Interactions between Lipid Vesicle Membranes and Single Amino Acid Fibrils: Probable Origin of Specific Neurological Disorders.

Amyloid fibrils are known to be responsible for several neurological disorders, like Alzheimer's disease (AD), Parkinson's disease (PD), etc. For decades, mostly proteins and peptide-based amyloid fibrils have been focused on, and the topic has acknowledged the rise, development, understanding of, and controversy, as well. However, the single amino acid based amyloid fibrils, responsible for several disorders, such as phenylketonuria, tyrosenimia type II, hypermethioninemia, etc., have gotten scientific attention lately. To understand the molecular level pathogenesis of such disorders originated due to the accumulation of single amino acid-based amyloid fibrils, interaction of these fibrils with phospholipid vesicle membranes is found to be an excellent cell-free in vitro setup. Based on such an in vitro setup, these fibrils show a generic mechanism of membrane insertion driven by electrostatic and hydrophobic effects inside the membrane that reduces the integral rigidity of the membrane. Alteration of such fundamental properties of the membrane, therefore, might be referred to as one of the prime pathological factors for the development of these neurological disorders. Hence, such interactions must be investigated in cellular and intracellular compartments to design suitable therapeutic modulators against fibrils.

Ultrafast Dynamics of the Medicinal Pigment Curcumin inside the Imidazolium Surface Active Ionic Liquid Containing Giant Vesicles and White Light Generation via Triple-FRET Technique.

The naturally occurring yellow polyphenolic medicinal pigment curcumin shows ultrafast dynamics in the excited states. These ultrafast dynamics are strongly influenced by the rigidity of the environments of the systems. The present investigation unveils the ultrafast excited-state intramolecular hydrogen atom transfer (ESIHT) (which is involved in the antioxidant mechanism) and the solvation dynamics of curcumin inside the imidazolium surface active ionic liquid (SAIL), 1-hexadecyl-3-methylimidazolium chloride ([C16mim]Cl) micelle, and giant vesicles after introducing sorbitan monoesters (Span 20 and Span 80) in the aqueous medium. Interestingly, the short hydrocarbon chain containing Span 20 forms smaller, less rigid vesicles, and the long hydrocarbon chain containing Span 80 forms larger, more rigid giant vesicles after being assembled with [C16mim]Cl. The ESIHT and the solvation dynamics are slower in Span 80, containing rigid vesicles, than that in Span 20, comprising less rigid vesicles. Finally, we have established a three-component fluorescence resonance energy transfer (Triple-FRET) system to generate white light (WL) in the micelle and giant vesicles. Here the hydrophobic dye 1,6-diphenyl-1,3,5-hexatriene (DPH) acts as the donor, and the hydrophilic anticancer drug doxorubicin hydrochloride (DOX) serves as the acceptor along with the intermediate donor, curcumin. At a specific combination of the concentrations of these dyes in a particular self-assembled system, WL is generated due to the triple-FRET phenomena.

Excited State Photophysics of Curcumin and its Modulation in Alkaline Non-Aqueous Medium.

Curcumin, a well-known medicinal pigment, has seen limited applications in biology despite having great potential as a therapeutic drug. Deprotonation is one of the possible ways to enhance solubility of curcumin in polar solvent. Here, we have explored the effect of deprotonation on the ultrafast dynamics of this biomolecule with the help of the time-resolved fluorescence spectroscopic measurements using the femtosecond fluorescence upconversion technique. The excited state photophysics of fully deprotonated curcumin significantly differs from that of neutral curcumin. We have observed that the completely deprotonated curcumin not only has higher quantum yield, but also higher excited state lifetime and slower solvation dynamics in comparison to neutral curcumin. We propose solvation dynamics and intramolecular charge transfer as the excited state processes associated with the radiative decay of the completely deprotonated molecule, while ruling out the possibility of excited state proton exchange or proton transfer. Our results are well supported by time-dependent density-functional theory calculations. Lastly, we have also demonstrated the possibility of modulating the ultrafast dynamics of fully deprotonated curcumin using non-aqueous alkaline binary solvent mixtures. We believe our results will provide significant physical insight towards unveiling the excited state dynamics of this molecule.

Insights into the Strong Emission Enhancement of Molecular Rotor Thioflavin T in Aqueous Cellulose Nanocrystal Dispersion: White Light Generation in Protein and Micellar Media.

Thioflavin t (THT) is a well-known molecular rotor extensively used to detect amyloid-like structures. But THT shows very weak emission in water. In this article, we have found that THT shows very strong emission in the presence of cellulose nanocrystals (CNCs). Steady-state and time-resolved emission techniques have been used to study the strong emission of THT in aqueous CNC dispersion. The time-resolved study showed that in the presence of CNCs, the lifetime increased by ∼1500 fold compared to pure water (<1 ps). To know the nature of interaction and also the reason for this increase in emission zeta potential, stimuli-dependent and temperature-dependent studies have been carried out. These studies proposed that electrostatic interaction is the main factor for this binding of THT with CNCs. Further, the addition of another anionic lipophilic dye, merocyanine 540 (MC540), with CNCs-THT in both BSA protein (CIE: 0.33, 0.32) and TX-100 micellar (4.5 mM) (CIE: 0.32, 0.30) solutions produced excellent white light emission. Lifetime decay and absorption studies proposed a possible fluorescence resonance energy transfer mechanism in this generation of white light emission.

Targeting genomic DNAs and oligonucleotide on base specificity: A comparative spectroscopic, computational and in vitro study.

Drug discovery in targeted nucleic acid therapeutics encompass several stages and rigorous challenges owing to less specificity of the DNA binders and high failure rate in different stages of clinical trials. In this perspective, we report newly synthesized ethyl 4-(pyrrolo[1,2-a]quinolin-4-yl)benzoate (PQN) with minor groove A-T base pair binding selectivity and encouraging in cell results. This pyrrolo quinolin derivative has shown excellent groove binding ability with three of our inspected genomic DNAs (cpDNA 73 % AT, ctDNA58% AT and mlDNA 28 % AT) with varying A-T and G-C content. Notably in spite of similar binding patterns PQN have strong binding preference with A-T rich groove of genomic cpDNA over the ctDNA and mlDNA. Spectroscopic experiments like steady state absorption and emission results have established the relative binding strengths (Kabs = 6.3 × 105 M-1, 5.6 × 104 M-1, 4.3 × 104 M-1 and Kemiss = 6.1 × 105 M-1, 5.7 × 104 M-1 and 3.5 × 104 M-1 for PQN-cpDNA, PQN-ctDNA and PQN-mlDNA respectively) whereas circular dichroism and thermal melting studies have unveiled the groove binding mechanism. Specific A-T base pair attachment with van der Waals interaction and quantitative hydrogen bonding assessment were characterized by computational modeling. In addition to genomic DNAs, preferential A-T base pair binding in minor groove was also observed with our designed and synthesized deca-nucleotide (primer sequences 5/-GCGAATTCGC-3/ and 3/-CGCTTAAGCG-5/). Cell viability assays (86.13 % in 6.58 µM and 84.01 % in 9.88 µM concentrations) and confocal microscopy revealed low cytotoxicity (IC50 25.86 µM) and efficient perinuclear localization of PQN. We propose PQN with excellent DNA-minor groove binding capacity and intracellular permeation properties, as a lead for further studies encompassing nucleic acid therapeutics.

Deciphering Dual Modes of Parkinsonian Biomolecules Derived Fluorescent Nanoparticles: Protein Specificity and White Light Generation.

Dopamine (DA) and 3,4-dihydroxy-l-phenylalanine (L-Dopa or DPA), a marker and medicine for the neurological disorder Parkinson's disease (PD), lead to the formation of polymeric fluorescent nanoparticles (F-Poly NPs or F-NPs or simply, NPs). The interaction study between proteins and NPs shows prominent interaction with strong specificity toward albumin type proteins for DPA derived and mixed NPs. Furthermore, encapsulation of the anticancer drug doxorubicin hydrochloride (Dox) inside the NP-protein conjugates results in excellent white light emission with pronounced specificity toward albumin proteins for F-PDPA and F-Mix NPs. Finally, the use of BSA protein fibril resulting in strong binding with NPs along with Dox assisted white light emission has also been studied.

Amyloids Formed by Nonaromatic Amino Acid Methionine and Its Cross with Phenylalanine Significantly Affects Phospholipid Vesicle Membrane: An Insight into Hypermethioninemia Disorder.

The incorrect metabolic breakdown of the nonaromatic amino acid methionine (Met) leads to the disorder called hypermethioninemia via an unknown mechanism. To understand the molecular level pathogenesis of this disorder, we prepared a DMPC lipid membrane, the mimicking setup of the cell membrane, and explored the effect of the millimolar level of Met on it. We found that Met forms toxic fibrillar aggregates that disrupt the rigidity of the membrane bilayer, and increases the dynamic response of water molecules surrounding the membrane as well as the heterogeneity of the membrane. Such aggregates strongly deform red blood cells. This opens the requirement to consider therapeutic antagonists either to resist or to inhibit the toxic amyloid aggregates against hypermethioninemia. Moreover, such disrupting effect on membrane bilayer and cytotoxicity along with deformation effect on RBC by the cross amyloids of Met and Phenylalanine (Phe) was found to be most virulent. This exclusive observation of the enhanced virulent effect of the cross amyloids is expected to be an informative asset to explain the coexistence of two amyloid disorders.

Formation of lipid tubules induced by a sugar-like molecule myo-inositol.

The sugar-like molecule myo-inositol (InOH) bears an uncanny structural resemblance to the pyranose form of the sugar D-glucose (DG). InOH and its derivatives play a pivotal role in cell biology; whereby its interaction with the model membrane needs to be studied. Here, we have demonstrated that lipid tubules are formed as a result of the above-said interactions and that these interactions can be prevented by using hydroxyl protected InOH derivatives. We have tried to elucidate the nature of the InOH-membrane interactions by comparing them with DG-membrane interactions and have proposed a mechanism for the same.

The Dietary Nutrient Trimethylamine N-Oxide Affects the Phospholipid Vesicle Membrane: Probable Route to Adverse Intake.

Trimethylamine N-oxide (TMAO), a choline-containing dietary supplement obtained from red meat, egg, and other animal resources, on excess accumulation is known to cause cardiovascular diseases (CVDs) like atherosclerosis. To understand the molecular mechanism of the pathogenesis of TMAO-induced CVDs, we have set up 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) membrane in water that mimicked the endothelial cell membrane-blood interface of the artery wall and investigated the effect of an elevated concentration of TMAO on the membrane. We found that TMAO exerts an "action at a distance" mechanism through electrostatic force of attraction that significantly alters various properties of the membrane, like hydrophobicity, lateral organization, and interfacial water dynamics, which elevates the rigidity of the membrane. Such an effect was found to be further amplified in the presence of known causes of CVDs, i.e., high content of cholesterol (Chol). Therefore, TMAO-induced membrane rigidity may restrict the intrinsic elasticity of an artery membrane, expected to be introducing "hardening of the arteries", which makes the membrane atherosclerotic.

Influence of a Polyneurotransmitter on DNA-Mediated Förster-Based Resonance Energy Transfer: A Path Leading to White Light Generation.

The physiologically important biomolecule, dopamine (DA), shows strong self-oxidation and aggregation behaviors, which have been controlled and modulated to result in fluorescent polydopamine (F-PDA) nanoparticles. On the other hand, the simultaneous binding of two diverse deoxyribonucleic acid (DNA) binding probes, 4',6-diamidino-2-phenylindole dihydrochloride (DAPI) and ethidium bromide (EtBr), has been elaborately established to follow the Förster-based resonance energy transfer (FRET) pathway. The comparative understanding of this DNA-mediated FRET in three media, phosphate buffer saline (PBS) of pH 7.4, DA, and F-PDA, has concluded that the FRET efficiency in the three media follows the order: PBS > DA > F-PDA. This controlled FRET in the fluorescent F-PDA matrix serves a pivotal role for efficient white light (WL) generation with excellent Commission Internationale de l'Eclairage (CIE) parameters that match well with that of pure WL emission. The obtained WL emission has been shown to be very specific with respect to concentrations of different participating components and the excitation wavelength of the illuminating source. Furthermore, the optical properties of the WL emitting solution have been observed to be retained excellently inside the well-known agarose gel matrix. Finally, the mechanistic pathway behind such a FRET-based WL generation has been established in detail, and to the best of our knowledge, the current study offers the first and only report that discloses the influence of a fluorescent polyneurotransmitter matrix for successful generation of WL emission.

Self-assembly of artificial sweetener aspartame adversely affects phospholipid membranes: plausible reason for its deleterious effects.

The prolonged intake of the artificial sweetener aspartame is known to have deleterious effects. Our biophysical experimentations indicate that aspartame forms self-assembled cytotoxic fibrillar etiologies that affect the intrinsic integrity of the phospholipid membrane bilayer through electrostatic interaction and hydrophobic insertion, thereby making the membrane less rigid and more heterogeneous.

Phenylalanine Interacts with Oleic Acid-Based Vesicle Membrane. Understanding the Molecular Role of Fibril-Vesicle Interaction under the Context of Phenylketonuria.

In the present contribution, on the basis of a spectroscopic and microscopic investigation, the characterization and photophysics of various assemblies of oleic acid/oleate solution at three pH values, namely, 8.28, 9.72, and 11.77, were explored. The variation in the dynamic response of aqua molecules in and around the assemblies has been interrogated by a picoseconds solvation dynamics experiment using a time-correlated single-photon counting setup employing coumarin-153 as a probe. On the one hand, the time-resolved fluorescence anisotropy measurement along with the fluorescence correlation spectroscopy experiment was executed to extract information regarding the comparison of the extent of the internal restricted confinement of these assemblies. On the other hand, an effort to investigate the cross-interaction between the self-assembled architectures of l-phenylalanine (l-Phe), responsible for phenylketonuria (PKU) disorder, and the oleic acid at the vesicle-forming pH established that the l-Phe fibrillar morphologies strongly alter the dynamic properties of the vesicle membrane formed by the oleic acid. Specifically, the interaction of the l-Phe assemblies with the oleic acid vesicle membrane is found to introduce the flexibility of the vesicle membrane and alter the hydration properties of the membrane. To track the fibril-induced alterations of the oleic acid vesicle properties, various spectroscopic and microscopic investigations were performed. The mutual reconciliation of the experimental outputs, therefore, portrays the state of the art, which accounts for the fibril-induced alterations of the properties of the oleic acid vesicle membrane, the mimicking setup of the cellular membrane, thereby informing us that alterations of such a property of the membrane should be taken into active consideration during the rational development of therapeutic modulators against disorders like PKU.

State of the Art and Perspectives on the Biofunctionalization of Fluorescent Metal Nanoclusters and Carbon Quantum Dots for Targeted Imaging and Drug Delivery.

The interface of nanobio science and cancer nanomedicine is one of the most important current frontiers in research, being full of opportunities and challenges. Ultrasmall fluorescent metal nanoclusters (MNCs) and carbon quantum dots (CQDs) have emerged as promising fluorescent nanomaterials due to their unique physicochemical and optical properties, facile surface functionalization, good photostability, biocompatibility, and aqueous dispersity. These characteristics make them advantageous over conventional fluorophores such as organic dye molecules and semiconductor quantum dots (QDs) for the detection, diagnosis, and treatment of various diseases including cancer. Recently, researchers have focused on the biofunctionalization strategy of the MNCs and CQDs which can tailor their physicochemical and biological properties and, in turn, can empower these biofunctionalized nanoprobes for diverse applications including imaging, drug delivery, theranostics, and other biomedical applications. In this invited feature article, we first discuss some fundamental structural and physicochemical characteristics of the fluorescent biocompatible quantum-sized nanomaterials which have some outstanding features for the development of multiplexed imaging probes, delivery vehicles, and cancer nanomedicine. We then demonstrate the diverse surface engineering of these fluorescent nanomaterials with reactive target specific functional groups which can help to construct multifunctional nanoprobes with improved targeting capabilities having minimal toxicity. The promising future of the biofunctionalized fluorescent quantum-sized nanomaterials in the field of bioanalytical and biomedical research is elaborately demonstrated, showing selected recent works with relevant applications. This invited feature article finally ends with a short discussion of the current challenges and future prospects of the development of these bioconjugated/biofunctionalized nanomaterials to provide insight into this burgeoning field of MNC- and CQD-based diagnostics and therapeutic applications.

A Comparative Study on DMSO-Induced Modulation of the Structural and Dynamical Properties of Model Bilayer Membranes.

Modulating the structures and properties of biomembranes via permeation of small amphiphilic molecules is immensely important, having diverse applications in cell biology, biotechnology, and pharmaceuticals, because their physiochemical and biological interactions lead to new pathways for transdermal drug delivery and administration. In this work, we have elucidated the role of dimethyl sulfoxide (DMSO), broadly used as a penetration-enhancing agent and cryoprotective agent on model lipid membranes, using a combination of fluorescence microscopy and time-resolved fluorescence spectroscopy. Spatially resolved fluorescence lifetime imaging microscopy (FLIM) has been employed to unravel how the fluidity of the DMSO-induced bilayer regulates the structural alteration of the vesicles. Moreover, we have also shown that the dehydration effect of DMSO leads to weakening of the hydrogen bond between lipid headgroups and water molecules and results in faster solvation dynamics as demonstrated by femtosecond time-resolved fluorescence spectroscopy. It has been gleaned that the water dynamics becomes faster because bilayer rigidity decreases in the presence of DMSO, which is also supported by time-resolved rotational anisotropy measurements. The enhanced diffusivity and increased membrane fluidity in the presence of DMSO are further ratified at the single-molecule level through fluorescence correlation spectroscopy (FCS) measurements. Our results indicate that while the presence of DMSO significantly affects the 1,2-dimyristoyl-rac-glycero-3-phosphocholine (DMPC) and 1,2-dipalmitoyl-rac-glycero-3-phosphatidylcholine (DPPC) bilayers, it has a weak effect on 1,2-dimyristoyl-sn-glycero-3-phospho-rac-glycerol (DMPG) vesicles, which might explain the preferential interaction of DMSO with the positively charged choline group present in DMPC and DPPC vesicles. The experimental findings have also been further verified with molecular dynamics simulation studies. Moreover, it has been observed that DMSO is likely to have a differential effect on heterogeneous bilayer membranes depending on the structure and composition of their headgroups. Our results illuminate the importance of probing the lipid structure and composition of cellular membranes in determining the effects of cryoprotective agents.

Modulation of Membrane Fluidity to Control Interfacial Water Structure and Dynamics in Saturated and Unsaturated Phospholipid Vesicles.

The structure and dynamics of interfacial water in biological systems regulate the biochemical reactions. But, it is still enigmatic how the behavior of the interfacial water molecule is controlled. Here, we have investigated the effect of membrane fluidity on the structure and dynamics of interfacial water molecules in biologically relevant phopholipid vesicles. This study delineates that modulation of membrane fluidity through interlipid separation and unsaturation not only mitigate membrane rigidity but also disrupt the strong hydrogen bond (H-bond) network around the lipid bilayer interface. As a result, a disorder in H-bonding between water molecules arises several layers beyond the first hydration shell of the polar headgroup, which essentially modifies the interfacial water structure and dynamics. Furthermore, we have also provided evidence of increasing transportation through these modulated membranes, which enhance the membrane mediated isomerization reaction rate.

Aging-Dependent Morphological Crystallinity Determines Membrane Activity of l-Phenylalanine Self-Assembles.

Amyloid polymorphism has emerged as an important topic of research in recent years to identify the particular species responsible for several neurodegenerative disorders, whereas the concept is overlooked in the case of the simplest building block, that is, l-phenylalanine (l-Phe) self-assembly. Here, we report the first evidence of l-Phe polymorphism and the conversion of metastable helical fibrillar to thermodynamically stable rodlike crystalline morphologies with increasing time and temperature. Furthermore, only the fibrillar l-Phe polymorph shows a significant modulation of the model membrane. In addition, the l-Phe molecules prefer to arrange in a multilayered rodlike fashion than in a lateral arrangement, which reduces the membrane binding ability of the l-Phe polymorph due to the decrease in the partial charge of the N-terminal of l-Phe units. The present work exemplifies a different approach to understanding l-Phe self-assembly and provides an effective strategy for the therapy of phenylketonuria by scrutinizing the discrete membrane activity of different l-Phe polymorphs.

Dynamics of the vesicles composed of fatty acids and other amphiphile mixtures: unveiling the role of fatty acids as a model protocell membrane.

Fundamental research at the interface of chemistry and biology has the potential to shine light on the question of how living cells can be synthesized from inanimate matter thereby providing plausible pathways for the emergence of cellular life. Compartmentalization of different biochemical reactions within a membrane bound water environment is considered an essential first step in any origin of life pathway. It has been suggested that fatty acid-based vesicles can be considered a model protocell having the potential for change via Darwinian evolution. As such, protocell models have the potential to assist in furthering our understanding of the origin of life in the laboratory. Fatty acids, both by themselves and in mixtures with other amphiphiles, can form different self-assembled structures depending on their surroundings. Recent studies of fatty acid-based membranes have suggested likely pathways of protocell growth, division and membrane permeabilisation for the transport of different nutrients, such as nucleotides across the membrane. In this review, different dynamic processes related to the growth and division of the protocell membrane are discussed and possible pathways for transition of the protocell to the modern cell are explored. These areas of research may lead to a better understanding of the synthesis of artificial cell-like entities and thus herald the possibility of creating new form of life distinct from existing biology. Graphical Abstract Table of Content (TOC) only.

Denaturant-Mediated Modulation of the Formation and Drug Encapsulation Responses of Gold Nanoparticles.

The extensive and diversified applications of the well-known plasmonic nanoparticle systems along with their easy and environment-friendly synthesis strategies drive us to investigate in-depth this important research field. In the current scenario, our present study deals with an important plasmonic nanomaterial, i.e., globular protein, and human serum albumin (HSA)-conjugated gold nanoparticle (HSA-Au NP) system. The well-known chemical denaturants, urea and guanidine hydrochloride (GdnHCl or GnHCl), are investigated to show detrimental effects toward the formation of gold nanoparticles; however, the effect of GdnHCl is observed to be much prominent compared to that of urea. The synthesized nanoparticle system is found to be highly biocompatible from the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)-based cytotoxicity assay, and therefore, the applications of encapsulation of the well-known anticancer drug molecule, doxorubicin hydrochloride (Dox), in the nanoparticle system are further studied. In this drug encapsulation study, drug-metal complexation between Dox and HAuCl4·3H2O has been discussed elaborately. Similar to the nanoparticle formation, the effects of denaturants on drug encapsulation have also been discovered, and interestingly, it has been observed that urea plays a positive role, whereas GdnHCl plays a negative or detrimental role toward drug encapsulation in the synthesized gold nanoparticle system. The detailed photophysical mechanisms behind the drug encapsulation in the synthesized plasmonic nanosystem at every stage have also been explored. Overall, this study will conclusively explain the influences of the extensively used chemical denaturants on the synthesis and drug encapsulation behaviors of a well-known protein-conjugated gold nanoparticle, and as a consequence, it can be highly useful and acceptable to the biomedical and pharmaceutical research communities.