Beta-secretase kinetics detected by MALDI TOF MS
| Název česky | Kinetika beta-sekretasy detekovaná pomocí MALDI TOF MS |
|---|---|
| Autoři | |
| Rok publikování | 2017 |
| Druh | Konferenční abstrakty |
| Fakulta / Pracoviště MU | |
| Citace | |
| Popis | Alzheimer’s disease is a serious neurodegenerative illness affecting around 44 million people worldwide. The crucial role in disease development possibly plays the ß-site APP-cleaving enzyme (ß-secretase, BACE). Its overexpression leads to the production of ?ß peptide (?ß42 rather than ?ß40 isoform) and to the accumulation of toxic amyloid plaques in brain [1]. ß-secretase activity can be measured by a commercial fluorescence resonance energy transfer (FRET) assay with fluorescently marked substrates, however, this approach is relatively insensitive and requires high concentrations of the enzyme. Alternative option is the detection of reaction products by mass spectrometry. Two distinctive techniques were established for this purpose, the first involving the capillary electrophoresis separation and electrospray mass spectrometry detection (CE-ESI MS). The advantages of this method are both sensitivity and preciseness, yet, it includes the time consuming separation step [2]. The separation may be completely omitted if using matrix-assisted laser desorption/ionization (MALDI), which employs UV-laser to desorb and generate ion molecules from the sample surface [3]. While MALDI is not perceived as quantitative in general, we showed that with suitable internal standard, it may represent easy and accurate method useful in enzymatic research. The Km for selected substrate and IC50 values for ß-secretase inhibitor were in a good agreement with results obtained by CE-ESI MS. |
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