ジャーナル: Front Mol Biosci / 年: 2017 タイトル: Electron Microscopy Structural Insights into CPAP Oligomeric Behavior: A Plausible Assembly Process of a Supramolecular Scaffold of the Centrosome. 著者: Ana L Alvarez-Cabrera / Sandra Delgado / David Gil-Carton / Gulnahar B Mortuza / Guillermo Montoya / Carlos O S Sorzano / Tang K Tang / Jose M Carazo / 要旨: Centrosomal P4.1-associated protein (CPAP) is a cell cycle regulated protein fundamental for centrosome assembly and centriole elongation. In humans, the region between residues 897-1338 of CPAP ...Centrosomal P4.1-associated protein (CPAP) is a cell cycle regulated protein fundamental for centrosome assembly and centriole elongation. In humans, the region between residues 897-1338 of CPAP mediates interactions with other proteins and includes a homodimerization domain. CPAP mutations cause primary autosomal recessive microcephaly and Seckel syndrome. Despite of the biological/clinical relevance of CPAP, its mechanistic behavior remains unclear and its C-terminus (the G-box/TCP domain) is the only part whose structure has been solved. This situation is perhaps due in part to the challenges that represent obtaining the protein in a soluble, homogeneous state for structural studies. Our work constitutes a systematic structural analysis on multiple oligomers of , using single-particle electron microscopy (EM) of negatively stained (NS) samples. Based on image classification into clearly different regular 3D maps (putatively corresponding to dimers and tetramers) and direct observation of individual images representing other complexes of CPAP (i.e., putative flexible monomers and higher-order multimers), we report a dynamic oligomeric behavior of this protein, where different homo-oligomers coexist in variable proportions. We propose that dimerization of the putative homodimer forms a putative tetramer which could be the structural unit for the scaffold that either tethers the pericentriolar material to centrioles or promotes procentriole elongation. A coarse fitting of atomic models into the NS 3D maps at resolutions around 20 Å is performed only to complement our experimental data, allowing us to hypothesize on the oligomeric composition of the different complexes. In this way, the current EM work represents an initial step toward the structural characterization of different oligomers of CPAP, suggesting further insights to understand how this protein works, contributing to the elucidation of control mechanisms for centriole biogenesis.
全体 : Putative homodimer of human CPAP (residues 897-1338)
全体
名称: Putative homodimer of human CPAP (residues 897-1338)
要素
複合体: Putative homodimer of human CPAP (residues 897-1338)
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超分子 #1: Putative homodimer of human CPAP (residues 897-1338)
超分子
名称: Putative homodimer of human CPAP (residues 897-1338) タイプ: complex / ID: 1 / 親要素: 0 詳細: Human CPAP (residues 897-1338) expresed in E. coli and purified by Immobilized Metal Affinity Chromatography (IMAC) and Size Exclusion Chromatography (SEC)