Exploring Molecular Energy Landscapes by Coupling the DFTB Potential with a Tree-Based Stochastic Algorithm: Investigation of the Conformational Diversity of Phthalates.

Exploring the global energy landscape of relatively large molecules at the quantum level is a challenging problem. In this work, we report the coupling of a nonredundant conformational space exploration method, namely, the robotics-inspired iterative global exploration and local optimization (IGLOO) algorithm, with the quantum-chemical density functional tight binding (DFTB) potential. The application of this fast and efficient computational approach to three close-sized molecules of the phthalate family (DBP, BBP, and DEHP) showed that they present different conformational landscapes. These differences have been rationalized by making use of descriptors based on distances and dihedral angles. Coulomb interactions, steric hindrance, and dispersive interactions have been found to drive the geometric properties. A strong correlation has been evidenced between the two dihedral angles describing the side-chain orientation of the phthalate molecules. Our approach identifies low-energy minima without prior knowledge of the potential energy surface, paving the way for future investigations into transition paths and states.

Dependence of lactose adsorption on the exposed crystal facets of metals: a comparative study of gold, silver and copper.

In this theoretical work, we investigated the adsorption of a lactose molecule on metal-based surfaces, with a focus on the influence of the nature of the metal and of the type of exposed crystal facet on the adsorbed structures and energetics. More precisely, we considered three flat crystallographic facets of three face-centered cubic metals (gold, silver, and copper). For the global exploration of the energy landscape, we employed a multi-stage procedure where high-throughput searches, using a stochastic method that performs global optimization by iterating local searches, are followed by a refinement of the most probable adsorption conformations of the molecule at the ab initio level. We predicted the optimal conformation of lactose on each of the nine metal-surface combinations, classified the many low-energy minima into possible adsorption modes, and analyzed the structural, electronic and energetic aspects of the lactose molecule on the surface, as well as their dependence on the type of metal and exposed crystal facet. We observed structural similarities between the various minimum-energy conformations of lactose in vacuum and on the surface, a rough correlation between adsorption and interaction energies of the molecule, and a small charge transfer between molecule and surface whose direction is metal-dependent. During adsorption, an electronic reorganization occurs at the metal-molecule interface only, without affecting the vacuum-pointing atoms of the lactose molecule. For all types of surfaces, lactose exhibits the weakest adsorption on silver substrates, while for each coinage metal the adsorption is strongest on the (110) crystal facet. This study demonstrates that the control of exposed facets can allow to modulate the interaction between metals and small saccharides.

Variations Among Crop Seasons and Inoculum Availability Have a Higher Impact on Grapevine Rust Epidemics than Training Systems or Plastic Cover.

Canopy management practices can be effective as part of the integrated management of grapevine diseases. This study aimed to determine whether training systems and plastic covers can contribute to prevent Asian grapevine leaf rust (AGLR) development. Additionally, the influence of crop season and inoculum availability on AGLR development was investigated. Six-season experiments were carried out to characterize 16 epidemics that developed from natural inoculum (NI) or artificial inoculum (AI) sources (NI + AI), conducted in different training systems and with or without the plastic cover. The Richards model was fitted to each AGLR disease progress curve to estimate and compare the onsets and intensities of epidemics using eight curve elements. Principal components analysis (PCA) identified the incidence progress rate, the area under the severity progress curve, final disease severity, time to disease onset, and time to reach the inflection point as the main descriptors for AGLR epidemics. The results showed that AGLR epidemic development was related mainly to differences in inoculum availability and climatic conditions throughout the seasons and to a lower extent to the training system and plastic cover. The earliest disease onset was observed in epidemics when the NI was supplemented with an AI source. Differences in AGRL intensity were correlated to accumulated precipitation, being less severe in autumn-winter than in the spring-summer season. The present findings provided a better understanding of the structure and the seasonal variation of AGLR in cultivar 'Niagara Rosada'. The strategies for reducing and/or delaying inoculum buildup among seasons are discussed.

Ten Challenges to Understanding and Managing the Insect-Transmitted, Xylem-Limited Bacterial Pathogen Xylella fastidiosa.

An unprecedented plant health emergency in olives has been registered over the last decade in Italy, arguably more severe than what occurred repeatedly in grapes in the United States in the last 140 years. These emergencies are epidemics caused by a stealthy pathogen, the xylem-limited, insect-transmitted bacterium Xylella fastidiosa. Although these epidemics spurred research that answered many questions about the biology and management of this pathogen, many gaps in knowledge remain. For this review, we set out to represent both the U.S. and European perspectives on the most pressing challenges that need to be addressed. These are presented in 10 sections that we hope will stimulate discussion and interdisciplinary research. We reviewed intrinsic problems that arise from the fastidious growth of X. fastidiosa, the lack of specificity for insect transmission, and the economic and social importance of perennial mature woody plant hosts. Epidemiological models and predictions of pathogen establishment and disease expansion, vital for preparedness, are based on very limited data. Most of the current knowledge has been gathered from a few pathosystems, whereas several hundred remain to be studied, probably including those that will become the center of the next epidemic. Unfortunately, aspects of a particular pathosystem are not always transferable to others. We recommend diversification of research topics of both fundamental and applied nature addressing multiple pathosystems. Increasing preparedness through knowledge acquisition is the best strategy to anticipate and manage diseases caused by this pathogen, described as "the most dangerous plant bacterium known worldwide."

New Approaches to Plant Pathogen Detection and Disease Diagnosis.

Detecting plant pathogens and diagnosing diseases are critical components of successful pest management. These key areas have undergone significant advancements driven by breakthroughs in molecular biology and remote sensing technologies within the realm of precision agriculture. Notably, nucleic acid amplification techniques, with recent emphasis on sequencing procedures, particularly next-generation sequencing, have enabled improved DNA or RNA amplification detection protocols that now enable previously unthinkable strategies aimed at dissecting plant microbiota, including the disease-causing components. Simultaneously, the domain of remote sensing has seen the emergence of cutting-edge imaging sensor technologies and the integration of powerful computational tools, such as machine learning. These innovations enable spectral analysis of foliar symptoms and specific pathogen-induced alterations, making imaging spectroscopy and thermal imaging fundamental tools for large-scale disease surveillance and monitoring. These technologies contribute significantly to understanding the temporal and spatial dynamics of plant diseases.

European Xylella fastidiosa Strains Can Cause Symptoms in Blueberry.

Strains of the bacterial pathogen Xylella fastidiosa subspecies multiplex (Xfm) and pauca (Xfp) isolated from symptomatic almond and olive plants in Spain and Italy were used in this study. Because of the risk of host jump and considering the importance of southern highbush blueberry production in Spain, we tested a small set of these strains for their potential to infect and cause disease symptoms in blueberries under greenhouse experiments. Xfm IVIA5901 (isolated from almonds in Alicante, Spain) caused symptoms similar to those caused by Xfm AlmaEm3 (isolated from blueberries in Georgia, U.S.A., and used as a reference strain capable of inducing severe symptoms in blueberry). Nevertheless, bacterial populations of Xfm IVIA5901 in planta were significantly lower than those of Xfm AlmaEm3. Xfm ESVL (isolated from almonds, Alicante, Spain) and Xfp XYL1961/18 (isolated from olives, Ibiza Island, Spain) caused limited symptoms, while Xfm XYL466/19 (isolated from wild olives, Mallorca Island, Spain) and Xfm XF3348 (isolated from almonds, Mallorca Island, Spain) and Xfp De Donno (isolated from olives, Puglia, Italy, and representative of the devastating olive quick decline syndrome) did not cause symptoms nor colonize blueberries. This study suggests that certain strains already found in Europe could infect blueberry if conditions conducive for a host jump in this region are met, such as proximity of blueberries to other infected hosts and presence of insect vectors that feed on these crops. Surveys on the presence of X. fastidiosa in blueberries in Spain and other European countries are needed to anticipate possible issues.

Unraveling the ecological niche signals of Verticillium dahliae: Insights from Mediterranean landscapes.

Verticillium wilt (VW), caused by the soil-borne plant pathogenic fungus Verticillium dahliae, is a major disease impacting olive crops globally. In view of the lack of effective post-infection treatments, exclusion and avoidance strategies are essential in disease management. Assessing the risks posed by this pathogen is essential to prevent the spread and to ensure selection of suitable sites for new plantations. This study aimed to elucidate the environmental factors driving V. dahliae establishment in the Andalusia region, in southern Spain, an emblematic Mediterranean landscape for olive cultivation. To this end, we explored ecological niche signals for this fungal pathogen by analyzing 62 environmental variables across 1.6 million hectares dedicated to olive and cotton cultivation, using a 15-yr survey data on VW incidence on presence-absence from both olive and cotton fields. To ensure robust identification of ecological niche signals, we employed randomization-based, non-parametric univariate tests to compare presence records with the broader sampling universe (including absence records). Our findings identified key environmental variables that are associated significantly with V. dahliae presence, including temperature range seasonality (including mean diurnal and annual ranges), summer temperature (maximum of the warmest month, mean of the warmest quarter), and moisture and water availability (near-surface humidity, potential evapotranspiration, vapor pressure) as core niche variables for V. dahliae. Our results replicated the pathogen's known distribution, identifying the Guadalquivir Valley as a particularly high-risk area in view of its mild winters and distinct rainy seasons, providing new insights into the specific environmental conditions that facilitate the pathogen's survival and spread. Furthermore, this study introduces a novel approach to niche modeling that prioritizes variables with consistent effects and significant impact on the presence and distribution of V. dahliae and identifies potential data artifacts. This approach enhances our understanding of ecological requirements in V. dahliae and informs targeted management strategies.

Analyzing the Thermal Characteristics of Three Lining Materials for Plantar Orthotics.

INTRODUCTION: The choice of materials for covering plantar orthoses or wearable insoles is often based on their hardness, breathability, and moisture absorption capacity, although more due to professional preference than clear scientific criteria. An analysis of the thermal response to the use of these materials would provide information about their behavior; hence, the objective of this study was to assess the temperature of three lining materials with different characteristics. MATERIALS AND METHODS: The temperature of three materials for covering plantar orthoses was analyzed in a sample of 36 subjects (15 men and 21 women, aged 24.6 ± 8.2 years, mass 67.1 ± 13.6 kg, and height 1.7 ± 0.09 m). Temperature was measured before and after 3 h of use in clinical activities, using a polyethylene foam copolymer (PE), ethylene vinyl acetate (EVA), and PE-EVA copolymer foam insole with the use of a FLIR E60BX thermal camera. RESULTS: In the PE copolymer (material 1), temperature increases between 1.07 and 1.85 °C were found after activity, with these differences being statistically significant in all regions of interest (p < 0.001), except for the first toe (0.36 °C, p = 0.170). In the EVA foam (material 2) and the expansive foam of the PE-EVA copolymer (material 3), the temperatures were also significantly higher in all analyzed areas (p < 0.001), ranging between 1.49 and 2.73 °C for EVA and 0.58 and 2.16 °C for PE-EVA. The PE copolymer experienced lower overall overheating, and the area of the fifth metatarsal head underwent the greatest temperature increase, regardless of the material analyzed. CONCLUSIONS: PE foam lining materials, with lower density or an open-cell structure, would be preferred for controlling temperature rise in the lining/footbed interface and providing better thermal comfort for users. The area of the first toe was found to be the least overheated, while the fifth metatarsal head increased the most in temperature. This should be considered in the design of new wearables to avoid excessive temperatures due to the lining materials.

MoMA-LoopSampler: a web server to exhaustively sample protein loop conformations.

SUMMARY: MoMA-LoopSampler is a sampling method that globally explores the conformational space of flexible protein loops. It combines a large structural library of three-residue fragments and a novel reinforcement-learning-based approach to accelerate the sampling process while maintaining diversity. The method generates a set of statistically likely loop states satisfying geometric constraints, and its ability to sample experimentally observed conformations has been demonstrated. This paper presents a web user interface to MoMA-LoopSampler through the illustration of a typical use-case. AVAILABILITY AND IMPLEMENTATION: MoMA-LoopSampler is freely available at: https://moma.laas.fr/applications/LoopSampler/. We recommend users to create an account, but anonymous access is possible. In most cases, jobs are completed within a few minutes. The waiting time may increase depending on the server load, but it very rarely exceeds an hour. For users requiring more intensive use, binaries can be provided upon request. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

The sequence context in poly-alanine regions: structure, function and conservation.

MOTIVATION: Poly-alanine (polyA) regions are protein stretches mostly composed of alanines. Despite their abundance in eukaryotic proteomes and their association to nine inherited human diseases, the structural and functional roles exerted by polyA stretches remain poorly understood. In this work we study how the amino acid context in which polyA regions are settled in proteins influences their structure and function. RESULTS: We identified glycine and proline as the most abundant amino acids within polyA and in the flanking regions of polyA tracts, in human proteins as well as in 17 additional eukaryotic species. Our analyses indicate that the non-structuring nature of these two amino acids influences the α-helical conformations predicted for polyA, suggesting a relevant role in reducing the inherent aggregation propensity of long polyA. Then, we show how polyA position in protein N-termini relates with their function as transit peptides. PolyA placed just after the initial methionine is often predicted as part of mitochondrial transit peptides, whereas when placed in downstream positions, polyA are part of signal peptides. A few examples from known structures suggest that short polyA can emerge by alanine substitutions in α-helices; but evolution by insertion is observed for longer polyA. Our results showcase the importance of studying the sequence context of homorepeats as a mechanism to shape their structure-function relationships. AVAILABILITY AND IMPLEMENTATION: The datasets used and/or analyzed during the current study are available from the corresponding author onreasonable request. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Primary cutaneous mucormycosis due to Rhizopus arrhizus in an immunosuppressed patient with paroxysmal nocturnal haemoglobinuria.

OBJECTIVE: We present a case of primary cutaneous mucormycosis in a patient with bone marrow failure secondary to paroxysmal nocturnal haemoglobinuria (PNH). CLINICAL CASE: A 60-year-old male patient with a history of PNH, complicated to a severe aplastic anaemia, presented to the emergency department complaining of papules on the lower limbs that rapidly turned into necrotic plaques within 2 months. Histopathological examination showed granulomatous and suppurative dermatitis with tissue necrosis and the presence of non-septate hyphae. Molecular identification was achieved by amplification and sequencing of the 18S-ITS1-5.8S-ITS2-28S rRNA region using the polymerase chain reaction. The sequence showed 100% identity with Rhizopus arrhizus. The patient received treatment with liposomal amphotericin B and surgical debridement. Nonetheless, the patient suffered from severe low red blood cells and platelets and also underwent septic shock; he died 6 days after admission to the hospital. CONCLUSION: Mucormycosis in the setting of immunosuppression is challenging. Upon suspicion of a diagnosis, immediate treatment is required. Adjunctive therapies may be considered; however, the case fatality rate remains high.

Plant growth-promoting and heavy metal-resistant Priestia and Bacillus strains associated with pioneer plants from mine tailings.

Open mine tailings dams are extreme artificial environments containing sizeable potentially toxic elements (PTEs), including heavy metals (HMs), transition metals, and metalloids. Furthermore, these tailings have nutritional deficiencies, including assimilable phosphorus sources, organic carbon, and combined nitrogen, preventing plant colonization. Bacteria, that colonize these environments, have mechanisms to tolerate the selective pressures of PTEs. In this work, several Priestia megaterium (formerly Bacillus megaterium), Bacillus mojavensis, and Bacillus subtilis strains were isolated from bulk tailings, anthills, rhizosphere, and endosphere of pioneer plants from abandoned mine tailings in Zacatecas, Mexico. Bacillus spp. tolerated moderate HMs concentrations, produced siderophores and indole-3-acetic acid (IAA), solubilized phosphates, and reduced acetylene in the presence of HMs. The strains harbored different PIB-type ATPase genes encoding for efflux pumps and Cation Diffusion Facilitator (CDF) genes. Moreover, nifH and nifD nitrogenase genes were detected in P. megaterium and B. mojavensis genomic DNA. They showed similarity with sequences of the beta-Proteobacteria species, which may represent likely horizontal transfer events. These Bacillus species precede the colonization of mine tailings by plants. Their phenotypic and genotypic features could be essential in the natural recovery of the sites by reducing the oxidative stress of HMs, fixing nitrogen, solubilizing phosphate, and accumulating organic carbon. These traits of the strains reflect the adaptations of Bacillus species to the mine tailings environment and could contribute to the success of phytoremediation efforts.

Complete Circularized Genome Resources of Seven Strains of Xylella fastidiosa subsp. fastidiosa Using Hybrid Assembly Reveals Unknown Plasmids.

Xylella fastidiosa is a vascular plant pathogenic bacterium native to the Americas that is causing significant epidemics and economic losses in olive and almonds in Europe, where it is a quarantine pathogen. Since its first detection in 2013 in Italy, mandatory surveys across Europe revealed the presence of the bacterium also in France, Spain, and Portugal. Combining Oxford Nanopore Technologies and Illumina sequencing data, we assembled high-quality complete genomes of seven X. fastidiosa subsp. fastidiosa strains isolated from different plants in Spain, the United States, and Mexico. Comparative genomic analyses discovered differences in plasmid content among strains, including plasmids that had been overlooked previously when using the Illumina sequencing platform alone. Interestingly, in strain CFBP8073, intercepted in France from plants imported from Mexico, three plasmids were identified, including two (plasmids pXF-P1.CFBP8073 and pXF-P2.CFBP8073) not previously described in X. fastidiosa and one (pXF5823.CFBP8073) almost identical to a plasmid described in a X. fastidiosa strain from citrus. Plasmids found in the Spanish strains here were similar to those described previously in other strains from the same subspecies and ST1 isolated in the Balearic Islands and the United States. The genome resources from this work will assist in further studies on the role of plasmids in the epidemiology, ecology, and evolution of this plant pathogen.

Impact of Xylella fastidiosa subspecies pauca in European olives.

Xylella fastidiosa is the causal agent of plant diseases that cause massive economic damage. In 2013, a strain of the bacterium was, for the first time, detected in the European territory (Italy), causing the Olive Quick Decline Syndrome. We simulate future spread of the disease based on climatic-suitability modeling and radial expansion of the invaded territory. An economic model is developed to compute impact based on discounted foregone profits and losses in investment. The model projects impact for Italy, Greece, and Spain, as these countries account for around 95% of the European olive oil production. Climatic suitability modeling indicates that, depending on the suitability threshold, 95.5 to 98.9%, 99.2 to 99.8%, and 84.6 to 99.1% of the national areas of production fall into suitable territory in Italy, Greece, and Spain, respectively. For Italy, across the considered rates of radial range expansion the potential economic impact over 50 y ranges from 1.9 billion to 5.2 billion Euros for the economic worst-case scenario, in which production ceases after orchards die off. If replanting with resistant varieties is feasible, the impact ranges from 0.6 billion to 1.6 billion Euros. Depending on whether replanting is feasible, between 0.5 billion and 1.3 billion Euros can be saved over the course of 50 y if disease spread is reduced from 5.18 to 1.1 km per year. The analysis stresses the necessity to strengthen the ongoing research on cultivar resistance traits and application of phytosanitary measures, including vector control and inoculum suppression, by removing host plants.

Analysis of the Localization of Schizosaccharomyces pombe Glucan Synthases in the Presence of the Antifungal Agent Caspofungin.

In recent years, invasive fungal infections have emerged as a common source of infections in immunosuppressed patients. All fungal cells are surrounded by a cell wall that is essential for cell integrity and survival. It prevents cell death and lysis resulting from high internal turgor pressure. Since the cell wall is not present in animal cells, it is an ideal target for selective invasive fungal infection treatments. The antifungal family known as echinocandins, which specifically inhibit the synthesis of the cell wall ß(13)glucan, has been established as an alternative treatment for mycoses. To explore the mechanism of action of these antifungals, we analyzed the cell morphology and glucan synthases localization in Schizosaccharomyces pombe cells during the initial times of growth in the presence of the echinocandin drug caspofungin. S. pombe are rod-shaped cells that grow at the poles and divide by a central division septum. The cell wall and septum are formed by different glucans, which are synthesized by four essential glucan synthases: Bgs1, Bgs3, Bgs4, and Ags1. Thus, S. pombe is not only a perfect model for studying the synthesis of the fungal ß(1-3)glucan, but also it is ideal for examining the mechanisms of action and resistance of cell wall antifungals. Herein, we examined the cells in a drug susceptibility test in the presence of either lethal or sublethal concentrations of caspofungin, finding that exposure to the drug for long periods at high concentrations (>10 µg/mL) induced cell growth arrest and the formation of rounded, swollen, and dead cells, whereas low concentrations (<10 µg/mL) permitted cell growth with a mild effect on cell morphology. Interestingly, short-term treatments with either high or low concentrations of the drug induced effects contrary to those observed in the susceptibility tests. Thus, low drug concentrations induced a cell death phenotype that was not observed at high drug concentrations, which caused transient fungistatic cell growth arrest. After 3 h, high concentrations of the drug caused the following: (i) a decrease in the GFP-Bgs1 fluorescence level; (ii) altered locations of Bgs3, Bgs4, and Ags1; and (iii) a simultaneous accumulation of cells with calcofluor-stained incomplete septa, which at longer times resulted in septation uncoupling from plasma membrane ingression. The incomplete septa revealed with calcofluor were found to be complete when observed via the membrane-associated GFP-Bgs or Ags1-GFP. Finally, we found that the accumulation of incomplete septa depended on Pmk1, the last kinase of the cell wall integrity pathway.

Statistical proofs of the interdependence between nearest neighbor effects on polypeptide backbone conformations.

Backbone dihedral angles ϕ and ψ are the main structural descriptors of proteins and peptides. The distribution of these angles has been investigated over decades as they are essential for the validation and refinement of experimental measurements, as well as for structure prediction and design methods. The dependence of these distributions, not only on the nature of each amino acid but also on that of the closest neighbors, has been the subject of numerous studies. Although neighbor-dependent distributions are nowadays generally accepted as a good model, there is still some controversy about the combined effects of left and right neighbors. We have investigated this question using rigorous methods based on recently-developed statistical techniques. Our results unambiguously demonstrate that the influence of left and right neighbors cannot be considered independently. Consequently, three-residue fragments should be considered as the minimal building blocks to investigate polypeptide sequence-structure relationships.

No evidence for motor-recovery-related cortical connectivity changes after stroke using resting-state fMRI.

It has been proposed that a form of cortical reorganization (changes in functional connectivity between brain areas) can be assessed with resting-state (rs) functional MRI (fMRI). Here, we report a longitudinal data set collected from 19 patients with subcortical stroke and 11 controls. Patients were imaged up to five times over 1 year. We found no evidence, using rs-fMRI, for longitudinal poststroke cortical connectivity changes despite substantial behavioral recovery. These results could be construed as questioning the value of resting-state imaging. Here, we argue instead that they are consistent with other emerging reasons to challenge the idea of motor-recovery-related cortical reorganization poststroke when conceived of as changes in connectivity between cortical areas.NEW & NOTEWORTHY We investigated longitudinal changes in functional connectivity after stroke. Despite substantial motor recovery, we found no differences in functional connectivity patterns between patients and controls, nor any changes over time. Assuming that rs-fMRI is an adequate method to capture connectivity changes between cortical regions after brain injury, these results provide reason to doubt that changes in cortico-cortical connectivity are the relevant mechanism for promoting motor recovery.

[Coordination between primary care and a geriatrics service, 20 years later]. / Coordinación entre atención primaria y un servicio de geriatría, 20 años después.

OBJECTIVE: To describe the evolution of the care and coordination activity of a specialized hospital team (Geriatric Home Care-AGD team) in support of Primary Care (PC) for the care of geriatric patients. LOCATION: Health-5 area of the Community of Madrid. PARTICIPANTS, INTERVENTIONS AND MAIN MEASUREMENTS: Evaluation of patients attended at home at the request of PC by AGD, in 1997-1999 and twenty years later, in 2017-2019. Sociodemographic, clinical, functional and mental variables were recorded. Also care data such as time to first visit, average length of stay, origin and destination of patients, reason for consultation and main reason for disability. RESULTS: 524 patients (58% of the total) requested from AP in 1997-1999 and 1196 (72.2% of the total) in 2017-2019 were attended. Currently we show an older patient, more physically and mentally disabled, with a higher prevalence of geriatric syndromes and greater social fragility. Dementia has increased as the main reason for incapacity, with an increase in Comprehensive Geriatric Assessment and clinical control as the main reasons for referral. CONCLUSIONS: in our setting, after 20 years, AGD continues to be an important support for PC in the complex approach to frail elderly patients confined to their homes, who are increasingly vulnerable from a clinical, functional, cognitive and social point of view. It contributes to the management of geriatric syndromes, deprescription, direct hospital admissions when needed without having to go to the emergency department, and a decrease in institutionalization, facilitating the maintenance of the patient at home.

Protein loops with multiple meta-stable conformations: A challenge for sampling and scoring methods.

Flexible regions in proteins, such as loops, cannot be represented by a single conformation. Instead, conformational ensembles are needed to provide a more global picture. In this context, identifying statistically meaningful conformations within an ensemble generated by loop sampling techniques remains an open problem. The difficulty is primarily related to the lack of structural data about these flexible regions. With the majority of structural data coming from x-ray crystallography and ignoring plasticity, the conception and evaluation of loop scoring methods is challenging. In this work, we compare the performance of various scoring methods on a set of eight protein loops that are known to be flexible. The ability of each method to identify and select all of the known conformations is assessed, and the underlying energy landscapes are produced and projected to visualize the qualitative differences obtained when using the methods. Statistical potentials are found to provide considerable reliability despite their being designed to tradeoff accuracy for lower computational cost. On a large pool of loop models, they are capable of filtering out statistically improbable states while retaining those that resemble known (and thus likely) conformations. However, computationally expensive methods are still required for more precise assessment and structural refinement. The results also highlight the importance of employing several scaffolds for the protein, due to the high influence of small structural rearrangements in the rest of the protein over the modeled energy landscape for the loop.

Interdomain linkers tailor the stability of immunoglobulin repeats in polyproteins.

Linkers in polyproteins are considered as mere spacers between two adjacent domains. However, a series of studies using single-molecule force spectroscopy have recently reported distinct thermodynamic stability of I27 in polyproteins with varying linkers and indicated the vital role of linkers in domain stability. A flexible glycine rich linker (-(GGG)n, n ≥ 3) featured unfolding at lower forces than the regularly used arg-ser (RS) based linker. Interdomain interactions among I27 domains in Gly-rich linkers were suggested to lead to reduced domain stability. However, the negative impact of inter domain interactions on domain stability is thermodynamically counter-intuitive and demanded thorough investigations. Here, using an array of ensemble equilibrium experiments and in-silico measurements with I27 singlet and doublets with two aforementioned linkers, we delineate that the inter-domain interactions in fact raise the stability of the polyprotein with RS linker. More surprisingly, a highly flexible Gly-rich linker has no interference on the stability of polyprotein. Overall, we conclude that flexible linkers are preferred in a polyprotein for maintaining domain's independence.