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LytM Fusion with SH3b-Like Domain Expands Its Activity to Physiological Conditions

View Article: PubMed Central - PubMed

ABSTRACT

Staphylococcus aureus remains one of the most common and at the same time the most dangerous bacteria. The spreading antibiotic resistance calls for intensification of research on staphylococcal physiology and development of new strategies for combating this threatening pathogen. We have engineered new chimeric enzymes comprising the enzymatically active domain (EAD) of autolysin LytM from S. aureus and the cell wall binding domain (CBD) from bacteriocin lysostaphin. They display potent activity in extended environmental conditions. Our results exemplify the possibility of exploring autolytic enzymes in engineering lysins with desired features. Moreover, they suggest a possible mechanism of autolysin physiological activity regulation by local ionic environments in the cell wall.

No MeSH data available.


Related in: MedlinePlus

Specific activity of the lysins against Staphylococcus aureus NCTC 8325-4 reference strain. (A) Enzymes activities were observed as a turbidity reduction after 1 hr in room temperature either in 1 × PBS or 50 mM glycine buffer, pH 8.0. (B) Lytic activity of the enzymes was determined by SYTOX® fluorescence assay performed in 1 × PBS or 50 mM glycine buffer, pH 8.0, at room temperature for 1 hr. PBS, phosphate-buffered saline.
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f2: Specific activity of the lysins against Staphylococcus aureus NCTC 8325-4 reference strain. (A) Enzymes activities were observed as a turbidity reduction after 1 hr in room temperature either in 1 × PBS or 50 mM glycine buffer, pH 8.0. (B) Lytic activity of the enzymes was determined by SYTOX® fluorescence assay performed in 1 × PBS or 50 mM glycine buffer, pH 8.0, at room temperature for 1 hr. PBS, phosphate-buffered saline.

Mentions: The LytM_EAD activity is very limited in physiological conditions (PBS),33 however, in low conductivity environment (glycine buffer), LytM_EAD demonstrated great bacteriolytic potential (Fig. 2A). This effect was even more notorious when the fluorometric assay was used. The specific activity of LytM_EAD was the highest among all tested enzymes in the glycine buffer and the lowest in PBS (Fig. 2B). These assays confirmed previous observations of strong inhibition of LytM_EAD bacteriolytic activity by ions present in the reaction environment,33 which might implicate at least one of the mechanisms of LytM activity regulation in vivo.


LytM Fusion with SH3b-Like Domain Expands Its Activity to Physiological Conditions
Specific activity of the lysins against Staphylococcus aureus NCTC 8325-4 reference strain. (A) Enzymes activities were observed as a turbidity reduction after 1 hr in room temperature either in 1 × PBS or 50 mM glycine buffer, pH 8.0. (B) Lytic activity of the enzymes was determined by SYTOX® fluorescence assay performed in 1 × PBS or 50 mM glycine buffer, pH 8.0, at room temperature for 1 hr. PBS, phosphate-buffered saline.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5036312&req=5

f2: Specific activity of the lysins against Staphylococcus aureus NCTC 8325-4 reference strain. (A) Enzymes activities were observed as a turbidity reduction after 1 hr in room temperature either in 1 × PBS or 50 mM glycine buffer, pH 8.0. (B) Lytic activity of the enzymes was determined by SYTOX® fluorescence assay performed in 1 × PBS or 50 mM glycine buffer, pH 8.0, at room temperature for 1 hr. PBS, phosphate-buffered saline.
Mentions: The LytM_EAD activity is very limited in physiological conditions (PBS),33 however, in low conductivity environment (glycine buffer), LytM_EAD demonstrated great bacteriolytic potential (Fig. 2A). This effect was even more notorious when the fluorometric assay was used. The specific activity of LytM_EAD was the highest among all tested enzymes in the glycine buffer and the lowest in PBS (Fig. 2B). These assays confirmed previous observations of strong inhibition of LytM_EAD bacteriolytic activity by ions present in the reaction environment,33 which might implicate at least one of the mechanisms of LytM activity regulation in vivo.

View Article: PubMed Central - PubMed

ABSTRACT

Staphylococcus aureus remains one of the most common and at the same time the most dangerous bacteria. The spreading antibiotic resistance calls for intensification of research on staphylococcal physiology and development of new strategies for combating this threatening pathogen. We have engineered new chimeric enzymes comprising the enzymatically active domain (EAD) of autolysin LytM from S. aureus and the cell wall binding domain (CBD) from bacteriocin lysostaphin. They display potent activity in extended environmental conditions. Our results exemplify the possibility of exploring autolytic enzymes in engineering lysins with desired features. Moreover, they suggest a possible mechanism of autolysin physiological activity regulation by local ionic environments in the cell wall.

No MeSH data available.


Related in: MedlinePlus