α 2 -macroglobulin associates with β-amyloid peptide and prevents fibril formation

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Abstract
We have used the yeast two-hybrid system to isolate cDNAs encoding proteins that specifically interact with the 42-aa β-amyloid peptide (Aβ), a major constituent of senile plaques in Alzheimer’s disease. The carboxy terminus of α2-macroglobulin (α2M), a proteinase inhibitor released in response to inflammatory stimuli, was identified as a strong and specific interactor of Aβ, utilizing this system. Direct evidence for this interaction was obtained by co-immunoprecipitation of α2M with Aβ from the yeast cell, and by formation of SDS-resistant Aβ complexes in polyacrylamide gels by using synthetic Aβ and purified α2M. The association of Aβ with α2M and various purified amyloid binding proteins was assessed by employing a method measuring protein–protein interactions in liquid phase. The dissociation constant by this technique for the α2M–Aβ association using labeled purified proteins was measured (Kd = 350 nM). Electron microscopy showed that a 1:8 ratio of α2M to Aβ prevented fibril formation in solution; the same ratio to Aβ of another acute phase protein, α1-antichymotrypsin, was not active in preventing fibril formation in vitro. These results were corroborated by data obtained from an in vitro aggregation assay employing Thioflavine T. The interaction of α2M with Aβ suggests new pathway(s) for the clearance of the soluble amyloid peptide.