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  • A potent enzybiotic against methicillin-resistant Staphylococcus aureus.

A potent enzybiotic against methicillin-resistant Staphylococcus aureus.

Virus genes (2020-05-06)
Jasjeet Kaur, Prashant Singh, Deepak Sharma, Kusum Harjai, Sanjay Chhibber
ABSTRACT

Staphylococcus aureus is one of the most dreadful infectious agents, responsible for high mortality and morbidity in both humans and animals. The increased prevalence of multidrug-resistant (MDR) Staphylococcus aureus strains has limited the number of available treatment options, which calls for the development of alternative and effective modalities against MDR S. aureus. Endolysins are bacteriophage-derived antibacterials, which attack essential conserved elements of peptidoglycan that are vital for bacterial survival, making them promising alternatives or complements to existing antibiotics for tackling such infections. For developing endolysin lysin-methicillin-resistant-5 (LysMR-5) as an effective antimicrobial agent, we evaluated its physical and chemical characteristics, and its intrinsic antibacterial activity against staphylococcal strains, including methicillin-resistant Staphylococcus aureus (MRSA). In this study, we cloned, expressed, and purified LysMR-5 from S. aureus phage MR-5. In silico analysis revealed that LysMR-5 harbors two catalytic and one cell wall-binding domain. Biochemical characterization and LC-MS analysis showed that both catalytic domains were active and had no dependence on divalent ions for their action, Zn2+ exerted a negative effect. The optimal lytic activity of the endolysin was at 37 °C/pH 7.0 and in the presence of ≥ 300 mM concentration of NaCl. Circular dichroism (CD) demonstrated a loss in secondary structure with an increase in temperature confirming the thermosensitive nature of endolysin. Antibacterial assays revealed that LysMR-5 was active against diverse clinical isolates of staphylococci. It showed high lytic efficacy against S. aureus ATCC 43300, as an endolysin concentration as low as 15 µg/ml was sufficient to achieve maximum lytic activity within 30 min and it was further confirmed by scanning electron microscopy. Our results indicate that rapid and strong bactericidal activity of LysMR-5 makes it a valuable candidate for eradicating multidrug-resistant S. aureus.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Muramic acid, ≥95% (TLC)
Sigma-Aldrich
L-Serine, ReagentPlus®, ≥99% (HPLC)
Sigma-Aldrich
Peptidoglycan from Staphylococcus aureus, cell wall component
Sigma-Aldrich
4-Phenylphenol, 97%
Millipore
Amicon® Ultra Centrifugal Filter, 10 kDa MWCO, sample volume 2 mL, regenerated cellulose membrane, MWCO 10 kDa