Enhancing colloidal stability of nanodiamond via surface modification with dendritic molecules for optical sensing in physiological environments.

Pre-treatment of diamond surface in low-temperature plasma for oxygenation and in acids for carboxylation was hypothesized to promote the branching density of the hyperbranched glycidol polymer. This was expected to increase the homogeneity of the branching level and suppress interactions with proteins. As a result, composite nanodiamonds with reduced hydrodynamic diameters that are maintained in physiological environments were anticipated. Surfaces of 140-nm-sized nanodiamonds were functionalized with oxygen and carboxyl groups for grafting of hyperbranched dendritic polyglycerol via anionic ring-opening polymerization of glycidol. The modification was verified with Fourier-transform infrared spectroscopy and X-ray photoelectron spectroscopy. Dynamic light scattering investigated colloidal stability in pH-diverse (2-12) solutions, concentrated phosphate-buffered saline, and cell culture media. Thermogravimetric analysis of nanodiamonds-protein incubations examined non-specific binding. Fluorescence emission was tested across pH conditions. Molecular dynamics simulations modeled interparticle interactions in ionic solutions. The hyperbranched polyglycerol grafting increased colloidal stability of nanodiamonds across diverse pH, high ionic media like 10 × concentrated phosphate-buffered saline, and physiological media like serum and cell culture medium. The hyperbranched polyglycerol suppressed non-specific protein adsorption while maintaining intensive fluorescence of nanodiamonds regardless of pH. Molecular modelling indicated reduced interparticle interactions in ionic solutions correlating with the improved colloidal stability.

A Novel Cryptic Clostridial Peptide That Kills Bacteria by a Cell Membrane Permeabilization Mechanism.

This work reports detailed characteristics of the antimicrobial peptide Intestinalin (P30), which is derived from the LysC enzyme of Clostridium intestinale strain URNW. The peptide shows a broader antibacterial spectrum than the parental enzyme, showing potent antimicrobial activity against clinical strains of Gram-positive staphylococci and Gram-negative pathogens and causing between 3.04 ± 0.12 log kill for Pseudomonas aeruginosa PAO1 and 7.10 ± 0.05 log kill for multidrug-resistant Acinetobacter baumannii KPD 581 at a 5 µM concentration. Moreover, Intestinalin (P30) prevents biofilm formation and destroys 24-h and 72-h biofilms formed by Acinetobacter baumannii CRAB KPD 205 (reduction levels of 4.28 and 2.62 log CFU/mL, respectively). The activity of Intestinalin is combined with both no cytotoxicity and little hemolytic effect against mammalian cells. The nuclear magnetic resonance and molecular dynamics (MD) data show a high tendency of Intestinalin to interact with the bacterial phospholipid cell membrane. Although positively charged, Intestinalin resides in the membrane and aggregates into small oligomers. Negatively charged phospholipids stabilize peptide oligomers to form water- and ion-permeable pores, disrupting the integrity of bacterial cell membranes. Experimental data showed that Intestinalin interacts with negatively charged lipoteichoic acid (logK based on isothermal titration calorimetry, 7.45 ± 0.44), causes membrane depolarization, and affects membrane integrity by forming large pores, all of which result in loss of bacterial viability. IMPORTANCE Antibiotic resistance is rising rapidly among pathogenic bacteria, becoming a global public health problem that threatens the effectiveness of therapies for many infectious diseases. In this respect, antimicrobial peptides appear to be an interesting alternative to combat bacterial pathogens. Here, we report the characteristics of an antimicrobial peptide (of 30 amino acids) derived from the clostridial LysC enzyme. The peptide showed killing activity against clinical strains of Gram-positive and Gram-negative pathogens. Experimental data and computational modeling showed that this peptide forms transmembrane pores, directly engaging the negatively charged phospholipids of the bacterial cell membrane. Consequently, dissipation of the electrochemical gradient across cell membranes affects many vital processes, such as ATP synthesis, motility, and transport of nutrients. This kind of dysfunction leads to the loss of bacterial viability. Our firm conviction is that the presented study will be a helpful resource in searching for novel antimicrobial peptides that could have the potential to replace conventional antibiotics.

Role of cholesterol in substrate recognition by [Formula: see text]-secretase.

[Formula: see text]-Secretase is an enzyme known to cleave multiple substrates within their transmembrane domains, with the amyloid precursor protein of Alzheimer's Disease among the most prominent examples. The activity of [Formula: see text]-secretase strictly depends on the membrane cholesterol content, yet the mechanistic role of cholesterol in the substrate binding and cleavage remains unclear. In this work, we used all-atom molecular dynamics simulations to examine the role of cholesterol in the initial binding of a direct precursor of [Formula: see text]-amyloid polypeptides by [Formula: see text]-secretase. We showed that in cholesterol-rich membranes, both the substrate and the enzyme region proximal to the active site induce a local membrane thinning. With the free energy methods we found that in the presence of cholesterol the substrate binds favorably to the identified exosite, while cholesterol depletion completely abolishes the binding. To explain these findings, we directly examined the role of hydrophobic mismatch in the substrate binding to [Formula: see text]-secretase, showing that increased membrane thickness results in higher propensity of the enzyme to bind substrates. Therefore, we propose that cholesterol promotes substrate binding to [Formula: see text]-secretase by increasing the membrane thickness, which leads to the negative hydrophobic mismatch between the membrane and binding partners.

Effect of chemical structure on complexation efficiency of aromatic drugs with cyclodextrins: The example of dibenzazepine derivatives.

It is widely believed that the hydrophobic effect governs the binding of guest molecules to cyclodextrins (CDs). However, it is also known that high hydrophobicity of guest molecules does not always translate to the formation of stable inclusion complexes with CDs. Indeed, a plethora of other factors can play a role in the efficiency of guest-CD interactions, rendering structure-based prediction of the complexation efficiency with CDs a non trivial task. In this combined experimental and computational study, we examine the major structural factors governing complexation efficiency of polycyclic aromatic drug-like compounds with natural CDs, using as an example iminostilbene and its N-substituted derivatives. We find that purely hydrophobic IS derivatives show negligible complexation efficiency with CDs and only IS with hydrophilic substituents form stable inclusion complexes in water. We show that the balance between the guest solubility and its affinity to CDs is critical for the effective formation of inclusion complexes. Finally, our results demonstrate that guest-host hydrogen bonds facilitate the formation of crystalline inclusion complexes with CDs.