A Rapid Lysostaphin Production Approach and a Convenient Novel Lysostaphin Loaded Nano-emulgel; As a Sustainable Low-Cost Methicillin-Resistant Staphylococcus aureus Combating Platform

Abstract

Staphylococcus aureus is a Gram-positive pathogen that is capable of infecting almost every organ in the human body. Alarmingly, the rapid emergence of methicillin-resistant S. aureus strains (MRSA) jeopardizes the available treatment options. Herein, we propose sustainable, low-cost production of recombinant lysostaphin (rLST), which is a native bacteriocin destroying the staphylococcal cell wall through its endopeptidase activity. We combined the use of E. coli BL21(DE3)/pET15b, factorial design, and simple Ni-NTA affinity chromatography to optimize rLST production. The enzyme yield was up to 50 mg/L culture, surpassing reported systems. Our rLST demonstrated superlative biofilm combating ability by inhibiting staphylococcal biofilms formation and detachment of already formed biofilms, compared to vancomycin and linezolid. Furthermore, we aimed at developing a novel rLST topical formula targeting staphylococcal skin infections. The phase inversion composition (PIC) method fulfilled this aim with its simple preparatory steps and affordable components. LST nano-emulgel (LNEG) was able to extend active LST release up to 8 h and cure skin infections in a murine skin model. We are introducing a rapid, convenient rLST production platform with an outcome of pure, active rLST incorporated into an effective LNEG formula with scaling-up potential to satisfy the needs of both research and therapeutic purposes.

Cloning and expression of <i>Staphylococcus simulan</i>s lysostaphin enzyme gene in <i>Bacillus subtilis</i> WB600

“…This is the first report on cloning and expression of the lysostaphin enzyme gene in B. subtilis WB600 using pWB980 expression system, which secretes a high level of r-lysostaphin enzyme several folds rather than the production of r-lysostaphin by pET and some other T7-based expression hosts E. coli BL21(DE3), BL21 star(DE3), BL21-A1, T7 Express, RV308(DE3), and HMS174(DE3). The optimum temperature and pH activity of r-lysostaphin were found at 37–40 °C and 8, respectively which is in concordance to the previous reports [31] , [32] .…”

Cloning and Expression of Staphylococcus Simulans Lysostaphin Enzyme Gene in Bacillus Subtilis&nbsp;WB600

“…This is the rst report on cloning and expression of the lysostaphin enzyme gene in B. subtilis WB600 using pWB980 expression system, which secretes a high level of r-lysostaphin enzyme several folds rather than the production of r-lysostaphin by pET and some other T7-based expression hosts E. coli BL21(DE3), BL21 star (DE3), BL21-A1, T7 Express, RV308(DE3), and HMS174(DE3). The optimum temperature and pH activity of r-lysostaphin were found at 37-40℃ and 8, respectively which is in concordance to the previous reports (29,30).…”

Staquorsin: A Novel Staphylococcus aureus Agr-Mediated Quorum Sensing Inhibitor Impairing Virulence in vivo Without Notable Resistance Development

“…In order to assess the role of staquorsin in biofilm formation and detachment pre- and post-exposure assays were performed as described before ( Nour El-Din et al, 2020 ) with slight modifications. Briefly, for biofilm formation, overnight cultures of staphylococcal cells were normalized to OD 600 of 1.0 and diluted 1:100 in fresh TSB.…”