Sensitive Fluorescence Polarization Technique for Rapid Screening of -Synuclein Oligomerization/Fibrillization Inhibitors
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- 作者：Kelvin C. Luk ; Edward G. Hyde ; John Q. Trojanowski ; Virginia M.-Y. Lee
- 刊名：Biochemistry
- 出版年：2007
- 出版时间：November 6, 2007
- 年：2007
- 卷：46
- 期：44
- 页码：12522 - 12529
- 全文大小：633K
- 年卷期：v.46,no.44(November 6, 2007)
- ISSN：1520-4995
文摘
Parkinson's disease (PD) is characterized by the accumulation of fibrillar -synuclein (-Syn) inclusions known as Lewy bodies (LBs) and Lewy neurites. Mutations in the -Syn gene or extracopies thereof cause familial PD or dementia with LBs (DLB) in rare kindreds, but abnormal accumulationsof wildtype -Syn also are implicated in the pathogenesis of sporadic PD, the most common movementdisorder. Insights into mechanisms underlying -Syn mediated neurodegeneration link -Syn oligomerization and fibrillization to the onset and progression of PD. Thus, inhibiting -Syn oligomer or fibrilformation is a compelling target for discovering disease modifying therapies for PD, DLB, and relatedsynucleinopathies. Although amyloid dyes recognize -Syn fibrils, efficient detection of soluble oligomersremains a challenge. Here, we report a novel fluorescence polarization (FP) technique for examining-Syn assembly by monitoring changes in its relative molecular mass during progression of normal -Synfrom highly soluble monomers to higher order multimers and thence insoluble amyloid fibrils. We reportthat FP is more sensitive than conventional amyloid dye methods for the quantification of mature fibrils,and that FP is capable of detecting oligomeric -Syn, allowing for rapid automated screening of potentialinhibitors of -Syn oligomerization and fibrillization. Furthermore, FP can be combined with an amyloiddye in a single assay that simultaneously provides two independent biophysical readouts for monitoring-Syn fibrillization. Thus, this FP method holds potential to accelerate discovery of disease modifyingtherapies for LB PD, DLB, and related neurodegenerative synucleinopathies.