The long term objective of this project is to learn the etiology and the pathogenesis of Alzheimer disease/senile dementia of the Alzheimer type (AD/SDAT) which constitutes one of the most important public health problems in our country since over two million people are affected.
The specific aim of this proposal is to determine the origin of paired helical filaments (PHF) which are the common structural element of the Alzheimer neurofibrillary tangles and the neuritic (senile) plaques, the two most characteristic brain lesions of AD/SDAT and in a much less concentration the normal aged humans. During the current grant period we have demonstrated that PHF and microtubule associated protein tau polypeptides comigrate on SDS-PAGE, and immunochemically crossreact with each other, tau is abnormally phosphorylated in PHF, by ELISA tau is a major component of PHF and both tau and PHF polypeptides are amino-terminally blocked. We now propose to determine the amino acid sequences of PHF and tau polypeptides by cleaving them into peptides with CNBr, trypsin and chymotrypsin, study their peptide maps, isolate the peptides by HPLC, sequence them by automated gas phase sequencing, obtain synthetic peptides corresponding to the sequences determined, raise antibodies to the synthetic peptides, confirm their localization to PHF immunocytochemically and by determining their crossreactivities with PHF polypeptides by Western blots and study their in vitro polymerization into fibrils. The techniques involved in this study are biochemical, immunological and morphological. Identification and characterization of the PHF polypeptide/s will be a major step towards learning the origin of PHF and may ultimately lead to an understanding of the disease mechanism. Immunochemical data strongly suggest that tau is a major protein of PHF. Amino acid sequence of tau and PHF polypeptides will demonstrate the presence and the amount of tau and any non-tau polypeptide/s in PHF. The in vitro polymerization of PHF/tau peptides might reveal the identity of the crucial amino acid sequence responsible for the formation of PHF.
