N-linked glycosylation of dimeric acetylcholinesterase in erythrocytes is essential for enzyme maturation and membrane targeting.
Acetylcholinesterase (AChE) is well-known for its cholinergic functions in the nervous system; however, this enzyme is also found in other tissues where its function is still not understood. AChE is synthesized through alternative splicing as splicing variants, with isoforms including read-through (AChE(R)), tailed (AChE(T)) and hydrophobic (AChE(H)). In human erythrocytes, AChE(H) is a glycophosphatidylinositol-linked dimer on the plasma membrane. Three N-linked glycosylation sites have been identified in the catalytic domain of human AChE. Here, we investigate the roles of glycosylation in assembly and trafficking of human AChE(H). In transfected fibroblasts, expression of AChE(H) was able to mimic the function of the dimeric form of AChE on the erythrocyte membrane. A glycan-depleted form was constructed by site-directed mutagenesis. By comparison with the wild-type AChE(H), the mutant had a much lower enzymatic activity and a much higher K(m) value. In addition, the mutant was dimerized in the endoplasmic reticulum, but was not trafficked to the Golgi apparatus. The results suggest that the glycosylation may affect AChE(H) enzymatic activity and trafficking, but not dimer formation. The present findings indicate the significance of N-glycosylation in controlling the biosynthesis of the AChE(H) dimer form.
Division of Life Science, Center for Chinese Medicine and State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science and Technology, Clear Water Bay Road, Hong Kong, China.
Wilson K W
WK
Luk
Vicky P
VP
Chen
Roy C Y
RC
Choi
Karl W K
KW
Tsim
D000110:Acetylcholinesterase / Q000097:blood* / Q000235:genetics; D015153:Blotting, Western; D002460:Cell Line; D019281:Dimerization; D004912:Erythrocytes / Q000201:enzymology*; D005455:Fluorescent Antibody Technique; D006031:Glycosylation; D006801:Humans; D016297:Mutagenesis, Site-Directed; D021381:Protein Transport; D013347:Subcellular Fractions / Q000201:enzymology
D000110:Acetylcholinesterase