Miracle-dz/UniFAR-FTUs huggingface dataset:
Expression, purification, and characterization of pyruvate kinase from Mycobacterium tuberculosis: A key allosteric regulatory enzyme

Background: The current research aims to isolate pyruvate kinase (Pyk) gene from Mycobacterium tuberculosis and expression of the gene (Rv1617) to obtain a purified enzyme. The enzyme activity and secondary structural features were assessed through biochemical assays and circular dichroism (CD) spectroscopy, respectively. Methods: The Pyk-encoding gene from the complete genome of M. tuberculosis was cloned, sequenced, and expressed in Escherichia coli BL21 (DE3). The enzyme was purified by nickel-nitrilotriacetic acid affinity chromatography and enzyme activity was determined by a lactose dehydrogenase-coupled assay system. Further, far ultraviolet CD spectra of the enzyme and the substrate bound enzyme were analyzed using a Jasco J712 spectrophotometer. Results: A single protein with an approximate molecular mass of 54 kDa was purified and a specific activity of 5.31 units/mg was determined from purified M. tuberculosis Pyk. The activity of the enzyme indicating a protein is defined by separate domain for each catalytic function. The secondary structure analysis of CD spectra of the recombinant Pyk has revealed a content of 17% α-helix, 34% β-sheet, and 49% turns in the enzyme. Conclusion: The growing evidence has impacted M. tuberculosis central carbon metabolism as a key determinant of the survival and pathogenicity in the host. The purified Pyk was observed to have increased enzyme activity in all steps of purification. Retention of Pyk activity indicates a possible catalytic role for the lower part of the glycolytic pathway. The overall results of the spectra obtained from the CD suggest that the substrate phosphoenolpyruvate and adenosine diphosphate binding to the enzyme can cause conformational changes resulting in the exposure or shielding of residues susceptible to modification.
Advancing a MEMS-Based 3D Cell Culture System for in vitro Neuro-Electrophysiological Recordings

In this work we present advances in three dimensional (3D) neuronal cell culture systems based on a reversible assembly of a microbioreactor with a microelectrode array (MEA) to create a MEMS-based 3D cell culture system for in vitro neuro-electrophysiological recordings. A batch of six molds were milled in poly (methyl methacrylate). The molds were used for soft lithography of polydimethylsiloxane (PDMS). In the center of the PDMS shape, a porous polyethersulfone (PES) cylindrical tube was press-fitted to form a growth barrier between the culture chamber inside the PES tube and the microfluidic channel surrounding the PES tube. A thin layer of partially cured PDMS was used to seal the bottom of the microbioreactor and provide reversible adhesion with the glass surface of a MEA. SH-SY5Y cells were successfully differentiated inside the microbioreactors in Matrigel and demonstrated extended neuronal networks over a height of at least 184 micrometers within the system. In previous microbioreactor designsvisibility was limited due to the closed top with the dispensing holes. The new open top design allows for a better evaluation of the cell culture by optical detection methods during the experiment. Electrophysiological activity was recorded within the microbioreactor using human induced pluripotent stem cell-derived cortical neurons cultured in Matrigel, in 3D, up until 21 days in vitro In summary, we present advances made in the design, the fabrication process and integration of microbioreactors with MEAs. Optical imaging capabilities improved significantly with an open top and the culture time was further extended from 7 to 21 DIV without leakage or degradation thanks to introducing PES as a barrier material and an enhanced assembly procedure. The latter facilitated a sufficient long-term culture for neurons to mature in an environment free from flow-induced stress and provided a proof of principle for. the recording of electrophysiological activity of cortical neurons cultured in 3D.
Novel broad spectrum virucidal molecules against enveloped viruses

Viral infections are an important cause of death worldwide. Unfortunately, there is still a lack of antiviral drugs or vaccines for a large number of viruses, and this represents a remarkable challenge particularly for emerging and re-emerging viruses. For this reason, the identification of broad spectrum antiviral compounds provides a valuable opportunity for developing efficient antiviral therapies. Here we report on a class of rhodanine and thiobarbituric derivatives displaying a broad spectrum antiviral activity against seven different enveloped viruses including an HSV-2 acyclovir resistant strain with favorable selectivity indexes. Due to their selective action on enveloped viruses and to their lipid oxidation ability, we hypothesize a mechanism on the viral envelope that affects the fluidity of the lipid bilayer, thus compromising the efficiency of virus-cell fusion and preventing viral entry.
Faecal immunochemical tests (FIT) versus colonoscopy for surveillance after screening and polypectomy: a diagnostic accuracy and cost-effectiveness study

Objective The English Bowel Cancer Screening Programme (BCSP) recommends 3 yearly colonoscopy surveillance for patients at intermediate risk of colorectal cancer (CRC) postpolypectomy (those with three to four small adenomas or one ≥10 mm). We investigated whether faecal immunochemical tests (FITs) could reduce surveillance burden on patients and endoscopy services. Design Intermediate-risk patients (60–72 years) recommended 3 yearly surveillance were recruited within the BCSP (January 2012–December 2013). FITs were offered at 1, 2 and 3 years postpolypectomy. Invitees consenting and returning a year 1 FIT were included. Participants testing positive (haemoglobin ≥40 µg/g) at years one or two were offered colonoscopy early; all others were offered colonoscopy at 3 years. Diagnostic accuracy for CRC and advanced adenomas (AAs) was estimated considering multiple tests and thresholds. We calculated incremental costs per additional AA and CRC detected by colonoscopy versus FIT surveillance. Results 74% (5938/8009) of invitees were included in our study having participated at year 1. Of these, 97% returned FITs at years 2 and 3. Three-year cumulative positivity was 13% at the 40 µg/g haemoglobin threshold and 29% at 10 µg/g. 29 participants were diagnosed with CRC and 446 with AAs. Three-year programme sensitivities for CRC and AAs were, respectively, 59% and 33% at 40 µg/g, and 72% and 57% at 10 µg/g. Incremental costs per additional AA and CRC detected by colonoscopy versus FIT (40 µg/g) surveillance were £7354 and £180 778, respectively. Conclusions Replacing 3 yearly colonoscopy surveillance in intermediate-risk patients with annual FIT could reduce colonoscopies by 71%, significantly cut costs but could miss 30%–40% of CRCs and 40%–70% of AAs. Trial registration number ISRCTN18040196; Results.
Major Motivators and Sociodemographic Features of Women Undergoing Labiaplasty.

Background Influencing factors to decide for labiaplasty are evolving and show diversity among different settings and countries. Updated underlying motivators may assist in the clinical decision-making and assessing outcomes from the patients' perspective. Objective Herein, the main motivators of women undergoing labiaplasty and sociodemographic features are discussed. Methods Seventy-one patients who were scheduled for labiaplasty were assessed with an open-ended, well-detailed questionnaire to determine the main motivational factors from the patients' perspective. Validated questionnaires to measure genital self-image, self-esteem, body appreciation, attitude toward having cosmetic surgery, sexual dysfunction, and body-dysmorphic disorder were performed. Results Half of the patients reported that they had an idea about the female genitalia (50.7%) and they were influenced through the media (47.9%). The majority of those (71.8%) stated that they did not have normal genitalia and considered labiaplasty more than 6 months ago (88.7%). The pornography consumption rate in the last month was 19.7% and was significantly related with lower genital self-image and self-esteem. The main motivation was found as having improvement in appearance (43.7%) and a better sexual life (26.8%). Patients reported aesthetic (52.1%), sexual (46.5%), and psychological (39.4%) reasons leading to their decision. Patient-reported 8-items out of 11-item motivators for having labiaplasty were statistically consistent with validated questionnaires. The self-esteem and body-appreciation of the patients were comparable with the literature. Genital self-image and sexual function were found to be impaired. Conclusions The main motivator for labiaplasty was found as improvement in genital appearance integrated with their aesthetic and sexual demands. Eleven-item motivators derived from the statements of women seeking labiaplasty were determined in this study. Sociodemographic features and main motivators can conveniently be used as normative data for this population.
Partner‐specific prediction of RNA‐binding residues in proteins: A critical assessment

RNA‐protein interactions play essential roles in regulating gene expression. While some RNA‐protein interactions are “specific”, that is, the RNA‐binding proteins preferentially bind to particular RNA sequence or structural motifs, others are “non‐RNA specific. ” Deciphering the protein‐RNA recognition code is essential for comprehending the functional implications of these interactions and for developing new therapies for many diseases. Because of the high cost of experimental determination of protein‐RNA interfaces, there is a need for computational methods to identify RNA‐binding residues in proteins. While most of the existing computational methods for predicting RNA‐binding residues in RNA‐binding proteins are oblivious to the characteristics of the partner RNA, there is growing interest in methods for partner‐specific prediction of RNA binding sites in proteins. In this work, we assess the performance of two recently published partner‐specific protein‐RNA interface prediction tools, PS‐PRIP, and PRIdictor, along with our own new tools. Specifically, we introduce a novel metric, RNA‐specificity metric (RSM), for quantifying the RNA‐specificity of the RNA binding residues predicted by such tools. Our results show that the RNA‐binding residues predicted by previously published methods are oblivious to the characteristics of the putative RNA binding partner. Moreover, when evaluated using partner‐agnostic metrics, RNA partner‐specific methods are outperformed by the state‐of‐the‐art partner‐agnostic methods. We conjecture that either (a) the protein‐RNA complexes in PDB are not representative of the protein‐RNA interactions in nature, or (b) the current methods for partner‐specific prediction of RNA‐binding residues in proteins fail to account for the differences in RNA partner‐specific versus partner‐agnostic protein‐RNA interactions, or both.
High Prevalence of Rodent-Borne Bartonella spp. in Urbanizing Environments in Sarawak, Malaysian Borneo.

Rodents are the most prominent animal host of Bartonella spp. , which are associated with an increasing number of human diseases worldwide. Many rodent species thrive in urban environments and live in close contact with people, which can lead to an increased human risk of infection from rodent-borne pathogens. In this study, we explored the prevalence and distribution of Bartonella spp. in rodents in urban, developing, and rural environments surrounding a growing city in Sarawak, Malaysian Borneo. We found that although Bartonella spp. infection was pervasive in most rodent species sampled, prevalence was highest in urban areas and infection was most commonly detected in the predominant indigenous rodent species sampled (Sundamys muelleri). Within the urban environment, parks and remnant green patches were significantly associated with the presence of both S. muelleri and Bartonella spp. , indicating higher localized risk of infection for people using these environments for farming, foraging, or recreation.
Role of gene body methylation in acclimatization and adaptation in a basal metazoan

Significance Whether epigenetics can facilitate persistence of acquired adaptive traits remains hotly debated. Here we use a primitive multicellular animal, a reef-building coral, to address this question for gene body methylation, which is a common epigenetic modification in organisms ranging from sunflowers to humans. We show that gene body methylation is altered by the environment in an unexpectedly simple way that reflects a shifting balance between expressions of housekeeping and environmentally responsive genes, which helps acclimatization. Whether these changes contribute to longer-term adaptation remains an open question: While the majority of fixed epigenetic differences between populations do not appear to be due to fixation of plastic changes, their transgenerational inheritance cannot be completely ruled out. Gene body methylation (GBM) has been hypothesized to modulate responses to environmental change, including transgenerational plasticity, but the evidence thus far has been lacking. Here we show that coral fragments reciprocally transplanted between two distant reefs respond predominantly by increase or decrease in genome-wide GBM disparity: The range of methylation levels between lowly and highly methylated genes becomes either wider or narrower. Remarkably, at a broad functional level this simple adjustment correlated very well with gene expression change, reflecting a shifting balance between expressions of environmentally responsive and housekeeping genes. In our experiment, corals in a lower-quality habitat up-regulated genes involved in environmental responses, while corals in a higher-quality habitat invested more in housekeeping genes. Transplanted fragments showing closer GBM match to local corals attained higher fitness characteristics, which supports GBM’s role in acclimatization. Fixed differences in GBM between populations did not align with plastic GBM changes and were mostly observed in genes with elevated FST, which suggests that they arose predominantly through genetic divergence. However, we cannot completely rule out transgenerational inheritance of acquired GBM states.
Characteristics of interactions at protein segments without non-local intramolecular contacts in the Protein Data Bank

The principle of three-dimensional protein structure formation is a long-standing conundrum in structural biology. A globular domain of a soluble protein is formed by a network of atomic contacts among amino acid residues, but regions without intramolecular non-local contacts are often observed in the protein structure, especially in loop, linker, and peripheral segments with secondary structures. Although these regions can play key roles for protein function as interfaces for intermolecular interactions, their nature remains unclear. Here, we termed protein segments without non-local contacts as floating segments and sought them in tens of thousands of entries in the Protein Data Bank. As a result, we found that 0.72% of residues are in floating segments. Regarding secondary structural elements, coil structures are enriched in floating segments, especially for long segments. Interactions with polypeptides and polynucleotides, but not chemical compounds, are enriched in floating segments. The amino acid preferences of floating segments are similar to those of surface residues, with exceptions; the small side chain amino acids, Gly and Ala, are preferred, and some charged side chains, Arg and His, are disfavored for floating segments compared to surface residues. Our comprehensive characterization of floating segments may provide insights into understanding protein sequence-structure-function relationships.
Characterisation of plasmid-mediated rmtB-1 in Enterobacteriaceae clinical isolates from São Paulo, Brazil

OBJECTIVES The emergence of 16S rRNA methyltranferases (16 RMTAses) has jeopardised the clinical use of aminoglycosides. RmtB is one of the most frequently reported in Gram-negatives worldwide. In this study, we aimed to estimate the frequency of 16S RMTAses encoding genes in Enterobacteriaceae isolated in a three-month period from a tertiary Brazilian hospital. METHODS All Gram-negatives classified as resistant to amikacin, gentamicin, and tobramycin by agar screening were selected for analysis. The presence of 16SRMTases encoding genes was verified by polymerase chain reaction (PCR). Antimicrobial susceptible profile was determined by broth microdilution. The genetic relationship among these isolates was accessed by pulsed field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Selected RmtB-producing isolates were characterised by whole genome sequencing (WGS) analysis. RESULTS Twenty-two of 1,052 (2.1%) Enterobacteriaceae were detected as producers of RmtB-1 [Klebsiella pneumoniae (n = 21) and Proteus mirabilis (n = 1)]. blaKPC-2 was identified among 20 RmtB-1-producing K. pneumoniae isolates that exhibited an identical PFGE and MLST (ST258) patterns. Two K. pneumoniae isolates, the A64216 (not harboring bla KPC-2), A64477 (harboring bla KPC-2) and one P. mirabilis isolate (A64421) were selected for WGS. rmtB-1 and bla KPC-2 genes were carried by distinct plasmids. While a plasmid belonging to the IncFIIk group harbored rmtB-1 in K. pneumoniae, this gene was carried by a non-typable plasmid in P. mirabilis. In the three analysed plasmids, rmtB-1 was inserted on a transposon, downstream a Tn2. CONCLUSION Our findings suggested that the rmtB-1 was harbored by plasmids distinct from those previously reported in Bolivia and China. It suggests that multiple mobilization events might have occurred in South America.
The hospital anxiety and depression scale, in patients with multiple sclerosis

Purpose Multiple sclerosis (MS) is a long-lasting disabling disease with psychological implications. Hospital Anxiety and Depression Scale (HADS) is a questionnaire developed to screen anxiety and depression among patients in hospital settings. The objective of this report was to study the metric properties of the Portuguese version of HADS regarding MS. Materials and methods This was a cross-sectional study. Three hundred and eighty individuals with a diagnosis of MS, according to the revised McDonald criteria, were recruited from an outpatient Neuroimmunology Clinic of the city of Porto. Participants had a mean age of 40.04 years (SD =10.86), 63.9% of them were female. HADS consists of two subscales – anxiety and depression – scored separately, with seven items each. Results The results of our study show the appropriateness of HADS for the identification of mood changes in people with MS and show good metric properties. Conclusion It seems to be a useful tool for the scrutiny of mood disorders in Portuguese patients with MS. The questionnaire is also sensitive to disease when compared to people with other severe illnesses, and with people without the disease.
A dyadic analysis of loneliness and health-related quality of life in Latinas with breast cancer and their informal caregivers

Abstract Objective: The primary aim of this investigation is to provide a novel dyadic test of a model of loneliness and health-related quality of life (HRQoL) in a sample of Latinas with breast cancer and their informal caregivers. Design: At baseline, dyads completed measures of loneliness and HRQoL. At a 3-month follow-up, they returned to complete the HRQoL measure. Associations were tested with the Actor–Partner Interdependence Model. Sample: About 234 Latinas with breast cancer diagnosed within the past year and their informal caregivers participated in the investigation. Findings: Loneliness was concurrently and negatively associated with HRQoL at baseline for both survivors and caregivers. Survivors’ baseline loneliness, controlling for their baseline HRQoL, negatively predicted their HRQoL at 3 months. Survivors’ HRQoL at baseline also predicted caregivers’ HRQoL at 3 months. Conclusion: Loneliness is a risk factor for declines in HRQoL among cancer survivors. Their caregivers are also at risk for degraded HRQoL when the survivor experiences compromised HRQoL. Loneliness complicates the HRQoL of the cancer survivor–caregiver dyad.
Ubiquitin-dependent switch during assembly of the proteasomal ATPases mediated by Not4 ubiquitin ligase

Significance In the proteasome, an essential protease of eukaryotes, the hexameric ATPases (Rpt1-Rpt6) inject ubiquitinated proteins into the proteolytic core particle. Individual Rpt proteins assemble into the hexameric Rpt ring through binding to specific chaperones: Nas2, Hsm3, Rpn14, and Nas6. Here, we show that Rpt ring assembly uses a ubiquitination-mediated control. Not4 ubiquitinates Rpt5 by competing against Nas2 first and then Hsm3 throughout Rpt ring assembly. Thus, Not4 selectively recognizes a Rpt ring that matures without these chaperones. Rpt5 ubiquitination blocks recruitment of Rpn1 ubiquitin receptor and Ubp6 deubiquitinase, halting proteasome assembly and its recognition of ubiquitinated substrates. Our findings reveal an assembly checkpoint where Not4 monitors chaperone actions during hexameric ATPase ring assembly, ensuring the accuracy of proteasome holoenzyme maturation. In the proteasome holoenzyme, the hexameric ATPases (Rpt1-Rpt6) enable degradation of ubiquitinated proteins by unfolding and translocating them into the proteolytic core particle. During early-stage proteasome assembly, individual Rpt proteins assemble into the hexameric “Rpt ring” through binding to their cognate chaperones: Nas2, Hsm3, Nas6, and Rpn14. Here, we show that Rpt ring assembly employs a specific ubiquitination-mediated control. An E3 ligase, Not4, selectively ubiquitinates Rpt5 during Rpt ring assembly. To access Rpt5, Not4 competes with Nas2 until the penultimate step and then with Hsm3 at the final step of Rpt ring completion. Using the known Rpt–chaperone cocrystal structures, we show that Not4-mediated ubiquitination sites in Rpt5 are obstructed by Nas2 and Hsm3. Thus, Not4 can distinguish a Rpt ring that matures without these chaperones, based on its accessibility to Rpt5. Rpt5 ubiquitination does not destabilize the ring but hinders incorporation of incoming subunits—Rpn1 ubiquitin receptor and Ubp6 deubiquitinase—thereby blocking progression of proteasome assembly and ubiquitin regeneration from proteasome substrates. Our findings reveal an assembly checkpoint where Not4 monitors chaperone actions during hexameric ATPase ring assembly, thereby ensuring the accuracy of proteasome holoenzyme maturation.
Intra- and Inter-Day Reliability of Body Composition Assessed by a Commercial Multifrequency Bioelectrical Impedance Meter

Abstract The purpose of this study was to examine the intra- and inter-day reliability of body composition measurements provided by a commercial multifrequency bioelectrical impedance meter. Eighteen healthy, well-trained students in physical education from the same ethnic group were assessed on four consecutive days, both in the morning and in the evening. Indexes provided by the device were gathered in four categories: tissular, metabolic, hydric and ionic blocks. There was no systematic bias between repeated measures, regardless of time of day. Relative reliability was high to very high in the morning (0.72<ICC<0.99) and moderate to very high in the afternoon (0.61<ICC<0.99). Absolute reliability varied substantially between indexes (1.5%<SEM<15%). The minimum difference considered as real was proportionally altered, since it ranged from 4.2 to 41.5%. In conclusion, body composition assessed with a multifrequency bioelectrical impedance meter requires a highly standardized protocol and adjusting the cut-off value for each parameter to ascertain during athlete follow-up that a real change has occurred. This assessment should preferably be scheduled in the morning in order to decrease these cut-off values.