Accumulation of aggregating b-amyloid peptide (Ab) has been implicated in Alzheimer's disease pathology (1). However, neither the toxic species of Ab nor the mechanism of neurotoxicity has been clearly resolved. Previous studies have shown that the ability of synthetic Ab to form transmembrane pores in vitro depends upon a glycine zipper motif located in the hydrophobic C-terminal region: Gly-XXX-Gly-XXX-Gly (2,3). We have engineered several transgenic C. elegans lines to express wild type Ab42 and Ab42 variants (G37L, N27G, I31G, M35G, G37F) in order to test the glycine zipper motif hypothesis. Substitutions in the glycine zipper region, particularly the G37L variant, were found to be significantly less toxic than wildtype Ab. We then engineered Ab variants with second site substitutions (N27G G37L, I31G G37L, or M35G G37L) that we predicted might restore toxicity based on structural models. Our results indicated that these second site substitutions did indeed restore Ab toxicity. We have previously shown that activation of the
skn-1/Nrf2 oxidative stress response pathway by exposure to 10 percent coffee extract strongly protects worms expressing wild type Ab (4). Interestingly, we found that coffee exposure did not further reduce toxicity in the Ab variants with glycine zipper substitutions, suggesting there may be some convergence between the protective mechanisms resulting from Ab sequence changes or activation of
skn-1 . We have also assayed the toxicity of the Ab variant peptides in Neuro 2a cells, primary hippocampal cultures, and rat cortical neurons. In all of the neuronal models G37L was less toxic when compared to wildtype Ab42. (1.) Tiraboschi, P., Hansen, L.A., Thal, L.J., Corey-Bloom, J, 2004 The importance of neuritic plaques and tangles to the development and evolution of AD. Neurology 62 (11): 1984-1989. (2.) Kim, S., Jeon, T.J., Oberia, A., Yang, D., Schmidt, J.J., Bowie, J.U, 2005 Transmembrane glycine zippers: Physiological and pathological roles in membrane proteins. Proc Natl Acad Sci (102): 14278-14283. (3.) Arispe, N., Pollared, H.B., Rojas, E,1995 Zn2+ interaction with Alzheimer amyloid b protein calcium channels. Proc Natl Acad Sci (93): 1710-1715. (4.) Dostal, V., Roberts, C.M., Link, C.D., 2010 Genetic mechanism of coffee extract protection in a Caenorhabditis elegans model of b-amyloid peptide toxicity. Genetics 186(3):857-66.