Nicastrin, the largest member among the four components of the γ-secretase complex, has been identified to be the substrate recognizer for the proteolytic activity of the complex. Here we report that full-length human nicastrin (hNCT) can be obtained by heterologous expression in E. coli. Milligram quantities of the target protein are purified in a two-step purification protocol using affinity chromatography followed by SEC. The FOS-choline 14 purified tetrameric hNCT exhibits a proper folding with 31% α-helix and 23% β-sheet content. Thermal stability studies reveal stable secondary and tertiary structure of the detergent purified hNCT. A physical interaction between nicastrin and the γ-secretase substrate APPC100 confirmed the functionality of hNCT as a substrate recognizer.

The data presented here are related to the research article entitled “Expression, purification, and preliminary characterization of human presenilin-2" [1].Human Presenilin-2 is the catalytic subunit of γ-secretase and a possible calcium leakage channel (Kimberly et al., 2000; Tu et al., 2006) [2], [3]. HisPS2 which was obtained by overexpression in E. coli strain C43 (DE3) was extracted by detergent solubilisation. The sample isolation efficiency by detergents and the protein identification by mass spectrometry and western blot are described.This data article describes the near and far UV circular dichroism measurements and the data deconvolution in terms of secondary structure at 4 and 98 °C. Also, a refolding spectrum is presented.The raw CD spectra used for deconvolution of the helix and stand segments and average length are deposited into Protein Circular Dichroism Data Bank with PCDDBid: CD0005962000 (4 °C far UV), CD0005963000 (98 °C far UV), CD0005964000 (back to 4 °C far UV) and CD0005965000 (4 °C near UV CD).

A B S T R A C TPresenilins (PS1 and PS2) exhibit similar γ-secretase-dependent and −independent functions with subtle variations. In this study, we established a cost-effective process to overexpress and purify full-length human PS2 in sufficient quantities and quality for structural studies. Upon optimization, milligram quantities of homogeneous trimeric hisPS2 were purified, which enabled the preliminary characterization of human hisPS2 zymogen. Far-UV and near-UV CD as well as fluorescence spectroscopy revealed that purified hisPS2 contained the expected secondary structure and was folded into a defined tertiary structure. Thermal stability analysis revealed a T m value of ∼55°C for secondary structure while cholesterol significantly increased the stability. The low melting temperature of ∼34°C for the tertiary structure was able to explain the purity and aggregation problems observed during purification. Additionally, the occurrence of calcium ions induced structural changes to different extents for PS2WT and PS2-D263A/D366A was observed, which is consistent with previous studies.

Presenilin (PS1 or PS2) functions as the catalytic subunit of γ-secretase, which produces the toxic amyloid beta peptides in Alzheimer’s disease (AD). The dependence of folding and structural stability of PSs on the lipophilic environment and mutation were investigated by far UV CD spectroscopy. The secondary structure content and stability of PS2 depended on the lipophilic environment. PS2 undergoes a temperature-dependent structural transition from α-helical to β-structure at 331 K. The restructured protein formed structures which tested positive in spectroscopic amyloid fibrils assays. The AD mutant PS1L266F, PS1L424V and PS1ΔE9 displayed reduced stability which supports a proposed ‘loss of function’ mechanism of AD based on protein instability. The exon 9 coded sequence in the inhibitory loop of the zymogen was found to be required for the modulation of the thermal stability of PS1 by the lipophilic environment.