title: The assembly of proline-rich membrane anchor (PRiMA)-linked acetylcholinesterase enzyme: glycosylation is required for enzymatic activity but not for oligomerization.
abstract: Acetylcholinesterase (AChE) anchors onto cell membranes by a transmembrane protein PRiMA (proline-rich membrane anchor) as a tetrameric form in vertebrate brain. The assembly of AChE tetramer with PRiMA requires the C-terminal "t-peptide" in AChE catalytic subunit (AChE(T)). Although mature AChE is well known N-glycosylated, the role of glycosylation in forming the physiologically active PRiMA-linked AChE tetramer has not been studied. Here, several lines of evidence indicate that the N-linked glycosylation of AChE(T) plays a major role for acquisition of AChE full enzymatic activity but does not affect its oligomerization. The expression of the AChE(T) mutant, in which all N-glycosylation sites were deleted, together with PRiMA in HEK293T cells produced a glycan-depleted PRiMA-linked AChE tetramer but with a much higher K(m) value as compared with the wild type. This glycan-depleted enzyme was assembled in endoplasmic reticulum but was not transported to Golgi apparatus or plasma membrane.
authors.affiliation: Division of Life Science and Center for Chinese Medicine, The Hong Kong University of Science and Technology, Hong Kong SAR, China.
authors.forename: Vicky P
authors.initials: VP
authors.lastname: Chen
authors.forename: Roy C Y
authors.initials: RC
authors.lastname: Choi
authors.forename: Wallace K B
authors.initials: WK
authors.lastname: Chan
authors.forename: K Wing
authors.initials: KW
authors.lastname: Leung
authors.forename: Ava J Y
authors.initials: AJ
authors.lastname: Guo
authors.forename: Gallant K L
authors.initials: GK
authors.lastname: Chan
authors.forename: Wilson K W
authors.initials: WK
authors.lastname: Luk
authors.forename: Karl W K
authors.initials: KW
authors.lastname: Tsim
mesh_terms: D000110:Acetylcholinesterase / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D055162:Biocatalysis; D002645:Chickens; D004795:Enzyme Stability; D058851:GPI-Linked Proteins / Q000737:chemistry / Q000378:metabolism; D006031:Glycosylation; D057809:HEK293 Cells; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D021381:Protein Transport; D011994:Recombinant Proteins / Q000378:metabolism
chemical_list: D058851:GPI-Linked Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D011134:Polysaccharides; D011994:Recombinant Proteins; C486718:prima1 protein, mouse; C548244:ACHE protein, human; D000110:Acetylcholinesterase