Mapping Free-Energy Landscapes to Decipher Amphiphilic Peptide Multistep Self-Assembly.

Correction to "Mapping Free-Energy Landscapes to Decipher Amphiphilic Peptide Multistep Self-Assembly".

Molecular mechanisms governing peptide nanodisc assembly and stability.

Apolipoprotein A-I mimetic 4F, an 18-residue amphipathic α-helix, can self-assemble with lipids to form peptide nanodiscs, yet the molecular determinants governing their assembly and stability remain poorly understood. Here, using coarse-grained molecular dynamics (CG-MD), we capture the de novo formation of 4F nanodiscs with DMPC and reveal a multistep assembly pathway involving nucleation, fusion, and ellipse-to-disc maturation. All-atom back-mapping shows that the nanodisc rim is structurally heterogeneous and stabilized by aromatic-acyl interactions, Lys headgroup anchoring, and inter-peptide electrostatic contacts. Lipid composition and temperature critically regulate nanodisc integrity: DMPC supports continuous peptide belts and long-term stability, whereas DPPC below its main phase transition temperature suppresses fusion and yields fragmented, non-uniform rims. These findings validate the ability of CG-MD to resolve nanodisc assembly mechanisms. Experimental measurements corroborate the simulations, demonstrating that 4F nanodiscs exhibit lower thermal resilience than MSP nanodiscs while retaining structural integrity at moderate temperatures. As a functional benchmark, MSP nanodiscs suppress the amyloid-binding thioflavin-T fluorescence signal associated with Aβ (1-40) fibrillar assembly, consistent with our previously reported findings for 4F nanodiscs and supporting the ability of amphipathic nanodisc rims to delay Aβ (1-40) aggregation. Together, these results establish a mechanistic framework and design principles for single-helix peptide nanodiscs and delineate the conditions under which they converge with or diverge from MSP-based scaffolds.

Temperature-Triggered Nanoscale Morphological Transitions in a Synthetic Intrinsically Disordered Protein.

Peptide self-assembly and liquid-liquid phase separation (LLPS), often mediated by intrinsically disordered regions (IDRs), are natural mechanisms that translate protein molecular features into complex nano- and mesoscale architectures. Although the thermodynamics and kinetics of these processes are well understood, synthetic materials integrating both functionalities remain rare. Inspired by the conserved IDR-assembly domain (AD) architecture of amyloidogenic proteins, we hypothesized that modular recombinant constructs combining LLPS-capable IDRs with β-sheet-forming ADs could generate materials with tunable structural properties. To test this, we engineered a library of elastin-like polypeptides (ELPs) fused to amphiphilic anionic or cationic amyloidogenic peptides, enabling systematic investigation of how sequence parameters─including ELP length, AD charge, and hydrophilicity─and environmental conditions, including temperature, pH, and salt concentration, influence material behavior. Our results reveal links between molecular design and emergent multiscale structures, including micelles and vesicles embedded within coacervates. This work provides a framework for designing hybrid proteins coupling LLPS and self-assembly.

Hydrophobic Effect in Self-Assembly of Metal-Bound Nucleotides: A Structural Study of Iron-Nucleotide Ternary Complexes

The influence of hydrophobic microenvironments on the self-assembly and structural dynamics of nucleotides remains fundamentally elusive at the molecular level. Here, we establish a robust biomimetic platform utilizing a Fe(III)-nucleoside monophosphate-TPA ternary complex, where the bulky TPA ligand serves as a tailored “hydrophobic umbrella”. High-resolution crystallographic investigations reveal that this localized confinement drives 5′-coordinated nucleotides to spontaneously assemble into discrete biological architectures, such as A-motifs and i-motifs. Translating these solid-state findings to aqueous solution, circular dichroism (CD) spectroscopy provides experimental validation of the supramolecular mechanism. Solvent-dependent titrations reveal a sharp, nonlinear sigmoidal phase transition, explicitly demonstrating that the TPA scaffold actively overrides the chaotic, isodesmic π–π stacking characteristic of free nucleotides. Instead, the rigid chiral locking of the assembly is cooperatively driven by the massive thermodynamic compressive forces of hydrophobic exclusion. Comprehensive computational modeling, including DFT, NCI, and Hirshfeld surface analyses, unequivocally maps and quantifies this driving force, isolating the explicit energetic stabilization provided by the dispersive hydrophobic shield. Furthermore, we resolve the paradoxical role of solvent within these confined cavities: while bulk water is aggressively repelled, a specifically localized primary hydration shell is strictly preserved. This inner hydration layer acts as a master thermodynamic determinant, mechanically locking the precise biomimetic sugar ring puckering (C3′-endo/C2′-exo) requisite for assembly. Collectively, these integrated structural, spectroscopic, and thermodynamic insights illuminate the physicochemical forces governing nucleic acid organization in restricted spaces, offering a definitive supramolecular model for biological molecular recognition.

Predictive Amphiphile Design Enables Control of Self-Assembled Nanoparticle-Cell Membrane Interactions.

Despite decades of development, the clinical translation of amphiphilic self-assembled nanoparticles remains limited by the lack of a predictive molecular design framework, owing to the complex, dynamic behavior of nanoparticles and cell membranes. Here, we introduce a building-block design strategy focused on amphiphile monomers rather than assembled nanoparticles, a critical distinction because nanoparticle morphology is not a fixed design parameter. This molecular framework links amphiphile chemical structure to programmable cellular interactions. Through predictive variation of headgroup and tail chemistries, we demonstrate that amphiphile design dictates nanoparticle fate at cell membranes. Fully fluorinated nanoparticles remain intact and adsorb onto the membrane surface, hydrocarbon nanoparticles rapidly dissociate and fuse, whereas semi-fluorinated nanoparticles exhibit tunable kinetic delays, enabling programmable responses such as stability at rigid interfaces but dissociation at fluid membranes. Crucially, we identify the free energy of transfer (ΔGtransfer)-the energetic difference between extracting an amphiphile from the nanoparticle and from the cell membrane-as a unified thermodynamic descriptor that accurately predicts behavior across simulations and experiments without requiring full-system modeling. Collectively, this predictive, chemistry-driven framework shifts the design paradigm from nanoparticle morphology to amphiphile chemistry, enabling the rational engineering of self-assembled nanocarriers for targeted drug delivery and responsive drug release applications.

Electrolyte-Controlled Self-Assembly of Water-Soluble Perylene Bisimides: A Kinetic and Stimulated Raman Study

Perylene bisimides (PBIs) are prime building blocks for functional supramolecular architectures, yet mapping their salt-induced self-assembly kinetics under physiological conditions remains challenging, due to rapid aggregation and overwhelming fluorescence. Here, we combine fluorescence stopped-flow kinetics and stimulated Raman scattering (SRS) spectroscopy to decipher the electrolyte-controlled polymerization of a spermine-functionalized PBI. We found that self-assembly obeys an anticooperative K2–K pathway, where nucleation yields rapid and salt-independent dimers. In contrast, polymer growth is strongly salt-dependent, with polymer dissociation rates decreasing as NaCl concentration increases. Crucially, chloride ions selectively stabilize the growing assemblies by drastically suppressing the monomer release, establishing a Michaelis–Menten-type saturation profile. To link these kinetic trajectories with molecular-scale structural rearrangements, SRS is used to effectively suppress the fluorescence background, revealing that the relative intensity of low-frequency Raman modes, associated with high-mass displacement and structurally delocalized in nature, are drastically reduced upon aggregation. This vibrational suppression underscores the role of packing constraints and excitonic pressure within the cofacial H-aggregates. By directly bridging macroscale kinetic pathways with microscopic vibrational dynamics, this work offers a generalizable framework to rationally design and tune water-soluble supramolecular materials for biomedical and optoelectronic applications.