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

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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

Activity of the enzymes against various bacterial strains was measured as OD595 decrease of tested bacterial strains incubated 1 hr in room temperature with 100 nM enzymes in 50 mM glycine buffer, pH 8.0, additionally supplemented with 100 mM NaCl for Chimera and lysostaphin.
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f3: Activity of the enzymes against various bacterial strains was measured as OD595 decrease of tested bacterial strains incubated 1 hr in room temperature with 100 nM enzymes in 50 mM glycine buffer, pH 8.0, additionally supplemented with 100 mM NaCl for Chimera and lysostaphin.

Mentions: One of the effects of fusing domains from various lysins is altered, usually extended specificity. LytM_EAD as well as Lss_CBD and its homologue from Staphylococcus capitis—EPK1, were reported to recognize and bind the same fragment of PGNs–pentaglycine crossbridges.22,25,37 We have confirmed that the specificity of those domains in the newly generated fusion enzymes has not been changed and Chimera sustained their activity against Staphylococcus strains, including MRSA. The enzyme is not active against other Gram-positive and Gram-negative bacteria. LytM_EAD, lysostaphin, and Chimera activities were tested against three reference MSSA strains, three MRSA strains, and selected Gram-negative bacteria, namely Pseudomonas aeruginosa, Escherichia coli, and Bacillus subtilis (Fig. 3). Engineered Chimera appeared slightly more effective in lysing Staphylococcus strains than the original enzymes.


LytM Fusion with SH3b-Like Domain Expands Its Activity to Physiological Conditions
Activity of the enzymes against various bacterial strains was measured as OD595 decrease of tested bacterial strains incubated 1 hr in room temperature with 100 nM enzymes in 50 mM glycine buffer, pH 8.0, additionally supplemented with 100 mM NaCl for Chimera and lysostaphin.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

f3: Activity of the enzymes against various bacterial strains was measured as OD595 decrease of tested bacterial strains incubated 1 hr in room temperature with 100 nM enzymes in 50 mM glycine buffer, pH 8.0, additionally supplemented with 100 mM NaCl for Chimera and lysostaphin.
Mentions: One of the effects of fusing domains from various lysins is altered, usually extended specificity. LytM_EAD as well as Lss_CBD and its homologue from Staphylococcus capitis—EPK1, were reported to recognize and bind the same fragment of PGNs–pentaglycine crossbridges.22,25,37 We have confirmed that the specificity of those domains in the newly generated fusion enzymes has not been changed and Chimera sustained their activity against Staphylococcus strains, including MRSA. The enzyme is not active against other Gram-positive and Gram-negative bacteria. LytM_EAD, lysostaphin, and Chimera activities were tested against three reference MSSA strains, three MRSA strains, and selected Gram-negative bacteria, namely Pseudomonas aeruginosa, Escherichia coli, and Bacillus subtilis (Fig. 3). Engineered Chimera appeared slightly more effective in lysing Staphylococcus strains than the original enzymes.

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