High level of aspartic acid-bond isomerization during the synthesis of an N-linked tau glycopeptide.
An increased degree of utilization of the potential N-glycosylation site in the fourth repeat unit of the human tau protein may be involved in the inability of tau to bind to the corresponding tubulin sequence(s) and in the subsequent development of the paired helical filaments of Alzheimer's disease. To model these processes, we synthesized the octadecapeptide spanning this region without sugar, and with the addition of an N-acetyl-glucosamine moiety. The carbohydrate-protected, glycosylated asparagine was incorporated as a building block during conventional Fmoc-solid phase peptide synthesis. While the crude non-glycosylated analog was obtained as a single peptide, two peptides with the identical, expected masses, in approximately equal amounts, were detected after the cleavage of the peracetylated glycopeptide. Surprisingly, the two glycopeptides switched positions on the reversed-phase high performance liquid chromatogram after removal of the sugar-protecting acetyl groups. Nuclear magnetic resonance spectroscopy and peptide sequencing identified the more hydrophobic deprotected peak as the target peptide, and the more hydrophilic deprotected peak as a peptide analog in which the aspartic acid-bond just preceding the glycosylated asparagine residue was isomerized resulting in the formation of a beta-peptide. The anomalous chromatographic behavior of the acetylated beta-isomer could be explained on the basis of the generation of an extended hydrophobic surface which is not present in any of the other three glycopeptide variants. Repetition of the syntheses, with altered conditions and reagents, revealed reproducibly high levels of aspartic acid-bond isomerization of the glycopeptide as well as lack of isomerization for the non-glycosylated parent analog. If similar increased aspartic acid-bond isomerization occurs in vivo, a protein modification well known to take place for both the amyloid deposits and the neurofibrillary tangles in Alzheimer's disease, this process may explain the aggregation of glycosylated tau into the paired helical filaments in the affected brains.
Hoffmann
R
R
The Wistar Institute, Philadelphia, PA 19104, USA.
Craik
D J
DJ
Bokonyi
K
K
Varga
I
I
Otvos
L
L
D000595:Amino Acid Sequence; D001224:Aspartic Acid / Q000737:chemistry*; D006023:Glycoproteins / Q000138:chemical synthesis* / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D007536:Isomerism; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis* / Q000378:metabolism*; D011487:Protein Conformation; D020539:Sequence Analysis, Protein; D016875:tau Proteins / Q000737:chemistry / Q000378:metabolism
D006023:Glycoproteins; D010446:Peptide Fragments; D016875:tau Proteins; D001224:Aspartic Acid