Human Amylin Research Articles

Beta Amyloid-Induced Depression of Hippocampal Long-Term Potentiation Is Mediated through the Amylin Receptor

Alzheimer's disease (AD) is characterized by accumulation of amyloid-β peptide (Aβ) in the brain regions that subserve memory and cognition. The amylin receptor is a potential target receptor for expression of the deleterious actions of soluble oligomeric Aβ species. We investigated whether the amylin receptor antagonist, AC253, neutralizes the depressant effects of Aβ(1-42) and human amylin on hippocampal long-term potentiation (LTP). Furthermore, we examined whether depressed levels of LTP observed in transgenic mice, which overexpress amyloid precursor protein (TgCRND8), could be restored with AC253. In mouse hippocampal brain slices, field EPSPs were recorded from the stratum radiatum layer of the CA1 area (cornu ammonis 1 region of the hippocampus) in response to electrical stimulation of Schaeffer collateral afferents. LTP was induced by 3-theta burst stimulation protocols. Aβ(1-42) (50 nM) and human amylin (50 nM), but not Aβ(42-1) (50 nM), depressed LTP evoked using both stimulation protocols. Preapplication of AC253 (250 nM) blocked Aβ- and human amylin-induced reduction of LTP without affecting baseline transmission or LTP on its own. In contrast to wild-type controls, where robust LTP is observed, 6- to 12-month-old TgCRND8 mice show blunted LTP that is significantly enhanced by application of AC253. Our data demonstrate that the effects of Aβ(1-42) and human amylin on LTP are expressed via the amylin receptor, and moreover, blockade of this receptor increases LTP in transgenic mice that show increased brain amyloid burden. Amylin receptor antagonists could serve as potentially useful therapeutic agents in AD.

A Single Pseudoproline and Microwave Solid Phase Peptide Synthesis Facilitates an Efficient Synthesis of Human Amylin 1–37

A chemical synthesis of the 37 residue polypeptide human amylin using microwave enhanced solid phase peptide chemistry is described. An optimised protocol used only a single pseudoproline derivative, a chemically pure resin and single couplings of all amino acids to deliver non-oxidised amylin in high yield. Oxidation of the crude peptide to form the disulfide bond was accomplished in 20 min using 2,2′-dipyridyl disulfide in dimethyl sulphoxide giving human amylin that was fully functional in a cAMP assay.

The role of two natural flavonoids on human amylin aggregation

Misfolded amylin, the human polypeptide hormone, forms amyloid deposits in pancreatic islets. These amyloid deposits contribute to the dysfunction of beta-cells and the loss of beta-cell mass in type 2 diabetes mellitus (T2DM). Inhibition of amylin fibrillization has been regarded as a potential therapeutic approach for T2DM. Using i¬‚uorescence spectroscopic analysis with thioi¬‚avin T, the role of two naturally occurring flavonoids named myricetin and epigallocatechin gallate on human amylin hormone fibrillization and destabilization of fibrillar aggregates were examined under near physiological conditions. The results showed that after 168 h incubation by shaker incubator in 37°C. Myricetin at 10 and 40 µM repressed amylin amyloid formation by 25.3 and 22.4%, respectively (p<0.05), and the similar values of epigallocatechin gallate inhibited the formation of β-sheet structure by 18.1 and 16.7%, respectively (p<0.05). The obtained data also confirmed that amyloidal sheet opening was induced by myricetin and epigallocatechin gallate significantly (p<0.05). Therefore, it was concluded that islet amyloid cytotoxicity to β-cells may be reduced by these two flavonoids, and these compounds should be key molecules for the development of the therapeutics for diabetes mellitus. Key words: Human islet amyloid polypeptide, hyperglycemia, myricetin, epigallocatechin gallate, flavonoids.

Fibril formation and toxicity of the non-amyloidogenic rat amylin peptide

Full-length native rat amylin 1–37 has previously been widely shown to be unable to form fibrils and to lack the toxicity of the human amylin form leading to its use as a non-amyloidogenic control peptide. A recent study has suggested that rat amylin 1–37 forms amyloidogenic β-sheet structures in the presence of the human amylin form and suggested that this property could promote toxicity. Using TEM analysis we show here fibril formation by synthetic rat amylin 1–37 and 8–37 peptides when the lyophilized HPLC purified peptides are initially dissolved in 20mM Tris–HCl. Dissolution of synthetic rat amylin 1–37 and 8–37 peptides in H2O or phosphate buffered saline failed to produce fibrils. Addition of 20mM Tris–HCl to synthetic rat amylin 1–37 and 8–37 peptides initially dissolved in H2O also failed to induce fibril formation. The rat amylin fibrils have a uniform structure and bind Congo red suggesting that they are amyloid fibrils. The rat amylin fibrils also bind catalase, which could be inhibited by Amyloid-β 31–35 and a catalase amyloid-β binding domain-like peptide (R9). The rat amylin 1–37 and 8–37 fibrils are toxic in both human pancreatic islet and neuronal cell culture systems. The toxicity of rat amylin fibrils can be inhibited by an amylin receptor antagonist (AC187) and a caspase inhibitor (zVAD-fmk) in a similar manner to previous observations for human amylin toxicity. Chemically induced rat amylin fibril formation of uniform structured fibrils provides a potentially novel anti-amyloid drug discovery tool.

Amyloid β (Aβ) Peptide Directly Activates Amylin-3 Receptor Subtype by Triggering Multiple Intracellular Signaling Pathways

The two age-prevalent diseases Alzheimer disease and type 2 diabetes mellitus share many common features including the deposition of amyloidogenic proteins, amyloid β protein (Aβ) and amylin (islet amyloid polypeptide), respectively. Recent evidence suggests that both Aβ and amylin may express their effects through the amylin receptor, although the precise mechanisms for this interaction at a cellular level are unknown. Here, we studied this by generating HEK293 cells with stable expression of an isoform of the amylin receptor family, amylin receptor-3 (AMY3). Aβ1-42 and human amylin (hAmylin) increase cytosolic cAMP and Ca(2+), trigger multiple pathways involving the signal transduction mediators protein kinase A, MAPK, Akt, and cFos. Aβ1-42 and hAmylin also induce cell death during exposure for 24-48 h at low micromolar concentrations. In the presence of hAmylin, Aβ1-42 effects on HEK293-AMY3-expressing cells are occluded, suggesting a shared mechanism of action between the two peptides. Amylin receptor antagonist AC253 blocks increases in intracellular Ca(2+), activation of protein kinase A, MAPK, Akt, cFos, and cell death, which occur upon AMY3 activation with hAmylin, Aβ1-42, or their co-application. Our data suggest that AMY3 plays an important role by serving as a receptor target for actions Aβ and thus may represent a novel therapeutic target for development of compounds to treat neurodegenerative conditions such as Alzheimer disease.

Effect of two herbal polyphenol compounds on human amylin amyloid formation and destabilization

Human Islet amyloid polypeptide gathering in the β-cell of the pancreas has been implicated in the pathology of type 2 diabetes. Inhibition of the formation of amylin fibrils could be therapeutic aims for the treatment of type 2 diabetes mellitus. In this study, the fluorimetric assay was used to examine the role of two herbal polyphenol compounds including Rosmarinic acid and Curcumin on amyloid formation and destabilization of human amylin amyloid under near-physiological circumstances. The results showed that after 192 h incubation by shaker incubator at 37°C, Rosmarinic acid at 10 and 40 µM repressed amylin amyloid formation by 18.6 and 19.3% respectively (P<0.05), and the similar values of Curcumin inhibited the formation of β-sheet structure by 17.2 and 29.9%, respectively (P<0.05). The obtained data also confirmed that amyloidal sheet opening was induced by Rosmarinic acid and Curcumin significantly (P<0.05). It was concluded that islet amyloid cytotoxicity to β-cells may be reduced by these two herbal extracts and these compounds should be key molecules for the development of the therapeutics for diabetes mellitus. Key words: Amylin, diabetes, rosmarinic acid, curcumin, amyloid.

Two-dimensional infrared spectroscopy reveals the complex behaviour of an amyloid fibril inhibitor

While amyloid formation has been implicated in the pathology of over twenty human diseases, the rational design of amyloid inhibitors is hampered by a lack of structural information about amyloid-inhibitor complexes. We use isotope labeling and two-dimensional infrared spectroscopy to obtain a residue-specific structure for the complex of human amylin, the peptide responsible for islet amyloid formation in type 2 diabetes, with a known inhibitor, rat amylin. Based on its sequence, rat amylin should block formation of the C-terminal β-sheet, but at 8 hours after mixing rat amylin blocks the N-terminal β-sheet instead. At 24 hours after mixing, rat amylin blocks neither β-sheet and forms its own β-sheet most likely on the outside of the human fibrils. This is striking because rat amylin is natively disordered and not previously known to form amyloid β-sheets. The results show that even seemingly intuitive inhibitors may function by unforeseen and complex structural processes.

Hyperamylinemia Contributes to Cardiac Dysfunction in Obesity and Diabetes

Rationale: Hyperamylinemia is common in patients with obesity and insulin resistance, coincides with hyperinsulinemia, and results in amyloid deposition. Amylin amyloids are generally considered a pancreatic disorder in type 2 diabetes. However, elevated circulating levels of amylin may also lead to amylin accumulation and proteotoxicity in peripheral organs, including the heart. Objective: To test whether amylin accumulates in the heart of obese and type 2 diabetic patients and to uncover the effects of amylin accumulation on cardiac morphology and function. Methods and Results: We compared amylin deposition in failing and nonfailing hearts from lean, obese, and type 2 diabetic humans using immunohistochemistry and Western blots. We found significant accumulation of large amylin oligomers, fibrils, and plaques in failing hearts from obese and diabetic patients but not in normal hearts and failing hearts from lean, nondiabetic humans. Small amylin oligomers were even elevated in nonfailing hearts from overweight/obese patients, suggesting an early state of accumulation. Using a rat model of hyperamylinemia transgenic for human amylin, we observed that amylin oligomers attach to the sarcolemma, leading to myocyte Ca 2+ dysregulation, pathological myocyte remodeling, and diastolic dysfunction, starting from prediabetes. In contrast, prediabetic rats expressing the same level of wild-type rat amylin, a nonamyloidogenic isoform, exhibited normal heart structure and function. Conclusions: Hyperamylinemia promotes amylin deposition in the heart, causing alterations of cardiac myocyte structure and function. We propose that detection and disruption of cardiac amylin buildup may be both a predictor of heart dysfunction and a novel therapeutic strategy in diabetic cardiomyopathy.

Effect of Sequence Variation on the Mechanical Response of Amyloid Fibrils Probed by Steered Molecular Dynamics Simulation

The mechanical failure of mature amyloid fibers produces fragments that act as seeds for the growth of new fibrils. Fragmentation may also be correlated with cytotoxicity. We have used steered atomistic molecular dynamics simulations to study the mechanical failure of fibrils formed by the amyloidogenic fragment of human amylin hIAPP20-29 subjected to force applied in a variety of directions. By introducing systematic variations to this peptide sequence in silico, we have also investigated the role of the amino-acid sequence in determining the mechanical stability of amyloid fibrils. Our calculations show that the force required to induce mechanical failure depends on the direction of the applied stress and upon the degree of structural order present in the β-sheet assemblies, which in turn depends on the peptide sequence. The results have implications for the importance of sequence-dependent mechanical properties on seeding the growth of new fibrils and the role of breakage events in cytotoxicity.

Polymeric particles for the controlled release of human amylin

Since its discovery the therapeutic use of the pancreatic hormone amylin has been limited due to its poor water solubility and propensity for amyloid aggregation. We have entrapped the human amylin protein in polymeric nanoparticles, using a single emulsion–solvent evaporation method and investigated its effectiveness in the controlled release of the peptide. Typical preparations composed of poly-ɛ-caprolactone had a mean particle size of approximately 200nm, low polydispersity index, high protein entrapment efficiency (80%) and process yield (90%), and spherical and smooth surfaces. These nanoparticles presented a controlled release in vitro for approximately 240h. Pharmacological evaluation in vivo by subcutaneous administration in fasting mice demonstrated the bioactivity and effectiveness of the released human amylin, resulting in reduced glycemia lasting for at least 36h. These features indicate the potential for the use of a confined particulate system in the therapeutic controlled and sustained release of human amylin.

Amyloid-Derived Peptide Forms Self-Assembled Monolayers on Gold Nanoparticle with a Curvature-Dependent β-Sheet Structure

Using a combination of Fourier transform infrared (FTIR) spectroscopy and solid-state nuclear magnetic resonance (SSNMR) techniques, the secondary structure of peptides anchored on gold nanoparticles of different sizes is investigated. The structure of the well-studied CALNN-capped nanoparticles is compared to the structure of nanoparticles capped with a new cysteine-terminated peptide, CFGAILSS. The design of that peptide is derived from the minimal amyloidogenic sequence FGAIL of the human islet polypeptide amylin. We demonstrate that CFGAILSS forms extended fibrils in solution. When constrained at a nanoparticle surface, CFGAILSS adopts a secondary structure markedly different from CALNN. Taking into account the surface selection rules, the FTIR spectra of CFGAILSS-capped gold nanoparticles indicate the formation of β-sheets which are more prominent for 25 nm diameter nanoparticles than for 5 nm nanoparticles. No intermolecular (13)C-(13)C dipolar coupling is detected with rotational resonance SSNMR for CALNN-capped nanoparticles, while CALNN is in a random coil configuration. Coupling is detected for CFGAILSS-capped gold nanoparticles, however, consistent with an intermolecular (13)C-(13)C distance of 5.0 ± 0.3 Å, in agreement with intermolecular hydrogen bonding in a parallel β-sheet structure.

Circulating Amylin Amyloid Oligomers Accumulate in the Heart and Induce Cardiomyocyte Dysfunction in a Rat Model of Type-2 Diabetes

Amylin amyloid deposition is generally considered a pancreatic disorder and a hallmark of type-2 diabetes. Recently, amylin amyloids were also found in failing hearts from obese and type-2 diabetic patients, suggesting a possible contribution to cardiac dysfunction. Here, we investigate the mechanism of cardiac amylin accumulation and consequent effects on myocyte structure and function in rats transgenic for human amylin. In this model, we observed that soluble amylin oligomers are released from the pancreas in the blood and accumulate in the heart. Amylin oligomers attach to the sarcolemma and raises the intracellular Ca2+ ([Ca2+]i) leading to myocyte dysfunction. In contrast, rats expressing same level of wild-type, non-amyloidogenic rat amylin showed normal cardiac myocyte structure and function. To test whether the rise of [Ca2+]i is an amylin oligomer-mediated effect, we measured Ca2+ transients in intact cardiac myocytes incubated with exogenous human/rat amylin. 50 μM of human amylin, which rapidly forms oligomers, increased substantially the amplitude of cardiac myocyte Ca2+ transients, while same concentration of rat amylin had no significant effects. Passive trans-sarcolemmal Ca2+ leak was substantially larger in myocytes incubated with human amylin vs. control, implying that amylin oligomers alter the structural integrity of the sarcolemma and increase sarcolemmal permeability to Ca2+. In conclusion, our data show that amylin oligomers circulate through the blood, accumulate in the heart, and alter cardiac myocyte function by disrupting Ca2+ homeostasis. The results suggest that circulating amylin oligomers should be targeted for pharmacological interventions to prevent heart dysfunction in patients with obesity and insulin resistance.

Self-assembly of human amylin-derived peptides studied by atomic force microscopy and single molecule force spectroscopy

The self-assembly of peptides and proteins into amyloid fibrils of nanometric thickness and up to several micrometres in length, a phenomenon widely observed in biological systems, has recently aroused a growing interest in nanotechnology and nanomedicine. Here we have applied atomic force microscopy and single molecule force spectroscopy to study the amyloidogenesis of a peptide derived from human amylin and of its reverse sequence. The spontaneous formation of protofibrils and their orientation along well-defined directions on graphite and DMSO-coated graphite substrates make the studied peptides interesting candidates for nanotechnological applications. The measured binding forces between peptides correlate with the number of hydrogen bonds between individual peptides inside the fibril structure according to molecular dynamics simulations.

Inhibition of the Mitochondrial Enzyme ABAD Restores the Amyloid-β-Mediated Deregulation of Estradiol

Alzheimer's disease (AD) is a conformational disease that is characterized by amyloid-β (Aβ) deposition in the brain. Aβ exerts its toxicity in part by receptor-mediated interactions that cause down-stream protein misfolding and aggregation, as well as mitochondrial dysfunction. Recent reports indicate that Aβ may also interact directly with intracellular proteins such as the mitochondrial enzyme ABAD (Aβ binding alcohol dehydrogenase) in executing its toxic effects. Mitochondrial dysfunction occurs early in AD, and Aβ's toxicity is in part mediated by inhibition of ABAD as shown previously with an ABAD decoy peptide. Here, we employed AG18051, a novel small ABAD-specific compound inhibitor, to investigate the role of ABAD in Aβ toxicity. Using SH-SY5Y neuroblastoma cells, we found that AG18051 partially blocked the Aβ-ABAD interaction in a pull-down assay while it also prevented the Aβ42-induced down-regulation of ABAD activity, as measured by levels of estradiol, a known hormone and product of ABAD activity. Furthermore, AG18051 is protective against Aβ42 toxicity, as measured by LDH release and MTT absorbance. Specifically, AG18051 reduced Aβ42-induced impairment of mitochondrial respiration and oxidative stress as shown by reduced ROS (reactive oxygen species) levels. Guided by our previous finding of shared aspects of the toxicity of Aβ and human amylin (HA), with the latter forming aggregates in Type 2 diabetes mellitus (T2DM) pancreas, we determined whether AG18051 would also confer protection from HA toxicity. We found that the inhibitor conferred only partial protection from HA toxicity indicating distinct pathomechanisms of the two amyloidogenic agents. Taken together, our results present the inhibition of ABAD by compounds such as AG18051 as a promising therapeutic strategy for the prevention and treatment of AD, and suggest levels of estradiol as a suitable read-out.

To determine the possible roles of two essential trace elements and ascorbic acid concerning amyloidal beta-sheet formation in diabetes mellitus

Amylin is a peptide hormone that is made and co-secreted along with insulin. Human amylin is the main component of amyloid beta-sheet found in the pancreas of majority of diabetic patients. Amyloidogenesis causes destruction of pancreatic β-cells. The subsequent lack of insulin leads to increased blood and urine glucose.In this article, the fluorimetric assay was used to examine the role of ascorbic acid and two essential trace elements including zinc and iron on beta-amyloid formation of human peptide of amylin hormone under near-physiological circumstances.Results obtained from in vitro study showed that after 120 h incubation by shaker incubator at 37°C, zinc element at 10 μM inhibited amylin 10 μM from amyloid fibril formation by 9.1% (p<0.05) while the similar value of iron element promoted the formation of β-sheet structure by 13.1% (p<0.05). The obtained data also demonstrated that ascorbic acid with concentration of 150 μM had inhibitory effects on formation of beta-amyloid sheet significantly (p<0.05). It may be concluded that if islet amyloid is cytotoxic to β-cells then zinc and ascorbic acid should protect these cells against degeneration in diabetic patients. Key words: Vitamin C, diabetes mellitus, amylin hormone, zinc, iron.

A simple and rapid method for identifying and semi‐quantifying peptide hormones in isolated pancreatic islets by direct‐tissue matrix‐assisted laser desorption ionization time‐of‐flight mass spectrometry

We describe a new, simple, robust and efficient method based on direct-tissue matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry that enables consistent semi-quantitation of peptide hormones in isolated pancreatic islets from normal and diabetic rodents. Prominent signals were measured that corresponded to all the main peptide hormones present in islet-endocrine cells: (α-cells) glucagon, glicentin-related polypeptide/GRPP; (β-cells) insulin I, insulin II, C-peptide I, C-peptide II, amylin; (δ-cells) somatostatin-14; and (PP-cells), and pancreatic polypeptide. The signal ratios coincided with known relative hormone abundances. The method demonstrated that severe insulin deficiency is accompanied by elevated levels of all non-β-cell-hormones in diabetic rat islets, consistent with alleviation of paracrine suppression of hormone production by non-β-cells. It was also effective in characterizing hormonal phenotype in hemizygous human-amylin transgenic mice that express human and mouse amylin in approx. equimolar quantities. Finally, the method demonstrated utility in basic peptide-hormone discovery by identifying a prominent new Gcg-gene-derived peptide (theoretical monoisotopic molecular weight 3263.5 Da), closely related to but distinct from GRPP, in diabetic islets. This peptide, whose sequence is HAPQDTEENARSFPASQTEPLEDPNQINE in Rattus norvegicus, could be a peptide hormone whose roles in physiology and metabolic disease warrant further investigation. This method provides a powerful new approach that could provide important new insights into the physiology and regulation of peptide hormones in islets and other endocrine tissues. It has potentially wide-ranging applications that encompass endocrinology, pharmacology, phenotypic analysis in genetic models of metabolic disease, and hormone discovery, and could also effectively limit the numbers of animals required for such studies.

Clustering and Internalization of Toxic Amylin Oligomers in Pancreatic Cells Require Plasma Membrane Cholesterol

Self-assembly of the human pancreatic hormone amylin into toxic oligomers and aggregates is linked to dysfunction of islet β-cells and pathogenesis of type 2 diabetes mellitus. Recent evidence suggests that cholesterol, an essential component of eukaryotic cells membranes, controls amylin aggregation on model membranes. However, the pathophysiological consequence of cholesterol-regulated amylin polymerization on membranes and biochemical mechanisms that protect β-cells from amylin toxicity are poorly understood. Here, we report that plasma membrane (PM) cholesterol plays a key role in molecular recognition, sorting, and internalization of toxic amylin oligomers but not monomers in pancreatic rat insulinoma and human islet cells. Depletion of PM cholesterol or the disruption of the cytoskeleton network inhibits internalization of amylin oligomers, which in turn enhances extracellular oligomer accumulation and potentiates amylin toxicity. Confocal microscopy reveals an increased nucleation of amylin oligomers across the plasma membrane in cholesterol-depleted cells, with a 2-fold increase in cell surface coverage and a 3-fold increase in their number on the PM. Biochemical studies confirm accumulation of amylin oligomers in the medium after depletion of PM cholesterol. Replenishment of PM cholesterol from intracellular cholesterol stores or by the addition of water-soluble cholesterol restores amylin oligomer clustering at the PM and internalization, which consequently diminishes cell surface coverage and toxicity of amylin oligomers. In contrast to oligomers, amylin monomers followed clathrin-dependent endocytosis, which is not sensitive to cholesterol depletion. Our studies identify an actin-mediated and cholesterol-dependent mechanism for selective uptake and clearance of amylin oligomers, impairment of which greatly potentiates amylin toxicity.

β-Amyloid protein (Aβ) and human amylin regulation of apoptotic genes occurs through the amylin receptor

Deposition of amyloid-beta (Aβ) protein, a 39-43 amino acid peptide, in the brain is a major pathological feature of Alzheimer's disease (AD). We have previously provided evidence that in primary cultures of rat basal forebrain and human fetal neurons (HFNs), neurotoxic effects of oligomeric Aβ are expressed through the amylin receptor. In this study, we utilized RT-PCR arrays to compare RNA expression levels of 84 markers for pro and anti- apoptotic signalling pathways following exposure of HFNs to either Aβ(1-42) (20μM) or human amylin (2μM). Oligomeric Aβ(1-42) or human amylin was applied to HFNs alone or after pre-treatment of cultures with the amylin receptor antagonist, AC253. Changes in RNA levels were then quantified and compared to each other in order to identify increases or decreases in gene expression of apoptotic markers. Applications of Aβ(1-42) or human amylin, but not the inactive inverse sequence Aβ(42-1) or rat amylin, resulted in a time-dependent marked increase in mediators of apoptosis including a 10- to 30-fold elevations in caspases 3, 6, 9, BID and XIAP levels. Amylin receptor antagonists, AC253 (10μM) or AC187 (10μM), significantly attenuated the induction of several pro-apoptotic mediators up-regulated following exposure to Aβ(1-42) or human amylin and increased the expression of several anti-apoptotic markers. These data allow us to identify key elements in the Aβ-induced apoptosis that are blocked by antagonism of the amylin receptor and further support the potential for amylin receptor blockade as a potential therapeutic avenue in AD.

2DIR spectroscopy of human amylin fibrils reflects stable β-sheet structure.

The aggregation of human amylin to form amyloid contributes to islet β-cell dysfunction in type 2 diabetes. Studies of amyloid formation have been hindered by the low structural resolution or relatively modest time resolution of standard methods. Two-dimensional infrared (2DIR) spectroscopy, with its sensitivity to protein secondary structures and its intrinsic fast time resolution, is capable of capturing structural changes during the aggregation process. Moreover, isotope labeling enables the measurement of residue-specific information. The diagonal line widths of 2DIR spectra contain information about dynamics and structural heterogeneity of the system. We illustrate the power of a combined atomistic molecular dynamics simulation and theoretical and experimental 2DIR approach by analyzing the variation in diagonal line widths of individual amide I modes in a series of labeled samples of amylin amyloid fibrils. The theoretical and experimental 2DIR line widths suggest a "W" pattern, as a function of residue number. We show that large line widths result from substantial structural disorder and that this pattern is indicative of the stable secondary structure of the two β-sheet regions. This work provides a protocol for bridging MD simulation and 2DIR experiments for future aggregation studies.

Determining the effects of two essential trace elements on polymerization of human amylin hormone

Determining the effects of two essential trace elements on polymerization of human amylin hormone