Structural study of metastable amyloidogenic protein oligomers by photo-induced cross-linking of unmodified proteins.
Oligomers of amyloidogenic proteins are believed to be key effectors of cytotoxicity and cause a variety of amyloid-related diseases. Dissociation or inhibition of formation of the toxic oligomers is thus an attractive strategy for the prevention and treatment of these diseases. In order to develop reagents capable of inhibiting protein oligomerization, the structures and mechanisms of oligomer formation must be understood. However, structural studies of oligomers are difficult because of the metastable nature of the oligomers and their existence in mixtures with monomers and other assemblies. A useful method for characterization of oligomer size distributions in vitro is photo-induced cross-linking of unmodified proteins (PICUP) (Fancy and Kodadek, 1999). By providing "snapshots" of dynamic oligomer mixtures, PICUP enables quantitative analysis of the relations between primary and quaternary structures, offering insights into the molecular organization of the oligomers. This chapter discusses the photochemical mechanism; reviews the scope, usefulness, and limitations of PICUP for characterizing metastable protein assemblies; and provides detailed experimental instructions for performing PICUP experiments.
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UCLA, Department of Neurology, David Geffen School of Medicine, Los Angeles, CA, USA.
D015082:2,2'-Dipyridyl / Q000031:analogs & derivatives / Q000737:chemistry; D000682:Amyloid / Q000737:chemistry*; D016229:Amyloid beta-Peptides / Q000737:chemistry; D002116:Calcitonin / Q000737:chemistry; D056831:Coordination Complexes; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D013007:Growth Hormone-Releasing Hormone / Q000737:chemistry; D010446:Peptide Fragments / Q000737:chemistry; D010777:Photochemistry; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide / Q000737:chemistry; D017510:Protein Folding; D020836:Protein Structure, Quaternary; D017975:Ruthenium Compounds
D015082:2,2'-Dipyridyl; D000682:Amyloid; D016229:Amyloid beta-Peptides; D002116:Calcitonin; D056831:Coordination Complexes; D003432:Cross-Linking Reagents; D013007:Growth Hormone-Releasing Hormone; D010446:Peptide Fragments; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; C078592:amyloid beta-protein (1-40); C547232:tris(2,2-bipyridine)-ruthenium(II); D017975:Ruthenium Compounds