Combining teaching and research: a BIP on geophysical and archaeological prospection of North Frisian medieval settlement patterns

We performed a research-oriented EU Erasmus+ Blended Intensive Program (BIP) with participants from four countries focused on North Frisian terp settlements from Roman Iron Age and medieval times. We show that the complex terp structure and environment can be efficiently prospected using combined magnetic and EMI mapping, and seismic and geoelectric profiling and drilling. We found evidence of multiple terp phases and a harbor at the Roman Iron Age terp of Tofting. In contrast, the medieval terp of Stolthusen is more simply constructed, probably uni-phase. The BIP proved to be a suitable tool for high-level hands-on education adding value to the research conducted in on-going projects

Structural analysis and aggregation propensity of pyroglutamate Aβ(3-40) in aqueous trifluoroethanol

A hallmark of Alzheimer's disease (AD) is the accumulation of extracellular amyloid-β (Aβ) plaques in the brains of patients. N-terminally truncated pyroglutamate-modified Aβ (pEAβ) has been described as a major compound of Aβ species in senile plaques. pEAβ is more resistant to degradation, shows higher toxicity and has increased aggregation propensity and β-sheet stabilization compared to non-modified Aβ. Here we characterized recombinant pEAβ(3-40) in aqueous trifluoroethanol (TFE) solution regarding its aggregation propensity and structural changes in comparison to its non-pyroglutamate-modified variant Aβ(1-40). Secondary structure analysis by circular dichroism spectroscopy suggests that pEAβ(3-40) shows an increased tendency to form β-sheet-rich structures in 20% TFE containing solutions where Aβ(1-40) forms α-helices. Aggregation kinetics of pEAβ(3-40) in the presence of 20% TFE monitored by thioflavin-T (ThT) assay showed a typical sigmoidal aggregation in contrast to Aβ(1-40), which lacks ThT positive structures under the same conditions. Transmission electron microscopy confirms that pEAβ(3-40) aggregated to large fibrils and high molecular weight aggregates in spite of the presence of the helix stabilizing co-solvent TFE. High resolution NMR spectroscopy of recombinantly produced and uniformly isotope labeled [U-15N]-pEAβ(3-40) in TFE containing solutions indicates that the pyroglutamate formation affects significantly the N-terminal region, which in turn leads to decreased monomer stability and increased aggregation propensity

Far-UV CD spectra of pEAβ(3–40) and Aβ(1–40).

<p>Peptides were dissolved in buffer (20–25% TFE in 50 mM potassium phosphate, pH 2.8). CD spectra were recorded at 20°C from 260 to 187 nm, accumulated 10 times and background corrected. (a) CD spectra of 25 μM pEAβ(3–40) in 25%, 23%, 22%, 21% and 20% TFE showed a shift from α-helical structure towards β-sheets with decreasing TFE concentrations. (b) The CD spectrum of 25 μM Aβ(1–40) indicated α-helices in 20% TFE while the spectrum of 25 μM pEAβ(3–40) in 20% TFE showed mainly β-sheet rich structures.</p

Chemical shift changes Δδ(¹H,¹⁵N) = ((10*Δδ1H^N)² + (Δδ¹⁵N)²)^1/2 between pEAβ(3–40) and Aβ(1–40) plotted as a function of the pEAβ(3–40) sequence.

<p>Pyroglutamate formation affects the N-terminal amino acids with decreasing effect towards the C-terminus and, interestingly, also the C-terminal V40.</p

Aggregation kinetics of pEAβ(3–40) and Aβ(1–40) in TFE.

<p>(a) 25 μM of monomerized pEAβ(3–40) were dissolved in buffer with various TFE contents (25%, 23%, 22%, 21% and 20% TFE in 50 mM potassium phosphate, pH 2.8) including 10 μM ThT. pEAβ(3–40) aggregated in 20% and 21% TFE but was significantly decreased in aqueous solution with higher TFE concentration. (b) 25 μM of monomerized pEAβ(3–40) and Aβ(1–40) were dissolved in buffer (20% TFE in 50 mM potassium phosphate, pH 2.8) including 10 μM ThT. An increase in ThT fluorescence was observed for pEAβ(3–40) but not for Aβ(1–40) within 72 h.</p

TEM image of pEAβ(3–40) in 50 mM potassium phosphate pH 2.8 containing 20% TFE.

<p>Monomerized pEAβ(3–40) (25 μM) was incubated for fibrillation at 20°C for five days and grids were prepared by negative staining. pEAβ(3–40) incubated in aqueous TFE solution formed large twisted fibrils up to several hundred nm in size which accumulate into large aggregates ranging from 1–5 μm in diameter.</p