Alzheimer鈥檚 Disease Amyloid 尾-Protein Mutations and Deletions That Define Neuronal Binding/Internalization as Early Stage Nonfibrillar/Fibrillar Aggregates and Late Stage Fibrils

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• 作者：Joseph F. Poduslo ; Kyle G. Howell ; Nicole C. Olson ; Marina Ramirez-Alvarado ; Karunya K. Kandimalla
• 刊名：Biochemistry
• 出版年：2012
• 出版时间：May 15, 2012
• 年：2012
• 卷：51
• 期：19
• 页码：3993-4003
• 全文大小：840K
• 年卷期：v.51,no.19(May 15, 2012)
• ISSN：1520-4995

文摘

Accumulation of amyloid 尾-protein (A尾) in neurons has been demonstrated to precede its formation as amyloid plaques in the extracellular space in Alzheimer鈥檚 disease (AD) patients. Consequently, intraneuronal A尾 accumulation is thought to be a critical first step in the fatal cascade of events that leads to neuronal degeneration in AD. Understanding the structural basis of neuronal binding and uptake of A尾 might lead to potential therapeutic targets that could block this binding and the subsequent neurodegeneration that leads to the pathogenesis of AD. Previously, we demonstrated that mutation of the two adjacent histidine residues of A尾40 (H13,14G) resulted in a significant decrease in its level of binding to PC12 cells and mouse cortical/hippocampal neurons. We now demonstrate that the weakened neuronal binding follows the mutation order of H13G < H14G < H13,14G, which suggests that the primary domain for neuronal binding of A尾40 involves histidine at position 13. A novel APP mutation (E693螖) that produced a variant A尾 lacking glutamate 22 (E22螖) in Japanese pedigrees was recently identified to have AD-type dementia without amyloid plaque formation but with extensive intraneuronal A尾 in transfected cells and transgenic mice expressing this deletion. Deletion of glutamate 22 of A尾40 resulted in a 6-fold enhancement of PC12 neuronal binding that was not decreased by the H13G mutation. The dose-dependent enhanced binding of E22螖 explains the high level of intraneuronal A尾 seen in this pedigree. Fluorescence anisotropy experiments at room temperature showed very rapid aggregation with increased tyrosine rigidity of A尾39E22螖, A尾41E22螖, and A尾42 but not A尾40. This rigidity was decreased but not eliminated by prior treatment with dimethyl sulfoxide. Surprisingly, all peptides showed an aggregated state when evaluated by transmission electron microscopy, with A尾39E22螖 having early stage fibrils, which was also verified by atomic force microscopy. This aggregation was not affected by centrifugation or pretreatment with organic solvents. The enhanced neuronal binding of A尾, therefore, results from aggregate binding to neurons, which requires H13 for A尾40 but not for E22螖 or A尾42. These latter proteins display increased tyrosine rigidity that likely masks the H13 residue, or alternatively, the H13 residue is not required for neuronal binding of these proteins as it is for A尾40. Late state fibrils also showed enhanced neuronal binding for E22螖 but not A尾40 with subsequent intraneuronal accumulation in lysosomes. This suggests that there are multiple pathways of binding/internalization for the different A尾 proteins and their aggregation states or fibrillar structure.