Photo-induced damage of amyloid  by the Chlorin e6 photosensitizer

Michal Richman, Chemistry, Bar-Ilan University, Ramat-Gan, Israel

Inhibition of amyloid  (Aβ) aggregation and destabilization of toxic Aβ oligomers are promising approaches for the prevention and therapy of Alzheimer's disease (AD). Compounds that block Aβ aggregation have been identified in many studies, for example, peptides containing a recognition element KLVFF (residues16-20 in Aβ) were developed to bind Aβ and disrupt its aggregation, thus protecting neurons from Aβ-associated toxicity. Herein, we utilize the high affinity of the hemin-like porphyrin, Chlorin e6 (Ce6), to A and its well-known photosensitizing property to selectively damage A upon illumination with visible light. The Thioflavin-T (ThT) assay as well as the transmission electron microscopy (TEM) studies demonstrated that Ce6 exhibits potent antiamyloidogenic activity in dark, however, brief illumination of the A-Ce6 mixture dramatically increases its activity. Ce6 also protected rat pheochromocytoma PC-12 cells from A toxicity, while illumination with visible light significantly increased this activity.

Using various immunoassays, circular dichroism spectroscopy, photoinduced cross-linking of unmodified proteins (PICUP) combined with SDS/PAGE, and NMR, we probed the mechanisms underlying Ce6 antiamyloidogenic activity. NMR spectroscopy indicated that Ce6 selectively binds the His residues in positions 13 and 14 of A. A significant loss of His residues was observed by NMR as well as amino acid analysis upon irradiation of the A-Ce6 mixture with light. Overall, the NMR and the amino acid analysis together with the PICUP results demonstrated that irradiation of Ce6 in the presence of Aβ induces intra-molecular cross-links and also modifies His residues in Aβ sequence, which can be responsible for its photo-induced antiamyloidogenic activity.