Synapse-Binding Subpopulations of A尾 Oligomers Sensitive to Peptide Assembly Blockers and scFv Antibodies

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• 作者：Pauline T. Velasco ; Marie C. Heffern ; Adriano Sebollela ; Izolda A. Popova ; Pascale N. Lacor ; Kevin B. Lee ; Xiaoxia Sun ; Benjamin N. Tiano ; Kirsten L. Viola ; Amanda L. Eckermann ; Thomas J. Meade ; William L. Klein
• 刊名：ACS Chemical Neuroscience
• 出版年：2012
• 出版时间：November 21, 2012
• 年：2012
• 卷：3
• 期：11
• 页码：972-981
• 全文大小：354K
• 年卷期：v.3,no.11(November 21, 2012)
• ISSN：1948-7193

文摘

Amyloid 尾42 self-assembly is complex, with multiple pathways leading to large insoluble fibrils or soluble oligomers. Oligomers are now regarded as most germane to Alzheimer鈥檚 pathogenesis. We have investigated the hypothesis that oligomer formation itself occurs through alternative pathways, with some leading to synapse-binding toxins. Immediately after adding synthetic peptide to buffer, solutions of A尾42 were separated by a 50 kDa filter and fractions assessed by SDS-PAGE silver stain, Western blot, immunoprecipitation, and capacity for synaptic binding. A尾42 rapidly assembled into aqueous-stable oligomers, with similar protein abundance in small (<50 kDa) and large (>50 kDa) oligomer fractions. Initially, both fractions were SDS-labile and resolved into tetramers, trimers, and monomers by SDS-PAGE. Upon continued incubation, the larger oligomers developed a small population of SDS-stable 10鈥?6mers, and the smaller oligomers generated gel-impermeant complexes. The two fractions associated differently with neurons, with prominent synaptic binding limited to larger oligomers. Even within the family of larger oligomers, synaptic binding was associated with only a subset of these species, as a new scFv antibody (NUsc1) immunoprecipitated only a small portion of the oligomers while eliminating synaptic binding. Interestingly, low doses of the peptide KLVFFA blocked assembly of the 10鈥?6mers, and this result was associated with loss of the smaller clusters of oligomers observed at synaptic sites. What distinguishes these smaller clusters from the unaffected larger clusters is not yet known. Results indicate that distinct species of A尾 oligomers are generated by alternative assembly pathways and that synapse-binding subpopulations of A尾 oligomers could be specifically targeted for Alzheimer鈥檚 therapeutics.

Keywords:

Alzheimer鈥檚 disease; amyloid beta; blocking peptide; therapeutics; single-chain variable fragment antibody; oligomerization