Observation of Mechanical and Kinetic Distinctions between Individual Isoleucine and Arginine Residues in a Peptide Dissociating from a Model Lipid Bilayer

Is the Bilayer Insertion of the Peptide KALP23 Under Kinetic or Thermodynamic Control?

Bilayer Elastic Properties Modulate Interface Energetics of Transmembrane β-Barrels.

Assembly energetics of transmembrane proteins occurs through the orchestrated molecular interplay of the protein's membrane partitioning free energy and the mechanical and physicochemical properties of the bilayer. While amphipathic residues help accommodate diametrically opposite forces and the steep energy gradient at the membrane-water interface, the magnitude to which the intrinsic curvature and elastic energy of the bilayer regulates protein thermodynamics at the interface remains poorly understood. Here, using PagP as a model β-barrel, we compare the per-residue energetics of an interface site across membranes with(out) distinct curvature stress, lipid packing, and headgroup chemistry. In vesicles, the measured interface energetics correlates well with known free energy scales. Interestingly, unlike vesicles, we show that planar bicelles impose fundamentally distinct membrane-protein coupling regimes: (i) PagP experiences altered folding with a stabilized membrane-associated intermediate and (ii) side chain transfer free energies are restructured in a residue-specific manner. Additionally, the energetics are unaffected by the presence of nonlamellar lipids. Our results reveal an unexpected dependence of partitioning energetics on both bilayer elastic energy and lipid packing defects, demonstrating how subtle changes in membrane physical properties could redirect protein folding pathways and thermodynamics. Our findings underscore the need for caution when extrapolating side chain transfer free energies from simplified model membranes to crowded, compositionally complex in vivo environments.

Distinct Molten-Globule Conformations in an Interacting Protein Domain Resolved by 19F NMR of Fluoroproline Residues.

This study illustrates how subtle residue-localized conformational bias can affect the overall protein conformational dynamics influencing protein-protein interactions that are important for cellular functions and related to diseases.

Lipid-Shell PARCH: A Physically Motivated Scale for Transmembrane Residue Hydropathy

Understanding how amino acid residues partition from water into lipid bilayers is fundamental to membrane protein folding, stability, and function. Existing hydrophobicity scales derive from isolated protein systems, organic solvent approximations, or computationally expensive free energy methods, each with significant limitations in capturing the full thermodynamic and topographic complexity of membrane protein environments. Here we present a lipid-shell modification to the protocol for assigning a residue’s character on a hydropathy (PARCH) scale, in which the protein’s first hydration shell is enclosed by a lipid boundary layer during thermal annealing. This modification preserves the core PARCH methodology─evaluating water retention around residues as a function of temperature─while imposing the chemical potential boundary condition appropriate for membrane-embedded proteins. Using the OmpLA host–guest system, we compute PARCH values (PVs) that align with the experimental water-to-bilayer transfer free energy scale of Moon and Fleming (PNAS, 108, 10174–10177, 2011) without calibration to that data. We further demonstrate that depth-dependent PV profiles for arginine and leucine mirror experimentally measured partition energies across six membrane positions. Finally, PVs for a tandem arginine double mutant reveal a per-residue cooperative hydration redistribution that provides microscopic insight into thermodynamic cooperativity previously measured experimentally. Together, these results establish the lipid-shell PARCH modification as a computationally affordable and physically meaningful approach to quantifying membrane hydropathy that is sensitive to residue identity, membrane depth, and cooperative hydration among tandem charged residues.

Benchmarking AI-generated structural ensembles of membrane proteins against physics-based modelling

It is demonstrated that BioEmu can generate plausible conformational ensembles for relatively large, six-and seven-pass membrane proteins, sampling rare states at a fraction of the computational cost of conventional MD simulations, suggesting that AI-based ensemble generation could provide an accessible approach for exploring membrane protein dynamics and complement conventional molecular modelling approaches.

Modulation of the phospholipid dynamics and bilayer water accessibility with the membrane-associated nucleation of β-amyloid (Aβ) peptides.

Membrane disruption along the amyloidogenic aggregation of β-amyloid (Aβ) peptides is considered a molecular mechanism for the Aβ-induced cell toxicity and death. Yet, the underlying structural basis for the harmful Aβ-membrane interactions that lead to disruption remains poorly understood. We have been utilizing solid-state nuclear magnetic resonance (ssNMR) spectroscopy to explore the intermediate states of membrane-associated Aβ aggregation, as well as their roles in membrane disruption process. Aligning with this general objective, complementary quantitative ssNMR spectroscopy focusing on the modulation of phospholipid dynamics in membrane bilayers will provide useful insight about how these intermediate states influence the physicochemical properties and architecture of membranes. In the current work, we systematically investigate how specific molecular motions of phospholipids change in the presence of membrane-associated 40- and 42-residue Aβ isoforms, within the time frame of nucleation processes. Physicochemical parameters, including the lipid headgroup and lateral diffusive motion correlation time, the lipid alkyl chain to headgroup 1H1H cross relaxation rate, and the H2O-assisted 1H13C cross polarization rate to lipid alkyl 13Cs, were monitored by various ssNMR spectroscopic approaches. The outcomes suggest a general rigidification of bilayers upon instant Aβ-bilayer interactions across different bilayer phospholipid compositions and Aβ isoforms, accompanied by increase of water accessibility to bilayer interiors. These observations, together with the knowledge of molecular structural evolution of Aβ within the same time frame, help to establish a molecular-level, schematic explanation of the membrane-associated Aβ nucleation process.