Understanding the metabolic burden of recombinant antibody production in Saccharomyces cerevisiae using a quantitative metabolomics approach

The cellular changes induced by heterologous protein expression in the yeast Saccharomyces cerevisiae have been analysed on many levels and found to be significant. However, even though high‐level protein production poses a metabolic burden, evaluation of the expression host at the level of the metabolome has often been neglected. We present a comparison of metabolite profiles of a wild‐type strain with those of three strains producing recombinant antibody variants of increasing size and complexity: an scFv fragment, an scFv–Fc fusion protein and a full‐length IgG molecule. Under producing conditions, all three recombinant strains showed a clear decrease in growth rate compared with the wild‐type strain and the severity of the growth phenotype increased with size of the protein. The levels of 76 intracellular metabolites were determined using a targeted (semi) quantitative mass spectrometry based approach. Based on unsupervised and supervised multivariate analysis of metabolite profiles, together with pathway activity profiling, the recombinant strains were found to be significantly different from each other and from the wild‐type strain. We observed the most prominent changes in metabolite levels for metabolites involved in amino acid and redox metabolism. Induction of the unfolded protein response was detected in all producing strains and is considered to be a contributing factor to the overall metabolic burden on the cells.     

A Novel Approach for Bolted T-Stub Connections

This paper reports an investigation into new connection types and their behaviors determined using full-scale experiments. T-shaped connections were created using the IPE standard profile. The aim of this study was to analyze the influence of T connections based on the IPE standard profile, height of beam to height of T-stub joint (H) of T-stub joints, and lengths (X) of T-stub joints on the behavior of steel connections, in order to provide the necessary data for improving Eurocode 3 and enable efficient use of residue IPE standard profiles and back to the consumption cycle. While the moment resistance values increased with an increase in H from H_min to H_max in model groups with X of 126 mm, and the energy dissipation increased with an increase in H from H_min to H_max and also with an increase in the lengths (X) of T-stub joints from 54 to 126 mm.     

Airborne bacterial assemblage in a zero carbon building: A case study

Currently, there is little information pertaining to the airborne bacterial communities of green buildings. In this case study, the air bacterial community of a zero carbon building (ZCB) in Hong Kong was characterized by targeting the bacterial 16S rRNA gene. Bacteria associated with the outdoor environment dominated the indoor airborne bacterial assemblage, with a modest contribution from bacteria associated with human skin. Differences in overall community diversity, membership, and composition associated with short (day‐to‐day) and long‐term temporal properties were detected, which may have been driven by specific environmental genera and taxa. Furthermore, time‐decay relationships in community membership (based on unweighted UniFrac distances) and composition (based on weighted UniFrac distances) differed depending on the season and sampling location. A Bayesian source‐tracking approach further supported the importance of adjacent outdoor air bacterial assemblage in sourcing the ZCB indoor bioaerosol. Despite the unique building attributes, the ZCB microbial assemblage detected and its temporal characteristics were not dissimilar to that of conventional built environments investigated previously. Future controlled experiments and microbial assemblage investigations of other ZCBs will undoubtedly uncover additional knowledge related to how airborne bacteria in green buildings may be influenced by their distinctive architectural attributes.     

GOLPH3 Regulates EGFR in T98G Glioblastoma Cells by Modulating Its Glycosylation and Ubiquitylation

Protein trafficking is altered when normal cells acquire a tumor phenotype. A key subcellular compartment in regulating protein trafficking is the Golgi apparatus, but its role in carcinogenesis is still not well defined. Golgi phosphoprotein 3 (GOLPH3), a peripheral membrane protein mostly localized at the trans-Golgi network, is overexpressed in several tumor types including glioblastoma multiforme (GBM), the most lethal primary brain tumor. Moreover, GOLPH3 is currently considered an oncoprotein, however its precise function in GBM is not fully understood. Here, we analyzed in T98G cells of GBM, which express high levels of epidermal growth factor receptor (EGFR), the effect of stable RNAi-mediated knockdown of GOLPH3. We found that silencing GOLPH3 caused a significant reduction in the proliferation of T98G cells and an unexpected increase in total EGFR levels, even at the cell surface, which was however less prone to ligand-induced autophosphorylation. Furthermore, silencing GOLPH3 decreased EGFR sialylation and fucosylation, which correlated with delayed ligand-induced EGFR downregulation and its accumulation at endo-lysosomal compartments. Finally, we found that EGF failed at promoting EGFR ubiquitylation when the levels of GOLPH3 were reduced. Altogether, our results show that GOLPH3 in T98G cells regulates the endocytic trafficking and activation of EGFR likely by affecting its extent of glycosylation and ubiquitylation.     

Hot corrosion studies of a salt-coated Ni-based superalloy under flowing wet air and sulfur vapor ambient

Corrosion of a salt-coated Ni-superalloy has been studied at 900°C under a wet air and sulfur vapor ambient. The corrosion thickness, after an incubation of ~60 hr, linearly increases with the corrosion time t and the onset of surface spallation occurred at t ≈ 60 hr. The corroded layer consists of a corrosion front dominated by Cr₃S₄ scales and linear precipitate structures, an inner corrosion layer dominated by Ni₃S₂ and NiO, and an outer corrosion layer dominated by Al₂O₃ networks surrounding the Ni₃S₂ and/or NiO scale structures. The corrosion mechanism is discussed based on the coexistence of H₂O, sulfur, and oxygen.     

Effect of low-pressure carburizing and plasma nitriding on mechanical properties and fatigue endurance limits of low alloy sintered steels

ABSTRACT

Thermochemical treatments like plasma nitriding or surface carburizing are commonly used to enhance surface hardness of steel components. An important difference between these treatments is the temperature at which they are carried out. In the present paper, the surface carburizing was carried out following a recently reported non-isothermal low pressure carburizing (LPC) treatment. In order to gain a comparative view of the effect of different treatments on the microstructure, microhardness, fatigue and impact properties, materials with distinct hardenability and widely used in the industrial production were evaluated. Tests were also carried out using industrially processed components aimed to an application demanding high wear resistance. The microstructural evolution during case hardening was studied by optical and electron microscopy.     

Managing risks in public-private partnership formation projects

While sourcing by means of Public-Private Partnerships has been lauded over recent years, increasingly risks appear to jeopardise public organisations' unique societal tasks. Integrated Risk Management has not yet been applied to public organisations getting involved in PPP in the sense of understanding risk management capabilities. This article explores risk awareness and risk management practices underpinning maintenance partnership formation by means of a dual case study of two PPP projects and a short industry survey. The results suggest that organisations face several “intolerable risks” linked to project governance and project management responsibilities: insufficient representation of qualified employees, absence of a shared performance system, assignment of responsibilities and decision-making authority, impractical or inappropriate partnership agreement, and timing of the partnership initiative. Cross-case analysis revealed the role of different levels of risk awareness and senior management involvement. Drawing on these findings, a framework for risk management for PPP formation projects is developed.     

Farm-scale carbon and nitrogen fluxes in pastoral dairy production systems using different nitrogen fertilizer regimes

Nutrient Cycling in Agroecosystems - The nitrogen (N) fertilizer application rate (kg ha?1 year?1) in pastoral dairy systems affects the flow of N through the soil,...     

Scrap leather valorization through composite fabrication using mustard oil resin and N-vinyl-2-pyrrolidone

Iranian Polymer Journal - Composites derived from petroleum sources like epoxy, unsaturated polyester and phenolics have limitations due to non-biodegradability toxicity which have harmful effects...