ИСТИНА

Staphylococcus aureus causes many serious visceral, skin, and respiratory diseases. About 90% of its today’s strains are resistant to multiple antibiotic drugs, but the use of lytic enzymes secreted by bacteriophages offers a viable alternative to antibiotic therapy. LysK is a staphylococcal bacteriophage endolysin from the phage K, a peptidoglycan hydrolase enzyme that can destroy (lyse) many staphylococcal strains including methicillin- and vancomycin resistant S. aureus (MRSA and VRSA). This work aims at developing approaches to produce nanozymes of the LysK enzyme for creating novel anti-infectives against Staphylococcus aureus. Potent drugs based on LysK enzyme should have high antimicrobial activity and be selective, non-toxic (for humans and animals). Materials: solution of the LysK recombinant enzyme, polycationic polymers, such as poly-L-lysines and their block-copolymers with polyethylene glycol. Methods: kinetic study, IR spectroscopy, dynamic light scattering, cytotoxicity investigation. We optimized conditions for LysK enzyme nanozymes formation. Nanozymes are complexes of LysK with polycationic polymers (poly-L-lysines (PLLs) of molecular weights 2.5, 9.6, and 55.2 kDa and block-copolymers (EG)114(Lys)10, (EG)114(Lys)30, (EG)23(Lys)30). The block-copolymers (EG)114(Lys)10, (EG)114(Lys)30, (EG)23(Lys)30 and 2.5, 9.6 and 55.2 kDa PLLs increased the activity of LysK. At physiological temperature (37°C) and NaCl (150 mM), LysK is the most stable in nanozymes with 55.2 kDa PLL and (EG)114(Lys)30 (the half-inactivation times increase from 0.5 hr for LysK alone to 4 – 10 and 15 – 25 hr, respectively). At 22°C, the stabilizing effect is the most prominent with 2.5 and 9.6 kDa PLLs and (EG)114(Lys)30 block-copolymer (the increase in half-inactivation time is as large as from 48 hours to 60, 30, and 34 days, respectively). At 4°C, LysK in complexes with these polymers remains almost inactivated for four months. Thus, we have optimized for the antimicrobial action of the enzyme and its storage through the choice of polycationic polymers and the conditions of their nanozymes formation with LysK.