RACEMIZATION OF ASP(23) RESIDUE AFFECTS THE AGGREGATION PROPERTIES OFALZHEIMER AMYLOID-BETA PROTEIN ANALOGS

The beta proteins in amyloid deposits of Alzheimer's disease have been found to be racemized and/or isomerized at their Asp residues (Roher, A. E., Lowenson, J. D., Clarke, S., Wolkow, C., Wang, R., Cotter, R. J., Reardon, I. M., Zurcher-Neely, H. A., Heinrikson, R. L., Ball, M. J., and Greenberg, B. D. (1993) J. Biol. Chem. 268, 3072-3083). To elu cidate the effect of racemization on the aggregation properties of bet a proteins, we synthesized four beta protein analogues in which D-Asp was substituted for L-Asp residues, i.e. normal beta 1-35, [D-Asp(7)]b eta 1-35, [D-Asp(23)]beta 135, and [D-Asp(7),D-Asp(23)]beta 1-35. The aggregation and fibril formation of the peptides were examined by mean s of spectrophotometry, sodium dodecyl sulfate-polyacrylamide gel elec trophoresis (SDS-PAGE), and electron microscopy. Of the four peptides, [D-Asp(23)]beta 1-35 showed the earliest increase in turbidity and ap pearance of a smear in SDS-PAGE. This was followed by [D-Asp(7),D-Asp( 23)]beta 1-35 and normal beta 1-35. [D-Asp(7)]beta 1-35 was considerab ly delayed in showing these signs of aggregation. Corresponding with t he increase in turbidity and the appearance of a smear in SDS-PAGE, fi bril formation was observed in electron microscopy. These results reve al that the aggregation properties of beta 1-35 peptides are affected by racemization of their Asp residues depending on their position. Rac emization at amino acid position 23 accelerated the peptide aggregatio n and fibril formation, while that at position 7 slowed down this reac tion. This suggests that the site specific racemization of p protein m ay be involved in the amyloid fibril formation in Alzheimer's disease.