Melittin is a prototype of the ubiquitous antimicrobial peptides that induce pores in membranes. It is commonly used as a molecular device for membrane permeabilization. Even at concentrations in the nanomolar range, melittin can induce transient pores that allow transmembrane conduction of atomic ions but not leakage of glucose or larger molecules. At micromolar concentrations, melittin induces stable pores allowing transmembrane leakage of molecules up to tens of kilodaltons, corresponding to its antimicrobial activities. Despite extensive studies, aspects of the molecular mechanism for pore formation remain unclear. To clarify the mechanism, one must know the states of the melittin-bound membrane before and after the process. By correlating experiments using giant unilamellar vesicles with those of peptide-lipid multilayers, we found that melittin bound on the vesicle translocated and redistributed to both sides of the membrane before the formation of stable pores. Furthermore, stable pores are formed only above a critical peptide-to-lipid ratio. The initial states for transient and stable pores are different, which implies different mechanisms at low and high peptide concentrations. To determine the lipidic structure of the pore, the pores in peptide–lipid multilayers were induced to form a lattice and examined by anomalous X-ray diffraction. The electron density distribution of lipid labels shows that the pore is formed by merging of two interfaces through a hole. The molecular property of melittin is such that it adsorbs strongly to the bilayer interface. Pore formation can be viewed as the bilayer adopting a lipid configuration to accommodate its excessive interfacial area.

Liposome development is of great interest owing to increasing requirements for efficient drug carriers. The structural features and thermal stability of such liposomes are crucial in drug transport and delivery. Reported here are the results of the structural characterization of PEGylated liposomes via small- and wide-angle X-ray scattering and an asymmetric flow field-flow fractionation (AF4) system coupled with differential refractive-index detection, multi-angle light scattering (MALS) and dynamic light scattering. This integrated analysis of the exemplar PEGylated liposome formed from hydrogenated soy phosphatidylcholine (HSPC) with the addition of cholesterol reveals an average hydrodynamic radius (R _h) of 52 nm with 10% polydispersity, a comparable radius of gyration (R _g) and a major liposome particle mass of 118 kDa. The local bilayer structure of the liposome is found to have asymmetric electronic density profiles in the inner and outer leaflets, sandwiched by two PEGylated outer layers ca 5 nm thick. Cholesterol was found to effectively intervene in lipid chain packing, resulting in the thickening of the liposome bilayer, an increase in the area per lipid and an increase in liposome size, especially in the fluid phase of the liposome. These cholesterol effects show signs of saturation at cholesterol concentrations above ca 1:5 cholesterol:lipid molar ratio.

Membrane active peptides directly target membranes rather than receptor proteins and perform their biological functions via cooperative action. To understand the mechanism of peptide–membrane interactions, biophysical characterization of the whole interaction process is essential. The characteristics determined by biophysical methods are briefly summarized and the limitations of these techniques discussed. Circular dichroism (CD), isothermal titration calorimetry (ITC), and fluorescence have been used to determine the binding isotherms of peptide binding to membranes in solution. Lamellar X-ray diffraction (LXD) and small single X-ray scattering (SAXS) have been used to probe peptide-induced thinning of membranes on a substrate and in solution, respectively. The aspiration method has been applied to monitor the area expansion induced by peptide binding to single giant unilamellar vesicles (GUVs). Oriented circular dichroism (OCD) has been used to detect orientation changes of peptides in membranes. The inner water columns of peptide-induced pores in membranes were detected by neutron scattering (NS), and pore structures were reconstructed using anomalous X-ray diffraction. Finally, the time evolution of pore formation induced by peptides binding to single GUVs was monitored by the aspiration method.

α-crystallin is a major structural protein in the eye lenses of vertebrates that is composed of two relative subunits, αA and αB crystallin, which function in maintaining lens transparency. As a member of the small heat-shock protein family (sHsp), α-crystallin exhibits chaperone-like activity to prevent the misfolding or aggregation of critical proteins in the lens, which is associated with cataract disease. In this study, high-purity αA and αB crystallin proteins were expressed from E. coli and purified by affinity and size-exclusion chromatography. The size-exclusion chromatography experiment showed that both αA and αB crystallins exhibited oligomeric complexes in solution. Here, we present the structural characteristics of α-crystallin proteins from low to high temperature by combining circular dichroism (CD) and small-angle X-ray scattering (SAXS). Not only the CD data, but also SAXS data show that α-crystallin proteins exhibit transition behavior on conformation with temperature increasing. Although their protein sequences are highly conserved, the analysis of their thermal stability showed different properties in αA and αB crystallin. In this study, taken together, the data discussed were provided to demonstrate more insights into the chaperone-like activity of α-crystallin proteins.

Antibiotic drug resistance is a serious issue for the treatment of bacterial infection. Understanding the resistance to antibiotics is a key issue for developing new drugs. We used penicillin and sulbactam as model antibiotics to study their interaction with model membranes. Cholesterol was used to target the membrane for comparison with the well‐known insertion model. Lamellar X‐ray diffraction (LXD) was used to determine membrane thickness using successive drug‐to‐lipid molar ratios. The aspiration method for a single giant unilamellar vesicle (GUV) was used to monitor the kinetic binding process of antibiotic–membrane interactions in an aqueous solution. Both penicillin and sulbactam are found positioned outside the model membrane, while cholesterol inserts perpendicularly into the hydrophobic region of the membrane in aqueous solution. This result provides structural insights for understanding the antibiotic–membrane interaction and the mechanism of antibiotics.