Exploring the Role of Amyloid‐Beta Precursor Protein (APP) O‐Glycosylation in Alzheimer’s Disease
Authors: Singh YashoNandini , Kent Thomas , Massetti Alessandra , Minond Dmitriy , Du Deguo , Cudic Maré 
Published: April 2020

ObjectiveTo synthesize both O‐glycosylated and non‐glycosylated APP peptide analogs bearing the Tn antigen on Thr663, Ser667, and Tyr681 in and around the Aβ42 region, to understand the role of O‐glycosylation on APP’s conformation, proteolytic stability and aggregation kinetics.AbstractThe amyloid‐β precursor protein (APP) is a transmembrane protein that can undergo proteolytic cleavage by three proteases, α‐, β‐ and γ‐secretases, to determine its fate in Alzheimer’s disease (AD) pathogenesis. The protein undergoes proteolysis by β‐ and γ‐secretases to produce amyloid‐β peptides, which is a hallmark of AD. However, only ten‐percent of the protein will follow this amyloidogenic pathway, whereas the remaining ninety‐percent will undergo proteolysis by α‐secretase and γ‐secretases, resulting in the non‐amyloidogenic pathway. Recently, it was found that the four threonine residues Thr633, Thr651, Thr652, and Thr659, in the vicinity of the β‐secretase cleavage site (Met671~Asp672) of APP, are modified by complex mucin‐type O‐glycans. Moreover, an increase of up to 2.5 times in Tyr681 glycosylation, located within the Aβ42 region of APP, was found in AD patients in comparison to the non‐AD patients. These findings suggest the possible role of O‐glycosylation in APP proteolytic processing. Therefore, we synthesized native and Swedish‐double‐mutated (Met671Asn and Asp672Leu) APP (glyco)peptides with O‐GalNAc and studying conformational changes, secretase activity, and aggregation kinetics using circular dichroism, enzyme kinetic and Thioflavin T assays, respectively. Our results show that the non‐glycosylated peptide analogs in water show characteristics of β‐sheet conformation, however, when there is a site‐specific O‐glycosylation on Thr, Ser or Tyr residues of their counterparts, there is a change in conformation that resembles a mixture of α‐helix and random coil. All peptides adopt an α‐helix in 50/50 (v/v) trifluoroethanol/water mixture. Furthermore, the level of β‐secretase activity significantly increases for the glycosylated analogs containing the Swedish mutation compared to their nonglycosylated counterparts. Lastly, the glycosylated analogs impact the protein’s aggregation kinetics by decreasing its lag phase, and this effect seems to be more pronounced in the Swedish mutation analogs. In conclusion, our results suggest that APP’s site‐specific O‐glycosylation can induce a conformational change in the protein and subsequently affect its proteolytic processing fate towards either amyloidogenic or non‐amyloidogenic pathway. Our long‐term goals are to assess the role of certain patterns of multiple glycosylation sites on Ser, Thr, and Tyr that are in close proximity to the relevant secretases cleavage sites to develop specific inhibitors of amyloid pathway‐stimulating secretases (β‐secretase) or activators of non‐amyloid pathway (α‐secretase).Support or Funding InformationNational Institute of Health Grant CA242351 to M. C.