Decoding the Structure–Function Relationship of the Muramidase Domain in E. coli O157.H7 Bacteriophage Endolysin: A Potential Building Block for Chimeric Enzybiotics
Bacteriophage endolysins are potential alternatives to conventional antibiotics for treating multidrug-resistant gram-negative bacterial infections. However, their structure–function relationships are poorly understood, hindering their optimization and application. In this study, we focused on the i...
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Veröffentlicht in: | The Protein Journal 2024-06, Vol.43 (3), p.522-543 |
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Sprache: | eng |
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Zusammenfassung: | Bacteriophage endolysins are potential alternatives to conventional antibiotics for treating multidrug-resistant gram-negative bacterial infections. However, their structure–function relationships are poorly understood, hindering their optimization and application. In this study, we focused on the individual functionality of the C-terminal muramidase domain of Gp127, a modular endolysin from
E. coli
O157:H7 bacteriophage PhaxI. This domain is responsible for the enzymatic activity, whereas the N-terminal domain binds to the bacterial cell wall. Through protein modeling, docking experiments, and molecular dynamics simulations, we investigated the activity, stability, and interactions of the isolated C-terminal domain with its ligand. We also assessed its expression, solubility, toxicity, and lytic activity using the experimental data. Our results revealed that the C-terminal domain exhibits high activity and toxicity when tested individually, and its expression is regulated in different hosts to prevent self-destruction. Furthermore, we validated the muralytic activity of the purified refolded protein by zymography and standardized assays. These findings challenge the need for the N-terminal binding domain to arrange the active site and adjust the gap between crucial residues for peptidoglycan cleavage. Our study shed light on the three-dimensional structure and functionality of muramidase endolysins, thereby enriching the existing knowledge pool and laying a foundation for accurate in
silico
modeling and the informed design of next-generation enzybiotic treatments. |
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ISSN: | 1572-3887 1875-8355 1875-8355 1573-4943 |
DOI: | 10.1007/s10930-024-10195-z |