Low Molecular Weight Ligands Research Articles

Induction of systemic resistance in tomato against Botrytis cinerea by N-decanoyl-homoserine lactone via jasmonic acid signaling

N-decanoyl-homoserine lactone activates plant systemic resistance against Botrytis cinerea in tomato plants, which is largely dependent on jasmonic acid biosynthesis and signal transduction pathways. Rhizosphere bacteria secrete N-acylated-homoserine lactones (AHLs), a type of specialized quorum-sensing signal molecule, to coordinate their population density during communication with their eukaryotic hosts. AHLs behave as low molecular weight ligands that are sensed by plants and promote the host's resistance against foliar pathogens. In this study, we report on N-decanoyl-homoserine lactone (DHL), which is a type of AHL that induces systemic immunity in tomato plants and protects the host organism against the necrotrophic fungus Botrytis cinerea. Upon DHL treatment, tomato endogenous jasmonic acid (JA) biosynthesis (rather than salicylic acid biosynthesis) and signal transduction were significantly activated. Strikingly, the DHL-induced systemic resistance against B. cinerea was blocked in the tomato JA biosynthesis mutant spr2 and JA signaling gene-silenced plants. Our findings highlight the role of DHL in systemic resistance against economically important necrotrophic pathogens and suggest that DHL-induced immunity against B. cinerea is largely dependent on the JA signaling pathway. Manipulation of DHL-induced resistance is an attractive disease management strategy that could potentially be used to enhance disease resistance in diverse plant species.

Calorimetric insights into the interaction of novel berberrubine derivatives with human telomeric G-quadruplex DNA sequence

Binding of low molecular weight ligands to G-quadruplex nucleic acid structures is known to inhibit telomere extension and prevent abnormal growth of cells leading to cancer. Such specialized DNA conformations have emerged as important targets for drug design and development. In this manuscript we present a detailed calorimetric investigation on the interaction of berberrubine (BBR) and its two synthetic analogues with telomeric G-quadruplex DNA using calorimetry techniques and thermal melting study. A stronger binding of the analogues in comparison with BBR was observed in the experiments. The 13-phenylalkyl derivative of berberrubine (BBR2) (K = 1.38 × 105 M−1) showed higher affinity than the 13-diphenylalkyl derivative of berberrubine (BBR1) (K = 9.70 × 104 M−1). Isothermal titration calorimetry study revealed an exothermic binding that was favoured by both enthalpy and entropy changes in BBR in contrast to the analogues where the binding was mostly enthalpy dominated. A 1:1 binding stoichiometry was revealed in all the systems. The negative standard molar heat capacity values in conjunction with enthalpy–entropy compensation (EEC) confirmed the presence of dominant multiple weak non-covalent interactions in their binding. Differential scanning calorimetry and optical melting study further revealed that BBR and its analogues stabilized quadruplex structure against thermal denaturation. The overall affinity pattern of the alkaloids towards telomeric G-quadruplex structures followed as BBR2 > BBR1 > BBR.

Bacterial cyclic β-(1,2)-glucans sequester iron to protect against iron-induced toxicity.

Cellular iron homeostasis is critical for survival and growth. Bacteria employ a variety of strategies to sequester iron from the environment and to store intracellular iron surplus that can be utilized in iron-restricted conditions while also limiting the potential for the production of iron-induced reactive oxygen species (ROS). Here, we report that membrane-derived oligosaccharide (mdo) glucan, an intrinsic component of Gram-negative bacteria, sequesters the ferrous form of iron. Iron-binding, uptake, and localization experiments indicated that both secreted and periplasmic β-(1,2)-glucans bind iron specifically and promote growth under iron-restricted conditions. Xanthomonas campestris and Escherichia coli mutants blocked in the production of β-(1,2)-glucan accumulate low amounts of intracellular iron under iron-restricted conditions, whereas they exhibit elevated ROS production and sensitivity under iron-replete conditions. Our results reveal a critical role of glucan in intracellular iron homeostasis conserved in Gram-negative bacteria.

Characterization of plasma labile heme in hemolytic conditions.

Extracellular hemoglobin, a byproduct of hemolysis, can release its prosthetic heme groups upon oxidation. This produces metabolically active heme that is exchangeable between acceptor proteins, macromolecules and low molecular weight ligands, termed here labile heme. As it accumulates in plasma labile heme acts in a pro‐oxidant manner and regulates cellular metabolism while exerting pro‐inflammatory and cytotoxic effects that foster the pathogenesis of hemolytic diseases. Here, we developed and characterized a panel of heme‐specific single domain antibodies (sdAbs) that together with a cellular‐based heme reporter assay, allow for quantification and characterization of labile heme in plasma during hemolytic conditions. Using these approaches, we demonstrate that when generated during hemolytic conditions labile heme is bound to plasma molecules with an affinity higher than 10−7 m and that 2–8% (~ 2–5 μm) of the total amount of heme detected in plasma can be internalized by bystander cells, termed here bioavailable heme. Acute, but not chronic, hemolysis is associated with transient reduction of plasma heme‐binding capacity, that is, the ability of plasma molecules to bind labile heme with an affinity higher than 10−7 m. The heme‐specific sdAbs neutralize the pro‐oxidant activity of soluble heme in vitro, suggesting that these maybe used to counter the pathologic effects of labile heme during hemolytic conditions. Finally, we show that heme‐specific sdAbs can be used to visualize cellular heme. In conclusion, we describe a panel of heme‐specific sdAbs that when used with other approaches provide novel insights to the pathophysiology of heme.

Label-free quantitative 1H NMR spectroscopy to study low-affinity ligand-protein interactions in solution: A contribution to the mechanism of polyphenol-mediated astringency.

Nuclear magnetic resonance (NMR) spectroscopy is well-established in assessing the binding affinity between low molecular weight ligands and proteins. However, conventional NMR-based binding assays are often limited to small proteins of high purity and may require elaborate isotopic labeling of one of the potential binding partners. As protein–polyphenol complexation is assumed to be a key event in polyphenol-mediated oral astringency, here we introduce a label-free, ligand-focused 1H NMR titration assay to estimate binding affinities and characterize soluble complex formation between proteins and low molecular weight polyphenols. The method makes use of the effects of NMR line broadening due to protein–ligand interactions and quantitation of the non-bound ligand at varying protein concentrations by quantitative 1H NMR spectroscopy (qHNMR) using electronic reference to access in vivo concentration (ERETIC 2). This technique is applied to assess the interaction kinetics of selected astringent tasting polyphenols and purified mucin, a major lubricating glycoprotein of human saliva, as well as human whole saliva. The protein affinity values (BC50) obtained are subsequently correlated with the intrinsic mouth-puckering, astringent oral sensation imparted by these compounds. The quantitative NMR method is further exploited to study the effect of carboxymethyl cellulose, a candidate “anti-astringent” protein binding antagonist, on the polyphenol–protein interaction. Consequently, the NMR approach presented here proves to be a versatile tool to study the interactions between proteins and low-affinity ligands in solution and may find promising applications in the discovery of bioactives.

Partial filling affinity capillary electrophoresis as a useful tool for fragment-based drug discovery: A proof of concept on thrombin

With the emergence of more challenging targets, a relatively new approach, fragment-based drug discovery (FBDD), proved its efficacy and gained increasing importance in the pharmaceutical industry. FBDD identifies low molecular-weight (MW) ligands (fragments) that bind to biologically important macromolecules, then a structure-guided fragment growing or merging approach is performed, contributing to the quality of the lead. However, to select the appropriate fragment to be evolved, sensitive analytical screening methods must be used to measure the affinity in the μM or even mM range. In this particular context, we developed a robust and selective partial filling affinity CE (ACE) method for the direct binding screening of a small fragment library in order to identify new thrombin inhibitors. To demonstrate the accuracy of our assay, the complex dissociation constants of three known thrombin inhibitors, namely benzamidine, p-aminobenzamidine and nafamostat were determined and found to be in good concordance with the previously reported values. Finally, the screening of a small library was performed and demonstrated the high discriminatory power of our method towards weak binders compared to classical spectrophotometric activity assay, proving the interest of our method in the context of FBDD.

Combined Docking with Classical Force Field and Quantum Chemical Semiempirical Method PM7.

Results of the combined use of the classical force field and the recent quantum chemical PM7 method for docking are presented. Initially the gridless docking of a flexible low molecular weight ligand into the rigid target protein is performed with the energy function calculated in the MMFF94 force field with implicit water solvent in the PCM model. Among several hundred thousand local minima, which are found in the docking procedure, about eight thousand lowest energy minima are chosen and then energies of these minima are recalculated with the recent quantum chemical semiempirical PM7 method. This procedure is applied to 16 test complexes with different proteins and ligands. For almost all test complexes such energy recalculation results in the global energy minimum configuration corresponding to the ligand pose near the native ligand position in the crystalized protein-ligand complex. A significant improvement of the ligand positioning accuracy comparing with MMFF94 energy calculations is demonstrated.

Bivalent kinetic binding model to surface plasmon resonance studies of antigen-antibody displacement reactions

Molecular and functional analysis of small molecule binding to protein can provoke insights into cellular signaling and regulatory systems as well as facilitate pharmaceutical drug discovery. In label free small molecule detection the displacement assay format can be applied. This is beneficial because displacement of high molecular weight receptors is detected instead of low molecular weight ligand as in classical binding analysis. Thus, detection limit is potentially lowered.Using the influenza haemagglutinin (HA) peptide binding to mono or bivalent anti-haemagglutinin peptide antibody displacement assay formats could be established. The exact time resolved analysis of binding and dissolution of ligand HA and anti-Haemagglutinin peptide antibody was achieved with surface plasmon resonance (SPR) spectroscopy.Mathematical models could be developed from kinetic equations of ligand binding to mono or bivalent antibodies. With this, an accurate simulation of the SPR results was reached. The simulation plot had to be exactly adjusted to the SPR results to determine all kinetic rate constants defining ligand and receptor binding kinetics. Large variations in receptor concentration gave almost identical rate constants in binding. It became obvious that rebinding is in any case not necessary to understand the binding kinetics of our model system HA/anti-HA. Maximum decline of SPR response could be used to determine ligand concentrations in analyte.

Interactions of aminoglycosides with RNAs and proteins via carbohydrate microarray

Aminoglycosides are broad-spectrum antibacterials to treat bacterial infections, especially gram-negative bacteria infections. However, aminoglycosides are losing efficacy because of the increase in antibiotic resistance and their inherent toxicity, attracting more interests in developing new aminoglycosides. Several clinically used aminoglycosides are mainly exerted by inhibition of protein synthesis through binding to bacterial rRNA. The bacterial ribosome RNA is the most currently exploited RNA drug target. Identification of new compounds that target RNAs is indispensable to fight with the growing threat that bacteria pose to human safety. In this work, we used carbohydrate microarrays to probe interactions of low molecular weight ligands with RNAs and proteins. Carbohydrate microarrays, comprising hundreds to thousands of different glycan structures on surfaces in a spatially discrete pattern, are sensitive and versatile tools to study the interactions between biological macromolecules. Herein, aminoglycosides have been immobilized onto the modified glass microscope slides and their interactions with RNAs and proteins are then measured through the labeled fluorescence. The results displayed that microarray can be used to detect the binding of aminoglycosides with three types of target molecules, including the small RNA oligonucleotide mimics of aminoglycoside binding sites in the ribosome (rRNA A-site mimics), the large group I ribozyme RNA (approximately 400 nucleotide) and certain proteins (toxicity-causing enzymes, such as DNA polymerase and phospholipase C). For rRNA A-site mimics, the fluorescence intensities of 16S rRNA is stronger than that of 18S rRNA, illustrating that as a screen technique, the microarray method can not only determine the binding affinity to RNA but also detect the specific binding to bacterial rRNA mimic. The ability to screen group I ribozyme RNA can be helpful to the discovery of new RNA therapeutic targets. Binding of immobilized aminoglycosides to toxicity-causing proteins (DNA polymerase and phospholipase C) is a new method to study of aminoglycoside toxicity. These studies lay the foundation for rapid identification of new RNA-binding ligands with strong and specific binding affinity for their desired targets.

Patterns of amino acid conservation in human and animal immunodeficiency viruses.

Due to their high genomic variability, RNA viruses and retroviruses present a unique opportunity for detailed study of molecular evolution. Lentiviruses, with HIV being a notable example, are one of the best studied viral groups: hundreds of thousands of sequences are available together with experimentally resolved three-dimensional structures for most viral proteins. In this work, we use these data to study specific patterns of evolution of the viral proteins, and their relationship to protein interactions and immunogenicity. We propose a method for identification of two types of surface residues clusters with abnormal conservation: extremely conserved and extremely variable clusters. We identify them on the surface of proteins from HIV and other animal immunodeficiency viruses. Both types of clusters are overrepresented on the interaction interfaces of viral proteins with other proteins, nucleic acids or low molecular-weight ligands, both in the viral particle and between the virus and its host. In the immunodeficiency viruses, the interaction interfaces are not more conserved than the corresponding proteins on an average, and we show that extremely conserved clusters coincide with protein-protein interaction hotspots, predicted as the residues with the largest energetic contribution to the interaction. Extremely variable clusters have been identified here for the first time. In the HIV-1 envelope protein gp120, they overlap with known antigenic sites. These antigenic sites also contain many residues from extremely conserved clusters, hence representing a unique interacting interface enriched both in extremely conserved and in extremely variable clusters of residues. This observation may have important implication for antiretroviral vaccine development. A Python package is available at https://bioinf.mpi-inf.mpg.de/publications/viral-ppi-pred/ voitenko@mpi-inf.mpg.de or kalinina@mpi-inf.mpg.de Supplementary data are available at Bioinformatics online.

StructMAn: annotation of single-nucleotide polymorphisms in the structural context.

The next generation sequencing technologies produce unprecedented amounts of data on the genetic sequence of individual organisms. These sequences carry a substantial amount of variation that may or may be not related to a phenotype. Phenotypically important part of this variation often comes in form of protein-sequence altering (non-synonymous) single nucleotide variants (nsSNVs). Here we present StructMAn, a Web-based tool for annotation of human and non-human nsSNVs in the structural context. StructMAn analyzes the spatial location of the amino acid residue corresponding to nsSNVs in the three-dimensional (3D) protein structure relative to other proteins, nucleic acids and low molecular-weight ligands. We make use of all experimentally available 3D structures of query proteins, and also, unlike other tools in the field, of structures of proteins with detectable sequence identity to them. This allows us to provide a structural context for around 20% of all nsSNVs in a typical human sequencing sample, for up to 60% of nsSNVs in genes related to human diseases and for around 35% of nsSNVs in a typical bacterial sample. Each nsSNV can be visualized and inspected by the user in the corresponding 3D structure of a protein or protein complex. The StructMAn server is available at http://structman.mpi-inf.mpg.de.

The biological inorganic chemistry of zinc ions

The solution and complexation chemistry of zinc ions is the basis for zinc biology. In living organisms, zinc is redox-inert and has only one valence state: Zn(II). Its coordination environment in proteins is limited by oxygen, nitrogen, and sulfur donors from the side chains of a few amino acids. In an estimated 10% of all human proteins, zinc has a catalytic or structural function and remains bound during the lifetime of the protein. However, in other proteins zinc ions bind reversibly with dissociation and association rates commensurate with the requirements in regulation, transport, transfer, sensing, signalling, and storage. In contrast to the extensive knowledge about zinc proteins, the coordination chemistry of the “mobile” zinc ions in these processes, i.e. when not bound to proteins, is virtually unexplored and the mechanisms of ligand exchange are poorly understood. Knowledge of the biological inorganic chemistry of zinc ions is essential for understanding its cellular biology and for designing complexes that deliver zinc to proteins and chelating agents that remove zinc from proteins, for detecting zinc ion species by qualitative and quantitative analysis, and for proper planning and execution of experiments involving zinc ions and nanoparticles such as zinc oxide (ZnO). In most investigations, reference is made to zinc or Zn2+ without full appreciation of how biological zinc ions are buffered and how the d-block cation Zn2+ differs from s-block cations such as Ca2+ with regard to significantly higher affinity for ligands, preference for the donor atoms of ligands, and coordination dynamics. Zinc needs to be tightly controlled. The interaction with low molecular weight ligands such as water and inorganic and organic anions is highly relevant to its biology but in contrast to its coordination in proteins has not been discussed in the biochemical literature. From the discussion in this article, it is becoming evident that zinc ion speciation is important in zinc biochemistry and for biological recognition as a variety of low molecular weight zinc complexes have already been implicated in biological processes, e.g. with ATP, glutathione, citrate, ethylenediaminedisuccinic acid, nicotianamine, or bacillithiol.

An Alternative Thiol-Reactive Dye to Analyze Ligand Interactions with the Chemokine Receptor CXCR2 Using a New Thermal Shift Assay Format

Integral membrane proteins (IMPs) play an important role in many cellular events and are involved in numerous pathological processes. Therefore, understanding the structure and function of IMPs is a crucial prerequisite to enable successful targeting of these proteins with low molecular weight (LMW) ligands early on in the discovery process. To optimize IMP purification/crystallization and to identify/characterize LMW ligand-target interactions, robust, reliable, high-throughput, and sensitive biophysical methods are needed. Here, we describe a differential scanning fluorimetry (DSF) screening method using the thiol-reactive BODIPY FL-cystine dye to monitor thermal unfolding of the G-protein-coupled receptor (GPCR), CXCR2. To validate this method, the seven-transmembrane protein CXCR2 was analyzed with a set of well-characterized antagonists. This study showed that the new DSF assay assessed reliably the stability of CXCR2 in a 384-well format. The analysis of 14 ligands with a potency range over 4 log units demonstrated the detection/characterization of LMW ligands binding to the membrane protein target. Furthermore, DSF results cross-validated with the label-free differential static light scattering (DSLS) thermal denaturation method. These results underline the potential of the BODIPY assay format as a general tool to investigate membrane proteins and their interaction partners.

Fine tuning of molecular and supramolecular properties of simple trianglimines – the role of the functional group

Readily available chiral, triangular poly-azamacrocycles can act as receptors, low molecular weight supergelators and ligands in asymmetric synthesis.

Steric stabilization of nanoparticles with grafted low molecular weight ligands in highly concentrated brines including divalent ions.

Whereas numerous studies of stabilization of nanoparticles (NPs) in electrolytes have examined biological fluids, the interest has grown recently in media with much higher ionic strengths including seawater and brines relevant to environmental science and subsurface oil and gas reservoirs. Given that electrostatic repulsion is limited at extremely high ionic strengths due to charge screening, we have identified ligands that are well solvated in concentrated brine containing divalent cations and thus provide steric stabilization of silica nanoparticles. Specifically, the hydrodynamic diameter of silica nanoparticles with grafted low molecular weight ligands, a diol ether, [3-(2,3-dihydroxypropoxy)propyl]-trimethoxysilane, and a zwitterionic sulfobetaine, 3-([dimethyl(3-trimethoxysilyl)propyl]ammonio)propane-1-sulfonate, is shown with dynamic light scattering to remain essentially constant, indicating lack of aggregation, at room temperature and up to 80 °C for over 30 days. An extended DLVO model signifies that steric stabilization is strongly dominant against van der Waals attraction for ∼10 nm particles given that these ligands are well solvated even in highly concentrated brine. In contrast, polyethylene glycol oligomers do not provide steric stabilization at elevated temperatures, even at conditions where the ligands are soluble, indicating complicating factors including bridging of the ether oxygens by divalent cations.

Total zinc quantification by inductively coupled plasma-mass spectrometry and its speciation by size exclusion chromatography–inductively coupled plasma-mass spectrometry in human milk and commercial formulas: Importance in infant nutrition

This paper summarises results of zinc content and its speciation in human milk from mothers of preterm and full-term infants at different stages of lactation and from synthetic formula milks. Human milk samples (colostrum, 7th, 14th, and 28th day after delivery) from Spanish and Brazilian mothers of preterm and full-term infants (and also formula milks) were collected. After adequate treatment of the sample, total Zn was determined, while speciation analysis of the Zn was accomplished by size exclusion chromatography coupled online with the ICP-MS.It is observed that total zinc content in human milk decreases continuously during the first month of lactation, both for preterm and full term gestations. All infant formulas analysed for total Zn were within the currently legislated levels. For Zn speciation analysis, there were no differences between preterm and full term human milk samples. Moreover Zn species elute mainly associated with immunoglobulins and citrate in human milk whey. Interestingly the speciation in formula milk whey turned out to be completely different as the observed Zn2+ was bound almost exclusively to low molecular weight ligands (citrate) and only comparatively very low amounts of the metal appeared to be associated with higher mass biomolecules (e.g. proteins).

Colon Targeted Liposomal Systems (CTLS): Theranostic Potential.

Colon targeted liposomal systems (CTLS) for the delivery of bioactives have been well addressed in therapeutic manifestations of colonic ailments. Number of approaches using various drug delivery systems for colon targeting has been worked out but CTLS are first time being lime lighted in this review. Although liposomes are not supposed to be suitable for colon targeting via oral route this review explicitly provides advances of CTLS using exploitable ligands such as peptides or proteins (e.g. RGD, NGR, fibronectin mimetic peptide, and transferrin), Sialyl Lewis X (SLX), low molecular weight ligand like folate, monoclonal antibodies, endostatin gene and sulfatide etc. Moreover, it is bringing forth the diagnostic (or imaging) potential of CTLS using (188)Re, (99)mTc, and (111)In, etc. This review presents nanotechnology based advances for liposome researchers engaged in design and development of colon targeted liposomes for theranostic exploration.

Roles of bacterial and mammalian siderophores in host-pathogen interactions

Iron is an essential nutriment for almost all forms of life, from bacteria to humans. Despite its key role in living organisms, iron becomes toxic at high concentrations. In the body, to circumvent this toxicity, almost all the intracellular iron is bound to proteins (especially to ferritin, a protein able to bind up to 4000 atoms of iron) and a small proportion (0.2% to 3%) to low molecular weight ligands (less than 2 kDa) constituting a free iron pool able to ensure the traffic of intracellular iron. A number of small molecules (citrate, phosphate, phospholipid, polypeptide) able to chelate iron, with variable affinities, have been known for a long time. In 2010, two teams have identified new mammal endogen chelators able to bind iron with similar chemical properties as bacterial siderophores. Recently, a few publications emphasized that most of the free iron present in the body cells is indeed linked to these siderophores, which play a key role in infected-host protection mechanisms during bacterial infections, through iron homeostasis and oxidative stress regulation.

Recognition- and reactivity-based fluorescent probes for studying transition metal signaling in living systems.

ConspectusMetals are essential for life, playing critical roles in all aspects of the central dogma of biology (e.g., the transcription and translation of nucleic acids and synthesis of proteins). Redox-inactive alkali, alkaline earth, and transition metals such as sodium, potassium, calcium, and zinc are widely recognized as dynamic signals, whereas redox-active transition metals such as copper and iron are traditionally thought of as sequestered by protein ligands, including as static enzyme cofactors, in part because of their potential to trigger oxidative stress and damage via Fenton chemistry. Metals in biology can be broadly categorized into two pools: static and labile. In the former, proteins and other macromolecules tightly bind metals; in the latter, metals are bound relatively weakly to cellular ligands, including proteins and low molecular weight ligands. Fluorescent probes can be useful tools for studying the roles of transition metals in their labile forms. Probes for imaging transition metal dynamics in living systems must meet several stringent criteria. In addition to exhibiting desirable photophysical properties and biocompatibility, they must be selective and show a fluorescence turn-on response to the metal of interest. To meet this challenge, we have pursued two general strategies for metal detection, termed “recognition” and “reactivity”. Our design of transition metal probes makes use of a recognition-based approach for copper and nickel and a reactivity-based approach for cobalt and iron. This Account summarizes progress in our laboratory on both the development and application of fluorescent probes to identify and study the signaling roles of transition metals in biology. In conjunction with complementary methods for direct metal detection and genetic and/or pharmacological manipulations, fluorescent probes for transition metals have helped reveal a number of principles underlying transition metal dynamics. In this Account, we give three recent examples from our laboratory and collaborations in which applications of chemical probes reveal that labile copper contributes to various physiologies. The first example shows that copper is an endogenous regulator of neuronal activity, the second illustrates cellular prioritization of mitochondrial copper homeostasis, and the third identifies the “cuprosome” as a new copper storage compartment in Chlamydomonas reinhardtii green algae. Indeed, recognition- and reactivity-based fluorescent probes have helped to uncover new biological roles for labile transition metals, and the further development of fluorescent probes, including ones with varied Kd values and new reaction triggers and recognition receptors, will continue to reveal exciting and new biological roles for labile transition metals.

A Cu(2+) specific metallohydrogel: preparation, multi-responsiveness and pillar[5]arene-induced morphology transformation.

A Cu(2+) specific metallohydrogel was constructed from a terpyridine-based low molecular weight ligand. The metallohydrogel showed multi-responsive gel-to-sol transitions and pillar[5]arene-induced nanofiber-to-vesicle transformation at the nanoscale.