The structural basis of pancreatic amyloid formation: isotope-edited spectroscopy in the solid state

Ashburn, Ted T.; Auger, Michèle; Lansbury, Peter T.

doi:10.1021/ja00028a073

Cited by 72 publications

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“…In fact, a parallel arrangement might be much more common for proteins/peptides with longer core regions, such as IAPP, A␤ (23), ␣-synuclein (22), tau (21), and as recently suggested for the yeast prion Ure2p (40). For shorter segments and peptide fragments of A␤ (41)(42)(43) and IAPP (18,44), however, anti-parallel ␤-sheet arrangements have been reported. Recently, the role of amphipathicity in determining the parallel/ anti-parallel organization of amyloid fibrils has been described (45).…”

Section: Discussionmentioning

confidence: 93%

Identifying Structural Features of Fibrillar Islet Amyloid Polypeptide Using Site-directed Spin Labeling

Jayasinghe

Langen

2004

Journal of Biological Chemistry

144

151

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Pancreatic amyloid deposits, composed primarily of the 37-residue islet amyloid polypeptide (IAPP), are a characteristic feature found in more than 90% of patients with type II diabetes. Although IAPP amyloid deposits are associated with areas of pancreatic islet ␤-cell dysfunction and depletion and are thought to play a role in disease, their structure is unknown. We used electron paramagnetic resonance spectroscopy to analyze eight spin-labeled derivatives of IAPP in an effort to determine structural features of the peptide. In solution, all eight derivatives gave rise to electron paramagnetic resonance spectra with sharp lines indicative of rapid motion on the sub-nanosecond time scale. These spectra are consistent with a rapidly tumbling and highly dynamic peptide. In contrast, spectra for the fibrillar form exhibit reduced mobility and the presence of strong intermolecular spin-spin interactions. The latter implies that the peptide subunits are ordered and that the same residues from neighboring peptides are in close proximity to one another. Our data are consistent with a parallel arrangement of IAPP peptides within the amyloid fibril. Analysis of spin label mobility indicates a high degree of order throughout the peptide, although the N-terminal region is slightly less ordered. Possible similarities with respect to the domain organization and parallelism of Alzheimer's amyloid ␤ peptide fibrils are discussed.Pancreatic amyloid deposits have been identified as a hallmark of non-insulin-dependent (type II) diabetes mellitus. More than 90% of patients with type II diabetes exhibit amyloid deposits in areas of ␤-cell dysfunction and death (1, 2). The major component of these deposits is the 37-residue islet amyloid polypeptide (IAPP) 1 (3,4), and multiple lines of investigation have provided experimental evidence implicating human IAPP amyloid deposits in type II diabetes (5-8).Although pancreatic amyloid deposits in a diabetic patient were first identified more than a century ago, structural features of these deposits were not described until recently. Electron microscopy (EM) studies of IAPP deposits revealed the presence of long fibrillar structures (9 -11). Fourier transform infrared and circular dichroism (CD) analysis of IAPP fibrils made from full-length as well as shorter fragments indicates the presence of significant amounts of ␤-sheet structure in the fibrillar form (12, 13). X-ray and electron diffraction studies using aligned fibrils have shown that the peptide chains are arranged in a cross-␤-conformation with the individual ␤-strands perpendicular to the fibril axis (14,15). Beyond these observations, little is known about the molecular details of IAPP in the fibrillar form.Amyloid deposits have been identified in a number of human diseases, such as Alzheimer's disease, Parkinson's disease, and familial amyloidotic polyneuropathy (16). Although the primary structure of amyloid forming proteins varies widely, most amyloid deposits appear to share common characteristics, such as the cross-␤-str...

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Section: Discussionmentioning

confidence: 93%

Identifying Structural Features of Fibrillar Islet Amyloid Polypeptide Using Site-directed Spin Labeling

Jayasinghe

Langen

2004

Journal of Biological Chemistry

144

151

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“…2B). Two macrocycles have sequences that overlap the N-terminal β-sheet of the fibrils (Mac [11][12][13][14][15][16][17] and Mac [15][16][17][18][19][20][21] ), two overlap the C-terminal sheet (Mac [26][27][28][29][30][31][32] and Mac [31][32][33][34][35][36][37], and one targets the FGAIL region under investigation (Mac [21][22][23][24][25][26][27] ). The sequences of the macrocycles are given in Fig.…”

Section: Resultsmentioning

confidence: 99%

“…3. Conformational free energy map for the dimerization of hIAPP in terms of order parameters Q [8][9][10][11][12][13][14][15][16] and Q [27][28][29][30][31][32][33][34][35] . Contour lines are drawn every 2.5 kJ/mol.…”

Section: Discussionmentioning

confidence: 99%

“…Early work revealed a correlation between the ability to form amyloid in vitro and the sequence in the 20-29 region (13,14). The fragment hIAPP [20][21][22][23][24][25][26][27][28][29] (SNNFGAILSS) readily forms amyloid fibrils, as do smaller fragments such as hIAPP [23][24][25][26][27] (13)(14)(15)(16)(17) and hIAPP [21][22][23][24][25][26][27] ; the latter has been characterized with X-ray crystallography (5). Molecular dynamics simulations corroborate the assembly of such fragments into β-sheet structures (18)(19)(20).…”

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confidence: 99%

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Mechanism of IAPP amyloid fibril formation involves an intermediate with a transient β-sheet

Buchanan

Dunkelberger

Tran

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

244

345

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Amyloid formation is implicated in more than 20 human diseases, yet the mechanism by which fibrils form is not well understood. We use 2D infrared spectroscopy and isotope labeling to monitor the kinetics of fibril formation by human islet amyloid polypeptide (hIAPP or amylin) that is associated with type 2 diabetes. We find that an oligomeric intermediate forms during the lag phase with parallel β-sheet structure in a region that is ultimately a partially disordered loop in the fibril. We confirm the presence of this intermediate, using a set of homologous macrocyclic peptides designed to recognize β-sheets. Mutations and molecular dynamics simulations indicate that the intermediate is on pathway. Disrupting the oligomeric β-sheet to form the partially disordered loop of the fibrils creates a free energy barrier that is the origin of the lag phase during aggregation. These results help rationalize a wide range of previous fragment and mutation studies including mutations in other species that prevent the formation of amyloid plaques.inhibitors | aggregation pathway | vibrational coupling T he misfolding of proteins into β-sheet-rich amyloid fibrils is associated with the pathology of more than 20 human diseases, including Alzheimer's, Parkinson, and Huntington diseases (1). Amyloid plaques are formed by masses of fibrils, but growing evidence suggests that the toxic species may be prefibrillar intermediates (2, 3). As a result, there is much interest in understanding the mechanism by which these proteins form fibrils and identifying intermediates in the aggregation pathway. However, obtaining structural information about intermediate species is difficult due to their transient nature. Solid-state NMR (ssNMR) and X-ray crystallography provide high-resolution structures of fibrils (4, 5) and optical techniques can track structural changes in real time (6, 7), but few techniques have both the structural and the temporal resolution to extract specific structural details about intermediates. Fragments have been trapped in intriguing oligomeric structures that may represent intermediate states (5,8) and transient secondary structures are known to exist from circular dichroism measurements and other experiments (9-11), but for full-length proteins it has been difficult to identify the specific residues that contribute to the secondary structure and thus understand their role in the aggregation mechanism. In this paper, we use 2D infrared (2D IR) spectroscopy and isotope labeling to monitor the structural evolution of the full-length human islet amyloid polypeptide (hIAPP or amylin), a 37-residue peptide implicated in type 2 diabetes. We observe the formation of a structured prefibrillar intermediate in a region that has long been known to influence aggregation, but that does not form well-ordered cross-β structure in the amyloid fibril. Its presence provides unique structural insights into the mechanism of amyloid aggregation and helps unify many seemingly inconsistent prior studies.Many studies on hIAPP have focused ...

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“…Recent transgenic models, e.g., the HIP rat, strongly support a role for IAPP in diabetic pathology (17). Residues 20-29 of IAPP, SNNFGAILSS, referred to here as IAPP [20][21][22][23][24][25][26][27][28][29] , have previously been shown to form amyloid independently of the rest of the sequence (18)(19)(20). Here, we study the kinetics of assembly of the cationic form of the peptide (amidated C terminus).…”

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confidence: 99%

Fiber-dependent amyloid formation as catalysis of an existing reaction pathway

Ruschak¹,

Miranker

2007

Proc. Natl. Acad. Sci. U.S.A.

203

251

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A central component of a number of degenerative diseases is the deposition of protein as amyloid fibers. Self-assembly of amyloid occurs by a nucleation-dependent mechanism that gives rise to a characteristic sigmoidal reaction profile. The abruptness of this transition is a variable characteristic of different proteins with implications to both chemical mechanism and the aggressiveness of disease. Because nucleation is defined as the rate-limiting step, we have sought to determine the nature of this step for a model system derived from islet amyloid polypeptide. We show that nucleation occurs by two pathways: a fiber-independent (primary) pathway and a fiber-dependent (secondary) pathway. We first show that the balance between primary and secondary contributions can be manipulated by an external interface. Specifically, in the presence of this interface, the primary mechanism dominates, whereas in its absence, the secondary mechanism dominates. Intriguingly, we determine that both the reaction order and the enthalpy of activation of the two nucleation processes are identical. We interrogate this coincidence by global analysis using a simplified model generally applicable to protein polymerization. A physically reasonable set of parameters can be found to satisfy the coincidence. We conclude that primary and secondary nucleation need not represent different processes for amyloid formation. Rather, they are alternative manifestations of the same, surfacecatalyzed nucleation event.amylin ͉ fibers ͉ islet amyloid polypeptide ͉ nucleation T he noncovalent, fibrous self-assembly of proteins, including hemoglobin S, actin, microtubules, and amyloid fibers, occurs by a nucleation-dependent mechanism (1-5). If polymer product formation is monitored as a function of time in a reaction where all of the protein is initially monodisperse, there is a lag phase in which little product is formed, followed by a period of rapid growth (2). Such observations are consistent with a rate-limiting step in which change occurs to a high-energy intermediate known as the nucleus. In many systems such as actin, collagen, and microtubules, the nucleus is modeled as an oligomeric species that is in a highly unfavorable equilibrium with monomer (1, 3-6). The nucleus may also be a high-energy conformation of monomer, such as that reported for polyglutamine (7). Regardless, the rate-limiting step involves a nucleus because it is defined as the species with the highest free energy and therefore lowest population (1, 4).This model of nucleation alone cannot account for the high apparent cooperativity of conversion observed in many systems. Historically, hemoglobin S was the first biological polymerization reaction to demonstrate a transition time that is much shorter than its preceding lag phase (8). This same phenomenon is commonly reported for amyloid systems including islet amyloid polypeptide (IAPP) from type II diabetes (9), A␤ from Alzheimer's (10), PrP from the mammalian prion (11), and Sup35 from the yeast prion (12, 13), as well as mode...

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The structural basis of pancreatic amyloid formation: isotope-edited spectroscopy in the solid state

Cited by 72 publications

References 0 publications

Identifying Structural Features of Fibrillar Islet Amyloid Polypeptide Using Site-directed Spin Labeling

Identifying Structural Features of Fibrillar Islet Amyloid Polypeptide Using Site-directed Spin Labeling

Mechanism of IAPP amyloid fibril formation involves an intermediate with a transient β-sheet

Fiber-dependent amyloid formation as catalysis of an existing reaction pathway

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