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Site-specific detection of protein secondary structure using 2D IR dihedral indexing: a proposed assembly mechanism of oligomeric hIAPP

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Abstract

Human islet amyloid polypeptide (hIAPP) aggregates into fibrils through oligomers that have been postulated to contain α-helices as well as β-sheets. We employ a site-specific isotope labeling strategy that is capable of detecting changes in dihedral angles when used in conjunction with 2D IR spectroscopy. The method is analogous to the chemical shift index used in NMR spectroscopy for assigning protein secondary structure. We introduce isotope labels at two neighbouring residues, which results in an increased intensity and positive frequency shift if those residues are α-helical versus a negative frequency shift in β-sheets and turns. The 2D IR dihedral index approach is demonstrated for hIAPP in micelles for which the polypeptide structure is known, using pairs of 13C18O isotope labels L12A13 and L16V17, along with single labeled control experiments. Applying the approach to aggregation experiments performed in buffer, we show that about 27–38% of hIAPP peptides adopt an α-helix secondary structure in the monomeric state at L12A13, prior to aggregation, but not at L16V17 residues. At L16V17, the kinetics are described solely by the monomer and fiber conformations, but at L12A13 the kinetics exhibit a third state that is created by an oligomeric intermediate. Control experiments performed with a single isotope label at A13 exhibit two-state kinetics, indicating that a previously unknown change in dihedral angle occurs at L12A13 as hIAPP transitions from the intermediate to fiber structures. We propose a mechanism for aggregation, in which helices seed oligomer formation via structures analogous to leucine rich repeat proteins.

Graphical abstract: Site-specific detection of protein secondary structure using 2D IR dihedral indexing: a proposed assembly mechanism of oligomeric hIAPP

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Publication details

The article was received on 30 Aug 2017, accepted on 02 Nov 2017 and first published on 03 Nov 2017


Article type: Edge Article
DOI: 10.1039/C7SC03789A
Citation: Chem. Sci., 2018, Advance Article
  • Open access: Creative Commons BY license
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    Site-specific detection of protein secondary structure using 2D IR dihedral indexing: a proposed assembly mechanism of oligomeric hIAPP

    M. Maj, J. P. Lomont, K. L. Rich, A. M. Alperstein and M. T. Zanni, Chem. Sci., 2018, Advance Article , DOI: 10.1039/C7SC03789A

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