Supramolecular citrate poly allylamine hydrochloride nanoparticles for citrate delivery and calcium oxalate nanocrystal dissolution.

HYPOTHESIS: Renal calculi (kidney stones) are mainly made by calcium oxalate and can cause different complications including malfunction of the kidney. The most important urinary stone inhibitors are citrate molecules. Unfortunately, the amount of citrate reaching the kidney after oral ingestion is low. We hypothesized that nanoparticles of polyallylamine hydrochloride (CIT-PAH) carrying citrate ions could simultaneously deliver citrates while PAH would complex oxalate triggering dissolution and removal of CaOx nanocrystals. EXPERIMENTS: We successfully prepared nanoparticles of citrate ions with polyallylamine hydrochloride (CIT-PAH), PAH with oxalate (OX-PAH) and characterize them by Small Angle X ray Scattering (SAXS), Transmission Electron Microscopy (TEM), Dynamic Light Scattering (DLS) and NMR. Dissolution of CaOx nanocrystals in presence of CIT-PAH have been followed with Wide Angle Xray Scattering (WAXS), DLS and Confocal Raman Microscopy. Raman spectroscopy was used to study the dissolution of crystals in synthetic urine samples. The release of citrate from CIT-PAH was followed by diffusion NMR. Molecular dynamics (MD) simulations were carried out to study the interaction of CIT and OX ions with PAH. FINDINGS: CIT-PAH nanoparticles dissolves CaOx nanocrystals as shown by NMR, DLS, TEM and WAXS in water and by Raman spectroscopy in artificial human urine. WAXS and Raman show that the crystal structure of CaOx disappears in the presence of CIT-PAH. DLS shows that the time required for CaOX dissolution will depend on the concentration of CIT-PAH NPs. NMR proves that citrate ions are released from the CIT PAH NPs during CaOX dissolution, MD simulations showed that oxalates exhibit a stronger interaction for PAH than citrate, explaining the removal of oxalate ions and replacement of the citrate in the polymer nanoparticles.

Gold nanoparticles decorated with monosaccharides and sulfated ligands as potential modulators of the lysosomal enzyme N-acetylgalactosamine-6-sulfatase (GALNS).

N-Acetylgalactosamine-6-sulfatase (GALNS) is an enzyme whose deficiency is related to the lysosomal storage disease Morquio A. For the development of effective therapeutic approaches against this disease, the design of suitable enzyme enhancers (i.e. pharmacological chaperones) is fundamental. The natural substrates of GALNS are the glycosaminoglycans keratan sulfate and chondroitin 6-sulfate, which mainly display repeating units of sulfated carbohydrates. With a biomimetic approach, gold nanoparticles (AuNPs) decorated with simple monosaccharides, sulfated ligands (homoligand AuNPs), or both monosaccharides and sulfated ligands (mixed-ligand AuNPs) were designed here as multivalent inhibitors of GALNS. Among the homoligand AuNPs, the most effective inhibitors of GALNS activity are the ß-D-galactoside-coated AuNPs. In the case of mixed-ligand AuNPs, ß-D-galactosides/sulfated ligands do not show better inhibition than the ß-D-galactoside-coated AuNPs. However, a synergistic effect is observed for α-D-mannosides in a mixed-ligand coating with sulfated ligands that reduced IC50 by one order of magnitude with respect to the homoligand α-D-mannoside-coated AuNPs. SAXS experiments corroborated the association of GALNS with ß-D-galactoside AuNPs. These AuNPs are able to restore the enzyme activity by almost 2-fold after thermal denaturation, indicating a potential chaperoning activity towards GALNS. This information could be exploited for future development of nanomedicines for Morquio A. The recent implications of GALNS in cancer and neuropathic pain make these kinds of multivalent bionanomaterials of great interest towards multiple therapies.

Effects of model membranes on lysozyme amyloid aggregation.

The study of the interaction between lipid membranes and amyloidogenic peptides is a turning point for understanding the processes involving the cytotoxicity of peptides involved in neurodegenerative diseases. In this work, we perform an experimental study of model membrane-lysozyme interaction to understand how the formation of amyloid fibrils can be affected by the presence of polar and zwitterionic phospholipid molecules (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine [POPC] and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol [POPG]). The study was conducted above and below the critical micellar concentration (CMC) using dynamic light scattering (DLS), atomic force microscopy (AFM), UV-Vis spectrophotometry, and the quartz crystal microbalance (QCM). Our results show that the presence of phospholipids appears to be a factor favoring the formation of amyloid aggregates. Spectrophotometric and DLS data revealed that the quantity of ß -structure increases in the presence of POPG and POPC at different concentrations. The presence of POPG and POPC increases the speed of the nucleation process, without altering the overall structures of the fibrillar final products.

A study of cyanidin/alginate complexation: Influence of pH in assembly and chiral properties.

Cyanidin 3-O-glucoside (CND) is a frequently-used anthocyanin that has excellent antioxidant properties but a limited bioavailability in bloodstream. Complexation of CND with alginate can improve its therapeutic outcome. Here we have studied the complexation of CND with alginate under a range of pH values from 2.5 to 5. CND is positively charged at low pH, and becomes neutral, and then negatively charged as pH increases. CND/alginate complexation was studied by dynamic light scattering, transmission electron microscopy, small angle X-ray scattering, STEM, UV-Vis spectroscopy and circular dichroism (CD). CND/alginate complexes at pH 4.0 and 5.0 form chiral fibres with a fractal structure. At these pH values, CD spectra show very intense bands, which are inverted compared with free CND. Complexation at lower pH results in disordered polymer structures and CD spectra show the same features as for CND in solution. Molecular dynamics simulations suggest the formation of parallel CND dimers through complexation with alginate at pH 3.0, while at pH 4.0 CND dimers form in a cross like arrangement.

Integrative effects of morphology, silicification, and light on diatom vertical movements.

Diatoms represent the most abundant and diversified class of primary producers in present oceans; their distinctive trait is the ability to incorporate silicic acid in a silica outer shell called frustule. Numerous adaptative functions are ascribed to frustules, including the control of vertical movements through the water column; this indirectly determines cell access to fundamental resources such as light and nutrients, and favors diatom escape from predators. At the same time, light guides phototroph movements in the water column by affecting cell density (e.g., by modulating Si deposition in diatoms, vacuole volume, and/or solution). We investigated how the tremendous diversity in morphology and silicification that characterizes the frustule and the crucial role of light in diatom spatial distribution govern diatom sinking capacity. To test their integrative effects, we acclimated four diatoms distinguished by frustule traits (Chaetoceros muelleri, Conticribra weissflogii, Phaeodactylum tricornutum, and Cylindrotheca fusiformis) to different light conditions and evaluated their physiological performance in terms of growth, elemental composition, morphological changes, and their in vivo sinking capacity. What emerged from this study was that silicification, more than other morphological characteristics, controls species vertical movements, while a higher energy availability enhances cell floating independently from the silica content.

Small-Angle Neutron Scattering Reveals the Nanostructure of Liposomes with Embedded OprF Porins of Pseudomonas aeruginosa.

The use of liposomes as drug delivery systems emerged in the last decades in view of their capacity and versatility to deliver a variety of therapeutic agents. By means of small-angle neutron scattering (SANS), we performed a detailed characterization of liposomes containing outer membrane protein F (OprF), the main porin of the Pseudomonas aeruginosa bacterium outer membrane. These OprF-liposomes are the basis of a novel vaccine against this antibiotic-resistant bacterium, which is one of the main hospital-acquired pathogens and causes each year a significant number of deaths. SANS data were analyzed by a specific model we created to quantify the crucial information about the structure of the liposome containing OprF, including the lipid bilayer structure, the amount of protein in the lipid bilayer, the average protein localization, and the effect of the protein incorporation on the lipid bilayer. Quantification of such structural information is important to enhance the design of liposomal delivery systems for therapeutic applications.

Impact of PEGylation on the degradation and pore organization in mesoporous silica nanoparticles: A study of the inner mesoporous structure in physiologically relevant ionic conditions.

The degradation of mesoporous silica nanoparticles (MSNs) in the biological milieu due to silica hydrolysis plays a fundamental role for the delivery of encapsulated drugs and therapeutics. However, little is known on the evolution of the pore arrangement in the MSNs in biologically relevant conditions. Small Angle X-ray scattering (SAXS) studies were performed on unmodified and PEGylated MSNs with a MCM-48 pore structure and average sizes of 140 nm, exposed to simulated body fluid solution (SBF) at pH 7.4 for different time intervals from 30 min to 24 h. Experiments were performed with silica concentrations below, at and over 0.14 mg/mL, the saturation concentration of silica in water at physiological temperature. At silica concentrations of 1 mg/mL (oversaturation), unmodified MSNs show variation in interpore distances over 6 h exposure to SBF, remaining constant thereafter. A decrease in radius of gyration is observed over the same time. Mesoporosity and radius of gyration of unmodified MSNs remain then unchanged up to 24 h. PEGylated MSNs at 1 mg/mL concentration show a broader diffraction peak but no change in the position of the peak is observed following 24 h exposure to SBF. PEGylated MSNs at 0.01 mg/mL show no diffraction peaks already after 30 min exposure to SBF, while at 0.14 mg/mL a small diffraction peak is present after 30 min exposure but disappears after 1 h.

Investigation on a MMACHC mutant from cblC disease: The c.394C>T variant.

The cblC disease is an inborn disorder of the vitamin B12 (cobalamin, Cbl) metabolism characterized by methylmalonic aciduria and homocystinuria. The clinical consequences of this disease are devastating and, even when early treated with current therapies, the affected children manifest symptoms involving vision, growth, and learning. The illness is caused by mutations in the gene codifying for MMACHC, a 282aa protein that transports and transforms the different Cbl forms. Here we present data on the structural properties of the truncated protein p.R132X resulting from the c.394C > T mutation that, along with c.271dupA and c.331C > T, is among the most common mutations in cblC. Although missing part of the Cbl binding domain, p.R132X is associated to late-onset symptoms and, therefore, it is supposed to retain residual function. However, to our knowledge structural-functional studies on c.394C > T mutant aimed at verifying this hypothesis are still lacking. By using a biophysical approach including Circular Dichroism, fluorescence, Small Angle X-ray Scattering, and Molecular Dynamics, we show that the mutant protein MMACHC-R132X retains secondary structure elements and remains compact in solution, partly preserving its binding affinity for Cbl. Insights on the fragile stability of MMACHC-R132X-Cbl are provided.

SAXS Reveals the Stabilization Effects of Modified Sugars on Model Proteins.

Many proteins are usually not stable under different stresses, such as temperature and pH variations, mechanical stresses, high concentrations, and high saline contents, and their transport is always difficult, because they need to be maintained in a cold regime, which is costly and very challenging to achieve in remote areas of the world. For this reason, it is extremely important to find stabilizing agents that are able to preserve and protect proteins against denaturation. In the present work, we investigate, by extensively using synchrotron small-angle X-ray scattering experiments, the stabilization effect of five different sugar-derived compounds developed at ExtremoChem on two model proteins: myoglobin and insulin. The data analysis, based on a novel method that combines structural and thermodynamic features, has provided details about the physical-chemical processes that regulate the stability of these proteins in the presence of stabilizing compounds. The results clearly show that some modified sugars exert a greater stabilizing effect than others, being able to maintain the active forms of proteins at temperatures higher than those in which proteins, in the absence of stabilizers, reach denatured states.

Taurine Stabilizing Effect on Lysozyme.

Taurine is an important organic osmolyte in mammalian cells, and it weakens inflammation and oxidative stress mediated injuries in some diseases. Recently, taurine has been demonstrated to play a therapeutic role against neurodegenerative disorders, although its parallel involvement in several biochemical mechanisms makes not clear taurine specific role in these diseases. Furthermore, the stabilizing effect of this molecule in terms of protein stability is known, but not deeply investigated. In this work we explore by Circular Dichroism the stabilizing impact of taurine in lysozyme thermal denaturation and its influence in lysozyme aggregation into amyloid fibrils. Taurine even at low concentration modifies protein-protein interactions in lysozyme native state, as revealed by Small Angle X-ray Scattering experiments, and alters the amyloid aggregation pattern without completely inhibiting it, as confirmed by UV/Vis spectroscopy with Congo Red and by Atomic Force Microscopy. Evaluation of the cytotoxicities of the amyloid fibrils grown in presence or in absence of taurine is investigated on SH-SY5Y neuroblastoma cells.

Novel Core-Shell Polyamine Phosphate Nanoparticles Self-Assembled from PEGylated Poly(allylamine hydrochloride) with Low Toxicity and Increased In Vivo Circulation Time.

An approach for reducing toxicity and enhancing therapeutic potential of supramolecular polyamine phosphate nanoparticles (PANs) through PEGylation of polyamines before their assembly into nanoparticles is presented here. It is shown that the number of polyethylene glycol (PEG) chains for polyamine largely influence physico-chemical properties of PANs and their biological endpoints. Poly(allylamine hydrochloride) (PAH) are functionalized through carbodiimide chemistry with three ratios of PEG molecules per PAH chain: 0.1, 1, and 10. PEGylated PAH is then assembled into PANs by exposing the polymer to phosphate buffer solution. PANs decrease size and surface charge with increasing PEG ratios as evidenced by dynamic light scattering and zeta potential measurements, with the ten PEG/PAH ratio PANs having practically zero charge. Small angle X-ray scattering (SAXS) proves that PEG chains form a shell around a polyamine core, which is responsible for the screening of positive charges. MTT experiments show that the screening of amine groups decreases nanoparticle toxicity, with the lowest toxicity for the 10 PEG/PAH ratio. Fluorescence correlation spectroscopy (FCS) proves less interaction with proteins for PEGylated PANs. Positron emission tomography (PET) imaging of 18 F labelled PANs shows longer circulation time in healthy mice for PEGylated PANs than non-PEGylated ones.

The Histidine Phosphocarrier Kinase/Phosphorylase from Bacillus Subtilis Is an Oligomer in Solution with a High Thermal Stability.

The histidine phosphocarrier protein (HPr) kinase/phosphorylase (HPrK/P) modulates the phosphorylation state of the HPr protein, and it is involved in the use of carbon sources by Gram-positive bacteria. Its X-ray structure, as concluded from crystals of proteins from several species, is a hexamer; however, there are no studies about its conformational stability, and how its structure is modified by the pH. We have embarked on the conformational characterization of HPrK/P of Bacillus subtilis (bsHPrK/P) in solution by using several spectroscopic (namely, fluorescence and circular dichroism (CD)) and biophysical techniques (namely, small-angle X-ray-scattering (SAXS) and dynamic light-scattering (DLS)). bsHPrK/P was mainly a hexamer in solution at pH 7.0, in the presence of phosphate. The protein had a high conformational stability, with an apparent thermal denaturation midpoint of ~70 °C, at pH 7.0, as monitored by fluorescence and CD. The protein was very pH-sensitive, precipitated between pH 3.5 and 6.5; below pH 3.5, it had a molten-globule-like conformation; and it acquired a native-like structure in a narrow pH range (between pH 7.0 and 8.0). Guanidinium hydrochloride (GdmCl) denaturation occurred through an oligomeric intermediate. On the other hand, urea denaturation occurred as a single transition, in the range of concentrations between 1.8 and 18 µM, as detected by far-UV CD and fluorescence.

The dimer-monomer equilibrium of SARS-CoV-2 main protease is affected by small molecule inhibitors.

The maturation of coronavirus SARS-CoV-2, which is the etiological agent at the origin of the COVID-19 pandemic, requires a main protease Mpro to cleave the virus-encoded polyproteins. Despite a wealth of experimental information already available, there is wide disagreement about the Mpro monomer-dimer equilibrium dissociation constant. Since the functional unit of Mpro is a homodimer, the detailed knowledge of the thermodynamics of this equilibrium is a key piece of information for possible therapeutic intervention, with small molecules interfering with dimerization being potential broad-spectrum antiviral drug leads. In the present study, we exploit Small Angle X-ray Scattering (SAXS) to investigate the structural features of SARS-CoV-2 Mpro in solution as a function of protein concentration and temperature. A detailed thermodynamic picture of the monomer-dimer equilibrium is derived, together with the temperature-dependent value of the dissociation constant. SAXS is also used to study how the Mpro dissociation process is affected by small inhibitors selected by virtual screening. We find that these inhibitors affect dimerization and enzymatic activity to a different extent and sometimes in an opposite way, likely due to the different molecular mechanisms underlying the two processes. The Mpro residues that emerge as key to optimize both dissociation and enzymatic activity inhibition are discussed.

Comprehensive Structural and Thermodynamic Analysis of Prefibrillar WT α-Synuclein and Its G51D, E46K, and A53T Mutants by a Combination of Small-Angle X-ray Scattering and Variational Bayesian Weighting.

The in solution synchrotron small-angle X-ray scattering SAXS technique has been used to investigate an intrinsically disordered protein (IDP) related to Parkinson's disease, the α-synuclein (α-syn), in prefibrillar diluted conditions. SAXS experiments have been performed as a function of temperature and concentration on the wild type (WT) and on the three pathogenic mutants G51D, E46K, and A53T. To identify the conformers that populate WT α-syn and the pathogenic mutants in prefibrillar conditions, scattering data have been analyzed by a new variational bayesian weighting method (VBWSAS) based on an ensemble of conformers, which includes unfolded monomers, trimers, and tetramers, both in helical-rich and strand-rich forms. The developed VBWSAS method uses a thermodynamic scheme to account for temperature and concentration effects and considers long-range protein-protein interactions in the framework of the random phase approximation. The global analysis of the whole set of data indicates that WT α-syn is mostly present as unfolded monomers and trimers (helical-rich trimers at low T and strand-rich trimers at high T), but not tetramers, as previously derived by several studies. On the contrary, different conformer combinations characterize mutants. In the α-syn G51D mutant, the most abundant aggregates at all the temperatures are strand-rich tetramers. Strand-rich tetramers are also the predominant forms in the A53T mutant, but their weight decreases with temperature. Only monomeric conformers, with a preference for the ones with the smallest sizes, are present in the E46K mutant. The derived conformational behavior then suggests a different availability of species prone to aggregate, depending on mutation, temperature, and concentration and accounting for the different neurotoxicity of α-syn variants. Indeed, this approach may be of pivotal importance to describe conformational and aggregational properties of other IDPs.

Gelling without Structuring: A SAXS Study of the Interactions among DNA Nanostars.

We evaluate, by means of synchrotron small-angle X-ray scattering, the shape and mutual interactions of DNA tetravalent nanostars as a function of temperature in both the gas-like state and across the gel transition. To this end, we calculate the form factor from coarse-grained molecular dynamics simulations with a novel method that includes hydration effects; we approximate the radial interaction of DNA nanostars as a hard-sphere potential complemented by a repulsive and an attractive Yukawa term; and we predict the structure factors by exploiting the perturbative random phase approximation of the Percus-Yevick equation. Our approach enables us to fit all the data by selecting the particle radius and the width and amplitude of the attractive potential as free parameters. We determine the evolution of the structure factor across gelation and detect subtle changes of the effective interparticle interactions, that we associate to the temperature and concentration dependence of the particle size. Despite the approximations, the approach here adopted offers new detailed insights into the structure and interparticle interactions of this fascinating system.

Trehalose Effect on the Aggregation of Model Proteins into Amyloid Fibrils.

Protein aggregation into amyloid fibrils is a phenomenon that attracts attention from a wide and composite part of the scientific community. Indeed, the presence of mature fibrils is associated with several neurodegenerative diseases, and in addition these supramolecular aggregates are considered promising self-assembling nanomaterials. In this framework, investigation on the effect of cosolutes on protein propensity to aggregate into fibrils is receiving growing interest, and new insights on this aspect might represent valuable steps towards comprehension of highly complex biological processes. In this work we studied the influence exerted by the osmolyte trehalose on fibrillation of two model proteins, that is, lysozyme and insulin, investigated during concomitant variation of the solution ionic strength due to NaCl. In order to monitor both secondary structures and the overall tridimensional conformations, we have performed UV spectroscopy measurements with Congo Red, Circular Dichroism, and synchrotron Small Angle X-ray Scattering. For both proteins we describe the effect of trehalose in changing the fibrillation pattern and, as main result, we observe that ionic strength in solution is a key factor in determining trehalose efficiency in slowing down or blocking protein fibrillation. Ionic strength reveals to be a competitive element with respect to trehalose, being able to counteract its inhibiting effects toward amyloidogenesis. Reported data highlight the importance of combining studies carried out on cosolutes with valuation of other physiological parameters that may affect the aggregation process. Also, the obtained experimental results allow to hypothesize a plausible mechanism adopted by the osmolyte to preserve protein surface and prevent protein fibrillation.

K vs. Na Effects on the Self-Assembly of Guanosine 5'-Monophosphate: A Solution SAXS Structural Study+.

The hierarchical process of guanosine (G) self-assembly, leading in aqueous solution and in the presence of metal cations to the formation of G-quadruplexes, represents an intriguing topic both for the biological correlation with telomerase activity and for the nano-technological applications, as demonstrated by the current measured in a quadruplex wire 100 nm long. Similar to G-rich DNA sequences and G-oligonucleotides, the guanosine 5'-monophosphate (GMP) self-aggregates in water to form quadruplexes. However, due to the absence of a covalent axial backbone, this system can be very useful to understand the chemical-physical conditions that govern the guanosine supramolecular aggregation. We have then investigated by in-solution Synchrotron Small Angle X-ray Scattering technique the role of different cations in promoting the quadruplex formation as a function of concentration and temperature. Results show how potassium, with its peculiar biological traits, favours the G-quadruplex elongation process in respect to other cations (Na + , but also NH 4 + and Li + ), determining the longest particles in solution. Moreover, the formation and the elongation of G-quadruplexes have been demonstrated to be controlled by both GMP concentration and excess cation content, even if they specifically contribute to these processes in different ways. The occurrence of condensed liquid crystalline phases was also detected, proving that excess cations play also unspecific effects on the effective charges on the G-quadruplex surface.

Biophysical Characterization of Membrane Phase Transition Profiles for the Discrimination of Outer Membrane Vesicles (OMVs) From Escherichia coli Grown at Different Temperatures.

Dynamic Light Scattering (DLS), Small Angle X-ray Scattering (SAXS) and Transmission Electron Microscopy (TEM) are physical techniques widely employed to characterize the morphology and the structure of vesicles such as liposomes or human extracellular vesicles (exosomes). Bacterial extracellular vesicles are similar in size to human exosomes, although their function and membrane properties have not been elucidated in such detail as in the case of exosomes. Here, we applied the above cited techniques, in synergy with the thermotropic characterization of the vesicles lipid membrane using a turbidimetric technique to the study of vesicles produced by Gram-negative bacteria (Outer Membrane Vesicles, OMVs) grown at different temperatures. This study demonstrated that our combined approach is useful to discriminate vesicles of different origin or coming from bacteria cultured under different experimental conditions. We envisage that in a near future the techniques employed in our work will be further implemented to discriminate complex mixtures of bacterial vesicles, thus showing great promises for biomedical or diagnostic applications.

Dynamics of the intrinsically disordered inhibitor IF7 of glutamine synthetase in isolation and in complex with its partner.

Glutamine synthetase (GS) catalyzes the ATP-dependent formation of glutamine from glutamate and ammonia. The activity of Synechocystis sp. PCC 6803 GS is regulated, among other mechanisms, by protein-protein interactions with a 65-residue-long, intrinsically disordered protein (IDP), named IF7. IDPs explore diverse conformations in their free states and, in some cases, in their molecular complexes. We used both nuclear magnetic resonance (NMR) at 11.7 T and small angle X-ray scattering (SAXS) to study the size and the dynamics in the picoseconds-to-nanosecond (ps-ns) timescale of: (i) isolated IF7; and (ii) the IF7/GS complex. Our SAXS findings, together with MD results, show: (i) some of the possible IF7 structures in solution; and, (ii) that the presence of IF7 affected the structure of GS in solution. The joint use of SAXS and NMR shows that movements of each amino acid of IF7 were uncorrelated with those of its neighbors. Residues of IF7 with the largest values of the relaxation rates (R1, R2 and ηxy), in the free and bound species, were mainly clustered around: (i) the C terminus of the protein; and (ii) Ala30. These residues, together with Arg8 (which is a hot-spot residue in the interaction with GS), had a restricted mobility in the presence of GS. The C-terminal region, which appeared more compact in our MD simulations of isolated IF7, seemed to be involved in non-native contacts with GS that help in the binding between the two macromolecules.

Metallo-responsive self-assembly of lipophilic guanines in hydrocarbon solvents: a systematic SAXS structural characterization.

Lipophilic guanines (LipoGs) in aprotic solvents undergo different self-assembly processes based on different H-bonded motifs. Cylindrical nanotubes made by π-π stacked guanine tetramers (G-quadruplexes) and flat, tape-like aggregates (G-ribbons) have been observed depending on the presence of alkali metal ions. To obtain information on the structural properties and stability of these LipoG aggregates, Small-Angle X-ray Scattering (SAXS) experiments have been performed in dodecane, both in the presence and in the absence of potassium ions. As a result, the occurrence of the two different metallo-responsive architectures (nanoribbons or columnar nanotubes) was confirmed and we reported here for the first time a systematic study on the dependence of the aggregate properties on composition, temperature and molecular unit structure. Even if dodecane was selected to favour LipoG solubility, a strong tendency to self-organize into ordered lyotropic phases was indeed detected.