BACKGROUND AND OBJECTIVESAntibiotics are one of the most important and influential medical inventions of the ۲۰th century. However, the number of infections caused by
multidrug-resistant (MDR) bacteria has been increasing since the beginning of the ۲۱st century.
Staphylococcus aureus is a pathogenic bacterium that is resistant to many antibiotics. With the increasing prevalence of antibiotic-resistant bacteria, phage endolysins are known as one of the promising alternatives to antibiotics. The desired chimeric
endolysin has two catalytic domains and one domain binding to the cell wall, and they target peptidoglycan, causing cell lysis. The process of designing, cloning, and initial expression of this enzyme has been done by the Iranian Research Organization for Science and Technology (IROST). In this study, we demonstrated that desired
endolysin has a high bactericidal activity against Staphylococcus aureus.MATERIALS AND METHODSThe desired plasmid is transformed into Bl۲۱ gold-competent cells through heat shock. Then protein expression was done for ۲۴ hours at ۱۵°C and finally, purification was done using HisPur™ Ni-NTA Resin by affinity chromatography and the protein concentration was determined. The purification was confirmed by the western blot analysis and the bactericidal activity of
endolysin was evaluated by
turbidity reduction assay (TRA).RESULTS AND DISCUSSIONPlate cultivation was done from the transformed Bl۲۱ gold and single colonies were observed in the plate containing TSA medium. Then the protein expression and purification were done and the desired ۴۶ kDa protein band was observed in SDS-PAGE, and the purified protein band was approved by Western Blot. The protein concentration was measured with a BCA kit and the purified protein was ۳۰۵ μg/ml. The bactericidal activity of
endolysin was evaluated by turbidity reduction assay. Due to having two catalytic domains that target bacterial peptidoglycan, chimeric
endolysin showed high bactericidal activity against
Staphylococcus aureus and was effective even at a low concentration of ۲.۳۸ μg/ml.CONCLUSIONAntimicrobial drugs and new treatment options are needed to deal with antibiotic resistance. These results show that a new chimeric
endolysin with higher activity and solubility can be created by domain swapping, which has great potential as an antimicrobial agent. Therefore, our strategy has significant potential for medical and biotechnological applications in combating
multidrug-resistant bacteria such as Staphylococcus aureus.