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Introduction: The ongoing COVID-19 pandemic situation caused by SARS-CoV-2 and variants of concern such as B.1.617.2 (Delta) and recently, B.1.1.529 (Omicron) is posing multiple challenges to humanity. The rapid evolution of the virus requires adaptation of diagnostic and therapeutic applications.
Objectives: In this study, we describe camelid heavy-chain-only antibodies (hcAb) as useful tools for novel in vitro diagnostic assays and for therapeutic applications due to their neutralizing capacity.
Methods: Five antibody candidates were selected out of a naïve camelid library by phage display and expressed as full length IgG2 antibodies. The antibodies were characterized by Western blot, enzyme-linked immunosorbent assays, surface plasmon resonance with regard to their specificity to the recombinant SARS-CoV-2 Spike protein and to SARS-CoV-2 virus-like particles. Neutralization assays were performed with authentic SARS-CoV-2 and pseudotyped viruses (wildtype and Omicron).
Results: All antibodies efficiently detect recombinant SARS-CoV-2 Spike protein and SARS-CoV-2 virus-like particles in different ELISA setups. The best combination was shown with hcAb B10 as catcher antibody and HRP-conjugated hcAb A7.2 as the detection antibody. Further, four out of five antibodies potently neutralized authentic wildtype SARS-CoV-2 and particles pseudotyped with the SARS-CoV-2 Spike proteins of the wildtype and Omicron variant, sublineage BA.1 at concentrations between 0.1 and 0.35 ng/mL (ND50).
Conclusion: Collectively, we report novel camelid hcAbs suitable for diagnostics and potential therapy.
Introduction: The ongoing COVID-19 pandemic situation caused by SARS-CoV-2 and variants of concern such as B.1.617.2 (Delta) and recently, B.1.1.529 (Omicron) is posing multiple challenges to humanity. The rapid evolution of the virus requires adaptation of diagnostic and therapeutic applications.
Objectives: In this study, we describe camelid heavy-chain-only antibodies (hcAb) as useful tools for novel in vitro diagnostic assays and for therapeutic applications due to their neutralizing capacity.
Methods: Five antibody candidates were selected out of a naïve camelid library by phage display and expressed as full length IgG2 antibodies. The antibodies were characterized by Western blot, enzyme-linked immunosorbent assays, surface plasmon resonance with regard to their specificity to the recombinant SARS-CoV-2 Spike protein and to SARS-CoV-2 virus-like particles. Neutralization assays were performed with authentic SARS-CoV-2 and pseudotyped viruses (wildtype and Omicron).
Results: All antibodies efficiently detect recombinant SARS-CoV-2 Spike protein and SARS-CoV-2 virus-like particles in different ELISA setups. The best combination was shown with hcAb B10 as catcher antibody and HRP-conjugated hcAb A7.2 as the detection antibody. Further, four out of five antibodies potently neutralized authentic wildtype SARS-CoV-2 and particles pseudotyped with the SARS-CoV-2 Spike proteins of the wildtype and Omicron variant, sublineage BA.1 at concentrations between 0.1 and 0.35 ng/mL (ND50).
Conclusion: Collectively, we report novel camelid hcAbs suitable for diagnostics and potential therapy.
Welfare beyond consumption
(2020)
In developed regions worldwide, so-called anti-consumers are increasingly resisting high-level consumption lifestyles or shifting to alternative forms of consumption. A general reduction in consumption levels is considered necessary to attain global sustainability goals. However, knowledge regarding the factors driving people to deliberately consume less and how anti-consumption affects individuals' well-being is limited. Against this background, this study considers the influence of human values and the well-being effects of two types of anti-consumption: voluntary simplicity and collaborative consumption. Based on representative data from the US (N = 1075) and Germany (N = 1070), the findings show that the two anti-consumption types do not reduce the well-being of individuals' but in some cases, even improve it, which suggests that lowering consumption can not only help protect environmental resources but also serve the greater good of society. In particular, this relationship holds among collaborative consumers with a strong need for cognition, i.e., a cognitive thinking style that involves a high level of decision control. According to the study results, opposite value orientations are the drivers of voluntary simplicity and collaborative consumption (i.e., a focus on self-transcendence versus self-enhancement). These findings are comparable in both countries; however, the strength of the effects differs.
Antisense transcription is common in naturally occurring genomes and is increasingly being used in synthetic genetic circuitry as a tool for gene expression control. Mutual influence on the expression of convergent genes can be mediated by antisense RNA effects and by transcriptional interference (TI). We aimed to quantitatively characterize long-range TI between convergent genes with untranslated intergenic spacers of increasing length. After controlling for antisense RNA-mediated effects, which contributed about half of the observed total expression inhibition, the TI effect was modeled. To achieve model convergence, RNA polymerase processivity and collision resistance were assumed to be modulated by ribosome trailing. The spontaneous transcription termination rate in regions of untranslated DNA was experimentally determined. Our modeling suggests that an elongating RNA polymerase with a trailing ribosome is about 13 times more likely to resume transcription than an opposing RNA polymerase without a trailing ribosome, upon head-on collision of the two.
We used single-molecule FRET in combination with other biophysical methods and molecular simulations to investigate the effect of temperature on the dimensions of unfolded proteins. With singlemolecule FRET, this question can be addressed even under nearnative conditions, where most molecules are folded, allowing us to probe a wide range of denaturant concentrations and temperatures. We find a compaction of the unfolded state of a small cold shock protein with increasing temperature in both the presence and the absence of denaturant, with good agreement between the results from single-molecule FRET and dynamic light scattering. Although dissociation of denaturant from the polypeptide chain with increasing temperature accounts for part of the compaction, the results indicate an important role for additional temperaturedependent interactions within the unfolded chain. The observation of a collapse of a similar extent in the extremely hydrophilic, intrinsically disordered protein prothymosin suggests that the hydrophobic effect is not the sole source of the underlying interactions. Circular dichroism spectroscopy and replica exchange molecular dynamics simulations in explicit water show changes in secondary structure content with increasing temperature and suggest a contribution of intramolecular hydrogen bonding to unfolded state collapse.
Environmental parameters such as rainfall, temperature and relative humidity can affect the composition of higher plant leaf wax. The abundance and distribution of leaf wax biomarkers, such as long chain n-alkanes, in sedimentary archives have therefore been proposed as proxies reflecting climate change. However, a robust palaeoclimatic interpretation requires a thorough understanding of how environmental changes affect leaf wax n-alkane distributions in living plants. We have analysed the concentration and chain length distribution of leaf wax n-alkanes in Acacia and Eucalyptus species along a 1500 km climatic gradient in northern Australia that ranges from subtropical to arid. We show that aridity affected the concentration and distribution of n-alkanes for plants in both genera. For both Acacia and Eucalyptus n-alkane concentration increased by a factor of ten to the dry centre of Australia, reflecting the purpose of the wax in preventing water loss from the leaf. Furthermore, Acacian-alkanes decreased in average chain length (ACL) towards the arid centre of Australia, whereas Eucalyptus ACL increased under arid conditions. Our observations demonstrate that n-alkane concentration and distribution in leaf wax are sensitive to hydroclimatic conditions. These parameters could therefore potentially be employed in palaeorecords to estimate past environmental change. However, our finding of a distinct response of n-alkane ACL values to hydrological changes in different taxa also implies that the often assumed increase in ACL under drier conditions is not a robust feature for all plant species and genera and as such additional information about the prevalent vegetation are required when ACL values are used as a palaeoclimate proxy.
This study aims to compare impacts of climate change on streamflow in four large representative African river basins: the Niger, the Upper Blue Nile, the Oubangui and the Limpopo. We set up the eco-hydrological model SWIM (Soil and Water Integrated Model) for all four basins individually. The validation of the models for four basins shows results from adequate to very good, depending on the quality and availability of input and calibration data.
For the climate impact assessment, we drive the model with outputs of five bias corrected Earth system models of Coupled Model Intercomparison Project Phase 5 (CMIP5) for the representative concentration pathways (RCPs) 2.6 and 8.5. This climate input is put into the context of climate trends of the whole African continent and compared to a CMIP5 ensemble of 19 models in order to test their representativeness. Subsequently, we compare the trends in mean discharges, seasonality and hydrological extremes in the 21st century. The uncertainty of results for all basins is high. Still, climate change impact is clearly visible for mean discharges but also for extremes in high and low flows. The uncertainty of the projections is the lowest in the Upper Blue Nile, where an increase in streamflow is most likely. In the Niger and the Limpopo basins, the magnitude of trends in both directions is high and has a wide range of uncertainty. In the Oubangui, impacts are the least significant. Our results confirm partly the findings of previous continental impact analyses for Africa. However, contradictory to these studies we find a tendency for increased streamflows in three of the four basins (not for the Oubangui). Guided by these results, we argue for attention to the possible risks of increasing high flows in the face of the dominant water scarcity in Africa. In conclusion, the study shows that impact intercomparisons have added value to the adaptation discussion and may be used for setting up adaptation plans in the context of a holistic approach.
Light-switchable proteins offer numerous opportunities as tools for manipulating biological systems with exceptional degrees of spatiotemporal control. Most designed light-switchable proteins currently in use have not been optimised using the randomisation and selection/screening approaches that are widely used in other areas of protein engineering. Here we report an approach for screening light-switchable DNA-binding proteins that relies on light-dependent repression of the transcription of a fluorescent reporter. We demonstrate that the method can be used to recover a known light-switchable DNA-binding protein from a random library.
A dual probe was investigated by UV-Vis, fluorescence, and ESR spectroscopy. It comprises the pyrene chromophore and the paramagnetic 2,2,6,6-tetramethylpiperidinyl-N-oxyl radical that are covalently linked together via an ester bridge. The dual probe was used to investigate molecular solvents of different polarity as well as ionic liquids bearing either imidazolium or pyrrolidinium cations and various anions, such as bis(trifluoromethylsulfonyl)imide, tetrafluoroborate, tris(pentafluoroethyl)trifluorophosphate, or dicyanamide. The dual probe does not show solvatochromism that is typical for some pyrenes. Furthermore, the dual probe is considerable less mobile compared to 2,2,6,6-tetramethylpiperidinyl-N-oxyl (TEMPO) without additional substituent as detected by ESR spectroscopy. This is caused by the bulky pyrenyl substituent bound at the dual probe resulting in a reduced mobility of the dual probe.
We have investigated transcriptional interference between convergent genes in E. coli and demonstrate substantial interference for inter-promoter distances of as far as 3 kb. Interference can be elicited by both strong σ70 dependent and T7 promoters. In the presented design, a strong promoter driving gene expression of a ‘forward’ gene interferes with the expression of a ‘reverse’ gene by a weak promoter. This arrangement allows inversely correlated gene expression without requiring further regulatory components. Thus, modulation of the activity of the strong promoter alters expression of both the forward and the reverse gene. We used this design to develop a dual selection system for conditional operator site binding, allowing positive selection both for binding and for non-binding to DNA. This study demonstrates the utility of this novel system using the Lac repressor as a model protein for conditional DNA binding, and spectinomycin and chloramphenicol resistance genes as positive selection markers in liquid culture. Randomized LacI libraries were created and subjected to subsequent dual selection, but mispairing IPTG and selection cues in respect to the wild-type LacI response, allowing the isolation of a LacI variant with a reversed IPTG response within three rounds of library generation and dual selection.