Ganglioside GM1 binding the N-terminus of amyloid precursor protein.
Secreted amyloid precursor protein (APPs) plays a role in several neuronal functions, including the promotion of synaptogenesis, neurite outgrowth and neuroprotection. Previous study has demonstrated that ganglioside GM1 inhibits the secretion of APPs; however the underlying mechanism remains unknown. Here we reported that GM1 can bind cellular full length APP and APPs secreted from APP(695) stably-transfected SH-SY5Y cells. To characterize the GM1-APP interaction further, we expressed and purified recombinant fragments of the N-terminal APP. Immunoprecipitation experiments revealed that GM1 was able to bind the recombinant APP(18-81) fragment. Moreover, the synthetic peptide APP(52-81) could inhibit the binding. Therefore, the binding site for GM1 appears to be located within residues 52-81 of APP. Furthermore, we found that only GM1, but not GD1a, GT1b and ceramide, binds APP-N-terminus, indicating that the specific binding depends on the sugar moiety of GM1. Fluorescent studies revealed a decrease in the intrinsic fluorescence intensity of the APP(52-81) peptide in phosphatidylcholine (PC)/GM1 vesicles. By using FTIR techniques, we found that the major secondary structure of the APP(52-81) peptide was altered in PC/GM1 vesicles. Our results demonstrate that GM1 binds the N-terminus of APP and induces a conformational change. These findings suggest that secreted APP is decreased by membrane GM1 binding to its precursor protein and provide a possible molecular mechanism to explain the involvement of GM1 in APP proteolysis and pathogenesis of Alzheimer's disease.
Zhang
Handi
H
Institute of Mental Health, Peking University, Key Laboratories for Mental Health, Ministry of Health, Beijing, China.
Ding
Jixin
J
Tian
Wenqiang
W
Wang
Lijun
L
Huang
Lixin
L
Ruan
Yan
Y
Lu
Tianlan
T
Sha
Yinlin
Y
Zhang
Dai
D
D016564:Amyloid beta-Protein Precursor / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002518:Ceramides / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005583:Fourier Analysis; D005677:G(M1) Ganglioside / Q000378:metabolism*; D005732:Gangliosides / Q000378:metabolism; D006801:Humans; D010713:Phosphatidylcholines / Q000378:metabolism; D058255:Protease Nexins; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013050:Spectrometry, Fluorescence / Q000379:methods; D053835:Unilamellar Liposomes / Q000378:metabolism
C545487:APP protein, human; D016564:Amyloid beta-Protein Precursor; D002518:Ceramides; D005732:Gangliosides; D010713:Phosphatidylcholines; D058255:Protease Nexins; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D053835:Unilamellar Liposomes; C021298:ganglioside, GD1a; D005677:G(M1) Ganglioside; C016481:trisialoganglioside GT1