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The fungal class D1 G-protein-coupled receptor (GPCR) Ste2 has a different arrangement of transmembrane helices compared with mammalian GPCRs and a distinct mode of coupling to the heterotrimeric G protein Gpa1–Ste2–Ste18 1 . In addition, Ste2 lacks conserved sequence motifs such as DRY, PIF and NPXXY, which are associated with the activation of class A GPCRs 2 . This suggested that the activation mechanism of Ste2 may also differ. Here we determined structures of Saccharomyces cerevisiae Ste2 in the absence of G protein in two different conformations bound to the native agonist α-factor, bound to an antagonist and without ligand. These structures revealed that Ste2 is indeed activated differently from other GPCRs. In the inactive state, the cytoplasmic end of transmembrane helix H7 is unstructured and packs between helices H1–H6, blocking the G protein coupling site. Agonist binding results in the outward movement of the extracellular ends of H6 and H7 by 6 Å. On the intracellular surface, the G protein coupling site is formed by a 20 Å outward movement of the unstructured region in H7 that unblocks the site, and a 12 Å inward movement of H6. This is a distinct mechanism in GPCRs, in which the movement of H6 and H7 upon agonist binding facilitates G protein coupling. Cryo-electron microscopy structures of ligand-free, agonist-bound and antagonist-bound Ste2 show that this class D1 G protein-coupled receptor has a distinct mechanism of activation compared with other receptor classes.

Recombinant human collagen (rhCol) holds broad potential in biomedical and industrial applications due to its high purity and low immunogenicity. However, large-scale production of structurally stable and functionally active rhCol remains challenging. A novel strategy integrating collagen sequence optimization and microbial prolyl-4-hydroxylase (P4H) screening was developed to enable efficient production of triple-helical rhCol in Komagataella phaffii . Five Type III collagen variants (ColP1 ~ ColP5) were rationally designed based on interchain salt-bridge engineering to improve structural stability and biological activity, with ColP2 showing superior expression and functionality. A systematic evaluation of four microbial P4Hs identified Bacillus megaterium P4H (BmP4H) as the most effective catalyst for proline hydroxylation, enabling stable triple-helix formation. Combined with strain optimization, promoter and signal peptide screening, and 5-L scale fermentation, this approach achieved a high rhCol yield of 2.54 g/L with confirmed triple-helical structure. These results demonstrate an integrated and scalable platform for high-level production of functional recombinant collagen, providing a promising foundation for its industrial and clinical applications. Key Points • Co-expression of BmP4H enables stable triple-helical collagen in yeast. • Strain X-33, promoter P AOX1 , and a-factor leader optimize collagen secretion. • Scale-up in 5L bioreactor achieves 2.54 g/L rhCol production.

Background β-Caryophyllene is a plant terpenoid with therapeutic and biofuel properties. Production of terpenoids through microbial cells is a potentially sustainable alternative for production. Adaptive laboratory evolution is a complementary technique to metabolic engineering for strain improvement, if the product-of-interest is coupled with growth. Here we use a combination of pathway engineering and adaptive laboratory evolution to improve the production of β-caryophyllene, an extracellular product, by leveraging the antioxidant potential of the compound. Results Using oxidative stress as selective pressure, we developed an adaptive laboratory evolution that worked to evolve an engineered β-caryophyllene producing yeast strain for improved production within a few generations. This strategy resulted in fourfold increase in production in isolated mutants. Further increasing the flux to β-caryophyllene in the best evolved mutant achieved a titer of 104.7 ± 6.2 mg/L product. Genomic analysis revealed a gain-of-function mutation in the a-factor exporter STE6 was identified to be involved in significantly increased production, likely as a result of increased product export. Conclusion An optimized selection strategy based on oxidative stress was developed to improve the production of the extracellular product β-caryophyllene in an engineered yeast strain. Application of the selection strategy in adaptive laboratory evolution resulted in mutants with significantly increased production and identification of novel responsible mutations.

Organic rich sedimentary rocks of the Late Cretaceous Muwaqqar Formation from the Lajjun outcrop in the Lajjun Sub-basin, Western Central Jordan were geochemically analyzed. This study integrates kerogen microscopy of the isolated kerogen from 10 oil shale samples with a new finding from unconventional geochemical methods [i.e., ultimate elemental (CHNS), fourier transform infrared spectroscopy and pyrolysis–gas chromatography (Py–GC)] to decipher the molecular structure of the analyzed isolated kerogen fraction and evaluate the kerogen composition and characteristics. The optical kerogen microscopy shows that the isolated kerogen from the studied oil shales is originated from marine assemblages [i.e., algae, bituminite and fluorescence amorphous organic matter] with minor amounts of plant origin organic matter (i.e., spores). This finding suggests that the studied kerogen is hydrogen-rich kerogen, and has the potential to generate high paraffinic oil with low wax content. The dominance of such hydrogen-rich kerogen (mainly Type II) was confirmed from the multi-geochemical ratios, including high hydrogen/carbon atomic of more than 1.30 and high A-factor of more than 0.60. This claim agrees with the molecular structure of the kerogen derived from Py–GC results, which suggest that the studied kerogen is mainly Type II-S kerogen exhibiting the possibility of producing high sulphur oils during earlier stages of diagenesis, according to bulk kinetic modeling. The kinetic models of the isolated kerogen fraction suggest that the kerogen conversion, in coincidence with a vitrinite reflectance range of 0.55–0.60%, commenced at considerably lower temperature value ranges between 100 and 106 °C, which have produced oils during the early stage of oil generation. The kinetic models also suggest that the commercial amounts of oil can generate by kerogen conversion of up to 50% during the peak stage of oil window (0.71–0.83%) at relatively low geological temperature values in the range of 122–138 °C. Therefore, further development of the Muwaqqar oil shale successions is highly approved in the shallowly buried stratigraphic succession in the Lajjun Sub-basin, Western Central Jordan.

The Leotiomycetes is a hugely diverse group of fungi, accommodating a wide variety of important plant and animal pathogens, ericoid mycorrhizal fungi, as well as producers of antibiotics. Despite their importance, the genetics of these fungi remain relatively understudied, particularly as they don’t include model taxa. For example, sexual reproduction and the genetic mechanisms that underly this process are poorly understood in the Leotiomycetes. We exploited publicly available genomic and transcriptomic resources to identify genes of the mating-type locus and pheromone response pathway in an effort to characterize the mating strategies and behaviors of 124 Leotiomycete species. Our analyses identified a putative a-factor mating pheromone in these species. This significant finding represents the first identification of this gene in Pezizomycotina species outside of the Sordariomycetes. A unique mating strategy was also discovered in Lachnellula species that appear to have lost the need for the primary MAT1-1-1 protein. Ancestral state reconstruction enabled the identification of numerous transitions between homothallism and heterothallism in the Leotiomycetes and suggests a heterothallic ancestor for this group. This comprehensive catalog of mating-related genes from such a large group of fungi provides a rich resource from which in-depth, functional studies can be conducted in these economically and ecologically important species.

Protein prenylation by farnesyltransferase (FTase) is often described as the targeting of a cysteine-containing motif (CaaX) that is enriched for aliphatic amino acids at the a 1 and a 2 positions, while quite flexible at the X position. Prenylation prediction methods often rely on these features despite emerging evidence that FTase has broader target specificity than previously considered. Using a machine learning approach and training sets based on canonical (prenylated, proteolyzed, and carboxymethylated) and recently identified shunted motifs (prenylation only), this study aims to improve prenylation predictions with the goal of determining the full scope of prenylation potential among the 8000 possible Cxxx sequence combinations. Further, this study aims to subdivide the prenylated sequences as either shunted (i.e., uncleaved) or cleaved (i.e., canonical). Predictions were determined for Saccharomyces cerevisiae FTase and compared to results derived using currently available prenylation prediction methods. In silico predictions were further evaluated using in vivo methods coupled to two yeast reporters, the yeast mating pheromone a -factor and Hsp40 Ydj1p, that represent proteins with canonical and shunted CaaX motifs, respectively. Our machine learning-based approach expands the repertoire of predicted FTase targets and provides a framework for functional classification.

Graphics are drawn about the sediment content, water flow, and water level of the Yellow River with time. A cluster scatter plot of sand content is made with changes over time. Using variance analysis of a factor, it is concluded that sediment content is affected by two variables month and year. By studying the changing pattern of water-sand flux, the timing diagram of the water and sediment flux of the Yellow River with time is drawn. The flux of water and sand is predicted for the next two years. According to the prediction results, the optimal sampling and monitoring scheme is developed.

Introduction: Any type of nutritional imbalance experienced during childhood will affect the health of an individual, both in their childhood and their adulthood. Several studies have proved that there is an association between cardiovascular disease (CVD) risk and endocrine and lipid markers at early stages of life. Objective: To establish the relationship between nutritional status (IGF-1 and serum levels of its binding proteins IGFBP-1, IGFBP-2 and IGFBP-3), and CVD risk markers in students aged 7 to 9 years. Materials and methods: Cross-sectional observational study conducted in 84 children attending two schools from Bogotá D.C. and Soacha, Colombia, to identify the relationship between possible variations in CVD risk markers and nutritional status. Sexual development stage, lipid profile, anthropometric data, blood sugar levels and IGF-1 and IGFBP serum levels of all participants were measured. Statistical analysis was conducted using the Pearson’s correlation coefficient, the analysis of variance (ANOVA), and the Kruskall-Wallis, Games-Howell and Dunnett’s tests. The confidence interval and statistical significance were 95% and p<0.05, respectively. Results:IGFBP-1 and IGFBP-2 levels proportionally decreased as weight increased. An inverse correlation between both proteins and triglyceride levels was found, as well as a direct correlation with HDL cholesterol levels. Conclusions: Alterations in CVD risk markers can be identified during childhood. If said alterations are timely detected, it is possible to adopt preventive and therapeutic actions such as the promotion of public policies aimed at preventing childhood overweight and obesity, which in turn will reduce the risk of developing cardiovascular disease in adulthood.

In first, the Ni-doped ZnO nanorods used as an appeal sonocatalyst was synthesized through co-precipitation method. Afterwards, the crystalline structure, functional groups, surface morphology, and elemental composition were characterized by a set of analysis. Removal of diazinon ((DZ) as a renowned pesticide) was investigated using sonocatalytic performance of US/Ni-doped ZnO system. In this empirical study, response surface methodology (RSM) based central composite design (CCD) was applied for optimization of operational factors. Under the optimum conditions such as initial pH = 5, initial DZ concentration = 15 mg L −1 , sonocatalyst dosage = 1 g L −1 , and in the presence of organic compounds (oxalic acid, humic acid, and folic acid) = 3 mg L −1 , the sonocatalytic degradation of DZ after 15 min was 82.29%. The F -value (6.64) and P -value (< 0.0001) for DZ degradation in the quadratic model imply the proposed model was significant. A -factor (pH) considers as a prominent factor owing to having the highest F -value. In addition, the sonocatalytic data in this study exhibited valid fitting for the first order kinetic model ( R 2  > 0.98). After six consecutive cycles, the Ni-doped ZnO nanorods could be recyclable for sonocatalytic degradation of DZ. The five main compounds produced during the US/Ni-doped ZnO embracing 2-isopropyl-6-methyl-4-pyrimidinol (IMP), diethyl phosphonate, diazoxon, hydroxyldiazinon, and diazinon methyl ketone are formed in the path of DZ degradation. OFAT style also revealed 99.99% of DZ degradation with 73.26% of mineralization rate in optimum status. The Ni-doped ZnO presented agreeable sonocatalytic facility in the refinement of real water and wastewater matrix. Finally, the results of toxicity evaluation ( Daphnia magna ) in the sonocatalytic degradation of DZ (by US/Ni-doped ZnO system) showed that the toxicity of the DZ solution lessened under US waves (LC 50 and TU 48 h equal to 36.472 and 2.741 volume percent, respectively).

BACKGROUND : Huntiella resides in the Ceratocystidaceae, a family of fungi that accommodates important plant pathogens and insect-associated saprotrophs. Species in the genus have either heterothallic or unisexual (a form of homothallism) mating systems, providing an opportunity to investigate the genetic mechanisms that enable transitions between reproductive strategies in related species. Two newly sequenced Huntiella genomes are introduced in this study and comparative genomics and transcriptomics tools are used to investigate the differences between heterothallism and unisexuality across the genus. RESULTS : Heterothallic species harbored up to seven copies of the a-factor pheromone, each of which possessed numerous mature peptide repeats. In comparison, unisexual Huntiella species had only two or three copies of this gene, each with fewer repeats. Similarly, while the heterothallic species expressed up to 12 copies of the mature α-factor pheromone, unisexual species had up to six copies. These significant differences imply that unisexual Huntiella species do not rely on a mating partner recognition system in the same way that heterothallic fungi do. CONCLUSION : While it is suspected that mating type-independent pheromone expression is the mechanism allowing for unisexual reproduction in Huntiella species, our results suggest that the transition to unisexuality may also have been associated with changes in the genes governing the pheromone pathway. While these results are specifically related to Huntiella, they provide clues leading to a better understanding of sexual reproduction and the fluidity of mating strategies in fungi more broadly.