|FILATOVA, LYUBOV - Lomonosov University|
|Foster Frey, Juli|
|PUGACHEV, VLADIMIRE - Lomonosov University|
|DMITRIEVA, NATALIA - Lomonosov University|
|CHUBAR, TATIANA - Lomonosov University|
|KLYACHKO, NATALIA - Lomonosov University|
|KABANOV, ALEXANDER - Lomonosov University|
Submitted to: Enzyme and Microbial Technology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/8/2015
Publication Date: 6/2/2015
Citation: Filatova, L.Y., Donovan, D.M., Foster Frey, J.A., Pugachev, V.G., Dmitrieva, N.F., Chubar, T.A., Klyachko, N.L., Kabanov, A.V. 2015. Bacteriophage phi11 lysin: physicochemical characterization and comparison with phage phi80a lysin. Enzyme and Microbial Technology. 73-74:51-58. DOI: 10.1016/j.enzmictec.2015.03.005.
Interpretive Summary: There is a need to develop novel antimicrobials to replace antibiotics that are no longer useful due to antibiotic resistance development in different disease-causing bacteria. One form of alternative being explored is the use of enzymes that degrade the bacterial cell wall and cause bacteria to lyse (be destroyed). It is important to expore the biophysical and biochemical properties of these enzymes as they might be sensitive to certain environmental conditions that antibiotics have not shown sensitivity. In this work, several environmental conditions were explored for storage and activity testing of two closely related enzymes. Thse enzymes kill numerous disease causing bacteria, including Staphylococus aureus. These results will help fellow scientists to identify conditions that are optimal for storage/use of these two enzymes (and potentially other related enzymes) and their predicted relative activity, pending the environment in which they are to be used.
Technical Abstract: Phage lytic enzymes are promising antimicrobial agents. Lysins of phage phi11 (LysPhi11) and phi80a (LysPhi80a) can lyse (destroy) biofilms and cells of antibiotic-resistant strains of Staphylococcus aureus. Stability of enzymes is one of the parameters making their practical use possible. The objectives of the study were to investigate the stability of phage phi11 and phi80a lysins in storage and functioning conditions, to identify optimum storage conditions and causes of inactivation. Stability of the recombinant LysPhi11 and LysPhi80a was studied using turbidimetry. CDspectroscopy, dynamic light scattering, and electrophoresis were used to identify causes of inactivation. At 37°C, pH 7.5 and concentration of NaCl not higher than 150 mM, LysPhi11 molecules contain a high percentage of random coils (43%). However, in spite of this the enzyme has high activity (0.4-0.8 OD600nms-1mg-1). In storage conditions (4°C and 22°C, pH 6.0-9.0, 10-500 mM NaCl) LysPhi11 is inactivated by a monomolecular mechanism. The optimum storage conditions for LysPhi11 (4°C, pH 6.0-7.5, 10 mM NaCl) were selected under which the time of the enzyme half-inactivation is 120-160 days. LysPhi80a stability is insufficient: at 37°C the enzyme loses half of its activity almost immediately; at 4°C and 22°C the time of half-inactivation of LysPhi80a varies in the range from several hours to 3 days. Despite the common properties in the manifestation of antistaphylococcal activity the kinetic behavior of the enzymes is different. LysPhi11 is a more promising candidate to be used as an antimicrobial agent.