doi: 10.1016/j.compchemeng.2018.02.013. Epub 2018 Feb 21.
Elucidating the multi-targeted anti-amyloid activity and enhanced islet amyloid polypeptide binding ofβ-wrapins
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- PMID:30405276
- PMCID: PMC6217933
- DOI: 10.1016/j.compchemeng.2018.02.013
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Elucidating the multi-targeted anti-amyloid activity and enhanced islet amyloid polypeptide binding ofβ-wrapins
Asuka A Orr et al. Comput Chem Eng..
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Abstract
β-wrapins are engineered binding proteins stabilizing theβ-hairpin conformations of amyloidogenic proteins islet amyloid polypeptide (IAPP), amyloid-β, andα-synuclein, thus inhibiting their amyloid propensity. Here, we use computational and experimental methods to investigate the molecular recognition of IAPP byβ-wrapins. We show that the multi-targeted, IAPP, amyloid-β, andα-synuclein, binding properties ofβ-wrapins originate mainly from optimized interactions betweenβ-wrapin residues and sets of residues in the three amyloidogenic proteins with similar physicochemical properties. Our results suggest that IAPP is a comparatively promiscuousβ-wrapin target, probably due to the low number of charged residues in the IAPPβ-hairpin motif. The sub-micromolar affinity ofβ-wrapin HI18, specifically selected against IAPP, is achieved in part by salt-bridge formation between HI18 residue Glu10 and the IAPP N-terminal residue Lys1, both located in the flexible N-termini of the interacting proteins. Our findings provide insights towards developing novel protein-based single- or multi-targeted therapeutics.
Keywords: Amylin; Amyloid-β; Intrinsically disordered proteins; Molecular dynamics; Protein aggregation; α-synuclein.
Figures
(A) Sequences of the investigatedβ-wrapin variants aligned to ZAβ3 and their corresponding dissociation constants for Aβ,α-syn, and IAPP reported here and in Mirecka et al. (2014a), Shaykhalishahi et al. (2015), and Mirecka et al. (2014b). The dissociation constant for ZAβ3 toα-syn was not detected (n.d.). (B, C, and D) Binding of IAPP to ZAβ3, AS69, and ZAβ3_A10E analyzed by SPR. Representative sensorgrams were recorded by injection of ZAβ3 (B), AS69 (C), or ZAβ3_A10E (D) at the indicated concentrations onto a flow cell with immobilized IAPP for 90 s, followed by washing with buffer for 600 s. Global fitting to a two-state interaction model is shown in black, yieldingKd values of 1.31μM, 1.23μM, or 620 nM for ZAβ3, AS69, or ZAβ3_A10E, respectively.
Free energy landscape (FEL) constructed from the 2D probability of principal components PC1 and PC2, calculated for the modeled region of the HI18:IAPP complex (residues 9 through 12 of HI18, subunit 1 and residues 1 through 9 of IAPP) derived from REMD simulations at 300 K. The global free energy minimum of the FEL is encircled in a black oval shape. Within the free energy minimum basin, the structures encompass a salt-bridge between subunit 1 residue Glu10 of HI18 with primarily the positively charged N-terminal domain of residues Lys1 of IAPP and, in a few structures, the side chain group of the same IAPP residue. The representative structure which was extracted from the FEL and used as a template and initial structure for the subsequent MD simulations is shown at the bottom of the FEL.
Molecular graphics images of HI18 in complex with IAPP. (A,B,C) Snapshots extracted from the MD simulations illustrating the increased mobility of the N-termini in comparison to the core of the complex. Glu10 of subunit 1 in HI18 forms salt-bridges with Lys1 and Arg11 of IAPP as well as hydrogen bonds with Thr9 of IAPP. The flexible N-termini are encircled with green dotted lines. Interactions between Glu10 of subunit 1 in HI18 and the N-terminus of IAPP remain prevalent throughout the MD simulations. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)
Molecular graphics images of HI18 in complex with IAPP. Polar interactions between the flexible N-termini enhance nonpolar interactions between IAPP and HI18, extending the hydrophobic surface shown in orange (β-wrapin subunit 1), red (β-wrapin subunit 2), cyan and blue (IAPP) surface representation, of the antiparallelβ-sheet in the structure.β-Wrapin subunits 1 and 2 are shown in red and orange tube representation, respectively, and IAPP is shown in cyan and blue tube representation. Red and blue labels indicate residues of subunit 1 of theβ-wrapin and IAPP, respectively. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)
Molecular graphics images of (A) HI18 and (B) ZAβ3_A10E in complex with IAPP. These twoβ-wrapins only differ at residue position 34, which is Ile in HI18 and Leu in ZAβ3_A10E. (A) Favorable interactions occurring in the HI18:IAPP complex are encapsulated in purple. (B) Weakened interactions occurring in the ZAβ3_A10E: IAPP complex are indicated with purple dotted lines between the two interacting residue pairs.β-Wrapin subunits 1 and 2 are shown in red and orange tube representation, respectively, and IAPP is shown in blue tube representation. Red, orange, and blue labels indicate residues of subunit 1 of theβ-wrapin, subunit 2 of theβ-wrapin, and IAPP, respectively. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)
Molecular graphics images of the common hydrophobic interactions of AS10 in complex with IAPP (panel A), Aβ (panel B), andα-syn (panel C). AS10 subunits 1 and 2 are shown in red and orange tube representation, respectively, and IAPP is shown in blue tube representation. The specified hydrophobic and aromatic interactions contribute significantly to the ability of AS10 to sequester all three of the amyloidogenic proteins. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)
Molecular graphics images of the intra-subunit polar interactions within the monomer subunits of the HI18:IAPP complex. HI18 subunits 1 and 2 are shown in red and orange tube representation, respectively, and IAPP is shown in blue tube representation. The specified hydrogen bond and salt-bridge interactions are indicated with black dotted lines. These polar interactions are present for allβ-wrapin:IAPP complexes investigated in this study, stabilizing the tertiary structure of the monomer subunits. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)
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