Kosmotrope-like hydration behavior of polyethylene glycol from microcalorimetry and binding isotherm measurements.

Polyethylene glycol (PEG) at various molecular weights (MWs) has been regarded as a wonder molecule in biomedical applications. For instance, PEG serves as a unique moiety for pegylation of "biobetter" drug development, PEG provides controlled-release and preserved activity of biologics, and PEG modified surface works as an antibiofouling surface. The primary characteristics of PEG molecules used in relevant applications have been attributed mainly to the hydration behavior in aqueous solutions. However, the effects on the solvation of solutes in solution caused by presenting PEG molecules as a cosolvent, as well as the thermodynamics aspect of the hydration behavior of PEG in solution, have not been well documented. The solvation behavior of solutes, such as protein, with PEG as a cosolvent, indicates the success of PEG applications, such as biofouling and controlled release. In this investigation, we examined the effects of a buffer solution containing PEG molecules on the solution behavior of solute and the interactions between solid surfaces with solutes. We adapted the study by selecting a lysozyme as a solute in a buffer solution with either ammonium sulfate (kosmotrope) or sodium chloride (chaotrope) and anionic resin (SP-Sepharose) as solid surfaces. The experiments primarily involved binding equilibrium measurements and thermodynamics analysis. The results revealed that, in both saline buffers, adding PEG increases the binding affinity between the lysozyme and the resin, similar to kosmotropic salt in the examined salt concentrations. The thermodynamics analyses involving microcalorimetric measurements show that the bindings are mainly driven by enthalpy, indicating that electrostatic interaction was the primary binding force under these experimental conditions. The variations of the enthalpy and entropy of the binding thermodynamics when adding PEG to different salt types in the buffer solution showed opposite behavior, and the results support the concept of kosmotrope-like behavior of PEG. The equilibrium and thermodynamics data demonstrate that PEG has a kosmotrope-like hydration behavior, and the extent of kosmotrope-like behavior depends on the molecular weight of PEG with the outcomes of various molecular weights of PEG being added to the binding solution. The results of this study provide essential knowledge for PEG as an additive (or cosolvent) in various research applications.

Coupling molecular dynamics simulations with experiments for the rational design of indolicidin-analogous antimicrobial peptides.

Antimicrobial peptides (AMPs) have attracted much interest in recent years because of their potential use as new-generation antibiotics. Indolicidin (IL) is a 13-residue cationic AMP that is effective against a broad spectrum of bacteria, fungi, and even viruses. Unfortunately, its high hemolytic activity retards its clinical applications. In this study, we adopted molecular dynamics (MD) simulations as an aid toward the rational design of IL analogues exhibiting high antimicrobial activity but low hemolysis. We employed long-timescale, multi-trajectory all-atom MD simulations to investigate the interactions of the peptide IL with model membranes. The lipid bilayer formed by the zwitterionic 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) was chosen as the model erythrocyte membrane; lipid bilayers formed from a mixture of POPC and the negatively charged 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol were chosen to model bacterial membranes. MD simulations with a total simulation time of up to 4 micros revealed the mechanisms of the processes of IL adsorption onto and insertion into the membranes. The packing order of these lipid bilayers presumably correlated to the membrane stability upon IL adsorption and insertion. We used the degree of local membrane thinning and the reduction in the order parameter of the acyl chains of the lipids to characterize the membrane stability. The order of the mixed 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol/POPC lipid bilayer reduced significantly upon the adsorption of IL. On the other hand, although the order of the pure-POPC lipid bilayer was perturbed slightly during the adsorption stage, the value was reduced more dramatically upon the insertion of IL into the membrane's hydrophobic region. The results imply that enhancing IL adsorption on the microbial membrane may amplify its antimicrobial activity, while the degree of hemolysis may be reduced through inhibition of IL insertion into the hydrophobic region of the erythrocyte membrane. In addition, through simulations, we identified the amino acids that are most responsible for the adsorption onto or insertion into the two model membranes. Positive charges are critical to the peptide's adsorption, whereas the presence of hydrophobic Trp8 and Trp9 leads to its deeper insertion. Combining the hypothetical relationships between the membrane disordering and the antimicrobial and hemolytical activities with the simulated results, we designed three new IL-analogous peptides: IL-K7 (Pro7-->Lys), IL-F89 (Trp8 and Trp9-->Phe), and IL-K7F89 (Pro7-->Lys; Trp8 and Trp9-->Phe). The hemolytic activity of IL-F89 is considerably lower than that of IL, whereas the antimicrobial activity of IL-K7 is greatly enhanced. In particular, the de novo peptide IL-K7F89 exhibits higher antimicrobial activity against Escherichia coli; its hemolytic activity decreased to only 10% of that of IL. Our simulated and experimental results correlated well. This approach-coupling MD simulations with experimental design-is a useful strategy toward the rational design of AMPs for potential therapeutic use.

Comparison of ultrasound and electromagnetic field effects on osteoblast growth.

This study compares the mechanisms of ultrasound (US) on osteoblast proliferation with those of pulsed electromagnetic field (PEMF), by different signal transduction pathway inhibitors. The cells were stimulated for 15 min under US or for 2 h under PEMF exposure. Twenty-four h after the beginning of stimulation, the cells were harvested and used for mitochondrial activity test (MTT) analysis. The results showed that there are different transduction pathways for US and PEMF stimulation that lead to an upgrade of osteoblast proliferation, although their pathways all lead to an increase in cytocolic Ca2+ and activation of calmodulin. These findings offer a biochemical mechanism to support the process of ultrasound and PEMF-induced enhanced healing of bone fractures.