiled coils. The N-terminal Leucine Zipper I (LZI) belongs to a section that binds dynein-DLIC and kinesin1-KHC, whilst the medial Leucine Zipper II (LZII) binds dynactin-p150glued and kinesin1-KLC. Structural data is available for the LZII, but the LZI section is still uncharacterized. Here we characterize the N-terminal part of JIP3/4 which consists of an RH1 (RILP homology 1) domain followed by the LZI coiled coil using bioinformatical, biophysical and structural approaches. The RH1-LZI tandem of JIP3 associates as a high affinity homodimer exhibiting elongated alpha-helical fold. 3D homology modelling of the RH1-LZI tandem reveals that the kinesin1-KHC binding site mainly overlaps with the RH1 domain. A sequence comparison search indicates that only one other protein family has RH1 domains similar to those of JIP3/4, the RILP (Rab-interacting lysosomal protein) family which consists of adaptor proteins linking Rab GTPases to cytoskeletal motors. RILPL2 is recruited through its RH1 domain by the myosin 5a motor. Here, we showed that the RH1 domain of JIP3 also interacts with myosin 5 A in vitro, highlighting JIP3/4 as possible myosin 5a adaptors. Finally, we propose that JIP3/4 and RILP family members define a unique RH1/RH2-architecture adaptor superfamily linking cytoskeletal motors and Rab GTPases.

Characterization of the binding mode of JNK-interacting protein 1 (JIP1) to kinesin-light chain 1 (KLC1).

JIP1 was first identified as scaffold protein for the MAP kinase JNK and is a cargo protein for the kinesin1 molecular motor. JIP1 plays significant and broad roles in neurons, mainly as a regulator of kinesin1-dependent transport, and is associated with human pathologies such as cancer and Alzheimer disease. JIP1 is specifically recruited by the kinesin-light chain 1 (KLC1) of kinesin1, but the details of this interaction are not yet fully elucidated. Here, using calorimetry, we extensively biochemically characterized the interaction between KLC1 and JIP1. Using various truncated fragments of the tetratricopeptide repeat (TPR) domain of KLC1, we narrowed down its JIP1-binding region and identified seven KLC1 residues critical for JIP1 binding. These isothermal titration calorimetry (ITC)-based binding data enabled us to footprint the JIP1-binding site on KLC1-TPR. This footprint was used to uncover the structural basis for the marginal inhibition of JIP1 binding by the autoinhibitory LFP-acidic motif of KLC1, as well as for the competition between JIP1 and another cargo protein of kinesin1, the W-acidic motif-containing alcadein-α. Also, we examined the role of each of these critical residues of KLC1 for JIP1 binding in light of the previously reported crystal structure of the KLC1-TPR:JIP1 complex. Finally, sequence search in eukaryotic genomes identified several proteins, among which is SH2D6, that exhibit a motif similar to the KLC1-binding motif of JIP1. Overall, our extensive biochemical characterization of the KLC:JIP1 interaction, as well as identification of potential KLC1-binding partners, improves the understanding of how this growing family of cargos is recruited to kinesin1 by KLC1.

Correction: Structural plasticity of the N-terminal capping helix of the TPR domain of kinesin light chain.

[This corrects the article DOI: 10.1371/journal.pone.0186354.].

Structural plasticity of the N-terminal capping helix of the TPR domain of kinesin light chain.

Kinesin1 plays a major role in neuronal transport by recruiting many different cargos through its kinesin light chain (KLC). Various structurally unrelated cargos interact with the conserved tetratricopeptide repeat (TPR) domain of KLC. The N-terminal capping helix of the TPR domain exhibits an atypical sequence and structural features that may contribute to the versatility of the TPR domain to bind different cargos. We determined crystal structures of the TPR domain of both KLC1 and KLC2 encompassing the N-terminal capping helix and show that this helix exhibits two distinct and defined orientations relative to the rest of the TPR domain. Such a difference in orientation gives rise, at the N-terminal part of the groove, to the formation of one hydrophobic pocket, as well as to electrostatic variations at the groove surface. We present a comprehensive structural analysis of available KLC1/2-TPR domain structures that highlights that ligand binding into the groove can be specific of one or the other N-terminal capping helix orientations. Further, structural analysis reveals that the N-terminal capping helix is always involved in crystal packing contacts, especially in a TPR1:TPR1' contact which highlights its propensity to be a protein-protein interaction site. Together, these results underline that the structural plasticity of the N-terminal capping helix might represent a structural determinant for TPR domain structural versatility in cargo binding.

Applicability and Effectiveness of Closed Reduction of Nasal Fractures under Local Anesthesia

Introduction: A significant portion of patients treated in emergency departments have nasal fracture. It is important that the otolaryngologist know how to treat such damage. Objectives To evaluate the effectiveness of nasal fracture reduction under local anesthesia and tolerance to the procedure. Methods: Twenty-four patients treated in the emergency department with closed reduction under local anesthesia were prospectively followed. Epidemiologic information and data regarding pain and complications during the management were noted. The degree of satisfaction was researched by visual analog scale. Results: The majority of patients were male (75%), and the most common cause of injury was motor vehicle accident. We found a significant association between time to reduction and referred pain during the procedure. In patients in whom the procedure was delayed (over 3 days), there was less pain, and those who bled during the procedure had a shorter average time to reduction than the group of patients who did not bleed. Most patients were very satisfied, with more than 95% of these willing to undergo the same process again, if necessary. Conclusions: The closed approach in the clinic under local anesthesia was effective and safe in restoration of the nose...

Applicability and Effectiveness of Closed Reduction of Nasal Fractures under Local Anesthesia.

Introduction A significant portion of patients treated in emergency departments have nasal fracture. It is important that the otolaryngologist know how to treat such damage. Objectives To evaluate the effectiveness of nasal fracture reduction under local anesthesia and tolerance to the procedure. Methods Twenty-four patients treated in the emergency department with closed reduction under local anesthesia were prospectively followed. Epidemiologic information and data regarding pain and complications during the management were noted. The degree of satisfaction was researched by visual analog scale. Results The majority of patients were male (75%), and the most common cause of injury was motor vehicle accident. We found a significant association between time to reduction and referred pain during the procedure. In patients in whom the procedure was delayed (over 3 days), there was less pain, and those who bled during the procedure had a shorter average time to reduction than the group of patients who did not bleed. Most patients were very satisfied, with more than 95% of these willing to undergo the same process again, if necessary. Conclusions The closed approach in the clinic under local anesthesia was effective and safe in restoration of the nose.

Desempenho de caprinos inteiros alimentados com diferentes quantidades de aguapé / Evaluate of performance of caprines whole feed with water hyacinth

Objetivo - Avaliar o desempenho ponderal de caprinos inteiros alimentados com níveis crescentes de aguapé na forma de volumoso.O aguapé (Eichhornia crassipes) é uma planta aquática de propagação agressiva e com potencial de uso bastante diversificado. Métodos - O experimento avaliou o efeito da adição de 0%, 20% e 40% de aguapé como volumoso sobre o ganho de peso de 10 caprinos inteiros da raça Saanen, com idade média de 70 dias, em quadrado latino 3X3 com três fases de 14 dias cada, alternadas por intervalos de cinco dias. O ganho de peso médio para os animais com 0% foi de 107,9 (± 45,5) g/dia; com 20% foi de 81,0 (± 44,2) g/dia; e com 40% foi de 111, 9 (± 51,6) g/dia. Resultados - Na análise dos resultados ficou apurado que os valores obtidos foram estatisticamente semelhantes para os três tratamentos. Conclusão - Não há diferença em adicionar até 40% de aguapé como volumoso na dieta de caprinos.

Objective - The aim of this study was to evaluate the performance of caprines whole feed with crescents levels of the water hyacinth with voluminous. The water hyacinth (Eichhornia crassipes) is an aquatic plant of aggressive propagation and with use potential much diversified. Methods - The experiment evaluated of the addition of 0%, 20% and 40% of water hyacinth as bulky on the earnings of weight of 10 caprine whole of the breed Saanen, with medium age of 70 days, in Latin square 3X3 with three stage of 14 days each, alternate for intervals of five days. The earnings of medium weight for the animal with 0% was of 107,9 (± 45,5) g/day; with 20% it was of 81,0 (±44,2) g/day and with 40% it was of 111,9 (± 51,6) g/day. Results - In the analysis of the experiment the results were similar in all tree treatments. Conclusion - There is no difference to add until 40% of water hyacinth on the caprines diet.

Parâmetros físico-químicos e cromatográficos em aguardentes de cana queimada e não queimada / Physicochemical and chromatographic parameters in sugar cane brandies from burnt and non-burnt cane

Este trabalho teve por objetivo avaliar as concentrações de furfural, álcoois superiores, ésteres, aldeídos, cobre, acidez volátil, metanol e grau alcoólico de aguardentes obtidas de cana, com e sem queima prévia. Foram utilizadas amostras de aguardente artesanal de cana queimada e não queimada, fermentadas com a mesma levedura e destiladas no mesmo destilador; amostras de produtor de aguardente industrial de cana queimada e amostras obtidas de outro produtor artesanal de cana não queimada. As análises físico-químicas e cromatográficas (CG) foram realizadas no Laboratório de Análise Físico-Química de Aguardente - LAFQA/DQI na Universidade Federal de Lavras. As concentrações de furfural apresentaram diferença significativa (P<0,01), entre os grupos de aguardentes artesanais, obtidas de cana-de-açúcar com e sem queima prévia, oriundas de um mesmo processo de produção, com médias de 1,48 mg.100mL-¹ etanol e 0,63 mg.100mL-¹ etanol, respectivamente, embora tenham estado abaixo do limite máximo (5,0 mg.100mL-¹ etanol) permitido. Os demais componentes das aguardentes estudadas não foram afetados significativamente pela queima prévia da cana-de-açúcar.

This work was intended to evaluate the concentrations of furfural, higher alcohols, esters, aldehydes, copper, volatile acidity methanol and alcoholic degree of sugar cane brandies obtained from sugar cane both with and without previous burning. Samples of artisanal brandies from burnt and non-burnt cane, fermented with the same yeast and distilled in the same still, samples of burnt cane from an industrial brandy producer and samples of non-burnt cane obtained from another artisanal producer were utilized. The physicochemical and chromatographic (CG) analyses were accomplished in the Sugar Cane Brandy Physicochemical Analysis Laboratory at the Federal University of Lavras. The concentrations of furfural showed significant differences (P<0.01) among the groups of artisanal brandies obtained from sugar cane with and without previous burning, coming from the same manufacturing process with means of 1.48 mg.100mL-1 ethanol and 0.63mg.100mL-1 ethanol, respectively, although they have been bellow the maximum limit (5.0 mg.100mL-1 ethanol) allowed. The other components of the brandies studied were not affected significantly by the previous burning of sugar cane.

Determinação das composições físico-químicas de cachaças do sul de minas gerais e de suas misturas / Determination of the physical-chemical composition of homemade cachaças produced in the South of Minas Gerais and their mixtures

Este trabalho teve por objetivo a determinação da composição físico-química de cachaças artesanais produzidas no sul do Estado de Minas Gerais e suas misturas. Foram analisados os teores de etanol, acidez volátil, aldeídos, cobre, ésteres, álcoois superiores totais e metanol, além dos álcoois propanol-1, isobutanol e 3-metil-butanol-1. Os resultados apresentam o perfil peculiar desta bebida, que apresentou teores satisfatórios de álcoois superiores, ésteres e aldeídos. O teor de cobre apresenta-se como preocupante já que algumas amostras excederam o limite de 5 mg.L-1. A produção de misturas foi estudada e esta apresenta-se como uma alternativa viável ao produtor e cooperativas.

The present study sought the physical-chemical composition of homemade cachaças produced in the south of the state Minas Gerais and of mixtures of the same cachaça samples. The ethanol, aldehyde, copper, higher alcohol, volatile acid and methanol concentrations were determined, in addition to the individual propanol, 2-methyl-1-propanol and 3-methyl-1-butanol concentrations. The results demonstrated the characteristic profile of this beverage, which presented satisfactory concentrations of higher alcohols, esters, and aldehydes. The copper concentrations of some samples exceeded the limit of 5 mg. L-1. The mixtures produced from some of the cachaça samples presented a viable alternative for the producers and cooperatives because the concentrations of the components were more adequate for exportation.

Iridociclite näo granulomatosa bilateral em um caso de leptospirose / Non- granulumatous bilateral acute iridocidytis in a young patient with leptospirosis

Ilustra-se um caso de iridociclite aguda näo granulomatosa bilateral em um paciente jovem com suspeita clínica e laboratorial de Leptospirose. É uma doença em ascençäo, porém pouco comentada em nosso meio oftalmológico