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Time-resolved studies define the nature of toxic IAPP intermediates, providing insight for anti-amyloidosis therapeutics.

Abedini A, Plesner A, Cao P, Ridgway Z, Zhang J, Tu LH, Middleton CT, Chao B, Sartori DJ, Meng F, Wang H, Wong AG, Zanni MT, Verchere CB, Raleigh DP, Schmidt AM - Elife (2016)

Bottom Line: These globally flexible, low order oligomers upregulate pro-inflammatory markers and induce reactive oxygen species.They do not bind 1-anilnonaphthalene-8-sulphonic acid and lack extensive β-sheet structure.Aromatic interactions modulate, but are not required for toxicity.

View Article: PubMed Central - PubMed

Affiliation: Diabetes Research Program, Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, New York University School of Medicine, New York, United States.

ABSTRACT
Islet amyloidosis by IAPP contributes to pancreatic β-cell death in diabetes, but the nature of toxic IAPP species remains elusive. Using concurrent time-resolved biophysical and biological measurements, we define the toxic species produced during IAPP amyloid formation and link their properties to induction of rat INS-1 β-cell and murine islet toxicity. These globally flexible, low order oligomers upregulate pro-inflammatory markers and induce reactive oxygen species. They do not bind 1-anilnonaphthalene-8-sulphonic acid and lack extensive β-sheet structure. Aromatic interactions modulate, but are not required for toxicity. Not all IAPP oligomers are toxic; toxicity depends on their partially structured conformational states. Some anti-amyloid agents paradoxically prolong cytotoxicity by prolonging the lifetime of the toxic species. The data highlight the distinguishing properties of toxic IAPP oligomers and the common features that they share with toxic species reported for other amyloidogenic polypeptides, providing information for rational drug design to treat IAPP induced β-cell death.

No MeSH data available.


Related in: MedlinePlus

Aromatic residues in the ensemble of toxic h-IAPP oligomers are solvent exposed.(A) Primary sequences of h-IAPP and p-cyano-phenylalanine variants; red X=cyanophenylalanine. (B) A structural model of the h-IAPP amyloid fibril. (C) Location of aromatic residues in h-IAPP which are replaced with p-cyano-phenylalanine in the h-IAPP variants. (D) Structure of the unnatural amino acid p-cyano-phenylalanine. (E) p-Cyano-phenylalanine fluorescence emission spectra reveal that aromatic side chains are solvent exposed in time-zero species (black, ····) and lag phase intermediates (blue, —), but are buried in amyloid fibrils (red, - - - -).DOI:http://dx.doi.org/10.7554/eLife.12977.040
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fig8: Aromatic residues in the ensemble of toxic h-IAPP oligomers are solvent exposed.(A) Primary sequences of h-IAPP and p-cyano-phenylalanine variants; red X=cyanophenylalanine. (B) A structural model of the h-IAPP amyloid fibril. (C) Location of aromatic residues in h-IAPP which are replaced with p-cyano-phenylalanine in the h-IAPP variants. (D) Structure of the unnatural amino acid p-cyano-phenylalanine. (E) p-Cyano-phenylalanine fluorescence emission spectra reveal that aromatic side chains are solvent exposed in time-zero species (black, ····) and lag phase intermediates (blue, —), but are buried in amyloid fibrils (red, - - - -).DOI:http://dx.doi.org/10.7554/eLife.12977.040

Mentions: We next probed the solvent exposure of the three aromatic residues of h-IAPP using the non-genetically coded fluorescent amino acid, p-cyano-phenylalanine (p-cyanoPhe) (Figure 8). p-CyanoPhe can be incorporated into proteins and used to follow amyloid formation (Marek et al., 2010a). Its fluorescence is high when the cyano-group is solvent exposed and hydrogen bonded, and low when it is not; the fluorescence is also quenched via FRET to Tyr with a Ro of 15Å. h-IAPP contains two Phe and one Tyr; thus, three analogs were prepared in which one aromatic residue was replaced by p-cyanoPhe at each position (Figure 8A-D). Fluorescence is high for unaggregated h-IAPP (time-zero) and is quenched in the amyloid fibrils. The fluorescence intensity of the lag phase intermediates is also high and shows only moderate differences from the value observed for each peptide at time-zero, but is much higher than the intensity observed from the amyloid fibrils (Figure 8E). The data indicate that Phe-15, Phe-23 and Tyr-37 are largely solvent exposed in the lag phase, and rule out a significant population of conformations in the ensemble in which the aromatic residues are buried, or in which the C-terminal Tyr forms persistent interactions with either Phe-15 or Phe-23. Again, these results are compatible with 2D IR studies undertaken at higher peptide concentrations, which postulate formation of β-sheet structure in the FGAIL region.10.7554/eLife.12977.040Figure 8.Aromatic residues in the ensemble of toxic h-IAPP oligomers are solvent exposed.


Time-resolved studies define the nature of toxic IAPP intermediates, providing insight for anti-amyloidosis therapeutics.

Abedini A, Plesner A, Cao P, Ridgway Z, Zhang J, Tu LH, Middleton CT, Chao B, Sartori DJ, Meng F, Wang H, Wong AG, Zanni MT, Verchere CB, Raleigh DP, Schmidt AM - Elife (2016)

Aromatic residues in the ensemble of toxic h-IAPP oligomers are solvent exposed.(A) Primary sequences of h-IAPP and p-cyano-phenylalanine variants; red X=cyanophenylalanine. (B) A structural model of the h-IAPP amyloid fibril. (C) Location of aromatic residues in h-IAPP which are replaced with p-cyano-phenylalanine in the h-IAPP variants. (D) Structure of the unnatural amino acid p-cyano-phenylalanine. (E) p-Cyano-phenylalanine fluorescence emission spectra reveal that aromatic side chains are solvent exposed in time-zero species (black, ····) and lag phase intermediates (blue, —), but are buried in amyloid fibrils (red, - - - -).DOI:http://dx.doi.org/10.7554/eLife.12977.040
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4940161&req=5

fig8: Aromatic residues in the ensemble of toxic h-IAPP oligomers are solvent exposed.(A) Primary sequences of h-IAPP and p-cyano-phenylalanine variants; red X=cyanophenylalanine. (B) A structural model of the h-IAPP amyloid fibril. (C) Location of aromatic residues in h-IAPP which are replaced with p-cyano-phenylalanine in the h-IAPP variants. (D) Structure of the unnatural amino acid p-cyano-phenylalanine. (E) p-Cyano-phenylalanine fluorescence emission spectra reveal that aromatic side chains are solvent exposed in time-zero species (black, ····) and lag phase intermediates (blue, —), but are buried in amyloid fibrils (red, - - - -).DOI:http://dx.doi.org/10.7554/eLife.12977.040
Mentions: We next probed the solvent exposure of the three aromatic residues of h-IAPP using the non-genetically coded fluorescent amino acid, p-cyano-phenylalanine (p-cyanoPhe) (Figure 8). p-CyanoPhe can be incorporated into proteins and used to follow amyloid formation (Marek et al., 2010a). Its fluorescence is high when the cyano-group is solvent exposed and hydrogen bonded, and low when it is not; the fluorescence is also quenched via FRET to Tyr with a Ro of 15Å. h-IAPP contains two Phe and one Tyr; thus, three analogs were prepared in which one aromatic residue was replaced by p-cyanoPhe at each position (Figure 8A-D). Fluorescence is high for unaggregated h-IAPP (time-zero) and is quenched in the amyloid fibrils. The fluorescence intensity of the lag phase intermediates is also high and shows only moderate differences from the value observed for each peptide at time-zero, but is much higher than the intensity observed from the amyloid fibrils (Figure 8E). The data indicate that Phe-15, Phe-23 and Tyr-37 are largely solvent exposed in the lag phase, and rule out a significant population of conformations in the ensemble in which the aromatic residues are buried, or in which the C-terminal Tyr forms persistent interactions with either Phe-15 or Phe-23. Again, these results are compatible with 2D IR studies undertaken at higher peptide concentrations, which postulate formation of β-sheet structure in the FGAIL region.10.7554/eLife.12977.040Figure 8.Aromatic residues in the ensemble of toxic h-IAPP oligomers are solvent exposed.

Bottom Line: These globally flexible, low order oligomers upregulate pro-inflammatory markers and induce reactive oxygen species.They do not bind 1-anilnonaphthalene-8-sulphonic acid and lack extensive β-sheet structure.Aromatic interactions modulate, but are not required for toxicity.

View Article: PubMed Central - PubMed

Affiliation: Diabetes Research Program, Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, New York University School of Medicine, New York, United States.

ABSTRACT
Islet amyloidosis by IAPP contributes to pancreatic β-cell death in diabetes, but the nature of toxic IAPP species remains elusive. Using concurrent time-resolved biophysical and biological measurements, we define the toxic species produced during IAPP amyloid formation and link their properties to induction of rat INS-1 β-cell and murine islet toxicity. These globally flexible, low order oligomers upregulate pro-inflammatory markers and induce reactive oxygen species. They do not bind 1-anilnonaphthalene-8-sulphonic acid and lack extensive β-sheet structure. Aromatic interactions modulate, but are not required for toxicity. Not all IAPP oligomers are toxic; toxicity depends on their partially structured conformational states. Some anti-amyloid agents paradoxically prolong cytotoxicity by prolonging the lifetime of the toxic species. The data highlight the distinguishing properties of toxic IAPP oligomers and the common features that they share with toxic species reported for other amyloidogenic polypeptides, providing information for rational drug design to treat IAPP induced β-cell death.

No MeSH data available.


Related in: MedlinePlus