title: Application of photochemical cross-linking to the study of oligomerization of amyloidogenic proteins.
abstract: Assembly of amyloidogenic proteins into toxic oligomers and fibrils is an important pathogenic feature of over 30 amyloid-related diseases. Understanding the structures and mechanisms involved in the assembly process is necessary for rational approaches geared at inhibiting formation of these toxic species. Here, we review the application of photo-induced cross-linking of unmodified proteins (PICUP) to two disease-related amyloidogenic proteins (1) islet amyloid polypeptide (IAPP), whose toxic oligomers are thought to cause the demise of pancreatic β-cells in type-2 diabetes mellitus and (2) α-synuclein, which aggregates into toxic oligomers and precipitates in Lewy bodies in Parkinson's disease. PICUP is an effective method allowing chemical "freezing" of dynamically changing oligomers and subsequent study of the oligomer size distribution that existed before cross-linking. The method has provided insights into the factors controlling early oligomerization, which could not be obtained by other means. We discuss sample preparation, experimental details, optimization of parameters, and troubleshooting.
authors.affiliation: Department of Neurology, David Geffen School of Medicine at UCLA, Los Angeles, CA, USA. dahabadalopes@ucla.edu
authors.forename: Dahabada H J
authors.initials: DH
authors.lastname: Lopes
authors.forename: Sharmistha
authors.initials: S
authors.lastname: Sinha
authors.forename: Clark
authors.initials: C
authors.lastname: Rosensweig
authors.forename: Gal
authors.initials: G
authors.lastname: Bitan
mesh_terms: D004591:Electrophoresis, Polyacrylamide Gel; D058228:Islet Amyloid Polypeptide / Q000737:chemistry* / Q000302:isolation & purification; D055668:Photochemical Processes / Q000528:radiation effects; D020005:Propanols / Q000494:pharmacology; D055503:Protein Multimerization / Q000187:drug effects / Q000528:radiation effects*; D017433:Protein Structure, Secondary; D016622:Silver Staining; D012995:Solubility; D051844:alpha-Synuclein / Q000737:chemistry* / Q000302:isolation & purification; D006845:Hydrocarbons, Fluorinated
chemical_list: D058228:Islet Amyloid Polypeptide; D020005:Propanols; D051844:alpha-Synuclein; C001337:hexafluoroisopropanol; D006845:Hydrocarbons, Fluorinated