A Chimeric LysK-Lysostaphin Fusion Enzyme Lysing Staphylococcus aureus Cells: a Study of Both Kinetics of Inactivation and Specifics of Interaction with Anionic Polymers

Filatova, L.Y.; Donovan, D.M.; Ishnazarova, N.T.; Foster-Frey, J.A.; Becker, S.C.; Pugachev, V.G.; Balabushevich, N.G.; Dmitrieva, N.F.; Klyachko, N.L.

Авторы: Filatova Lyubov Y., Donovan David M., Ishnazarova Nadiya T., Foster-Frey Juli A., Becker Stephen C., Pugachev Vladimir G., Balabushevich Nadezda G., Dmitrieva Natalia F., Klyachko Natalia L.
Журнал: Applied Biochemistry and Biotechnology
Том: 180
Номер: 3
Год издания: 2016
Издательство: Humana Press, Inc.
Местоположение издательства: United States
Первая страница: 544
Последняя страница: 557
DOI: 10.1007/s12010-016-2115-7
Аннотация: A staphylolytic fusion protein (chimeric enzyme K-L) was created, harboring three unique lytic activities comprised of the LysK CHAP endopeptidase, and amidase domains, and the lysostaphin glycyl-glycine endopeptidase domain. To assess the potential of possible therapeutic applications the kinetic behavior of chimeric enzyme K-L was investigated. As a protein antimicrobial, with potential antigenic properties, the biophysical effect of including chimeric enzyme K-L in anionic polymer matrices that might help reduce the immunogenicity of the enzyme was tested. Chimeric enzyme K-L reveals a high lytic activity under the following optimal (opt) conditions: pHopt 6.0-10.0, topt 20-30°C, NaClopt 400-800 mM. At the working temperature of 37°C chimeric enzyme K-L is inactivated by a monomolecular mechanism and possesses a high half-inactivation time of 12.7±3.0 hrs. At storage temperatures of 22°C and 4°C a complex mechanism (combination of monomolecular and bimolecular mechanisms) is involved in the chimeric enzyme K-L inactivation. The optimal storage conditions under which the enzyme retains 100% activity after 140 days of incubation (4°C, the enzyme concentration of 0.8 mg/mL, pH 6.0 or 7.5) were established. Chimeric enzyme K-L is included in complexes with block-copolymers of poly-L-glutamic acid and polyethylene glycol, while the enzyme activity and stability are retained, thus suggesting methods to improve the application of this fusion as an effective antimicrobial agent.
Добавил в систему: Филатова Любовь Юрьевна

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