Tandem Repeats in Genes, Proteins, and Disease pp 21-40

Part of the Methods in Molecular Biology book series (MIMB, volume 1017) | Cite as

Atomic Force Microscopy Assays for Evaluating Polyglutamine Aggregation in Solution and on Surfaces

  • Kathleen A. Burke
  • Justin Legleiter
Protocol

Abstract

Mutations which cause an expansion of CAG triplet repeats encoding polyglutamine (polyQ) are responsible for the subsequent misfolding of specific proteins that contribute directly to the pathogenesis of at least nine neurodegenerative disorders, including Huntington’s disease (HD) and the spinocerebellar ataxias (SCAs). Expansion of polyQ tracts results in the aggregation of these proteins, potentially through a variety of precursor aggregates, into fibrillar structures. There may also be a variety of aggregates formed that are off-pathway to the formation of fibrils. Here, detailed protocols for analyzing the aggregation of mutant huntingtin (htt) fragments (associated with HD) and synthetic polyQ peptides with atomic force microscopy (AFM) are described. Ex situ AFM can be used to characterize htt aggregate formation and morphology. In situ AFM allows for tracking the formation and fate of individual polyQ peptide aggregates on surfaces. The interaction of htt with a variety of surfaces, including lipid bilayers, can also be investigated. Furthermore, the mechanical impact of htt on lipid surfaces can be studied using specialized AFM techniques. Methods to analyze AFM images of htt aggregates are also presented.

Key words

Atomic force microscopy Polyglutamine Huntington’s disease Oligomers Fibrils 

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Copyright information

© Springer Science+Business Media, New York 2013

Authors and Affiliations

  • Kathleen A. Burke
    • 1
  • Justin Legleiter
    • 1
    • 2
  1. 1.The C. Eugene Bennett Department of ChemistryWest Virginia UniversityMorgantownUSA
  2. 2.WVnano Initiative and the Center for NeurosciencesWest Virginia UniversityMorgantownUSA

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