Recognition of the minimal epitope of monoclonal antibody Tau-1 depends upon the presence of a phosphate group but not its location.
The major constituent of the paired helical filaments (PHFs) of Alzheimer's disease is the abnormally phosphorylated form of the microtubule-associated protein, tau. Monoclonal antibody (mAb) Tau-1 is used extensively to stain normal human tau, and tau isolated from the brains of Alzheimer's disease patients after dephosphorylation. We used a panel of 6 synthetic peptides to localize the minimal epitope of Tau-1 between amino acids 192-204. All 4 serine residues within this fragment were later phosphorylated individually by chemical methods, and it was shown that none of the peptides carrying a single phosphate group were recognized by the antibody. The serines included those that are probably not transformed in AD and consequently, conclusions drawn about malfunctioning kinase activity, based on Tau-1 immunoreactivity, can be extremely misleading. The recognition was restored at a decreased level when one of the serines was replaced by an alanine residue. mAb AT8 was made by immunizing with the PHFs and was reported to recognize the same region of the protein in a phosphorylated form. AT8 did not, however, cross-react with any of the singly phosphorylated peptides, indicating that the recognition site of the two antibodies are not entirely complementary or the binding to AT8 needs multiple phosphorylation of the antigen. The abolished recognition of the phosphorylated peptides cannot be attributed to a conformational change due to phosphorylation, since all peptides exhibited reverse-turn secondary structures, as indicated by circular dichroism (CD) spectroscopy. Anti-tau mAbs may distinguish between phosphorylated and non-phosphorylated forms of epitopes regardless of the location of the phosphate group.
Szendrei
G I
GI
Wistar Institute of Anatomy and Biology, Philadelphia, Pennsylvania 19104.
Lee
V M
VM
Otvos
L
L
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology*; D002942:Circular Dichroism; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology / Q000302:isolation & purification; D010710:Phosphates / Q000737:chemistry / Q000276:immunology*; D010766:Phosphorylation; D016875:tau Proteins / Q000737:chemistry / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D010455:Peptides; D010710:Phosphates; D016875:tau Proteins