Publikace detail

Relationship of PHF6 hexapeptide motif and abnormal sticky behaviour: complication in tau protein analysis at peptide level

Autoři: Hromádková Lenka | Kupčík Rudolf | Jankovičová Barbora | Bílková Zuzana | Řípová Daniela

Rok: 2015

Druh publikace: ostatní - přednáška nebo poster

Strana od-do: nestránkováno

Tituly:

Jazyk	Název	Abstrakt	Klíčová slova
eng	Relationship of PHF6 hexapeptide motif and abnormal sticky behaviour: complication in tau protein analysis at peptide level	Tau protein belongs to intrinsically disordered proteins. In Alzheimer disease, tau undergoes modifications leading to the misfolding and formation of paired helical filaments (PHF). Our aim was to optimize the epitope extraction protocol enabling identification of linear immunodominant epitopes using tau digested by trypsin. Within the eluted fragments containing epitopes specifically reactive with antibodies bound to solid phase, additionally fragment 299-HVPGGGSVQIVYKPVDLSK-317 was detected by mass spectrometry (MS). This fragment comprising minimal interaction motif responsible for the PHF formation, hexapeptide PHF6 (306-VQIVYK-311), distinctly complicated the MS spectra interpretation. The conventional approaches to suppress the nonspecific sorption were not effective to preclude or at least to reduce the ‘‘sticky’’ fragment sorption. Therefore we carried out protein cleavage by two immobilized enzymes. Chymotrypsin was selected as an additional protease due to the specific cleavage site between Y310 and K311. This newly introduced approach led to suppression of nonspecific sorption without affecting specific epitope–antibody interactions in the epitope extraction protocol.	PHF6 hexapeptide; tau protein; mass spectrometry; epitope extraction

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