Mechanism Of Action Of Magnolol Against Staphylococcus Aureus

Staphylococcus aureus (S. aureus) is one of the most important pathogens in both hospitals and communities, and it causes numerous diseases in humans, ranging from minor skin and wound infections to life-threatening diseases, such as endocarditis, osteomyelitis and septicemia. With the increase in the incidence of drug-resistant S. aureus, some strains have developed resistance to various antibiotics. Prudent use of antimicrobial drugs for minimizing the further emergence and dissemination of antimicrobial resistance is certainly a daunting long-term challenge.Natural medicines are important resource of material for prevention and treatment of human disease. Many of them show remarkable biological activities. As traditional Chinese medicine, Magnolia officinalis had been used for thousands of years. Magnolol (MOL), the major component isolated from the stem barks of Magnolia officinalis, has been shown to possess a variety of pharmacological activities, including antiinflammatory, antimicrobial, anti-allergic and anti-asthmatic activities. However, only few data could be found in the literature regarding the molecular mechanism of MOL action aganist S.aureus. This study investigated effects of MOL on autolysis, release ofα-haemolysin and adhesion of S.aureus.Autolysis enzymes degrade peptidoglycan saccules, resulting in cell lysis. In this study, we used Triton X-100-induced autolysis assays, quantitative bacteriolytic assays and zymographic analysis to evaluate autolysis inhibition caused by MOL. This interesting result motivated us to further research its mechanism of action.Using an Affymetrix GeneChip, we analysed the global transcriptional patterns of S. aureus in response to subinhibitory concentrations of MOL. Global gene expression profile can be analyzed quickly and systematically in the whole genome level of gene expression by use of DNA microarray. Microarray provides a powerful tool in modern science and medical diagnostics. Transcriptional profiles generated by GeneChip analysis of bacteria can provide valuable information for investigating differential gene expression in response to antimicrobial agents. By this technology, the bio-effects of drugs would be related with the changes of genes which then will be helpful to further studies on drug. GeneChip analysis revealed that a large number of genes (550) were differentially regulated in response to sub-inhibitory concentrations of MOL.210 genes showed significant increases and 340 genes showed significant decreases in transcription.Herein, We analyzed the expression levels of 143 genes involved in autolysis or cell wall biosynthesis or known as related regulators on a microarray. Notablely, this study in the international areas for the first time analyzed the expression profile of MOL effected on S.aureus.Decreased atl, slel and cidA transcript levels were consistent with the induced expression of their autolytic repressors (?)rgA, (?)rgB, arlR and sarA in the MOL-treated strain compared to the control strain, which individually or collectively may contribute to the autolysis-inhibited phenotype, according to previous studies. Furthermore, a small subset of those transcripts was evaluated by real-time RT-PCR.Like other Gram-positive bacteria, the pathogenicity of S.aureus is largely dependent upon extracellular virulence factors, including both secreted and surface proteins. Virulence factor is regulated by a number of extracellular and intracellular signals, such as the two-component systems. Expression of 35 virulence-associated genes were differentially regulated in response to MOL. Most of the genes were downregulated. We found that expression of the iron-regulated surface determinant system genes were inhibited by MOL. In pathogenic gram-positive bacteria, the isd system was found to encode factors responsible for haemoglobin binding and passage of haeme-iron to the cytoplasm, where it acts as an essential nutrient. Isd appears to act as an import apparatus that uses cell wall-anchored proteins to transport haeme-iron across the bacterial envelope.The exoproteins of S. aureus can cause a number of diseases. One of the most important extracellular proteins is a-hemolysin. The toxin can cause pore formation of a wide range of human cells. Furthermore, it can also induce pro-inflammatory changes in mammalian cells. The consequent cellular damage may contribute to manifestations of the sepsis syndrome. In the present study, MOL was shown to inhibit a-toxin expression by S. aureus via hemolysin assays, western blot analysis, and real-time RT-PCR. Our results suggest that the structure of MOL may be used as a basic structure for the development of drugs aimed at bacterial virulence factors.The initial step in pathogenesis of infections is often cell adhesion. Adhesion is then followed by bacterial invasion and colonization of the host tissue and is often a prerequisite for successful colonization of epithelia. Following the completion of the human genome sequence, much attention is now shifting towards the functions of proteins. The structure study was a main approaches to explain their function.To study the molecular mechanism of MOL, we performed clone, expression and purificatin of ClfB, SdrD, SdrE. Crystals were successfully grown using the method of hanging-drop vapour-diffusion. Finally, we obtained the high quality crystals of SdrD, SdrE and solved the structures of rClfB fragment in complex with FgAa/CK10. The seleno-methionine derivatives of ClfB proteins were purified similarly. Native and Se-SAD data were collected at the Shanghai Synchrotron Radiation Facility (SSRF). The crystal structure was solved by molecular replacement.The current study was undertaken to characterize the interaction of ClfB and ligands to define in detail the binding mechanism of it. We solved the high resolution crystal structure, and revealed the real Dock Lock Latch (DLL) mechanism. Base on carefully analysis of binding cleft and the feature of the peptides a small motif was proposed. Surface plasmon resonance technology was used to analyse the affinity between ClfB mutants and its ligand. The affinity change of mutants and ligand correspond with their structure difference.By comparison of the two crystal structure of peptides binding to ClfB adhesive domains we found they adopt similar binding mode and share similar binding motif also. Therefore it will be meaningful for identification of putative substrates of the surface adhesive protein ClfB and for further understanding of the infection progress of S. aureus. Our findings also provide important leads toward the development new therapeutic agents capable of eradicating S. aureus carriage in individuals ai interfering staphylococcal infection efficiently.