Homocysteine thiolactone and H2 O2 induce amino acid modifications and alter the fibrillation propensity of the Aß25-35 peptide.

Of the proteinaceous ß-sheet-rich amyloid fibrillar structures, the Aß25-35 peptide, a component of the full-length Aß involved in Alzheimer's disease, has similar toxicity to the parent peptide. In this study, the effects of homocysteine thiolactone (HCTL) and hydrogen peroxide (H2 O2 ) on the conformation and fibrillation propensity of the Aß25-35 peptide were investigated. Both HCTL and H2 O2 induced amino acid modifications along with alteration in aggregation propensity. Methionine (Met)-35 was oxidized by H2 O2 and aggregation was attenuated following the increased hydrophilicity of the peptide due to sulfoxide/sulfone formation. The HCTL-modified lysine (Lys-28) residue destabilizes the structure of the peptide, which leads to fibrillation. Our studies provide important information regarding the relationship between amino acid modifications and the amyloid fibrillation process.

Permeation of flavonoid loaded human serum albumin nanoparticles across model membrane bilayers.

The rapid growth in the applications of nanoparticles (NPs) in biomedical and pharmaceutical fields requires an understanding of the interactions with the lipid bilayer membrane for further in vivo studies. Zwitterionic 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), negatively charged 1,2-dioleoyl-sn-glycero-3-phospho-l-serine (DOPS) and positively charged 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) have been used to prepare model lipid membranes and the ability of flavonoid loaded nanoparticles to cross the membranes investigated. The lipid vesicles have been prepared by a freeze-thaw method followed by an extrusion technique and characterised by dynamic light scattering (DLS) and high-resolution transmission electron microscopy (HRTEM). The synthesized model lipid membranes exhibited a bilayer spherical type of morphology with an average diameter of less than 150 nm. A calcein leakage assay and fluorescence anisotropy measurement indicated that the membranes are permeable to the flavonoid (fisetin/morin/epicatechin) loaded human serum albumin nanoparticles. This implies that drug/compound encapsulated nanoparticles are able to effectively cross the lipid bilayer thus permitting the design and development of new compounds that may be encapsulated for safe and potential use in biomedical applications.

Multiplexed Covalent Patterns on Double-Reactive Porous Coating.

We have conceptualized and demonstrated an approach based on the combination of hydrophobicity, a substrate-independent dip coating as porous material with double residual chemical reactivities for implementing multiplexed, miniaturized and unclonable bulk-infused patterns of different fluorophores following distinct reaction pathways. The embedded hydrophobicity (∼102°) restricted the unwanted spreading of beaded aqueous ink on the coating. The constructions of micropatterns on porous dip-coating via ink-jet printing or microchannel cantilever spotting offered orthogonal read-out and remained readable even after removal of the exterior of the coating.

Nanoencapsulation as a Promising Platform for the Delivery of the Morin-Cu(II) Complex: Antibacterial and Anticancer Potential.

Nanoencapsulation has emerged as a promising approach for the effective delivery of poorly aqueous soluble compounds. The current study focuses on the preparation of human serum albumin (HSA)-based nanoparticles (NPs) and poly lactic-co-glycolic acid (PLGA)-based nanoparticles for effective delivery of the morin-Cu(II) complex. The NPs were analyzed based on different parameters such as particle size, surface charge, morphology, encapsulation efficiency, and in vitro release properties. The average particle sizes were found to be 214 ± 6 nm for Mor-Cu-HSA-NPs and 185 ± 7.5 nm for Mor-Cu-PLGA-NPs. The release of the morin-Cu(II) complex from both the NPs (Mor-Cu-HSA-NPs and Mor-Cu-PLGA-NPs) followed a biphasic behavior, which comprises an early burst release followed by a sustained and controlled release. The resulting NPs also exhibit free radical scavenging activity confirmed by a standard antioxidant assay. The antibacterial activities of the NPs were investigated using a disk diffusion technique, and it was observed that both the NPs showed better antibacterial activity than morin and the morin-Cu(II) complex. The anticancer activities of the prepared NPs were examined on MDA-MB-468 breast cancer cell lines using a cytotoxicity assay, and the mode of cell death was visualized using fluorescence microscopy. Our results revealed that NPs kill the cancer cells with greater efficiency than free morin and the morin-Cu(II) complex. Thus, both HSA-based NPs and PLGA-based NPs can act as promising delivery systems for the morin-Cu(II) complex and can be utilized for further biomedical applications.

Abrasion tolerant, non-stretchable and super-water-repellent conductive & ultrasensitive pattern for identifying slow, fast, weak and strong human motions under diverse conditions.

The conversion of mechanical deformation into electrical signals is a widely used principle for various relevant applications. Facile & scalable fabrication, ultrahigh-sensitivity, low-response time and uninterrupted performance under severe conditions are hallmarks of an efficient strain-sensor that would be suitable for realistic application. In the past, various approaches were introduced to achieve high gauge factor-mainly associated with a large tensile deformation. But, in reality, a flexible strain sensor that displays a high gauge factor at low applied strain and remains efficient under practically relevant diverse and challenging conditions would be more appropriate for unambiguous and effective monitoring of human motions and other relevant applications. But, a low-strain sensor with ultrahigh sensitivity and durability is yet to be introduced in the literature. Here, a metal-free, chemically reactive and conductive ink is unprecedentedly introduced following a 1,4-conjugate addition reaction. Furthermore, a strategic integration of a chemically reactive porous paper with the prepared conductive ink allowed the development of a chemically reactive and conductive interface that allowed desired post covalent modification with selected alkylamines under ambient conditions. Taking advantage of the spatially selective deposition of the prepared ink on chemically recative paper and the ability of post covalent modification of the prepared ink, an abrasion tolerant superhydrophobic & conductive patterned interface was developed for achieving a low-strain (below 0.2%) based flexible strain sensor with an ultrahigh sensitivity (gauge factor ∼18 300) and low response time (8 ms). The external low-strain induced cracks on the flexible & durable superhydrophobic and conductive patterned interface provided a facile basis for real-time & wireless monitoring of slow, fast, weak and strong human motions & expressions-under diverse conditions, including continuous aqueous exposures, physical abrasions etc.

Exploring the Inhibitory and Antioxidant Effects of Fullerene and Fullerenol on Ribonuclease A.

Fullerene-protein interaction studies have been a key topic of investigation in recent times, but the lower water solubility of fullerene somewhat limits its application in the biological system. In this work, we have compared the activities of fullerene and its water-soluble hydrated form, that is fullerenol, on ribonuclease A (RNase A) under physiological conditions (pH 7.4). The interaction studies of fullerene and fullerenol with protein suggest that the binding depends on the hydrophobic interactions between the protein and the ligand. In addition, fullerene and fullerenol slow down the ribonucleolytic activity of RNase A through noncompetitive and mixed types of inhibition, respectively. This precisely gives the idea about the ligand-binding sites in RNase A, which has further been explored using docking studies. Both these nanoparticles show a reduction in dityrosine formation in RNase A caused due to oxidative stress and also prevent RNase A dimer formation to different extents depending on their concentration.

Microwave-radiation-induced molecular structural rearrangement of hen egg-white lysozyme.

We have investigated the nonthermal effect of 10 GHz/22 dBm microwave radiation on hen egg-white lysozyme (HEWL) over different irradiation times, ranging from 2 min to 1 h. To ensure a control over the radiation parameters, a pair of microwave rectangular waveguides is used to irradiate the samples. Optical spectroscopic measurements, which include UV-visible absorption spectroscopy, Raman spectroscopy, and far UV CD spectroscopy, reveal the exposure of the buried tryptophan (Trp) residues of the native molecule between 15 and 30 min of radiation. The higher duration of the perturbation leads to a compact structure of the protein and Trp residues are buried again. Interestingly, we do not find any change in the secondary structure of the protein even for 1 h duration of radiation. The relaxation dynamics of the irradiated molecules also has been discussed. We have shown that the molecules relax to their native configuration in 7-8 h after the radiation field is turned off. The structural rearrangement over the above timescale has further been probed by a model calculation, based on a modified Langevin equation. Our coarse-grained simulation approach utilizes the mean of atomic positions and net atomic charge of each amino acid of native HEWL to mimic the initial conformation of the molecule. The modified positions of the residues are then calculated for the given force fields. The simulation results reveal the nonmonotonous change in overall size of the molecule, as observed experimentally. The radiation parameters used in our experiments are very similar to those of some of the electronic devices we often come across. Thus, we believe that the results of our studies on a simple protein structure may help us in understanding the effect of radiation on complex biological systems as well.

Probing the inhibitory potency of epigallocatechin gallate against human γB-crystallin aggregation: Spectroscopic, microscopic and simulation studies.

Aggregation of human ocular lens proteins, the crystallins is believed to be one of the key reasons for age-onset cataract. Previous studies have shown that human γD-crystallin forms amyloid like fibres under conditions of low pH and elevated temperature. In this article, we have investigated the aggregation propensity of human γB-crystallin in absence and presence of epigallocatechin gallate (EGCG), in vitro, when exposed to stressful conditions. We have used different spectroscopic and microscopic techniques to elucidate the inhibitory effect of EGCG towards aggregation. The experimental results have been substantiated by molecular dynamics simulation studies. We have shown that EGCG possesses inhibitory potency against the aggregation of human γB-crystallin at low pH and elevated temperature.

Gallic acid induced dose dependent inhibition of lysozyme fibrillation.

Amyloidosis is primarily characterized by the deposition of misfolded protein aggregates. Although the natural polyphenols have long been known as effective amyloid inhibitors, the mechanistic details of their inhibitory actions still remain unclear. Our present study explores the inhibition mechanism of polyphenols by studying the anti-amyloidogenic property of gallic acid (GA), the smallest structural unit of tea polyphenols, on hen egg white lysozyme (HEWL) at physiological pH. Using various spectroscopic techniques such as UV-vis, fluorescence, circular dichroism and dynamic light scattering, and microscopic techniques such as TEM and FESEM, it has been shown that GA potentially inhibits the self-aggregation process in a concentration dependent manner. Gel electrophoresis studies suggest that the o-dihydroxy moiety of GA is oxidized into the quinone moiety and H2O2 in the system under the experimental conditions. The quinone binds near the hydrophobic region of HEWL and restricts hydrophobic exposure. Cyclic voltammetry studies reveal that the Met residues of HEWL are oxidized by H2O2 to highly polar sulfoxide-modified side chains. The partially unfolded intermediates formed under the denaturing conditions employed remain in contact with the solvent thus preventing further aggregation.

Inhibition of Human Serum Albumin Fibrillation by Two-Dimensional Nanoparticles.

The formation and deposition of amyloid fibrils have been linked to the pathogenesis of numerous debilitating neurodegenerative disorders. Serum albumins serve as good model proteins for understanding the molecular mechanisms of protein aggregation and fibril formation. Graphene-based nanotherapeutics appear to be promising candidates for designing inhibitors of protein fibrillation. The inhibitory effect of graphene oxide (GO) nanoparticles on the fibrillation of human serum albumin (HSA) in an in vitro mixed solvent system has been investigated. The methods used include ThT fluorescence, ANS binding, Trp fluorescence, circular dichroism, fluorescence microscopy, field-emission scanning electron microscopy, and high-resolution transmission electron microscopy. It was observed that GO inhibits HSA fibrillation and forms agglomerates with ß-sheet rich prefibrillar species. Binding of GO prevents the formation of mature fibrils with characteristic cross-ß sheet but does not promote refolding to the native state.

Protection of human γB-crystallin from UV-induced damage by epigallocatechin gallate: spectroscopic and docking studies.

The transparency of the human eye lens depends on the solubility and stability of the structural proteins of the eye lens, the crystallins. Although the mechanism of cataract formation is still unclear, it is believed to involve protein misfolding and/or aggregation of proteins due to the influence of several external factors such as ultraviolet (UV) radiation, low pH, temperature and exposure to chemical agents. In this article, we report the study of UV induced photo-damage (under oxidative stress) of recombinant human γB-crystallin in vitro in the presence of the major green tea polyphenol, (-)-epigallocatechin gallate (EGCG). We have shown that EGCG has the ability to protect human γB-crystallin from oxidative stress-induced photo-damage.

Hydrophobic tail length plays a pivotal role in amyloid beta (25-35) fibril-surfactant interactions.

The amyloid ß-peptide fragment comprising residues 25-35 (Aß25-35 ) is known to be the most toxic fragment of the full length Aß peptide which undergoes fibrillation very rapidly. In the present work, we have investigated the effects of the micellar environment (cationic, anionic, and nonionic) on preformed Aß25-35 fibrils. The amyloid fibrils have been prepared and characterized by several biophysical and microscopic techniques. Effects of cationic dodecyl trimethyl ammonium bromide (DTAB), cetyl trimethylammonium bromide (CTAB), anionic sodium dodecyl sulfate (SDS), and nonionic polyoxyethyleneoctyl phenyl ether (Triton X-100 or TX) on fibrils have been studied by Thioflavin T fluorescence, UV-vis spectroscopy based turbidity assay and microscopic analyses. Interestingly, DTAB and SDS micelles were observed to disintegrate prepared fibrils to some extent irrespective of their charges. CTAB micelles were found to break down the fibrillar assembly to a greater extent. On the other hand, the nonionic surfactant TX was found to trigger the fibrillation process. The presence of a longer hydrophobic tail in case of CTAB is assumed to be a reason for its higher fibril disaggregating efficacy, the premise of their formation being largely attributed to hydrophobic interactions. Proteins 2016; 84:1213-1223. © 2016 Wiley Periodicals, Inc.

Interfacial force-driven pattern formation during drying of Aß (25-35) fibrils.

Pattern formation during evaporation of biofluids finds significant applications in the biomedical field for disease identification. Aß (25-35) is the smallest peptide in the amyloid peptide family that retains the toxicity of a full length peptide responsible for Alzheimer's disease and is chosen here as the model solute. Drying experiments on substrates of varying wettability exhibit unique drying patterns of Aß (25-35) fibrils visualized through fluorescence microscopy and transmission electron microscopy. The unique pattern formations can be interpreted as manifestations of the changes in the self-pinning mechanism with changes in wettability, which in some cases resembles the well-known coffee ring effect. Additionally, the delicate balance between the drag and capillary forces has been perturbed by initiating controlled rates of evaporation and probing their effects on the fibril patterning.