title: Spectroscopic studies on native and protofibrillar insulin.
abstract: The structure of insulin in amyloid fibrillar form has been recently shown as a well folded conformation using cryoelectron microscopy [Jimenez, J.L., Nettleton, E.J., Bouchard, M., Robinson, C.V., Dobson, C.M., Saibil H.R., 2002. The protofilament structure of insulin amyloid fibrils. Proc. Natl. Acad. Sci. USA. 99 9196-9201.]. Most of the amyloid aggregates elicit maximum toxicity in the protofibrillar (PF) intermediate state. Here, we describe PF intermediates of insulin are made-up monomers with flexible conformers. We also show protofibrils have three-dimensionally extended hydrophobic cavity to bind with 1-anilinonaphthalene-8-sulphonate (ANS) molecules. Energy transfer measurement revealed that ANS dye binding site of PF is within the range of FRET distance of insulin tyrosine residues. Significant proportion of beta-sheet, helical, and turn structures in the PF form indicate conformational dynamics in the folded chain of insulin in the PF assembled form. Though the conformational flexibility is noticeably present in the assembly, addition of GdnHCl could completely unfold PF into disordered structure suggesting structural "zipping" in the PF form. We have also shown that helical conformer inducing solvent 2,2,2-trifluoroethanol (TFE) could dissociate the PF aggregate indicating possible involvement of beta-sheets in contributing to PF stability.
authors.affiliation: Bioorganic and Neurochemistry Laboratory, Central Leather Research Institute, Adyar, Chennai 600 020, India.
authors.forename: J
authors.initials: J
authors.lastname: Murali
authors.forename: R
authors.initials: R
authors.lastname: Jayakumar
mesh_terms: D000682:Amyloid / Q000737:chemistry*; D000817:Anilino Naphthalenesulfonates; D001665:Binding Sites; D031541:Fluorescence Resonance Energy Transfer; D007328:Insulin / Q000737:chemistry*; D011487:Protein Conformation; D017510:Protein Folding
chemical_list: D000682:Amyloid; D000817:Anilino Naphthalenesulfonates; D007328:Insulin; C027132:1-anilino-8-naphthalenesulfonate