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

作者：Poduslo Joseph F^*; Howell Kyle G; Olson Nicole C; Ramirez Alvarado Marina; Kandimalla Karunya K

来源：Biochemistry, 2012, 51(19): 3993-4003.

DOI：10.1021/bi300275g

Accumulation of amyloid beta-protein (A beta) in neurons has been demonstrated to precede its formation as amyloid plaques in the extracellular space in Alzheimer's disease (AD) patients. Consequently, intraneuronal A beta 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 beta 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 beta 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 beta 40 involves histidine at position 13. A novel APP mutation (E693 Delta) that produced a variant A beta lacking glutamate 22 (E22 Delta) in Japanese pedigrees was recently identified to have AD-type dementia without amyloid plaque formation but with extensive intraneuronal A beta in transfected cells and transgenic mice expressing this deletion. Deletion of glutamate 22 of A beta 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 Delta explains the high level of intraneuronal A beta seen in this pedigree. Fluorescence anisotropy experiments at room temperature showed very rapid aggregation with increased tyrosine rigidity of A beta 39E22 Delta, A beta 41E22 Delta, and A beta 42 but not A beta 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 beta 39E22 Delta 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 beta therefore, results from aggregate binding to neurons, which requires H13 for A beta 40 but not for E22 Delta or A beta 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 beta 40. Late state fibrils also showed enhanced neuronal binding for E22 Delta but not A beta 40 with subsequent intraneuronal accumulation in lysosomes. This suggests that there are multiple pathways of binding/internalization for the different A beta proteins and their aggregation states or fibrillar structure.

• 出版日期2012-5-15