• Bulletin of the Korean Chemical Society
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• v.33 no.3
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• pp.848-854
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• 2012

We have studied the oligomerization of a fibril-forming segment of ${\alpha}$-Synulcein using a replica exchange molecular dynamics (REMD) simulation. The simulation was performed with trimers and tetramers of a 12 amino acid residue stretch (residues 71-82) of ${\alpha}$-Synulcein. From extensive REMD simulations, we observed the spontaneous formation of both trimer and tetramer, demonstrating the self-aggregating and fibril-forming properties of the peptides. Secondary structure profile and clustering analysis illustrated that antiparallel ${\beta}$-sheet structures are major species corresponding to the global free energy minimum. As the size of the oligomer increases from a dimer to a tetramer, conformational stability is increased. We examined the evolution of simple order parameters and their free energy profiles to identify the process of aggregation. It was found that the degree of aggregation increased as time passed. Tetramer formation was slower than trimer formation and a transition in order parameters was observed, indicating the full development of tetramer conformation which is more stable than that of the trimer. The shape of free energy surface and change of order parameter distributions indicate that the oligomer formation follows a dock-and-lock process.

• Bulletin of the Korean Chemical Society
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• v.34 no.2
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• pp.665-668
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• 2013

• Bulletin of the Korean Chemical Society
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• v.30 no.8
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• pp.1845-1850
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• 2009

We have performed replica-exchange molecular dynamics (REMD) simulations on the dimer formation of fibrilforming segments of $\alpha$-Synuclein (residues 71 - 82) using implicit solvation models with two kinds of force fields- AMBER parm99SB and parm96. We observed spontaneous formation of dimers from the extensive simulations, demonstrating the self-aggregating and fibril forming properties of the peptides. Secondary structure profile and clustering analysis showed that dimers with antiparallel $\beta$-sheet conformations, stabilized by well-defined hydrogen boding, are major species corresponding to global free energy minimum. Parallel dimers with partial $\beta$-sheets are found to be off-pathway intermediates. The relative instability of the parallel arrangements is due to the repulsive interactions between bulky and polar side chains as well as weaker backbone hydrogen bonds.

• Molecular & Cellular Toxicology
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• v.4 no.2
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• pp.157-163
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• 2008

Formation of ${\beta}$-amyloid $(A{\beta})$ fibrils has been identified as one of the major characteristics of Alzheimer's disease (AD). Inhibition of $A{\beta}$ fibril formation in the CNS would be attractive therapeutic targets for the treatment of AD. Several small compounds that inhibit amyloid formation or amyloid neurotoxicity in vitro have been known. Citrate has surfactant function effect because of its molecular structure having high anionic charge density, in addition to the well-known antibacterial and antioxidant properties. Therefore, we hypothesized that citrate might have the inhibitory effect against $A{\beta}$ fibril formation in vitro and have the protective effect against $A{\beta}$-induced neurotoxicity in PC12 cells. We examined the effect of citrate against the formation of $A{\beta}$ fibrils by measuring the intensity of fluorescence in thioflavin-T (Th-T) assay of between $A{\beta}_{25-35}$ groups treated with citrate and the control with $A{\beta}_{25-35}$ alone. The neuroprotective effect of citrate against $A{\beta}$-induced toxicity in PC12 cells was investigated using the WST-1 assay. Fluorescence spectroscopy showed that citrate inhibited dose-dependently the formation of $A{\beta}$ fibrils from ${\beta}$-amyloid peptides. The inhibition percentages of $A{\beta}$ fibril formation by citrate (1, 2.5, and 5 mM) were 31%, 60%, and 68% at 7 days, respectively in thioflavin-T (Th-T) assay. WST-1 assay revealed that the toxic effect of $A{\beta}_{25-35}$ was reduced, in a dose-dependent manner to citrate. The percentages of neuroprotection by citrate (1, 2.5, and 5 mM) against $A{\beta}-induced$ toxicity were 19%, 31 %, and 34%, respectively. We report that citrate inhibits the formation of $A{\beta}$ fibrils in vitro and has neuroprotective effect against $A{\beta}$-induced toxicity in PC12 cells. Neuroprotective effects of citrate against $A{\beta}$ might be, to some extent, attributable to its inhibition of $A{\beta}$ fibril formation. Although the mechanism of anti-amyloidogenic activity is not clear, the possible mechanism is that citrate might have two effects, salting-in and surfactant effects. These results suggest that citrate could be of potential therapeutic value in Alzheimer's disease.

• BMB Reports
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• v.42 no.9
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• pp.541-551
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• 2009

Amyloidogenesis defines a condition in which a soluble and innocuous protein turns to insoluble protein aggregates known as amyloid fibrils. This protein suprastructure derived via chemically specific molecular self-assembly process has been commonly observed in various neurodegenerative disorders such as Alzheimer's, Parkinson's, and Prion diseases. Although the major culprit for the cellular degeneration in the diseases remains unsettled, amyloidogenesis is considered to be etiologically involved. Recent recognition of fibrillar polymorphism observed mostly from in vitro amyloidogeneses may indicate that multiple mechanisms for the amyloid fibril formation would be operated. Nucleation-dependent fibrillation is the prevalent model for assessing the self-assembly process. Following thermodynamically unfavorable seed formation, monomeric polypeptides bind to the seeds by exerting structural adjustments to the template, which leads to accelerated amyloid fibril formation. In this review, we propose another in vitro model of amyloidogenesis named double-concerted fibrillation. Here, two consecutive assembly processes of monomers and subsequent oligomeric species are responsible for the amyloid fibril formation of $\alpha$-synuclein, a pathological component of Parkinson's disease, following structural rearrangement within the oligomers which then act as a growing unit for the fibrillation.

• Journal of the Korean Magnetic Resonance Society
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• v.15 no.2
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• pp.175-186
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• 2011

Amyloid fibrils have long been known to be the well known ${\alpha}$-helix to ${\beta}$-sheet transition characterizing the conversion of cellular to scrapie forms of the prion protein. A very short sequence of Yeast prion-like protein, GNNQQNY (SupN), is responsible for aggregation that induces diseases. KSI-fused tandem repeats of SupN vector are constructed and used to express SupN peptide in Escherichia coli (E.Coli). A method for a production, purification, and cleavage of tandem repeats of recombinant isotopically enriched SupN in E. coli is described. This method yields as much as 20 mg/L of isotope-enriched fusion proteins in minimal media. Synthetic SupN peptides and $^{13}C$ Gly labeled SupN peptides are studied by Congo Red staining, Birefringence and transmission electron microscopy to characterize amyloid fibril formation. To get a better understanding of aggregation-structure relationship of 7 residues of Yeast prion-like protein, the change of a conformational structure will be studied by $^{13}C$ solid-state nmr spectroscopy as powder of both amorphous and fibrillar forms.

• Proceedings of the Korean Society of Applied Pharmacology
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• 2006.04a
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• pp.123-135
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• 2006

Alzheimer's disease (AD) is the most common form of dementia in the aging population and is clinically characterized by a progressive loss of cognitive abilities. Pathologically, it is defined by the appearance of senile plaques - extracellular insoluble, congophilic protein aggregates composed of amyloid $\beta$ (A$\beta$) and neurofibrillary tangles (NFTs) - inyracellular lesions consisting of paired helical filaments from hyperphosphorylated cytoskeletal tau protein as described by Alois Alzheimer a century ago. These hallmarks still serve as the major criteria for a definite diagnosis of the disease. Consequently, one of the key strategy for drug development in this disease area focuses on reducing the concentration of cerebral A$\beta$ plaque by using substances that inhibit A$\beta$ fibril formation. We focused on developing inhibitors by synthesizing several kinds of aromatic molecules. The synthetic compounds were initially screened to evaluate the effective compound by tioflavin T fluorescence assay. The selected effective compounds were tested cytotoxicity and protective effect from A$\beta$-induced neuronal toxicity by cell based MTT assay with HT22 hippocampal neurons. The BBB permeability on effectors was also tested in in vitro co-culture model(HUVEC/C6 cell line). The behavior test wea carried out in mutant APP/PS1 transgenic mouse model of Alzheimer's disease. And inhibition of A$\beta$ fibril formation by the effective compound was monitored with transmitted electron microscopic images.

• Bulletin of the Korean Chemical Society
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• v.35 no.12
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• pp.3542-3546
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• 2014

Relationship between molecular freedom of amyloidogenic protein and its self-assembly into amyloid fibrils has been evaluated with ${\alpha}$-synuclein, an intrinsically unfolded protein related to Parkinson's disease, by restricting its structural plasticity through an end-to-end disulfide bond formation between two newly introduced cysteine residues on the N- and C-termini. Although the resulting circular form of ${\alpha}$-synuclein exhibited an impaired fibrillation propensity, the restriction did not completely block the protein's interactive core since co-incubation with wild-type ${\alpha}$-synuclein dramatically facilitated the fibrillation by producing distinctive forms of amyloid fibrils. The suppressed fibrillation propensity was instantly restored as the structural restriction was unleashed with ${\beta}$-mercaptoethanol. Conformational flexibility of the accreting amyloidogenic protein to pre-existing seeds has been demonstrated to be critical for fibrillar extension process by exerting structural adjustment to a complementary structure for the assembly.

• BMB Reports
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• v.46 no.4
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• pp.195-200
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• 2013

To understand the corneal regeneration induced by bevacizumab, we investigated the structure changes of stroma and basement membrane regeneration. A Stick soaked in 0.5 N NaOH onto the mouse cornea and 2.5 mg/ml of bevacizumab was delivered into an alkali-burned cornea (2 ${\mu}l$) by subconjunctival injections at 1 hour and 4 days after injury. At 7 days after injury, basement membrane regeneration was observed by transmission electron microscope. Uneven and thin epithelial basement membrane, light density of hemidesmosomes, and edematous collagen fibril bundles are shown in the alkali-burned cornea. Injured epithelial basement membrane and hemidesmosomes and edematous collagen fibril bundles resulting from alkali-burned mouse cornea was repaired by bevacizumab treatment. This study demonstrates that bevacizumab can play an important role in wound healing in the cornea by accelerating the reestablishment of basement membrane integrity that leads to barriers for scar formation.

• Polymer(Korea)
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• v.27 no.4
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• pp.313-322
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• 2003

The morphology of poly(ethylene terephthalate) (PET)/poly(m-xylene adipamide) (MXD-6) blends, which was prepared by adding compatibilizer and interchange reaction agent, was investigated. The morphological change in the stretched blend films was also studied. The stretched film showed a dispersed MXD-6 fibril. This fibril became finer with increasing draw ratio (DR). The addition of compatibilizer and interchange reaction agent had no effect on the improvement of interfacial adhesion but caused a defect between the continuous phase and the dispersed phase, leading to the formation of irregular fibril. The change in the superstructure of blends with composition and draw ratio was examined with light scattering (LS). The H$\sub$v/ LS patterns showed a double-cross type pattern consisting of a broad rod-like pattern and a sharp cross streak. On the basis of the model calculation of the H$\sub$v/ pattern, it was found that the appearance of the double-cross type pattern was attributed to the stacking of crystals oriented along the draw direction. The crystals were gradually oriented to the stretching direction with draw ratio. As a result, the high level of orientation was obtained fur the sample of draw ratio is 6.0.