title: Backbone Engineering within a Latent β-Hairpin Structure to Design Inhibitors of Polyglutamine Amyloid Formation.
abstract: Candidates for the toxic molecular species in the expanded polyglutamine (polyQ) repeat diseases range from various types of aggregates to "misfolded" monomers. One way to vet these candidates is to develop mutants that restrict conformational landscapes. Previously, we inserted two self-complementary β-hairpin enhancing motifs into a short polyQ sequence to generate a mutant, here called "βHP," that exhibits greatly improved amyloid nucleation without measurably enhancing β-structure in the monomer ensemble. We extend these studies here by introducing single-backbone H-bond impairing modificationsαN-methyl Gln or l-Pro at key positions within βHP. Modifications predicted to allow formation of a fully H-bonded β-hairpin at the fibril edge while interfering with H-bonding to the next incoming monomer exhibit poor amyloid formation and act as potent inhibitors in trans of simple polyQ peptide aggregation. In contrast, a modification that disrupts intra-β-hairpin H-bonding within βHP, while also aggregating poorly, is ineffective at inhibiting amyloid formation in trans. The inhibitors constitute a dynamic version of the edge-protection negative design strategy used in protein evolution to limit unwanted protein aggregation. Our data support a model in which polyQ peptides containing strong β-hairpin encouraging motifs only rarely form β-hairpin conformations in the monomer ensemble, but nonetheless take on such conformations at key steps during amyloid formation. The results provide insights into polyQ solution structure and fibril formation while also suggesting an approach to the design of inhibitors of polyQ amyloid growth that focuses on conformational requirements for fibril and nucleus elongation.
authors.affiliation: Department of Structural Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15260, USA.
authors.forename: Karunakar
authors.initials: K
authors.lastname: Kar
authors.affiliation: Department of Chemistry, Kenneth P. Dietrich School of Arts and Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA.
authors.forename: Matthew A
authors.initials: MA
authors.lastname: Baker
authors.affiliation: Department of Chemistry, Kenneth P. Dietrich School of Arts and Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA.
authors.forename: George A
authors.initials: GA
authors.lastname: Lengyel
authors.affiliation: Department of Structural Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15260, USA.
authors.forename: Cody L
authors.initials: CL
authors.lastname: Hoop
authors.affiliation: Department of Structural Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15260, USA.
authors.forename: Ravindra
authors.initials: R
authors.lastname: Kodali
authors.affiliation: Department of Structural Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15260, USA.
authors.forename: In-Ja
authors.initials: IJ
authors.lastname: Byeon
authors.affiliation: Department of Chemistry, Kenneth P. Dietrich School of Arts and Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA.
authors.forename: W Seth
authors.initials: WS
authors.lastname: Horne
authors.affiliation: Department of Structural Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15260, USA.
authors.forename: Patrick C A
authors.initials: PC
authors.lastname: van der Wel
authors.affiliation: Department of Structural Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15260, USA. Electronic address: rwetzel@pitt.edu.
authors.forename: Ronald
authors.initials: R
authors.lastname: Wetzel
mesh_terms: D000595:Amino Acid Sequence; D016229:Amyloid beta-Peptides / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D010455:Peptides / Q000037:antagonists & inhibitors / Q000737:chemistry*; D015202:Protein Engineering; D054730:Protein Interaction Domains and Motifs; D017433:Protein Structure, Secondary; D015203:Reproducibility of Results
chemical_list: D016229:Amyloid beta-Peptides; D010455:Peptides; C097188:polyglutamine
keywords: MAS-NMR; dock-and-lock; elongation; nucleation; trpzip