Investigating in vitro amyloid peptide 1-42 aggregation: Impact of higher molecular weight stable adducts

The self-assembly of amyloid peptides (A?), in particular A?-42, into oligomers and fibrils is one of the main pathological events related to Alzheimer's disease. Recent studies have demonstrated the ability of carbon monoxide-releasing molecules (CORMs) to protect neurons and astrocytes from A?- toxicity. In fact, CORMs are able to carry and release controlled levels of CO and are known to exert a wide range of anti-inflammatory and anti-apoptotic activities at physiologically relevant concentrations. In order to investigate the direct effects of CORMs on A?-42, we studied the reactivity of CORM-2 and CORM-3 with A?- in vitro and the potential inhibition of its aggregation by mass spectrometry (MS), as well as fluorescence and circular dichroism spectroscopies. The application of an electrospray ionization-MS (ESI-MS) method allowed the detection of stable A?-/CORMs adducts, involving the addition of the Ru(CO) portion of CORMs at histidine residues on the A?- skeleton. Moreover, CORMs showed anti-aggregating properties through formation of stable adducts with A?- as demonstrated by a thioflavin T fluorescence assay and MS analysis. As further proof, comparison of the CD spectra of A?- recorded in the absence and in the presence of CORM-3 at a 1:1 molar ratio showed the ability of CORM-3 to stabilize the peptide in its soluble, unordered conformation, thereby preventing its misfolding and aggregation. This multimethodological investigation revealed novel interactions between A?- and CORMs, contributing new insights into the proposed neuroprotective mechanisms mediated by CORMs and disclosing a new strategy to divert amyloid aggregation and toxicity.