title: The dye SYPRO orange binds to amylin amyloid fibrils but not pre-fibrillar intermediates.
abstract: Amyloid deposition underlies a broad range of diseases including multiple neurodegenerative diseases, systemic amyloidosis and type-2 diabetes. Amyloid sensitive dyes, particularly thioflavin-T, are widely used to detect ex-vivo amyloid deposits, to monitor amyloid formation in vitro and to follow the kinetics of amyloid self-assembly. We show that the dye SYPRO-orange binds to amyloid fibrils formed by human amylin, the polypeptide responsible for islet amyloid formation in type-2 diabetes. No fluorescence enhancement is observed in the presence of pre-fibrillar species or in the presence of non-amyloidogenic rat amylin. The kinetics of human amylin amyloid formation can be monitored by SYPRO-orange fluorescence and match the time course determined with thioflavin-T assays. Thus, SYPRO-orange offers an alternative to thioflavin-T assays of amylin amyloid formation. The implications for the interpretation of SYPRO-orange-based assays of protein stability and protein-ligand interactions are discussed.
authors.affiliation: Department of Chemistry, Stony Brook University, Stony Brook, New York, 11794-3400.
authors.forename: Amy G
authors.initials: AG
authors.lastname: Wong
authors.affiliation: Department of Chemistry, Stony Brook University, Stony Brook, New York, 11794-3400. daniel.raleigh@stonybrook.edu.|Graduate Program in Biochemistry and Structural Biology, Stony Brook University, Stony Brook, New York, 11794. daniel.raleigh@stonybrook.edu.
authors.forename: Daniel P
authors.initials: DP
authors.lastname: Raleigh
mesh_terms: D000682:Amyloid / Q000737:chemistry*; D052160:Benzothiazoles; D005456:Fluorescent Dyes / Q000737:chemistry*; D006801:Humans; D058228:Islet Amyloid Polypeptide / Q000737:chemistry*; D007700:Kinetics; D013844:Thiazoles / Q000737:chemistry
chemical_list: D000682:Amyloid; D052160:Benzothiazoles; D005456:Fluorescent Dyes; D058228:Islet Amyloid Polypeptide; D013844:Thiazoles; C009462:thioflavin T
keywords: SYPRO-orange; amylin; amyloid; islet amyloid polypeptide; thioflavin-T