Evaluation of the Binding Mechanism of Human Defensin 5 in a Bacterial Membrane: A Simulation Study

Human α-defensin 5 (HD5) is a host-defense peptide exhibiting broad-spectrum antimicrobial activity. The lipopolysaccharide (LPS) layer on the Gram-negative bacterial membrane acts as a barrier to HD5 insertion. Therefore, the pore formation and binding mechanism remain unclear. Here, the binding mechanisms at five positions along the bacterial membrane axis were investigated using Molecular Dynamics. (MD) simulations. We found that HD5 initially placed at positions 1 to 3 moved up to the surface, while HD5 positioned at 4 and 5 remained within the membrane interacting with the middle and inner leaflet of the membrane, respectively. The arginines were key components for tighter binding with 3-deoxy-d-manno-octulosonic acid (KDO), phosphates of the outer and inner leaflets. KDO appeared to retard the HD5 penetration.

The penetration of Human Defensin 5 (HD5) through bacterial outer membrane: Simulation studies

Human α-defensin 5 (HD5) is one of cationic antimicrobial peptides which plays a crucial role in an innate immune system in human body. HD5 shows the killing activity against a broad spectrum of pathogenic bacteria by making a pore in a bacterial membrane and penetrating into a cytosol. Nonetheless, its pore-forming mechanisms remain unclear. Thus, in this work, the constant-velocity steered molecular dynamics (SMD) simulation was used to simulate the permeation of a dimeric HD5 into a gram-negative LPS membrane model. Arginine-rich HD5 is found to strongly interact with a LPS surface. Upon arrival, arginines on HD5 interact with lipid A head groups and then drag these charged moieties down into a hydrophobic core resulting in the formation of water-filled pore. Although all arginines are found to interact with a membrane, R13 and R32 appear to play a dominant role in the HD5 adsorption on a gram-negative membrane. Furthermore, one chain of a dimeric HD5 is required for HD5 adhesion. The interactions of arginine-Lipid A head groups play a major role in adhering a cationic HD5 on a membrane surface and retarding a HD5 passage in the meantime.

Lower human defensin 5 in elderly people compared to middle-aged is associated with differences in the intestinal microbiota composition: the DOSANCO Health Study

Recently, aging is considered a risk factor for various diseases. Although changes in the intestinal microbiota along with aging are thought to associate with the increased disease risk, mechanisms that cause age-related transition of the intestinal microbiota remain unknown. This study aims to clarify relationships between the amount of human defensin 5 (HD5), a Paneth cell α-defensin, which is known to regulate the intestinal microbiota, and age-related differences of the intestinal microbiota composition. Fecal samples from 196 healthy Japanese (35 to 81 years old) were collected and measured HD5 concentration. HD5 concentration in the elderly group (age > 70 years old) was significantly lower than the middle-aged group (age ≤ 70 years old). Furthermore, individual age was negatively correlated with HD5 concentration (r = − 0.307, p < 0.001). In β-diversity, the intestinal microbiota of the elderly showed a significantly different composition compared to the middle-aged. At the genus level, relative abundance of Collinsella, Alistipes, Peptococcaceae; unassigned, Lactobacillus, Lactococcus, Weissella, Christensenellaceae R-7 group, Megasphaera, and [Eubacterium] eligens group was significantly higher, and Lachnospiraceae; unassigned, Blautia, Anaerostipes, Fusicatenibacter, Dorea, and Faecalibacterium was significantly lower in the elderly compared to the middle-aged. In addition, HD5 concentration was negatively correlated with Alistipes, Peptococcaceae; unassigned, and Christensenellaceae R-7 group and positively correlated with Lachnospiraceae; unassigned and Dorea. These results provide novel insights into the immunosenescence of enteric innate immunity, indicating low HD5 is suggested to contribute to the age-related differences in the intestinal microbiota and may relate to increased risk of diseases in elderly people.

A Simplified Derivative of Human Defensin 5 with Potent and Efficient Activity against Multidrug-Resistant Acinetobacter baumannii

ABSTRACT The increasing incidence of multidrug-resistant Acinetobacter baumannii (MDRAb) infections worldwide has necessitated the development of novel antibiotics. Human defensin 5 (HD5) is an endogenous peptide with a complex architecture and antibacterial activity against MDRAb. In the present study, we attempted to simplify the structure of HD5 by removing disulfide bonds. We found that the Cys2-4 bond was most indispensable for HD5 to inactivate MDRAb, although the antibacterial activity of the derivative was significantly attenuated. We then replaced the noncationic and nonhydrophobic residues with electropositive Arg to increase the antibacterial activity of HD5 derivative that contains a Cys2-4 bond, obtaining another derivative termed HD5d5. The in vitro antibacterial assay and irradiation-wound-infection animal experiment both showed that HD5d5 was much more effective than HD5 at eliminating MDRAb. Further investigations revealed that HD5d5 efficiently bound to outer membrane lipid A and penetrated membranes, leading to bacterial collapse and peptide translocation. Compared to HD5, more HD5d5 molecules were located in the cytoplasm of MDRAb, and HD5d5 was more efficient at reducing the activities of superoxide dismutase and catalase, causing the accumulation of reactive oxygen species that are detrimental to microbes. In addition, HD5 failed to suppress the pathogenic outer membrane protein A of Acinetobacter baumannii (AbOmpA) at concentrations up to 50 μg/ml, whereas HD5d5 strongly bound to AbOmpA and exhibited a dramatic toxin-neutralizing ability, thus expanding the repertoire of drugs that is available to treat MDRAb infections.