Types Of Fibrils Research Articles

Transmission electron microscopy characterization of fluorescently labelled amyloid β 1-40 and α-synuclein aggregates

BackgroundFluorescent tags, including small organic molecules and fluorescent proteins, enable the localization of protein molecules in biomedical research experiments. However, the use of these labels may interfere with the formation of larger-scale protein structures such as amyloid aggregates. Therefore, we investigate the effects of some commonly used fluorescent tags on the morphologies of fibrils grown from the Alzheimer's disease-associated peptide Amyloid β 1-40 (Aβ40) and the Parkinson's disease-associated protein α-synuclein (αS).ResultsUsing transmission electron microscopy (TEM), we verify that N-terminal labeling of Aβ40 with AMCA, TAMRA, and Hilyte-Fluor 488 tags does not prevent the formation of protofibrils and amyloid fibrils of various widths. We also measure the two-photon action cross-section of Aβ40 labelled with Hilyte Fluor 488 and demonstrate that this tag is suitable for use with two-photon fluorescence techniques. Similarly, we find that Alexa Fluor 488 labelling of αS variant proteins near either the N or C terminus (position 9 or 130) does not interfere with the formation of amyloid and other types of αS fibrils. We also present TEM images of fibrils grown from αS C-terminally labelled with enhanced green fluorescent protein (EGFP). Near neutral pH, two types of αS-EGFP fibrils are observed via TEM, while denaturation of the EGFP tag leads to the formation of additional species.ConclusionsWe demonstrate that several small extrinsic fluorescent tags are compatible with studies of amyloid protein aggregation. However, although fibrils can be grown from αS labelled with EGFP, the conformation of the fluorescent protein tag affects the observed aggregate morphologies. Thus, our results should assist researchers with label selection and optimization of solution conditions for aggregation studies involving fluorescence techniques.

Structural polymorphism in amyloids: new insights from studies with Y145Stop prion protein fibrils.

The C-terminally-truncated human prion protein variant Y145Stop (or PrP23-144), associated with a familial prion disease, provides a valuable model for studying the fundamental properties of protein amyloids. In previous solid-state NMR experiments, we established that the β-sheet core of the PrP23-144 amyloid is composed of two β-strand regions encompassing residues ∼113-125 and ∼130-140. The former segment contains a highly conserved hydrophobic palindrome sequence, (113)AGAAAAGA(120), which has been considered essential to PrP conformational conversion. Here, we examine the role of this segment in fibrillization of PrP23-144 using a deletion variant, Δ113-120 PrP23-144, in which the palindrome sequence is missing. Surprisingly, we find that deletion of the palindrome sequence affects neither the amyloidogenicity nor the polymerization kinetics of PrP23-144, although it does alter amyloid conformation and morphology. Using two-dimensional and three-dimensional solid-state NMR methods, we find that Δ113-120 PrP23-144 fibrils contain an altered β-core extended N-terminally to residue ∼106, encompassing residues not present in the core of wild-type PrP23-144 fibrils. The C-terminal β-strand of the core, however, is similar in both fibril types. Collectively, these data indicate that amyloid cores of PrP23-144 variants contain "essential" (i.e. nucleation-determining) and "nonessential" regions, with the latter being "movable" in amino acid sequence space. These findings reveal an intriguing new mechanism for structural polymorphism in amyloids and suggest a potential means for modulating the physicochemical properties of amyloid fibrils without compromising their polymerization characteristics.

Disulfide bonds reduce the toxicity of the amyloid fibrils formed by an extracellular protein.

Disulfide bonds reduce the toxicity of the amyloid fibrils formed by an extracellular protein.

Unusual structural transition of antimicrobial VP1 peptide

VP1 peptide, an active domain of m-calpain enzyme with antimicrobial activity is found to undergo an unusual conformational transition in trifluoroethanol (TFE) solvent. The nature of, and time dependent variations in, circular dichroism associated with the amide I vibrations, suggest that VP1 undergoes self-aggregation forming anti-parallel β-sheet structure in TFE. Transmission electron micrograph (TEM) images revealed that β-sheet aggregates formed by VP1 possess fibril-like assemblies.

Destruction of Amyloid Fibrils of Keratoepithelin Peptides by Laser Irradiation Coupled with Amyloid-specific Thioflavin T

Mutations in keratoepithelin are associated with blinding ocular diseases, including lattice corneal dystrophy type 1 and granular corneal dystrophy type 2. These diseases are characterized by deposits of amyloid fibrils and/or granular non-amyloid aggregates in the cornea. Removing the deposits in the cornea is important for treatment. Previously, we reported the destruction of amyloid fibrils of β(2)-microglobulin K3 fragments and amyloid β by laser irradiation coupled with the binding of an amyloid-specific thioflavin T. Here, we studied the effects of this combination on the amyloid fibrils of two 22-residue fragments of keratoepithelin. The direct observation of individual amyloid fibrils was performed in real time using total internal reflection fluorescence microscopy. Both types of amyloid fibrils were broken up by the laser irradiation, dependent on the laser power. The results suggest the laser-induced destruction of amyloid fibrils to be a useful strategy for the treatment of these corneal dystrophies.

Modulating α-synuclein misfolding and fibrillation in vitro by agrochemicals

Correspondence: Vladimir N Uversky Department of Molecular Medicine, University of South Florida, 12901 Bruce B Downs Blvd, MDC 7, Tampa, FL 33612-4799, USA Email vuversky@helth.usf.edu Abstract: A combination of spectroscopic techniques including atomic force microscopy (AFM) and transmission electron microscopy (TEM), was used to analyze the effect of chemically distinct agrochemicals (pesticides, herbicides, and fungicides) on the in vitro misfolding and aggregation of a presynaptic intrinsically disordered protein α-synuclein. Despite their differences in chemical properties, almost all the compounds screened affected the α-synuclein fibrillation in a concentration-dependent manner. The morphology of the aggregated α-synuclein was characterized by AFM and TEM techniques. In addition to typical fibrils abundantly found at the equilibrium phase, this analysis revealed the existence of a noticeable nonfibrillar fraction where α-synuclein was present as protofilaments, small oligomers, and large oligomeric species. The aggregated α-synuclein samples were separated into soluble and insoluble fractions by ultracentrifugation and subjected to structural and morphological characterization. Attenuated total reflectance Fourier transform infrared spectroscopic analysis showed that the insoluble α-synuclein fractions possessed a high content of ordered β-structure, whereas the β-structure content of the supernatant pool populated by oligomeric species was noticeably lower. This study provides evidence that chemically distinct agrochemicals can directly interact with α-synuclein to induce structural changes and affect the fibrillation process of this important protein.

Polymorphism of Amyloid Fibrils Formed by a Short Peptide from Yeast Prion Protein Sup35: AFM and Tip Enhanced Raman Scattering Study

Misfolding of prion protein and its subsequent aggregation is the cause of various prion related diseases. Prions are composed of aggregates of a misfolded prion protein. Although primary structure of prion protein is the same, its infectious form, amyloid fibrils, exist as multiple strains. The ability of prions to generate multiple strains poses an immediate health threat, which renders studies of this phenomenon very important. The strains are thought to represent structurally different prion protein molecules packed into amyloid fibrils. The biophysical properties of the fibrils, such as fragility, represent a major mechanism of prion amplification. Here, we demonstrate that variations in environmental conditions such as pH, salt concentration, temperature and mechanical stress (stirring) produces a variety of fibrillar polymorphs for a short peptide CGNNQQNY from yeast prion protein Sup35. The fibrils differ by their length and diameter as well as their ability to bundle together. We have used Tip-Enhanced Raman Scattering (TERS) in combination with AFM to study underlying conformational peculiarities of peptides within individual aggregates. Two types of fibrils were investigated, one formed in water and another one at pH 5.6. These conditions produce morphologically distinct fibrils in terms of their length and diameter. They also exhibited different kinetics of aggregation. The Raman spectra obtained with TERS also revealed the peptide conformational differences between these two types of fibrils. The observed differences are mostly manifested in the positions of the characteristic amide bands (I and III), suggesting that peptides in these two types of fibrils have different conformational states. This study demonstrates potentials of such a combined method as TERS/AFM for structural analysis of individual protein aggregates.

Detection of AA-type amyloid protein in labial salivary glands

Among the diverse forms of amyloidosis, secondary type is the most frequent one. Diagnosis of amyloid deposition is based on the identification of the fibrillary protein amyloid by means of Congo Red (CR) or crystal violet (CV) stains, but these techniques do not differentiate between the different types of amyloid fibrils. The aim of this study was to identify by immunofluorescence (IF) AA amyloid a pathological fibrillar low-molecular-weight protein formed by cleavage of serum amyloid A (SAA) protein in labial salivary gland (LSG) biopsies from patients with secondary amyloidosis. 98 LSG were studied, 65 were from patients with secondary amyloidosis and 33 from subjects with chronic inflammatory diseases without evidence of this anomaly. All sections were stained with hematoxylin and eosin (H &E), CV, CR and IF using anti-AA antibodies. Positive and negative controls were used for all techniques. CV and CR demonstrated that the amyloid substance was found mainly distributed periductally (93.8%), followed by periacinar and perivascular locations (p <0.001); however, the IF demonstrated that amyloid AA substance predominates in the periacinar area (73.8%), followed by periductal and perivascular locations (p <0.001). IF has a sensitivity of 83%, 100% of specificity, 100% of predictive positive value and 75% of predictive negative value. The results of this study confirm the efficacy of the LSG biopsy as a highly reliable method for diagnosis of secondary amyloidosis.

Intrinsic Linear Heterogeneity of Amyloid β Protein Fibrils Revealed by Higher Resolution Mass-per-length Determinations

Amyloid β proteins spontaneously form fibrils in vitro that vary in their thermodynamic stability and in morphological characteristics such as length, width, shape, longitudinal twist, and the number of component filaments. It is vitally important to determine which variant best represents the type of fibril that accumulates in Alzheimer disease. In the present study, the nature of morphological variation was examined by dark-field and transmission electron microscopy in a preparation of seeded amyloid β protein fibrils that formed at relatively low protein concentrations and exhibited remarkably high thermodynamic stability. The number of filaments comprising these fibrils changed frequently from two to six along their length, and these changes only became apparent when mass-per-length (MPL) determinations are made with sufficient resolution. The MPL results could be reproduced by a simple stochastic model with a single adjustable parameter. The presence of more than two primary filaments could not be discerned by transmission electron microscopy, and they had no apparent relationship to the longitudinal twist of the fibrils. However, the pitch of the twist was strongly affected by the pH of the negative stain. We conclude that highly stable amyloid fibrils may form in which a surprising amount of intrinsic linear heterogeneity may be obscured by MPL measurements of insufficient resolution, and by the negative stains used for imaging fibrils by electron microscopy.

Variation in amount of wild-type transthyretin in different fibril and tissue types in ATTR amyloidosis

Familial transthyretin (TTR) amyloidosis is caused by a mutation in the TTR gene, although wild-type (wt) TTR is also incorporated into the amyloid fibrils. Liver transplantation (LT) is the prevailing treatment of the disease and is performed in order to eliminate the mutant TTR from plasma. The outcome of the procedure is varied; especially problematic is a progressive cardiomyopathy seen in some patients, presumably caused by continued incorporation of wtTTR. What determines the discrepancy in outcome is not clear. We have previously shown that two structurally distinct amyloid fibrils (with or without fragmented ATTR) are found among ATTRV30M patients. In this study, we investigated the proportion of wtATTR in cardiac and adipose amyloid from patients having either fibril type. It was found that cardiac amyloid more easily incorporates wtTTR than adipose amyloid, offering a potential explanation for the vulnerability of cardiac tissue for continued amyloidosis after LT. In cardiac tissue, fibrils with fragmented ATTR contained a higher wt proportion than fibrils without, suggesting that continued incorporation of wtTTR after LT, perhaps, can take place more easily in these patients. In adipose tissue, a rapid increase in wt proportion after LT indicates that a rather fast turnover of the deposits must occur. A difference in wt proportion between the fibril types was seen post-LT but not pre-LT, possibly caused by differences in turnover rate. Conclusively, this study further establishes the basic dissimilarities between the two fibril types and demonstrates that their role in LT outcome needs to be further investigated.

The nature of amyloid-like glucagon fibrils.

Protein aggregation and formation of amyloid fibrils is a phenomenon usually associated with proteotoxicity and degenerative diseases, such as type 2 diabetes, Alzheimer's disease, and prion diseases. However, several protein and peptide hormones are known to have a high propensity to form amyloid-like fibrils in vitro raising concerns about safety and stability of pharmaceutical protein solutions. Comprehensive understanding of the aggregation mechanisms is an important prerequisite to the design of strategies to prevent fibril formation. Detailed kinetic, spectroscopic, and morphological studies have revealed that glucagon can form several types of fibrils that differ at the level of molecular packing of the peptide. Each type forms through distinct nucleation-dependent aggregation pathways influenced by solution conditions and can be self-propagated by seeding. An increasing number of functional amyloid-like structures have been discovered in nature, and it has recently been proposed that an amyloid-like state of glucagon may be utilized by the pancreatic α-cells as in vivo storage form. This article reviews the current state of our knowledge about the nature of the different types of amyloid-like glucagon fibrils, the mechanisms by which they form, and discusses implications for formulation strategies and the safety of glucagon pharmaceuticals.

Amyloid structure – one but not the same: the many levels of fibrillar polymorphism

Many proteins and peptides can form amyloid-like structures both in vivo and in vitro. Although strikingly similar fibrillar structures can be observed across a variety of amino acid sequences, the fibrils formed often exhibit a stunning wealth of polymorphisms at the level of electron or atomic force microscopy. This appears to violate the Anfinsen principle seen for globular proteins, where each protein sequence codes for just one well-defined fold. To a large extent, polymorphism reflects variable packing of a single protofilament structure in the mature fibrils. However, we and others have recently demonstrated that polymorphism can also reflect real structural differences in the molecular packing of the polypeptide chains leading to several possible protofilament structures and diverse mature fibrillar structures. Glucagon has been a particularly useful model system for studying the fibrillogenesis mechanisms that lead to the formation of structural polymorphism, thanks to its single tryptophan residue and the availability of large quantities at pharmaceutical-grade quality. Combinations of structural investigations and seed extension experiments have revealed the reproducible formation of at least five different self-propagating fibril types from subtle variations in growth conditions. These reflect the underlying complexity of the peptide conformational landscape and provide a link to natively disordered proteins, where structure is dictated by context in the form of different binding partners. Here we review some of the latest advances in the study of glucagon fibrillar polymorphism and their implications for mechanisms of fibril formation in general.

In search of an arrhythmic cause of coronary artery disease: the electrophysiologist's next frontier?

This editorial refers to ‘Cavotricuspid isthmus dependent flutter is associated with an increased incidence of occult coronary artery disease’ by J.P. de Bono et al., on page 1774 de Bono et al. 1 report the coronary angiogram results on 138 patients undergoing ablation for typical atrial flutter (AFL, n ¼ 37) or atrial fibrillation (AF, n ¼ 61), in comparison to a control group of patients undergoing ablation for supraventricular tachycardia (SVT) or idiopathic right ventricular outflow tract tachycardia (RVOT-VT) (n ¼ 40). The demographic data for these groups were statistically similar, and the groups were evenly matched for age and coronary disease risk factors. The authors observed that patients undergoing ablation for typical AFL had a significantly higher incidence of occult (i.e. asymptomatic) coronary artery disease (CAD, P ¼ 0.005) compared with those undergoing ablation for AF, SVT, or RVOT-VT. Specifically, 54% of the AFL patients had coronary atheroma, compared with 26% of the patients with AF and 21% of the patients with SVT or RVOT-VT. No difference was noted in the incidence of coronary atheroma in patients with AF compared with those with SVT or RVOT-VT (P ¼ 0.68). However, the majority of the CAD detected was mild non-obstructive plaque, 75% in AFL patients, 44% in AF patients, and 67% in patients with SVT or RVOT-VT. The authors conclude that there may be a mechanistic link between the right atrial substrate underlying typical AFL and that it may be responsible for a higher incidence of CAD in these patients, when compared with similar age- and CAD risk factormatched patients with AF, SVT, or RVOT-VT. What specific mechanistic link this might be, however, is not determined by this strictly observational report. The authors note that there is a close relationship between AF and AFL, with pulmonary vein ectopy commonly triggering both AF and AFL. 2 However, although not all patients with AFL have AF, most patients with AF have AFL at one time or another. 2 Thus, the authors note that patients with typical AFL probably have a substrate that ‘specifically allows initiation and maintenance of AF’. This has been previously shown to be true, with slower conduction velocity noted in the right atrium in the medial cavo-tricuspid isthmus (CTI) and inferior septum compared with other areas in the atrium, and slower conduction in the CTI in patients with AFL compared with those with SVT. 3,4 The mechanism responsible for this slower conduction velocity is not known 3,4 and has never been specifically studied. Thus, the findings of this study are intriguing, suggesting that underlying CAD could be a possible causative factor, leading to ischaemia and fibrosis of the right atrium. Unfortunately, it appears that the CAD in most patients with AFL in this study was very mild, and not likely to cause ischaemia. Therefore, an alternative explanation must be sought if this phenomenon is true, and not just a chance observation due to the small number of patients included in this study.

Formation of abnormally large-sized tubular amyloid β aggregates on a nanostructured gold surface

The effect of the properties of a nanostructured gold surface (nano-Au surface) on the aggregation of Amyloid β(1–40) (Aβ40) was investigated. A nano-Au surface, in the form of immobilized nanoparticles, was prepared by using a thermal evaporator, resulting in the formation of nanosized clusters with sizes less than 10 nm. When Aβ40 was incubated with the nano-Au surface, abnormally large-sized tubular aggregates were formed on the surface and typical fibril formation was suppressed in the solution. This abnormally large tubular structure represents a novel type of Aβ40 aggregate. In the absence of the nano-Au surface, the diameters of the Aβ40 fibrils were less than 10 nm. However, the height of the tubular aggregates formed on a nano-Au surface was 80–100 nm. Such large-sized aggregates of Aβ40 have not been reported in previous studies dealing with interactions of suspended nanoparticles with proteins. This can be attributed to differences in the aggregation mechanism between immobilized and suspended nanoparticles. The formation of Aβ40 aggregates by nano-Au surface will provide the possible mechanism for abnormal fibril formation.

Magic angle spinning NMR analysis of beta2-microglobulin amyloid fibrils in two distinct morphologies.

Beta(2)-microglobulin (beta(2)m) is the major structural component of amyloid fibrils deposited in a condition known as dialysis-related amyloidosis. Despite numerous studies that have elucidated important aspects of the fibril formation process in vitro, and a magic angle spinning (MAS) NMR study of the fibrils formed by a small peptide fragment, structural details of beta(2)m fibrils formed by the full-length 99-residue protein are largely unknown. Here, we present a site-specific MAS NMR analysis of fibrils formed by the full-length beta(2)m protein and compare spectra of fibrils prepared under two different conditions. Specifically, long straight (LS) fibrils are formed at pH 2.5, while a very different morphology denoted as worm-like (WL) fibrils is observed in preparations at pH 3.6. High-resolution MAS NMR spectra have allowed us to obtain (13)C and (15)N resonance assignments for 64 residues of beta(2)m in LS fibrils, including part of the highly mobile N-terminus. Approximately 25 residues did not yield observable signals. Chemical shift analysis of the sequentially assigned residues indicates that these fibrils contain an extensive beta-sheet core organized in a non-native manner, with a trans-P32 conformation. In contrast, WL fibrils exhibit more extensive dynamics and appear to have a smaller beta-sheet core than LS fibrils, although both cores seem to share some common elements. Our results suggest that the distinct macroscopic morphological features observed for the two types of fibrils result from variations in structure and dynamics at the molecular level.

Laser-induced Propagation and Destruction of Amyloid β Fibrils

The amyloid deposition of amyloid beta (Abeta) peptides is a critical pathological event in Alzheimer disease (AD). Preventing the formation of amyloid deposits and removing preformed fibrils in tissues are important therapeutic strategies against AD. Previously, we reported the destruction of amyloid fibrils of beta(2)-microglobulin K3 fragments by laser irradiation coupled with the binding of amyloid-specific thioflavin T. Here, we studied the effects of a laser beam on Abeta fibrils. As was the case for K3 fibrils, extensive irradiation destroyed the preformed Abeta fibrils. However, irradiation during spontaneous fibril formation resulted in only the partial destruction of growing fibrils and a subsequent explosive propagation of fibrils. The explosive propagation was caused by an increase in the number of active ends due to breakage. The results not only reveal a case of fragmentation-induced propagation of fibrils but also provide insights into therapeutic strategies for AD.

Molecular mechanism of Thioflavin-T binding to amyloid fibrils

Intense efforts to detect, diagnose, and analyze the kinetic and structural properties of amyloid fibrils have generated a powerful toolkit of amyloid-specific molecular probes. Since its first description in 1959, the fluorescent dye Thioflavin-T (ThT) has become among the most widely used "gold standards" for selectively staining and identifying amyloid fibrils both in vivo and in vitro. The large enhancement of its fluorescence emission upon binding to fibrils makes ThT a particularly powerful and convenient tool. Despite its widespread use in clinical and basic science applications, the molecular mechanism for the ability of ThT to recognize diverse types of amyloid fibrils and for the dye's characteristic fluorescence has only begun to be elucidated. Here, we review recent progress in the understanding of ThT-fibril interactions at an atomic resolution. These studies have yielded important insights into amyloid structures and the processes of fibril formation, and they also offer guidance for designing the next generation of amyloid assembly diagnostics, inhibitors, and therapeutics.

DMA analysis and wood bonding of PVAc latex reinforced with cellulose nanofibrils

Suspensions of commercial refined beech pulp (RBP) were further processed through mechanical disintegration (MD-RBP), chemical modification (CM-RBP) and through chemical modification followed by mechanical disintegration (CM-MD-RBP). Nanocomposites were prepared by compounding a poly(vinyl acetate) (PVAc) latex adhesive with increasing contents of the different types of nanofibrils, and the resulting nanocomposites were analyzed by dynamic mechanical analysis (DMA). Also, the suitability of using the CM-RBP fibrils to formulate PVAc adhesives for wood bonded assemblies with improved heat resistance was studied. The presence of cellulose nanofibrils had a strong influence on the viscoelastic properties of PVAc latex films. For all nanocomposites, increasing amounts of cellulose nanofibrils (treated or untreated) led to increasing reinforcing effects in the glassy state, but especially in the PVAc and PVOH glass transitions. This reinforcement primarily resulted from interactions between the cellulose fibrils network and the hydrophilic PVOH matrix that led to the complete disappearance of the PVOH glass transition (tan δ peak) for some fibril types and contents. At any given concentration in the PVOH transition, the CM-MD-RBP nanofibrils provided the highest reinforcement, followed by the MD-RBP, CM-RBP and the untreated RBP. Finally, the use of the CM-RBP fibrils to prepare PVAc reinforced adhesives for wood bonding was promising since, even though they generally performed worse in dry and wet conditions, the boards showed superior heat resistance (EN 14257) and passed the test for durability class D1.

Low Resolution Structure of a Membrane-Permeabilizing Oligomer of α-Synuclein: the Basis for a High-Throughput Screening of Compounds against α-Synuclein Aggregation

α-synuclein is a 140-residue natively unfolded protein, whose aggregation is implicated in the development of Parkinson's Disease. It is thought that the cytotoxic species is not the mature fibril, but rather a prefibrillar aggregate which has membrane-permeabilizing properties. We have used Small Angle X-ray Scattering (SAXS) to determine the low-resolution structures of the different species formed during α-synuclein fibrillation in a non-invasive fashion. In addition to the starting monomer-dimer equilibrium and two bona fide fibril types accumulating towards the end of the aggregation process, we have identified a wreath-shaped oligomeric state which has a very distinct central hole.

Thermodynamic Instability of A Self-Assembled 16-Residue Alanine-Based Oligopeptide in Aqueous Media: Hydrogel, Fibril, and Beaded Filament Formation

Oligoalanines with greater than ca. 14 residues, which have been doped with charged residues (e.g. lysine), usually adopt α-helical conformations in aqueous media.1 In contrast, Ac-(AAKA)4-NH2 aggregates instantaneously when dissolved in aqueous media at concentrations ranging from 70 μM to 7 mM. Evidence from UV circular dichroism (UV-CD) and FTIR spectroscopies suggest a mostly β-sheet like conformation. Kinetics studies of the initially formed conformations suggest a transition from a mostly β-like to a mostly PPII-like conformation, in contrast to typical fibril formation/growth studies. AFM images of freshly prepared samples indicate a porous, tissue-like architecture. Addition of salts (e.g. NaCl) stabilizes this architecture, resulting in the formation of a macroscopic hydrogel. β-sheet and hydrogel formation by such an amino acid sequence is quite unusual, as it does not obey typical rules required for peptide hydrogel formation. In particular, there are no alternating complimentary charges, nor alternating hydrophilic and hydrophobic residues. Without the added stabilization provided by the addition of salts, however, the peptide transitions from a mostly β-sheet-like structure to a PPII-like conformation, possibly via triple helix formation, as evidenced by both electronic and vibrational CD. AFM images suggest the formation of a heterogeneous mixture of assemblies, including bead-like filaments as well as networks of well-defined, intertwined fibrils. Potential applications for peptide hydrogels include drug delivery devices and tissue engineering scaffolds. In this regard, release studies of model peptides which have been trapped within the peptide hydrogel will be presented, as well as rheological characterization of the hydrogel as a function of salt and peptide concentration.[1] Scholtz, J. M.; Marqusee, S.; Baldwin, R. L.; York, E. J.; Stewart, J. M.; Santoro, M.; Bolen. D. W. Natl. Acad. Sci. U.S.A.1991, 88, 2854-2860.