Recombinant Fungi Proteinase K

The microbiota influences host health through several mechanisms, including protecting it from pathogen colonization. Staphylococcus epidermidis is one of the most frequently found species in the skin microbiota, and its presence can limit the development of pathogens such as Staphylococcus aureus. S. aureus causes diverse types of infections ranging from skin abscesses to bloodstream infections. Given the increasing prevalence of S. aureus drug-resistant strains, it is imperative to search for new strategies for treatment and prevention. Thus, we investigated the activity of molecules produced by a commensal S. epidermidis isolate against S. aureus biofilms. We showed that molecules present in S. epidermidis cell-free conditioned media (CFCM) caused a significant reduction in biofilm formation in most S. aureus clinical isolates, including all 4 agr types and agr-defective strains, without any impact on growth. S. epidermidis molecules also disrupted established S. aureus biofilms and reduced the antibiotic concentration required to eliminate them. Preliminary characterization of the active compound showed that its activity is resistant to heat, protease inhibitors, trypsin, proteinase K, and sodium periodate treatments, suggesting that it is not proteinaceous. RNA sequencing revealed that S. epidermidissecreted molecules modulate the expression of hundreds of S. aureus genes, some of which are associated with biofilm production. Biofilm formation is one of the main virulence factors of S. aureus and has been associated with chronic infections and antimicrobial resistance. Therefore, molecules that can counteract this virulence factor may be promising alternatives as novel therapeutic agents to control S. aureus infections. IMPORTANCE S. aureus is a leading agent of infections worldwide, and its main virulence characteristic is the ability to produce biofilms on surfaces such as medical devices. Biofilms are known to confer increased resistance to antimicrobials and to the host immune responses, requiring aggressive antibiotic treatment and removal of the infected surface. Here, we investigated a new source of antibiofilm compounds, the skin microbiome. Specifically, we found that a commensal strain of S. epidermidis produces molecules with antibiofilm activity, leading to a significant decrease of S. aureus biofilm formation and to a reduction of previously established biofilms. The molecules potentiated the activity of antibiotics and affected the expression of hundreds of S. aureus genes, including those associated with biofilm formation. Our research highlights the search for compounds that can aid us in the fight against S. aureus infections.

Withaferin-A is a withanolide, predominantly present in Ashwagandha (Withania somnifera). It has been shown to possess anticancer activity in a variety of human cancer cells in vitro and in vivo. Molecular mechanism of such cytotoxicity has not yet been completely understood. Withaferin-A and Withanone were earlier shown to activate p53 tumor suppressor and oxidative stress pathways in cancer cells. 2,3-dihydro-3 beta-methoxy analogue of Withaferin-A (3 beta mWi-A) was shown to lack cytotoxicity and well tolerated at higher concentrations. It, on the other hand, protected normal cells against oxidative, chemical and UV stresses through induction of anti-stress and pro-survival signaling. We, in the present study, investigated the effect of Wi-A and 3 beta mWi-A on cell migration and metastasis signaling. Whereas Wi-A binds to vimentin and heterogeneous nuclear ribonucleoprotein K (hnRNP-K) with high efficacy and downregulates its effector proteins, MMPs and VEGF, involved in cancer cell metastasis, 3 beta mWi-A was ineffective. Consistently, Wi-A, and not 3 beta mWi-A, caused reduction in cytoskeleton proteins (Vimentin, N-Cadherin) and active protease (u-PA) that are essential for three key steps of cancer cell metastasis (EMT, increase in cell migration and invasion).