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A Simple Protocol for the Determination of Lysostaphin Enzymatic Activity

1
N.F. Gamaleya National Research Center for Epidemiology and Microbiology, Ministry of Health of the Russian Federation, 123098 Moscow, Russia
2
All-Russia Research Institute of Agricultural Biotechnology, Russian Academy of Sciences, 127550 Moscow, Russia
3
Faculty of Bioengineering and Bioinformatics, M.V. Lomonosov Moscow State University, 119234 Moscow, Russia
4
A.N. Belozersky Institute of Physical and Chemical Biology, M.V. Lomonosov Moscow State University, 119992 Moscow, Russia
*
Author to whom correspondence should be addressed.
Antibiotics 2020, 9(12), 917; https://doi.org/10.3390/antibiotics9120917
Received: 26 October 2020 / Revised: 4 December 2020 / Accepted: 15 December 2020 / Published: 17 December 2020
(This article belongs to the Section Novel Antimicrobial Agents)
Antibacterial lysins are enzymes that hydrolyze bacterial peptidoglycan, which results in the rapid death of bacterial cells due to osmotic lysis. Lysostaphin is one of the most potent and well-studied lysins active against important nosocomial pathogen Staphylococcus aureus. Similarly to most other lysins, lysostaphin is composed of enzymatic and peptidoglycan-binding domains, and both domains influence its antibacterial activity. It is thus desirable to be able to study the activity of both domains independently. Lysostaphin cleaves pentaglycine cross-bridges within the staphylococcal peptidoglycan. Here, we report the protocol to study the catalytic activity of lysostaphin on the isolated pentaglycine peptide that is based on the chromogenic reaction of peptide amino groups with ninhydrin. Unlike previously reported assays, this protocol does not require in-house chemical synthesis or specialized equipment and can be readily performed in most laboratories. We demonstrate the use of this protocol to study the effect of EDTA treatment on the lysostaphin enzymatic activity. We further used this protocol to determine the catalytic efficiency of lysostaphin on the isolated pentaglycine and compared it to the apparent catalytic efficiency on the whole staphylococcal cells. These results highlight the relative impact of enzymatic and peptidoglycan-binding domains of lysostaphin on its bacteriolytic activity. View Full-Text
Keywords: lysostaphin; pentaglycine; enzymatic activity; catalytic efficiency; protocol; assay; lysin; Staphylococcus aureus lysostaphin; pentaglycine; enzymatic activity; catalytic efficiency; protocol; assay; lysin; Staphylococcus aureus
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MDPI and ACS Style

Grishin, A.V.; Konstantinova, S.V.; Vasina, I.V.; Shestak, N.V.; Karyagina, A.S.; Lunin, V.G. A Simple Protocol for the Determination of Lysostaphin Enzymatic Activity. Antibiotics 2020, 9, 917. https://doi.org/10.3390/antibiotics9120917

AMA Style

Grishin AV, Konstantinova SV, Vasina IV, Shestak NV, Karyagina AS, Lunin VG. A Simple Protocol for the Determination of Lysostaphin Enzymatic Activity. Antibiotics. 2020; 9(12):917. https://doi.org/10.3390/antibiotics9120917

Chicago/Turabian Style

Grishin, Alexander V., Svetlana V. Konstantinova, Irina V. Vasina, Nikita V. Shestak, Anna S. Karyagina, and Vladimir G. Lunin 2020. "A Simple Protocol for the Determination of Lysostaphin Enzymatic Activity" Antibiotics 9, no. 12: 917. https://doi.org/10.3390/antibiotics9120917

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