title: Clues for divergent, polymorphic amyloidogenesis through dissection of amyloid forming steps of bovine carbonic anhydrase and its critical amyloid forming stretch.
abstract: Certain amino acid stretches are considered 'critical' to trigger amyloidogenesis in a protein. Synthetic peptides corresponding to these stretches are often used as experimental mimics for studying the amyloidogenesis of their parent protein. Here we provide evidence that such simple extrapolation is misleading. We scrutinized each step of amyloid progression of full length bovine carbonic anhydrase (BCA) and compared it with the amyloidogenic process of its critical peptide stretch 201-227 (PepB). We found that under similar solution conditions amyloidogenesis of BCA followed surface-catalyzed secondary nucleation, whereas, that of PepB followed classical nucleation-dependent pathway. AFM images showed that while BCA formed short, thick and branched fibrils, PepB formed thin, long and unbranched fibrils. Structural information obtained by ATR-FTIR spectroscopy suggested parallel arrangement of intermolecular β-sheet in BCA amyloids in contrast to PepB amyloids which arranged into antiparallel β sheets. Amyloids formed by BCA were unable to seed the fibrillation of PepB and vice versa. Even the intermediates formed during lag phase revealed contrasting FTIR and far UV CD signature, hydrophobicity, morphology and cell cytotoxicity. Thus, we propose that sequences other than critical amyloidogenic stretches may significantly influence the initiation, polymerization and final fibrillar morphology of amyloid forming protein. The results have been discussed in light of primary sequence mediated amyloid polymorphism and its importance in the rational design of amyloid nanomaterials possessing desired physico-chemical properties.
authors.affiliation: Kusuma School of Biological Sciences, Indian Institute of Technology Delhi, Hauz Khas, New Delhi 110016, India.
authors.forename: Dushyant Kumar
authors.initials: DK
authors.lastname: Garg
authors.affiliation: Kusuma School of Biological Sciences, Indian Institute of Technology Delhi, Hauz Khas, New Delhi 110016, India. Electronic address: bkundu@bioschool.iitd.ac.in.
authors.forename: Bishwajit
authors.initials: B
authors.lastname: Kundu
mesh_terms: D000595:Amino Acid Sequence; D000682:Amyloid / Q000737:chemistry* / Q000648:ultrastructure; D000818:Animals; D002256:Carbonic Anhydrases / Q000737:chemistry* / Q000648:ultrastructure; D002417:Cattle; D018625:Microscopy, Atomic Force; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D020539:Sequence Analysis, Protein; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D000682:Amyloid; D002256:Carbonic Anhydrases
keywords: Amyloid fibril; Amyloid polymorphism; Bovine carbonic anhydrase; Critical amyloid stretch; FTIR; Secondary nucleation