The assembly of proline-rich membrane anchor (PRiMA)-linked acetylcholinesterase enzyme: glycosylation is required for enzymatic activity but not for oligomerization |
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Authors: | Chen Vicky P Choi Roy C Y Chan Wallace K B Leung K Wing Guo Ava J Y Chan Gallant K L Luk Wilson K W Tsim Karl W K |
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Institution: | Division of Life Science and Center for Chinese Medicine, The Hong Kong University of Science and Technology, Hong Kong SAR, China. |
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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. |
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Keywords: | Acetylcholinesterase Glycosylation Membrane Trafficking Protein Assembly Subcellular Fractionation PRiMA Tetramer |
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