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Solution nuclear magnetic resonance (NMR) spectroscopy is a powerful technique for characterizing protein–protein interactions. NMR-monitored titration proves to be effective in determining dissociation constants, particularly for Kd values in the micromolar to millimolar range. In conventional NMR titrations, planning for optimal titration conditions requires a prior estimate of Kd. In addition, a highly concentrated ligand stock solution is often required, posing challenges when the ligand exhibits limited solubility and stability at elevated concentrations. To overcome these constraints, we propose a simulation-guided "swapping" protocol for NMR titrations. Guided by simulations of the binding curves, two samples, one with zero and the other with maximum ligand concentration, but both containing identical protein concentrations, initiate the titration. Using a "swapping" strategy, intermediate ligand concentrations in between those of the two initial samples are generated without the need of a concentrated ligand stock, while maintaining constant protein concentration. More importantly, this protocol facilitates estimation of Kd by early titration points and allows on-the-fly optimization of the titration points. The proposed approach enhances the efficiency of NMR titrations and provides a straightforward means to optimize the experimental conditions for the titrations. [ABSTRACT FROM AUTHOR]

p97/VCP is an essential AAA+ ATPase involved in diverse cellular activities by interacting with an array of protein adaptors that recruit p97 for specific tasks. p47 is one of the adaptors that targets p97 for membrane remodeling by forming a stable complex with p97 through multivalent interactions. Here, we report a pair of previously unidentified interactions between the N-terminal part of p47 (residues 1–94) and the N-terminal domain (NTD) of p97. Using nuclear magnetic resonance (NMR) spectroscopy, we identify two binding sites on p47, one located on the ubiquitin-associated (UBA) domain and the other on the intrinsically disordered linker, that interact with the same basic patch on p97 NTD, driven by electrostatic forces. Reciprocal NMR titration experiments between p47 (residues 1–94) and p97 NTD reveal that these interactions are relatively weak in nature with dissociation constants on the order of hundreds of micromolar to millimolar in trans. Structural models of the two interactions are developed based on NMR chemical shift perturbations, which reveal details of the tentative binding interfaces. Our findings provide new insights into the mechanism by which ubiquitinated substrates are delivered from p47 to p97 for unfolding. [ABSTRACT FROM AUTHOR]

Aim/purpose – This paper aims to introduce and describe a new paradigm (model) of evolutionary dynamism of family business potential in the succession process and its empirical simulation in family enterprises from culturally close “post-socialistic” countries: the Czech Republic and Poland.

Background: Malaria has become a serious threat in many developing countries, including Indonesia. Malaria infection without proper treatment can cause death. Plasmodium berghei (P. berghei) is a malaria parasite that mostly infects rodents, and is often used as a model for in vivo antimalarial research in mice. Giving the right vaccine can be one of the efforts to control and eliminate malaria.Objectives: The purpose of this article is to review the protein coding for P. berghei as a vaccine candidate. Review: Antibodies are components of the immune system that play a role in the humoral immune response. Expression of Pb51, P47, PbVaC, PSOP25 genes in P. berghei are selected antigens against malaria parasites, has the ability to block malaria through chemotherapy and immunoprophylaxis interventions efficiently as a potential malaria vaccine candidate. Conclusion: The protein coding for P. berghei has good antigenicity as a candidates for malaria vaccine. The mechanism of blocking the transmission of parasites to mosquitoes, induction of the functional immune system and the effectiveness of malaria vaccines require further studies. [ABSTRACT FROM AUTHOR]

The study of financial literacy and locus of control in the creative economy is a relatively new area of research. However, there is a growing body of evidence that suggests that these factors can have a significant impact on the performance of businesses in this sector. Therefore, the purpose of this study is to determine if training, locus of control, and financial literacy have a direct influence on the performance of creative MSMEs; and to determine whether training has an indirect effect on the performance of creative MSMEs through financial literacy. A convenience sample of 115 respondents served as the study's main source of data in the municipality (Yogyakarta) and four regencies (Sleman, Bantul, Kulonprogo, Gunung Kidul). The data were analyzed using SPSS 22, path analysis and Sobel's test. The results revealed that training, the internal position of control, and financial literacy positively affected performance. However, the position of the external control did not affect the performance. In addition, the training had a positive effect on financial literacy and affected performance through financial literacy. These results indicate that financial understanding, training and individual self-perception have become critical elements to enhance company performance because they enable actors to manage their businesses according to objectives.

Botulinum neurotoxins (BoNTs) are causative agents of the life-threatening disease botulism. They are naturally produced by species of the bacteria Clostridium botulinum as stable and non-covalent complexes, in which the BoNT molecule is assembled with several auxiliary non-toxic proteins. Some BoNT serotypes, represented by the well-studied BoNT serotype A (BoNT/A), are produced by Clostridium strains that carry the ha gene cluster, which encodes four neurotoxin-associated proteins (NTNHA, HA17, HA33, and HA70) that play an important role to deliver and protect BoNTs in the gastrointestinal tract during oral intoxication. In contrast, BoNT/E- and BoNT/F-producing strains carry a distinct gene cluster that encodes five proteins (NTNHA, P47, OrfX1, OrfX2, and OrfX3, termed the orfX cluster). The structures and functions of these proteins remain largely unknown. Here, we report the crystal structure of P47 resolved at 2.8 Å resolution. Surprisingly, P47 displays a structural topology that is similar to bactericidal/permeability-increasing (BPI) like proteins, which were previously identified only in eukaryotes. The similarity of a hydrophobic cleft of P47 with the phospholipid-binding groove of BPI suggests that P47 might be involved in lipid association to exert its function. Consistently, P47 associates and induces aggregation of asolectin-containing liposomes in a protein- and lipid-concentration dependent manner. These findings laid the foundation for future structural and functional studies of the potential roles of P47 and OrfX proteins in facilitating oral intoxication of BoNTs.

Dominant mutations in p97/VCP (valosin-containing protein) cause a rare multisystem degenerative disease with varied phenotypes that include inclusion body myopathy, Paget's disease of bone, frontotemporal dementia, and amyotrophic lateral sclerosis. p97 disease mutants have altered N-domain conformations, elevated ATPase activity, and altered cofactor association. We have now discovered a previously unidentified disease-relevant functional property of p97 by identifying how the cofactors p37 and p47 regulate p97 ATPase activity. We define p37 as, to our knowledge, the first known p97-activating cofactor, which enhances the catalytic efficiency (kcat/Km) of p97 by 11-fold. Whereas both p37 and p47 decrease the Km of ATP in p97, p37 increases the kcat of p97. In contrast, regulation by p47 is biphasic, with decreased kcat at low levels but increased kcat at higher levels. By deleting a region of p47 that lacks homology to p37 (amino acids 69-92), we changed p47 from an inhibitory cofactor to an activating cofactor, similar to p37. Our data suggest that cofactors regulate p97 ATPase activity by binding to the N domain. Induced conformation changes affect ADP/ATP binding at the D1 domain, which in turn controls ATPase cycling. Most importantly, we found that the D2 domain of disease mutants failed to be activated by p37 or p47. Our results show that cofactors play a critical role in controlling p97 ATPase activity, and suggest that lack of cofactor-regulated communication may contribute to p97-associated disease pathogenesis.

Existing research on resilience varies across fields on analysis and presents scattered and diverse definitions. For example, the literature suggests that resilience in organisations is influenced by factors of various levels, occurring at both the individual and organisational level. However, there are currently few insights into how these different levels of analysis interact with each other. Therefore, focusing on resilience in the context of entrepreneurship, this paper aims to explore the relationship between the employee-level resilience dimensions (cognitive, behavioural and contextual) and entrepreneurs' perceived performance, and the moderating role of entrepreneur resilience. A survey has been conducted with 195 entrepreneurs managing small family firms. Results confirm the hypothesis that the dimensions of employee-level resilience affect performance positively when the entrepreneur has a strong propensity towards personal resilience. Entrepreneurs may use these insights to increase awareness of their actions towards achieving organisational resilience and to implement practices aimed at increasing employee-level resilience. [ABSTRACT FROM AUTHOR]

The role of citizens' collective action for the emergence and consolidation of democracy is generally analysed within bottom-up theories. However, top-down theories show that elites might impede or promote both democracy and collective action through a set of strategies which are often unobserved and vary over time. Democratic persistence and change require then to be assessed in a dynamic framework which considers both citizens and elites' strategies. For such reason, on a large sample of countries in the period 1971–2014, we jointly estimate the probability of collective action and democracy using a Structural Dynamic Model. This allows us to account for the dynamic nature of the two political phenomena under investigation by controlling for their persistence, for initial conditions and time-varying unobserved heterogeneity. We find that collective action matters for the emergence of democracy but not for its consolidation which seems to be related to more structural economic factors. [ABSTRACT FROM AUTHOR]

An effective vaccine to reduce malaria transmission is central to control and ultimately achieve disease eradication. Recently, we demonstrated that antibodies targeting the Plasmodium falciparum surface protein P47 (Pfs47) reduce parasite transmission to Anopheles gambiae mosquitoes. Here, Plasmodium berghei (Pb) was used as a model to assess the in vivo efficacy of a P47-targeted transmission blocking vaccine (Pbs47). Mice were immunized following a prime/boost regimen and infected with P. berghei. The effect of immunization on infectivity to mosquitoes was evaluated by direct feeding on P. berghei-infected mice. The key region in Pbs47 where antibody binding confers protection was mapped, and the immunogenicity of this protective antigen was enhanced by conjugation to a virus-like particle. Passive immunization with 100 and 50 μg/mL of anti-Pbs47 IgG reduced oocyst density by 77 and 67%, respectively. Furthermore, affinity purified Pbs47-specific IgG significantly reduced oocyst density by 88 and 77%, respectively at doses as low as 10 and 1 μg/mL. These studies suggest that P47 is a promising transmission blocking target and show that antibodies to the same specific region in Pfs47 and Pbs47 confer protection.

This paper argues that the traditional social contract that underlies the free market economy has run its course and needs to be replaced by a new contract, based on a new conception of the “empowering economy.” Whereas different social contracts are relevant to different societies, all these contracts have some features in common, addressing some basic human needs that are common to all. These are needs that every thriving society must satisfy. In the presence of current global problems – such as climate change and financial crises – satisfying these needs can also generate the popular approval for multilateral agreements to tackle these problems. The paper identifies three inconvenient truths for the existing social contract: (i) economic performance involves more than material prosperity, (ii) free markets naturally generate inequality, and (iii) human progress rests primarily on cooperation. In response, the paper proposes a new social contract that can be promoted through three policy approaches: (1) policy that focuses not just on material prosperity, but also on personal empowerment and social solidarity, (2) automatic stabilizers that reduce inequalities of economic power and (3) policy that develops the human capabilities of cooperation.

VCP/p97, an enzyme critical to proteostasis, is regulated through interactions with protein adaptors targeting it to specific cellular tasks. One such adaptor, p47, forms a complex with p97 to direct lipid membrane remodeling. Here, we use NMR and other biophysical methods to study the structural dynamics of p47 and p47–p97 complexes. Disordered regions in p47 are shown to be critical in directing intra-p47 and p47–p97 interactions via a pair of previously unidentified linear motifs. One of these, an SHP domain, regulates p47 binding to p97 in a manner that depends on the nucleotide state of p97. NMR and electron cryomicroscopy data have been used as restraints in molecular dynamics trajectories to develop structural ensembles for p47–p97 complexes in adenosine diphosphate (ADP)- and adenosine triphosphate (ATP)-bound conformations, highlighting differences in interactions in the two states. Our study establishes the importance of intrinsically disordered regions in p47 for the formation of functional p47–p97 complexes. [ABSTRACT FROM AUTHOR]

An effective vaccine to reduce malaria transmission is central to control and ultimately achieve disease eradication. Recently, we demonstrated that antibodies targeting the Plasmodium falciparum surface protein P47 (Pfs47) reduce parasite transmission to Anopheles gambiae mosquitoes. Here, Plasmodium berghei (Pb) was used as a model to assess the in vivo efficacy of a P47-targeted transmission blocking vaccine (Pbs47). Mice were immunized following a prime/boost regimen and infected with P. berghei. The effect of immunization on infectivity to mosquitoes was evaluated by direct feeding on P. berghei -infected mice. The key region in Pbs47 where antibody binding confers protection was mapped, and the immunogenicity of this protective antigen was enhanced by conjugation to a virus-like particle. Passive immunization with 100 and 50 μg/mL of anti-Pbs47 IgG reduced oocyst density by 77 and 67%, respectively. Furthermore, affinity purified Pbs47-specific IgG significantly reduced oocyst density by 88 and 77%, respectively at doses as low as 10 and 1 μg/mL. These studies suggest that P47 is a promising transmission blocking target and show that antibodies to the same specific region in Pfs47 and Pbs47 confer protection. [ABSTRACT FROM AUTHOR]

Autophagy is a conserved cellular process, and its dysfunction is implicated in cancer and other diseases. Here, we employ an in vivo CRISPR screen targeting genes implicated in the regulation of autophagy to identify the Nsfl1c gene encoding p47 as a suppressor of human epidermal growth factor receptor 2 (HER2)+ breast cancer metastasis. p47 ablation specifically increases metastasis without promoting primary mammary tumor growth. Analysis of human breast cancer patient databases and tissue samples indicates a correlation of lower p47 expression levels with metastasis and decreased survival. Mechanistic studies show that p47 functions in the repair of lysosomal damage for autophagy flux and in the endosomal trafficking of nuclear factor κB essential modulator for lysosomal degradation to promote metastasis. Our results demonstrate a role and mechanisms of p47 in the regulation of breast cancer metastasis. They highlight the potential to exploit p47 as a suppressor of metastasis through multiple pathways in HER2+ breast cancer cells. [Display omitted] • In vivo CRISPR screen reveals p47 as a suppressor of HER2+ breast cancer metastasis • p47 depletion reduces NEMO endosomal trafficking and increases NF-κB signaling • p47 ablation impairs lysosome repair and autophagy flux and increases metastasis • Lower p47 expression correlates to increased metastasis in human breast cancer Hao et al. identify p47 as a suppressor of HER2+ breast cancer metastasis with an in vivo CRISPR-Cas9 screen. They show that p47 plays a role in the repair of lysosomal damage for autophagy flux and in endosomal trafficking of NEMO for lysosomal degradation to restrain HER2+ breast cancer metastasis. [ABSTRACT FROM AUTHOR]

Valosin-containing protein (VCP) was found to play a vital protective role against cardiac stresses. Genetic mutations of VCP are associated with human dilated cardiomyopathy. However, the essential role of VCP in the heart during the physiological condition remains unknown since the VCP knockout in mice is embryonically lethal. We generated a cardiac-specific dominant-negative VCP transgenic (DN-VCP TG) mouse to determine the effects of impaired VCP activity on the heart. Using echocardiography, we showed that cardiac-specific overexpression of DN-VCP induced a remarkable cardiac dilation and progressively declined cardiac function during the aging transition. Mechanistically, DN-VCP did not affect the endogenous VCP (EN-VCP) expression but significantly reduced cardiac ATPase activity in the DN-VCP TG mouse hearts, indicating a functional inhibition. DN-VCP significantly impaired the aging-related cytoplasmic/nuclear shuffling of EN-VCP and its co-factors in the heart tissues and interrupted the balance of the VCP-cofactors interaction between the activating co-factors, ubiquitin fusion degradation protein 1 (UFD-1)/nuclear protein localization protein 4 (NPL-4) complex, and its inhibiting co-factor P47, leading to the binding preference with the inhibitory co-factor, resulting in functional repression of VCP. This DN-VCP TG mouse provides a unique functional-inactivation model for investigating VCP in the heart in physiological and pathological conditions.

The scaffold protein Tks4 is a member of the p47phox-related organizer superfamily. It plays a key role in cell motility by being essential for the formation of podosomes and invadopodia. In addition, Tks4 is involved in the epidermal growth factor (EGF) signaling pathway, in which EGF induces the translocation of Tks4 from the cytoplasm to the plasma membrane. The evolutionarily-related protein p47phox and Tks4 share many similarities in their N-terminal region: a phosphoinositide-binding PX domain is followed by two SH3 domains (so called “tandem SH3”) and a proline-rich region (PRR). In p47phox, the PRR is followed by a relatively short, disordered C-terminal tail region containing multiple phosphorylation sites. These play a key role in the regulation of the protein. In Tks4, the PRR is followed by a third and a fourth SH3 domain connected by a long (~420 residues) unstructured region. In p47phox, the tandem SH3 domain binds the PRR while the first SH3 domain interacts with the PX domain, thereby preventing its binding to the membrane. Based on the conserved structural features of p47phox and Tks4 and the fact that an intramolecular interaction between the third SH3 and the PX domains of Tks4 has already been reported, we hypothesized that Tks4 is similarly regulated by autoinhibition. In this study, we showed, via fluorescence-based titrations, MST, ITC, and SAXS measurements, that the tandem SH3 domain of Tks4 binds the PRR and that the PX domain interacts with the third SH3 domain. We also investigated a phosphomimicking Thr-to-Glu point mutation in the PRR as a possible regulator of intramolecular interactions. Phosphatidylinositol-3-phosphate (PtdIns(3)P) was identified as the main binding partner of the PX domain via lipid-binding assays. In truncated Tks4 fragments, the presence of the tandem SH3, together with the PRR, reduced PtdIns(3)P binding, while the presence of the third SH3 domain led to complete inhibition.

Data Envelopment Analysis (DEA) is a deterministic method for the aggregation of multidimensional measures and subsequent efficiency analysis. Due to its inherent determinism, however, it reacts sensitively to outliers in datasets. Existing methods for identifying such outliers have two main disadvantages. First, from a more conceptional point of view, a uniform definition of an outlier is missing. Second, there are technical disadvantages of each method. For instance, arbitrarily limited values have to be set by the user, like the amount of efficiency value from which on a decision making unit is regarded as an outlier. This paper initially presents a definition of outliers, which explicitly takes the specifics of DEA into account. Based on this definition, an approach for identifying outliers in DEA is introduced which explicitly tackles the technical disadvantages and takes them into account in the developed algorithm. The plausibility of this approach is validated on the basis of empirical examples from performance measurement at the university level. [ABSTRACT FROM AUTHOR]

The botulinum neurotoxin (BoNT) has been extensively researched over the years in regard to its structure, mode of action, and applications. Nevertheless, the biological roles of four proteins encoded from a number of BoNT gene clusters, i.e., OrfX1-3 and P47, are unknown. Here, we investigated the diversity of orfX-p47 gene clusters using in silico analytical tools. We show that the orfX-p47 cluster was not only present in the genomes of BoNT-producing bacteria but also in a substantially wider range of bacterial species across the bacterial phylogenetic tree. Remarkably, the orfX-p47 cluster was consistently located in proximity to genes coding for various toxins, suggesting that OrfX1-3 and P47 may have a conserved function related to toxinogenesis and/or pathogenesis, regardless of the toxin produced by the bacterium. Our work also led to the identification of a putative novel BoNT-like toxin gene cluster in a Bacillus isolate. This gene cluster shares striking similarities to the BoNT cluster, encoding a bont/ntnh-like gene and orfX-p47, but also differs from it markedly, displaying additional genes putatively encoding the components of a polymorphic ABC toxin complex. These findings provide novel insights into the biological roles of OrfX1, OrfX2, OrfX3, and P47 in toxinogenesis and pathogenesis of BoNT-producing and non-producing bacteria.

This paper examines the effect of various types of bank capital on the profitability and efficiency of conventional and Islamic banks. Our results show that higher quality forms of capital improve the profitability and efficiency for both systems although the results are stronger for conventional banks. The capital effect is more pronounced for large, too-big-to-fail, and highly capitalized banks. The results are robust across various subsamples, alternative profitability and efficiency measures, and different estimation techniques. [ABSTRACT FROM AUTHOR]

Abstract Apoptosis is an essential type of programmed cell death. Previous studies have demonstrated that a wide range of natural-derived anticancer agents induce apoptosis by trigging oxidative stress. Artemisia argyi is a traditional Chinese herb for treating diverse diseases including dyspepsia, arthroncus, and anaphylactic disease. In this study, sesquiterpene lactone 3 (SL3), a bioactive ingredient isolated from Artemisia argyi was found to show obvious inhibitory effect on two gastric carcinoma cells. Mechanism study revealed that SL3 promoted the membrane translocation of p47, activated nicotinamide adenine dinucleotide (NADPH) oxidase, and evaluated intracellular reactive oxygen species production, leading to the activation of mitochondria-dependent caspase apoptosis pathway. Collectively, these findings show that SL3 is a promising anticancer candidate against gastric carcinoma by activating NADPH oxidase/reactive oxygen species/mitochondrial pathway. [ABSTRACT FROM AUTHOR]

I expound a game theoretical model of the constitution of patron-client relations and patronage appointments. For all parameter values, the game has open access equilibriums, where patronage does not occur. For some parameter values, it has also a limited access equilibrium, where patronage does occur. I prove open and limited access equilibriums are not Pareto-comparable to one another. Defining social welfare in utilitarian terms, I prove also that social welfare is maximised in a particular open access equilibrium, but that a limited access equilibrium can be a second-best solution, when it exists. [ABSTRACT FROM AUTHOR]

This article deals with innovation performance and its measurement. The objective is to present the measuring of innovation performance as it is implemented in today’s Czech business environment. It begins with a comprehensive theory and the definition of essential terms. The theoretical part is followed by the analysis of the current state of the issue from the perspective of enterprises in the Czech Republic. The analysis consists of four own primary questionnaire surveys. The research outputs reveal weaknesses in current approaches to innovation effectiveness measurement. Our enterprises use all possible means to increase productivity and achieve operational excellence. However, they tend to neglect the important area of research and development. It has been proven, with help of questionnaire surveys, that many enterprises still do not measure innovation performance despite the importance of innovation as an engine of growth. Only a few organisations appear to have an effective system for measuring their overall innovation performance. [ABSTRACT FROM PUBLISHER]