G protein-coupled receptor endocytosis generates spatiotemporal bias in ß-arrestin signaling.

The stabilization of different active conformations of G protein-coupled receptors is thought to underlie the varying efficacies of biased and balanced agonists. Here, profiling the activation of signal transducers by angiotensin II type 1 receptor (AT1R) agonists revealed that the extent and kinetics of ß-arrestin binding exhibited substantial ligand-dependent differences, which were lost when receptor internalization was inhibited. When AT1R endocytosis was prevented, even weak partial agonists of the ß-arrestin pathway acted as full or near-full agonists, suggesting that receptor conformation did not exclusively determine ß-arrestin recruitment. The ligand-dependent variance in ß-arrestin translocation was much larger at endosomes than at the plasma membrane, showing that ligand efficacy in the ß-arrestin pathway was spatiotemporally determined. Experimental investigations and mathematical modeling demonstrated how multiple factors concurrently shaped the effects of agonists on endosomal receptor-ß-arrestin binding and thus determined the extent of functional selectivity. Ligand dissociation rate and G protein activity had particularly strong, internalization-dependent effects on the receptor-ß-arrestin interaction. We also showed that endocytosis regulated the agonist efficacies of two other receptors with sustained ß-arrestin binding: the V2 vasopressin receptor and a mutant ß2-adrenergic receptor. In the absence of endocytosis, the agonist-dependent variance in ß-arrestin2 binding was markedly diminished. Our results suggest that endocytosis determines the spatiotemporal bias in GPCR signaling and can aid in the development of more efficacious, functionally selective compounds.

ArreSTick motif controls ß-arrestin-binding stability and extends phosphorylation-dependent ß-arrestin interactions to non-receptor proteins.

The binding and function of ß-arrestins are regulated by specific phosphorylation motifs present in G protein-coupled receptors (GPCRs). However, the exact arrangement of phosphorylated amino acids responsible for establishing a stable interaction remains unclear. We employ a 1D sequence convolution model trained on GPCRs with established ß-arrestin-binding properties. With this approach, amino acid motifs characteristic of GPCRs that form stable interactions with ß-arrestins can be identified, a pattern that we name "arreSTick." Intriguingly, the arreSTick pattern is also present in numerous non-receptor proteins. Using proximity biotinylation assay and mass spectrometry analysis, we demonstrate that the arreSTick motif controls the interaction between many non-receptor proteins and ß-arrestin2. The HIV-1 Tat-specific factor 1 (HTSF1 or HTATSF1), a nuclear transcription factor, contains the arreSTick pattern, and its subcellular localization is influenced by ß-arrestin2. Our findings unveil a broader role for ß-arrestins in phosphorylation-dependent interactions, extending beyond GPCRs to encompass non-receptor proteins as well.

A dual role of lysophosphatidic acid type 2 receptor (LPAR2) in nonsteroidal anti-inflammatory drug-induced mouse enteropathy.

Lysophosphatidic acid (LPA) is a bioactive phospholipid mediator that has been found to ameliorate nonsteroidal anti-inflammatory drug (NSAID)-induced gastric injury by acting on lysophosphatidic acid type 2 receptor (LPAR2). In this study, we investigated whether LPAR2 signaling was implicated in the development of NSAID-induced small intestinal injury (enteropathy), another major complication of NSAID use. Wild-type (WT) and Lpar2 deficient (Lpar2-/-) mice were treated with a single, large dose (20 or 30 mg/kg, i.g.) of indomethacin (IND). The mice were euthanized at 6 or 24 h after IND treatment. We showed that IND-induced mucosal enteropathy and neutrophil recruitment occurred much earlier (at 6 h after IND treatment) in Lpar2-/- mice compared to WT mice, but the tissue levels of inflammatory mediators (IL-1ß, TNF-α, inducible COX-2, CAMP) remained at much lower levels. Administration of a selective LPAR2 agonist DBIBB (1, 10 mg/kg, i.g., twice at 24 h and 30 min before IND treatment) dose-dependently reduced mucosal injury and neutrophil activation in enteropathy, but it also enhanced IND-induced elevation of several proinflammatory chemokines and cytokines. By assessing caspase-3 activation, we found significantly increased intestinal apoptosis in IND-treated Lpar2-/- mice, but it was attenuated after DBIBB administration, especially in non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice. Finally, we showed that IND treatment reduced the plasma activity and expression of autotaxin (ATX), the main LPA-producing enzyme, and also reduced the intestinal expression of Lpar2 mRNA, which preceded the development of mucosal damage. We conclude that LPAR2 has a dual role in NSAID enteropathy, as it contributes to the maintenance of mucosal integrity after NSAID exposure, but also orchestrates the inflammatory responses associated with ulceration. Our study suggests that IND-induced inhibition of the ATX-LPAR2 axis is an early event in the pathogenesis of enteropathy.

[How did the survival of acute myeloid leukemia change over the last ten years in our unit?] / Hogyan változott az akut myeloid leukaemiás betegek túlélése a terápiás lehetoségek bovülésével az elmúlt 10 évben klinikánkon?

INTRODUCTION: Acute myeloid leukemia (AML) is a hematological malignancy with high mortality rate. The treatment is especially challenging in patients older than 65 years, which is the large majority of those. For patients unfit for intensive chemotherapy regimens, only palliative cytoreduction and basic supportive care used to be the options in our unit. However, from 2018, the azacitidine-venetoclax combination has been a new therapeutic alternative. This treatment resulted in marked survival benefit in clinical trials, however, its impact on the daily clinical practice and the entire patient population is unclear. OBJECTIVE: Our goal was to evaluate how the application of azacitidine-venetoclax changed the treatment and survival of AML patients in our practice. METHOD: We retrospectively analyzed the available clinical data of all AML patients treated consecutively between January 1, 2011 and December 31, 2021 at the 3rd Department of Internal Medicine (from 2020 onward called Department of Internal Medicine and Hematology), examining their treatment depending on the time period of therapy (2011-2017 and 2018-2021). Patients with acute promyelocytic leukemia were excluded. RESULTS: 423 patients were diagnosed during this period. The number of cases showed a marked increase: in the first 7 years of our study, 184 patients were diagnosed, while this rose to 239 during the subsequent 4 years. The median age of patients was 67.6 years, with more than 60% of patients aged over 65. An improving trend can be observed in the overall survival: between 2011 and 2017, the median overall survival was 4.8 ± 0.9 months, while between 2018 and 2021, it was 8.3 ± 1.4 months (p = 0.051). Moreover, in the case of patients over 65 there was a significant overall survival improvement: 3.1 ± 0.5 vs. 4.9 ± 0.6 months (p = 0,01). The main factor behind this improvement could be that a large proportion of over 65 patients previously only fit for supportive care could now be treated with azacitidine-venetoclax: the percentage of actively treated patients grew from 57.1% to 75.3% in the second period. CONCLUSION: The survival of patients unfit for curative therapy and older than 65 showed a steady increase which can be attributed to the introduction of new therapeutic alternatives. Orv Hetil. 2023; 164(45): 1787-1794.

Food safety knowledge and climate in the university canteens of three European countries.

The association of food safety knowledge and climate with gender, education level, length of employment, food safety training, and professional role was measured using a 15-item food safety climate survey and a 20-item food safety questionnaire on a sample of 263 employees from 19 small and medium-sized university canteens in Croatia, Hungary, and Portugal. The relationship between knowledge and climate and the demographic determinants of both variables were examined. Food safety knowledge was inadequate (45.5% of correct responses), while perceptions of food safety, as measured by the food safety climate survey, were positive (2.69 out of a maximum of 3.00). The perception of resources in canteens was the least favorable across all countries. Leaders did not exhibit better food safety knowledge or perceptions. Food safety climate and knowledge were significantly positively correlated and influenced by training. Perceptions of food safety compared to employee knowledge levels indicated that some employees were overly optimistic about food safety risks. Therefore, food safety knowledge and food safety climate should be assessed in parallel, and both could be improved through ongoing training of employees, especially leaders.

Hybrid Prediction-Driven High-Throughput Sustainability Screening for Advancing Waste-to-Dimethyl Ether Valorization.

Assessing the prospective climate preservation potential of novel, innovative, but immature chemical production techniques is limited by the high number of process synthesis options and the lack of reliable, high-throughput quantitative sustainability pre-screening methods. This study presents the sequential use of data-driven hybrid prediction (ANN-RSM-DOM) to streamline waste-to-dimethyl ether (DME) upcycling using a set of sustainability criteria. Artificial neural networks (ANNs) are developed to generate in silico waste valorization experimental results and ex-ante model the operating space of biorefineries applying the organic fraction of municipal solid waste (OFMSW) and sewage sludge (SS). Aspen Plus process flowsheeting and ANN simulations are postprocessed using the response surface methodology (RSM) and desirability optimization method (DOM) to improve the in-depth mechanistic understanding of environmental systems and identify the most benign configurations. The hybrid prediction highlights the importance of targeted waste selection based on elemental composition and the need to design waste-specific DME synthesis to improve techno-economic and environmental performances. The developed framework reveals plant configurations with concurrent climate benefits (-1.241 and -2.128 kg CO2-eq (kg DME)-1) and low DME production costs (0.382 and 0.492 € (kg DME)-1) using OFMSW and SS feedstocks. Overall, the multi-scale explorative hybrid prediction facilitates early stage process synthesis, assists in the design of block units with nonlinear characteristics, resolves the simultaneous analysis of qualitative and quantitative variables, and enables the high-throughput sustainability screening of low technological readiness level processes.

Cannabinoid receptor type 1 (CB1R) inhibits hypothalamic leptin signaling via ß-arrestin1 in complex with TC-PTP and STAT3.

Molecular interactions between anorexigenic leptin and orexigenic endocannabinoids, although of great metabolic significance, are not well understood. We report here that hypothalamic STAT3 signaling in mice, initiated by physiological elevations of leptin, is diminished by agonists of the cannabinoid receptor 1 (CB1R). Measurement of STAT3 activation by semi-automated confocal microscopy in cultured neurons revealed that this CB1R-mediated inhibition requires both T cell protein tyrosine phosphatase (TC-PTP) and ß-arrestin1 but is independent of changes in cAMP. Moreover, ß-arrestin1 translocates to the nucleus upon CB1R activation and binds both STAT3 and TC-PTP. Consistently, CB1R activation failed to suppress leptin signaling in ß-arrestin1 knockout mice in vivo, and in neural cells deficient in CB1R, ß-arrestin1 or TC-PTP. Altogether, CB1R activation engages ß-arrestin1 to coordinate the TC-PTP-mediated inhibition of the leptin-evoked neuronal STAT3 response. This mechanism may restrict the anorexigenic effects of leptin when hypothalamic endocannabinoid levels rise, as during fasting or in diet-induced obesity.

Extensive comparison of methods for removal of organic halogen compounds from pharmaceutical process wastewaters with life cycle, PESTLE, and multi-criteria decision analyses.

Recycling and disposing wastewater from the pharmaceutical industry are of utmost importance in mitigating chemical waste generation, where unmanaged hazardous waste fluxes could cause massive environmental damage. Air stripping, steam stripping, distillation, and incineration offer significant emission reduction potentials for pharmaceutical applications; however, selecting specific process units is a complicated task due to the high number of influencing screening criteria. The mentioned chemical processes are modelled with the Aspen Plus program. This study examines the environmental impacts of adsorbable organic halogens (AOX) containing pharmaceutical process wastewater disposal by conducting life cycle impact assessments using the Product Environmental Footprint (PEF), IMPACT World + Endpoint V1.01, and Recipe 2016 Endpoint (H) V1.06 methods. The results show that the distillation-based separation of AOX compounds is characterized by the most favourable climate change impact and outranks the PEF single score of air stripping, steam stripping, and incineration by 6.3%, 29.1%, 52.0%, respectively. The energy-intensive distillation technology is further evaluated by considering a wide selection of energy sources (i.e., fossil fuel, nuclear, solar, wind onshore, and wind offshore) using PESTLE (Political, Economic, Social, Technological, Legal, Environmental) analysis combined with multi-criteria decision support to determine the most beneficial AOX disposal scenario. The best overall AOX regeneration performance and lowest climate change impact (7.25 × 10-3 kg CO2-eq (1 kg purified wastewater)-1) are obtained by supplying variable renewable electricity from onshore wind turbines, reaching 64.87% carbon emission reduction compared to the baseline fossil fuel-based process alternative.

[Venetoclax-based salvage in multiple myeloma with (11;14) translocation after suboptimal response to first-line therapy]. / Venetoclaxalapú mentokezelés (11;14)-transzlokációs myeloma multiplexben az elso vonalbeli kezelésre adott nem megfelelo válasz esetén.

INTRODUCTION: Multiple myeloma is one of the most common hematologic malignancies, with approximately 400 patients diagnosed in Hungary annually. Novel therapies emerging in the last decade have made a great impact on most patients' survival, however, those responding poorly to standard first-line therapy and failing to proceed to stem cell transplantation face a dire prognosis. Venetoclax, a selective Bcl-2 inhibitor has been shown very effective in the treatment of relapsed/refractory t(11;14) patients, but there are only a few studies about its safety and efficacy when used as salvage in the second line. OBJECTIVE: The aim of our study was to analyze the data of t(11;14) patients treated with venetoclax salvage at our clinic and to evaluate its efficacy. METHOD: Between 2017 and 2021, 13 patients received venetoclax therapy at our clinic after suboptimal response to frontline treatment, whose data we analyzed retrospectively. RESULTS: Adverse prognostic markers were very prevalent in our group, 4 of our patients had del(17p), 5 had amp(1q21) and 6 had stage 3. Nevertheless, all 13 patients responded well to venetoclax therapy, with 6 reaching very good partial response and 7 complete response. All eligible patients (10) could proceed to transplantation. After median 38 months follow up, neither median progression-free, nor median overall survival was reached, since only 3 patients progressed and 1 died. CONCLUSION: We have shown that when salvage is needed for t(11;14) patients who respond suboptimally to standard frontline therapy, venetoclax is a remarkably good option. Orv Hetil. 2023; 164(23): 894-899.

Soluble B-cell maturation antigen as a monitoring marker for multiple myeloma.

Objective: Response to treatment in multiple myeloma (MM) is routinely measured by serum and urine M-protein and free light chain (FLC), as described by the International Myeloma Working Group (IMWG) consensus statement. A non-negligible subgroup of patients however present without measurable biomarkers, others become oligo or non-secretory during recurrent relapses. The aim of our research was to evaluate soluble B-cell maturation antigen (sBCMA) as a monitoring marker measured concurrent with the standard monitoring in MM patients at diagnosis, at relapse and during follow up, in order to establish its potential usefulness in oligo and non-secretory disease. Method: sBCMA levels were measured in 149 patients treated for plasma cell dyscrasia (3 monoclonal gammopathy of unknown significance, 5 smoldering myeloma, 7 plasmacytoma, 8 AL amyloidosis and 126 MM) and 16 control subjects using a commercial ELISA kit. In 43 newly diagnosed patients sBCMA levels were measured at multiple timepoints during treatment, and compared to conventional IMWG response and progression free survival (PFS). Results: sBCMA levels among control subjects were significantly lower than among newly diagnosed or relapsed MM patients [20.8 (14.7-38.7) ng/mL vs. 676 (89.5-1,650) and 264 (20.7-1,603) ng/mL, respectively]. Significant correlations were found between sBCMA and the degree of bone marrow plasma cell infiltration. Out of the 37 newly diagnosed patients who have reached partial response or better per IMWG criteria, 33 (89%) have had at least a 50% drop in sBCMA level by therapy week 4. Cohorts made similarly to IMWG response criteria-achieving a 50% or 90% drop in sBCMA levels compared to level at diagnosis-had statistically significant differences in PFS. Conclusion: Our results confirmed that sBCMA levels are prognostic at important decision points in myeloma, and the percentage of BCMA change is predictive for PFS. This highlights the great potential use of sBCMA in oligo- and non-secretory myeloma.

Methods to assess the effectiveness and acceptance of information and communication technology-based assistive technology for older adults: a scoping review.

An aging society is a growing challenge for families, social and rehabilitation service providers, and economies. Information and communication technology-based assistive technology can bolster the independence of older adults (65 years and above) and reduce their burden on caregivers. Currently, there is no unified methodology to assess the effectiveness and acceptance of these technologies. The present study undertakes a scoping review to (1) identify and characterize the methods for assessing the acceptability and usability of information and communication technology-based assistive technologies, (2) explore the advantages and disadvantages of the assessment methods, (3) determine the possibilities of combining the assessment methods and (4) define the most commonly used assessment method and set of outcome measures. The literature was searched in MEDLINE, Scopus, IEEE Cochrane and Web of Science bibliographic databases using the keywords defined by reviewers for articles in English published between 2011 and 2021. Of the 1696 matches, 31 met the inclusion criteria. It was found that a combination of different assessment methods was common in outcome measurements. Of the 31 studies, assessment methods were combined in 21 studies and multiple questionnaires were used in 11 studies. The most common technique of outcome measurement was the use of questionnaires (81%), conducting interviews (48%) and recording usability-performance measures (39%). The advantages and disadvantages of the assessment methods could not be determined in the selected studies in this scoping review.

Spectral and Redox Properties of a Recombinant Mouse Cytochrome b561 Protein Suggest Transmembrane Electron Transfer Function.

Cytochrome b561 proteins (CYB561s) are integral membrane proteins with six trans-membrane domains, two heme-b redox centers, one on each side of the host membrane. The major characteristics of these proteins are their ascorbate reducibility and trans-membrane electron transferring capability. More than one CYB561 can be found in a wide range of animal and plant phyla and they are localized in membranes different from the membranes participating in bioenergization. Two homologous proteins, both in humans and rodents, are thought to participate-via yet unidentified way-in cancer pathology. The recombinant forms of the human tumor suppressor 101F6 protein (Hs_CYB561D2) and its mouse ortholog (Mm_CYB561D2) have already been studied in some detail. However, nothing has yet been published about the physical-chemical properties of their homologues (Hs_CYB561D1 in humans and Mm_CYB561D1 in mice). In this paper we present optical, redox and structural properties of the recombinant Mm_CYB561D1, obtained based on various spectroscopic methods and homology modeling. The results are discussed in comparison to similar properties of the other members of the CYB561 protein family.

An Unexpected Enzyme in Vascular Smooth Muscle Cells: Angiotensin II Upregulates Cholesterol-25-Hydroxylase Gene Expression.

Angiotensin II (AngII) is a vasoactive peptide hormone, which, under pathological conditions, contributes to the development of cardiovascular diseases. Oxysterols, including 25-hydroxycholesterol (25-HC), the product of cholesterol-25-hydroxylase (CH25H), also have detrimental effects on vascular health by affecting vascular smooth muscle cells (VSMCs). We investigated AngII-induced gene expression changes in VSMCs to explore whether AngII stimulus and 25-HC production have a connection in the vasculature. RNA-sequencing revealed that Ch25h is significantly upregulated in response to AngII stimulus. The Ch25h mRNA levels were elevated robustly (~50-fold) 1 h after AngII (100 nM) stimulation compared to baseline levels. Using inhibitors, we specified that the AngII-induced Ch25h upregulation is type 1 angiotensin II receptor- and Gq/11 activity-dependent. Furthermore, p38 MAPK has a crucial role in the upregulation of Ch25h. We performed LC-MS/MS to identify 25-HC in the supernatant of AngII-stimulated VSMCs. In the supernatants, 25-HC concentration peaked 4 h after AngII stimulation. Our findings provide insight into the pathways mediating AngII-induced Ch25h upregulation. Our study elucidates a connection between AngII stimulus and 25-HC production in primary rat VSMCs. These results potentially lead to the identification and understanding of new mechanisms in the pathogenesis of vascular impairments.

Utilizing the 1H-15N NMR Methods for the Characterization of Isomeric Human Milk Oligosaccharides.

Human milk oligosaccharides (HMOs) are structurally complex unconjugated glycans that are the third largest solid fraction in human milk after lactose and lipids. HMOs are in the forefront of research since they have been proven to possess beneficial health effects, especially on breast-fed neonates. Although HMO research is a trending topic nowadays, readily available analytical methods suitable for the routine investigation of HMOs are still incomplete. NMR spectroscopy provides detailed structural information that can be used to indicate subtle structural differences, particularly for isomeric carbohydrates. Herein, we propose an NMR-based method to identify the major isomeric HMOs containing GlcNAc and/or Neu5Ac building blocks utilizing their amide functionality. Experimental conditions were optimized (H2O:D2O 9:1 v/v solvent at pH 3.0) to obtain 1H-15N HSQC and 1H-15N HSQC-TOCSY NMR spectra of the aforementioned building blocks in HMOs. Four isomeric HMO pairs, LNT/LNnT, 3'SL/6'SL, LNFP II/LNFP III, and LSTa/LSTb, were investigated, and complete NMR resonance assignments were provided. In addition, 1H and 15N NMR resonances were found to be indicative of various linkages, thereby facilitating the distinction of isomeric tri-, tetra-, and pentasaccharide HMOs. The rapid growth of HMO products (from infant formulas and dietary supplements to cosmetics) undoubtedly requires expanding the range of applicable analytical methods. Thus, our work provides a 15N NMR-based method to advance this challenging field of carbohydrate analysis.

Life Cycle, PESTLE and Multi-Criteria Decision Analysis of Membrane Contactor-Based Nitrogen Recovery Process.

Nitrogen is one of the most critical nutrients in the biosphere, and it is an essential nutrient for plant growth. Nitrogen exists in the atmosphere vastly as a gaseous form, but only reactive nitrogen is usable for plants. It is a valuable resource and worth recovering in the wastewater sector. The aim of this work was to prepare a comprehensive environmental analysis of a novel membrane contactor-based process, which is capable of highly efficient nitrogen removal from wastewater. Life cycle assessment (LCA), PESTLE and multi-criteria decision analysis (MCDA) were applied to evaluate the process. The EF 3.0 method, preferred by the European Commission, IMPACT World+, ReCiPe 2016 and IPCC 2021 GWP100 methods were used with six different energy resources-electricity high voltage, solar, nuclear, heat and power and wind energy. The functional unit of 1 m3 of water product was considered as output and "gate-to-gate" analysis was examined. The results of our study show that renewable energy resources cause a significantly lower environmental load than traditional energy resources. TOPSIS score was used to evaluate the alternatives in the case of MCDA. For the EU region, the most advantageous option was found to be wind energy onshore with a score of 0.76, and the following, nuclear, was 0.70.

Effects of Energy Intensification of Pressure-Swing Distillation on Energy Consumption and Controllability.

The aim of process integration is the efficient use of energy and natural resources. However, process integration can result in a more precise process operation, that is, it influences controllability. Pressure-swing distillation processes are designed for the separation of azeotropic mixtures, but their inherent heat integration option can be utilized to significantly reduce their energy consumption. One maximum-boiling and three minimum-boiling azeotropes are considered to study and compare the nonintegrated and integrated alternatives with the tool of mathematical modeling where ASPEN Plus and MATLAB software are used. The results show that the heat-integrated alternatives result in 32-45% energy savings that are proportional to the emission reduction and the consumption of natural resources. As far as the operability is concerned, the heat-integrated alternatives show worse controllability features than the nonintegrated base case. This can be due to the loss of one controllability degree of freedom. This recommends using more sophisticated control structures for the sake of safe operation if process integration is applied.

A Technology-Based Intervention to Support Older Adults in Living Independently: Protocol for a Cross-National Feasibility Pilot.

Innovative technologies can support older adults with or without disabilities, allowing them to live independently in their environment whilst monitoring their health and safety conditions and thereby reducing the significant burden on caregivers, whether family or professional. This paper discusses the design of a study protocol to evaluate the acceptance, usability, and efficiency of the SAVE system, a custom-developed information technology-based elderly care system. The study will involve older adults (aged 65 or older), professional and lay caregivers, and care service decision-makers representing all types of users in a care service scenario. The SAVE environmental sensors, smartwatches, smartphones, and Web service application will be evaluated in people's homes situated in Romania, Italy, and Hungary with a total of 165 users of the three types (cares, elderly, and admin). The study design follows the mixed method approach, using standardized tests and questionnaires with open-ended questions and logging all the data for evaluation. The trial is registered to the platform ClinicalTrials.gov with the registration number NCT05626556. This protocol not only guides the participating countries but can be a feasibility protocol suitable for evaluating the usability and quality of similar systems.

The genome loading model for the origin and maintenance of sex in eukaryotes.

Understanding why sexual reproduction-which involves syngamy (union of gametes) and meiosis-emerged and how it has subsisted for millions of years remains a fundamental problem in biology. Considered as the essence of sex, meiotic recombination is initiated by a DNA double-strand break (DSB) that forms on one of the pairing homologous chromosomes. This DNA lesion is subsequently repaired by gene conversion, the non-reciprocal transfer of genetic information from the intact homolog. A major issue is which of the pairing homologs undergoes DSB formation. Accumulating evidence shows that chromosomal sites where the pairing homologs locally differ in size, i.e., are heterozygous for an insertion or deletion, often display disparity in gene conversion. Biased conversion tends to duplicate insertions and lose deletions. This suggests that DSB is preferentially formed on the "shorter" homologous region, which thereby acts as the recipient for DNA transfer. Thus, sex primarily functions as a genome (re)loading mechanism. It ensures the restoration of formerly lost DNA sequences (deletions) and allows the efficient copying and, mainly in eukaryotes, subsequent spreading of newly emerged sequences (insertions) arising initially in an individual genome, even if they confer no advantage to the host. In this way, sex simultaneously repairs deletions and increases genetic variability underlying adaptation. The model explains a remarkable increase in DNA content during the evolution of eukaryotic genomes.

Quinone binding site in a type VI sulfide:quinone oxidoreductase.

Monotopic membrane-bound flavoproteins, sulfide:quinone oxidoreductases (SQRs), have a variety of physiological functions, including sulfide detoxification. SQR enzymes are classified into six groups. SQRs use the flavin adenine dinucleotide (FAD) cofactor to transfer electrons from sulfide to quinone. A type VI SQR of the photosynthetic purple sulfur bacterium, Thiocapsa roseopersicina (TrSqrF), has been previously characterized, and the mechanism of sulfide oxidation has been proposed. This paper reports the characterization of quinone binding site (QBS) of TrSqrF composed of conserved aromatic and apolar amino acids. Val331, Ile333, and Phe366 were identified near the benzoquinone ring of enzyme-bound decylubiquinone (dUQ) using the TrSqrF homology model. In silico analysis revealed that Val331 and Ile333 alternately connected with the quinone head group via hydrogen bonds, and Phe366 and Trp369 bound the quinones via hydrophobic interactions. TrSqrF variants containing alanine (V331A, I333A, F366A) and aromatic amino acid (V331F, I333F, F366Y), as well as a C-terminal α-helix deletion (CTD) mutant were generated. These amino acids are critical for quinone binding and, thus, catalysis. Spectroscopic analyses proved that all mutants contained FAD. I333F replacement resulted in the lack of the charge transfer complex. In summary, the interactions described above maintain the quinone molecule's head in an optimal position for direct electron transfer from FAD. Surprisingly, the CTD mutant retained a relatively high level of specific activity while remaining membrane-anchored. This is a unique study because it focuses on the QBS and the oxidative stage of a type VI sulfide-dependent quinone reduction. KEY POINTS: • V331, I333, F366, and W369 were shown to interact with decylubiquinone in T. roseopersicina SqrF • These amino acids are involved in proper positioning of quinones next to FAD • I333 is essential in formation of a charge transfer complex from FAD to quinone.

Improving green hydrogen production from Chlorella vulgaris via formic acid-mediated hydrothermal carbonisation and neural network modelling.

This study investigates the formic acid-mediated hydrothermal carbonisation (HTC) of microalgae biomass to enhance green hydrogen production. The effects of combined severity factor (CSF) and feedstock-to-suspension ratio (FSR) are examined on HTC gas formation, hydrochar yield and quality, and composition of the liquid phase. The hydrothermal conversion of Chlorella vulgaris was investigated in a CSF and FSR range of -2.529 and 2.943; and 5.0 wt.% - 25.0 wt.%. Artificial neural networks (ANNs) were developed based on experimental data to model and analyse the HTC process. The results show that green hydrogen formation can be increased up to 3.04 mol kg-1 by applying CSF 2.433 and 12.5 wt.% FSR reaction conditions. The developed ANN model (BR-2-11-9-11) describes the hydrothermal process with high testing and training performance (MSEz = 1.71E-06 & 1.40E-06) and accuracy (R2 = 0.9974 & R2 = 0.9781). The enhanced H2 yield indicates an effective alternative green hydrogen production scenario at low temperatures using high-moisture-containing biomass feedstocks.