Biosynthesis of man-β-G1cNAc-G1cNAc-pyrophosphoryl-polyprenol by a solubilized enzyme from aorta

The particulate enzyme fraction from pig aorta was treated with Triton X-100 or Nonidet P-40 to yield a soluble enzyme preparation. This solubilized enzyme catalyzed the transfer of mannose from GDP-[¹⁴C]mannose, but not from [¹⁴C]mannosyl-phosphoryl-polyprenol, to G1cNAc-G1cNAc-pyrophosphoryl-polyprenol to form the trisaccharide-lipid, Man-β-GlcNAc-GlcNAc-pyrophosphoryl-polyprenol. The trisaccharide-lipid formed in these reactions was isolated by solvent fractionation and was subjected to mild acid hydrolysis to release the [¹⁴C]trisaccharide. Essentially all of the radioactivity was released from this trisaccharide as mannose upon treatment with β-mannosidase while α-mannosidase had no effect.     

Monoclonal antibody production in hollow fiber bioreactors using serum-free medium

Murine hybridoma cells that produce monoclonal antibody directed against human fibronectin have been cultured in VITAFIBER II and VITAFIBER V hollow fiber bioreactors using defined, serum-free WRC 935 medium. During a two-week growth period, following inoculation of the bioreactors, the cells proliferated to an extent where the bioreactor was filled with cultured cells. Using a 5 sq. ft. VITAFIBER V bioreactor, over 15 grams of antibody were produced during the 40 days of the experiment. This antibody was greater than 95% IgG. During the production period, this packed mass of cells produced 579 +/- 15 mg IgG per day. Because the medium is formulated for air equilibration and high cell densities, WRC 935 medium is especially useful for production of gram quantities of monoclonal antibodies using continuous feed hollow fiber bioreactor cell culture systems.     

COPII collar defines the boundary between ER and ER exit site and does not coat cargo containers

COPII and COPI mediate the formation of membrane vesicles translocating in opposite directions within the secretory pathway. Live-cell and electron microscopy revealed a novel mode of function for COPII during cargo export from the ER. COPII is recruited to membranes defining the boundary between the ER and ER exit sites, facilitating selective cargo concentration. Using direct observation of living cells, we monitored cargo selection processes, accumulation, and fission of COPII-free ERES membranes. CRISPR/Cas12a tagging, the RUSH system, and pharmaceutical and genetic perturbations of ER-Golgi transport demonstrated that the COPII coat remains bound to the ER–ERES boundary during protein export. Manipulation of the cargo-binding domain in COPII Sec24B prohibits cargo accumulation in ERES. These findings suggest a role for COPII in selecting and concentrating exported cargo rather than coating Golgi-bound carriers. These findings transform our understanding of coat proteins’ role in ER-to-Golgi transport.     

Structure-based design and synthesis of potent benzothiazole inhibitors of interleukin-2 inducible T cell kinase (ITK)

Inhibition of the non-receptor tyrosine kinase ITK, a component of the T-cell receptor signalling cascade, may represent a novel treatment for allergic asthma. Here we report the structure-based optimization of a series of benzothiazole amides that demonstrate sub-nanomolar inhibitory potency against ITK with good cellular activity and kinase selectivity. We also elucidate the binding mode of these inhibitors by solving the X-ray crystal structures of several inhibitor-ITK complexes.     

Comparative life cycle sustainability assessment of urban water reuse at various centralization scales

Purpose

Population growth and urbanization lead to increasing water demand, putting significant pressure on natural water sources. The rising amounts of domestic wastewater (WW) in urban areas may be treated to serve as an alternative water source that may alleviate this pressure. This study examines sustainability of utilizing reclaimed domestic wastewater in urban households for toilet flushing and garden irrigation. It models a city characterized by water scarcity, using a coal-based electricity mix.

Methods

Four approaches were compared: (0) Business-as-usual (BAU) alternative, where the central WW treatment plant effluent is discharged to nature; (1) central WW treatment and urban reuse of the effluent produced; (2) semi-distributed greywater treatment and reuse, at cluster scale; (3) Distributed greywater treatment and reuse, at building scale. Environmental life cycle assessment (LCA), social LCA (S-LCA), and life cycle costing (LCC) were applied to the system model of the above scenarios, with seawater desalination as the source for potable water. System boundaries include water supply, WW collection, and treatment facilities. Analytical hierarchy process (AHP), a multi-criteria decision analysis (MCDA) methodology, was integrated into the life cycle sustainability assessment (LCSA) framework as a means for weighting sustainability criteria through judgment elicitation from a panel of 20 experts.

Results and discussion

Environmentally and socially, the two distributed alternatives perform better in most impact categories. Socially, semi-distributed (cluster scale) reuse is somewhat advantageous over the fully distributed alternative (building scale), due to the benefits of community engagement. Economically, the cluster-level scenario is the most preferable, while the building-scale scenario is the least preferable. A hierarchical representation of the problem’s criteria was constructed, according to the principals of AHP. Each criterion was weighted and those of extreme low importance were eliminated, while maintaining the integrity of the experts’ judgments. Weighted and aggregated sustainability scores revealed that cluster level reclamation, under modeled conditions, is the most sustainable option and the BAU scenario is the least sustainable. The other two alternatives, centralized and fully distributed reclamation, obtained similar intermediate scores.

Conclusions

Distributed urban water reuse was found to be more sustainable than current practice. Different alternative solutions are advantageous in different ways, but overall, the reclamation and reuse of greywater at the cluster level seems to be the best option among the three reuse options examined in this assessment. AHP proved an effective method for aggregating the multiple sustainability criteria. The hierarchical view maintains transparency of all local weights while leading to the final weight vector.

     

Epigenetic metaphors: an interdisciplinary translation of encoding and decoding

Looking at the new and often disputed science of epigenetics, we examined the challenges faced by scientists when they communicate scientific research to the public. We focused on the use of metaphors to illustrate notions of epigenetics and genetics. We studied the “encoding” by epigeneticists and “decoding” in focus groups with diverse backgrounds. We observed considerable overlap in the dominant metaphors favored by both researchers and the lay public. However, the groups differed markedly in their interpretations of which metaphors aided understanding or not. We conclude by discussing the role of metaphors and their interpretations in the context of a shift from pre-deterministic genomic metaphors to more active, dynamic and nuanced epigenetic metaphors. These reflections on the choice of metaphors and differences in encoding/decoding are important for science communication and scientific boundary-maintenance.     

The sentineling—Foraging trade‐off in dominant and subordinate Arabian babblers

Many cooperative breeders forage under predation risks, sentineling is a central activity, and groupmates have to balance between sentineling and foraging. The optimal balance between sentinel activity and foraging may differ among dominant and subordinate individuals, as dominants are more efficient foragers. Two theoretical models pertain to this balance and predict when individuals with different foraging abilities should switch between the two activities on the basis of their energetic state. In one of these models, individuals must attain a critical energetic level by dusk to pass the night, and in the second model fitness is monotonically increasing with the energetic state. We tested these models in the cooperatively breeding Arabian babbler, Turdoides squamiceps. We measured the length of sentinel bouts and the gaps between them both in natural conditions and following experimental feeding. Following feeding ad libitum, subordinates expanded their sentinel bouts significantly more than dominants in comparison with natural conditions. These findings are consistent with the first model, but not with the second. In the experiment, we measured the mass of mealworms consumed by each individual following a sentinel bout relative to its body mass. This ratio was larger for subordinates, indicating that they ended their sentinel bouts at a lower energetic state than dominants. This finding is consistent with the second model, but not with the first. Immediately after eating ad libitum, in 62% of the cases the first behavior performed by the babblers was a new sentinel bout, but in 17% it was a mutual interaction with a groupmate, indicating that social interactions also play a role in the trade‐off vis‐à‐vis sentinel activity.     

Glycerol triacetate as solvent and acyl donor in the production of isoamyl acetate with Candida antarctica lipase B

Glycerol triacetate was successfully used as a green solvent and as the acyl donor in the transesterification of isoamyl alcohol to produce isoamyl acetate using free and immobilized Candida antarctica lipase B. Immobilized lipase was more catalytically active than free lipase and could be easily separated from the reaction mixture by filtration. In addition, it was found that increasing either the reaction temperature or the enzyme to substrate ratio increased the conversion of isoamyl alcohol. Using triacetin as the solvent also enabled the separation of product by simple extraction with petroleum ether and catalyst recycling.     

Fracture mechanics analysis of vertical root fracture from condensation of gutta-percha

A two-dimensional fracture mechanics analysis of vertical root fracture (VRF) in single-canal roots from apical condensation of gutta-percha (gp) is developed. The resulting analytic relation for apical load causing VRF agrees with major trends reported in in-vitro tests on roots subjected to either continuous or, the more clinically relevant, repeating vertical condensation of gp. The model explicitly exposes the role of root canal morphology and dentin fracture toughness on VRF. Ovoid and irregular canals are prone to fracture while the effect of mean root canal radius is modest. Canal taper and instrumentation details may affect VRF only marginally and indirectly. The model predicts dentinal cracks to occur following root canal instrumentation and obturation, which may pose long-term threats to tooth integrity.