Catalytic diversity and cell wall binding repeats in the phage‐encoded endolysins

Broendum, Sebastian S.; Buckle, Ashley M.; McGowan, Sheena

doi:10.1111/mmi.14134

Cited by 78 publications

(81 citation statements)

References 108 publications

(174 reference statements)

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“…However, there are structural and functional differences between endolysins targeting Gram-positive and Gram-negative hosts [3,4]. Several "modes" of action are characteristic for endolysins, including the ability to hydrolyze either or both glycosidic and amide (including peptide) bonds [3,5].…”

Section: Introductionmentioning

confidence: 99%

Modulation of Endolysin LysECD7 Bactericidal Activity by Different Peptide Tag Fusion

et al. 2020

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The use of recombinant endolysins is a promising approach for antimicrobial therapy capable of counteracting the spread of antibiotic-resistant strains. To obtain the necessary biotechnological product, diverse peptide tags are often fused to the endolysin sequence to simplify enzyme purification, improve its ability to permeabilize the bacterial outer membrane, etc. We compared the effects of two different types of protein modifications on endolysin LysECD7 bactericidal activity in vitro and demonstrated that it is significantly modulated by specific permeabilizing antimicrobial peptides, as well as by widely used histidine tags. Thus, the tags selected for the study of endolysins and during the development of biotechnological preparations should be used with the appropriate precautions to minimize false conclusions about endolysin properties. Further, modifications of LysECD7 allowed us to obtain a lytic enzyme that was largely devoid of the disadvantages of the native protein and was active over the spectra of conditions, with high in vitro bactericidal activity not only against Gram-negative, but also against Gram-positive, bacteria. This opens up the possibility of developing effective antimicrobials based on N-terminus sheep myeloid peptide of 29 amino acids (SMAP)-modified LysECD7 that can be highly active not only during topical treatment but also for systemic applications in the bloodstream and tissues.

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Section: Introductionmentioning

confidence: 99%

Modulation of Endolysin LysECD7 Bactericidal Activity by Different Peptide Tag Fusion

et al. 2020

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“…Many cysteine peptidases containing the NlpC/P60 domain were characterized to date (1,2,24,26,30,(33)(34)(35), several of which displaying a four-domain organization similar to PnpA. However, until now, only the three-dimensional structure of DvLysin from Desulfovibrio vulgaris was reported, with a N-terminal "c-clip" or "N_NLPC_P60" stabilizing domain, two SH3b domains and a C-terminal NlpC/p60 cysteine peptidase domain (26).…”

Section: Discussionmentioning

confidence: 99%

“…NlpC/P60containing peptidases are involved in the catalysis of the N-acetylmuramate-L-alanine or D-γglutamyl-mesodiaminopimelate linkages, with four major groups identified so far: (i) P60-like, (ii) AcmB/LytN-like, (iii) YaeF/Poxvirus G6R, and (iv) lecithin retinol acyltransferase (LRAT)like (24). The NlpC/P60 domain is structurally similar to a primitive papain-like peptidase (24)(25)(26)(27)(28)(29) and can be found alone or fused to other domains, with or without catalytic functions, to form multifunctional proteins (1,2,24,26,(30)(31)(32)(33)(34)(35). Several of these domains, such as the SH3 (sarcoma homology 3) domain (31,32,35), are involved in anchoring hydrolases to cell wall components, allowing their appropriate concentration and positioning for the formation of an efficient enzyme-substrate complex (1).…”

Section: Introductionmentioning

confidence: 99%

A secreted NlpC/P60 endopeptidase fromPhotobacterium damselaesubsp.piscicidacleaves the peptidoglycan of potentially competing bacteria

Lisboa

Pereira

Rifflet

et al. 2020

Preprint

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ABSTRACTPeptidoglycan (PG) is a major component of the bacterial cell wall, forming a mesh-like structure enwrapping the bacteria that is essential for maintaining structural integrity and providing support for anchoring other components of the cell envelope. PG biogenesis is highly dynamic and requires multiple enzymes, including several hydrolases that cleave glycosidic or amide bonds in the PG. Here, it is described the structural and functional characterization of an NlpC/P60-containing peptidase from Photobacterium damselae subsp. piscicida (Phdp), a Gram-negative bacterium that causes high mortality of warm-water marine fish with great impact for the aquaculture industry. PnpA (PhotobacteriumNlpC-like Protein A) has a four-domain structure with a hydrophobic and narrow access to the catalytic center and specificity for the γ-D-glutamyl-meso-diaminopimelic acid bond. However, PnpA does not cleave the PG of Phdp and neither PG of several Gram-negative and Gram-positive bacterial species. Interestingly, it is secreted by the Phdp type II secretion system and degrades the PG of Vibrio anguillarum and V. vulnificus. This suggests that PnpA is used by Phdp to gain an advantage over bacteria that compete for the same resources or to obtain nutrients in nutrient-scarce environments. Comparison of the muropeptide composition of PG susceptible and resistant to the catalytic activity of PnpA, showed that the global content of muropeptides is similar, suggesting that susceptibility to PnpA is determined by the three-dimensional organization of the muropeptides in the PG.IMPORTANCEPeptidoglycan (PG) is a major component of the bacterial cell wall formed by long chains of two alternating sugars interconnected by short peptides, originating a mesh-like structure that enwraps the bacterial cell. Although PG provides structural integrity and support for anchoring other components of the cell envelope, it is constantly being remodeled through the action of specific enzymes that cleave or joint its components. Here, it is shown that Photobacterium damselae subsp. piscicida, a bacterium that causes high mortality in warm-water marine fish, produces PnpA, an enzyme that is secreted into the environment and is able to cleave the PG of potentially competing bacteria, either for gaining competitive advantages and/or to get nutrients. The specificity of PnpA to the PG of some bacteria and its inability to cleave others may be explained by differences in the structure of the PG mesh and not by different muropeptide composition.

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“…Cocktails of multiple distinct phages, each with complementary infective ranges against isolates of any one target species, may be combined to both provide broad strain coverage and reduce the potential for evolving resistance. Another antimicrobial strategy is based on a large family of phage-encoded peptidoglycan hydrolases called endolysins (7)(8)(9)(10). Endolysins normally act at the end of the phage life cycle by translocating from the cytoplasm to the cell wall compartment and cleaving the peptidoglycan matrix, resulting in osmotic lysis and progeny virion release.…”

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confidence: 99%

The PlyB Endolysin of Bacteriophage vB_BanS_Bcp1 Exhibits Broad-Spectrum Bactericidal Activity against Bacillus cereus Sensu Lato Isolates

Schuch

Pelzek

Nelson

et al. 2019

Appl Environ Microbiol

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Lytic bacteriophages (or phages) drive bacterial mortality by elaborating exquisite abilities to bind, breach, and destroy bacterial cell membranes and subjugate critical bacterial cell functions. These antimicrobial activities make phages ideal candidates to serve as, or provide sources of, biological control measures for bacterial pathogens. In this study, we isolated the Myoviridae phage vB_BanS_Bcp1 (here referred to as Bcp1) from landfill soil, using a Bacillus anthracis host. The antimicrobial activities of both Bcp1 and its encoded endolysin, PlyB, were examined across different B. cereus sensu lato group species, including B. cereus sensu stricto, Bacillus thuringiensis, and Bacillus anthracis, with pathogenic potential in humans and multiple different uses in biotechnological applications. The Bcp1 phage infected only a subset (11 to 66%) of each B. cereus sensu lato species group tested. In contrast, functional analysis of purified PlyB revealed a potent bacteriolytic activity against all B. cereus sensu lato isolates tested (n ϭ 79). PlyB was, furthermore, active across broad temperature, pH, and salt ranges, refractory to the development of resistance, bactericidal as a single agent, and synergistic with a second endolysin, PlyG. To confirm the potential for PlyB as an antimicrobial agent, we demonstrated the efficacy of a single intravenous treatment with PlyB alone or combination with PlyG in a murine model of lethal B. anthracis infection. Overall, our findings show exciting potential for the Bcp1 bacteriophage and the PlyB endolysin as potential new additions to the antimicrobial armamentarium. IMPORTANCE Organisms of the Bacillus cereus sensu lato lineage are ubiquitous in the environment and are responsible for toxin-mediated infections ranging from severe food poisoning (B. cereus sensu stricto) to anthrax (Bacillus anthracis). The increasing incidence of many of these infections, combined with the specter of antibiotic resistance, has created a need for novel antimicrobials with potent activity, including bacteriophages (or phages) and phage-encoded products (i.e., endolysins). In this study, we describe a broadly infective phage, Bcp1, and its encoded endolysin, PlyB, which exhibited a rapidly bacteriolytic effect against all B. cereus sensu lato isolates tested with no evidence of evolving resistance. Importantly, PlyB was highly efficacious in a mouse model of lethal bacteremia with B. anthracis. Both the Bcp1 phage and the PlyB endolysin represent novel mechanisms of action compared to antibiotics, with potential applications to address the evolving problem of antimicrobial resistance.

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Catalytic diversity and cell wall binding repeats in the phage‐encoded endolysins

Cited by 78 publications

References 108 publications

Modulation of Endolysin LysECD7 Bactericidal Activity by Different Peptide Tag Fusion

Modulation of Endolysin LysECD7 Bactericidal Activity by Different Peptide Tag Fusion

A secreted NlpC/P60 endopeptidase fromPhotobacterium damselaesubsp.piscicidacleaves the peptidoglycan of potentially competing bacteria

The PlyB Endolysin of Bacteriophage vB_BanS_Bcp1 Exhibits Broad-Spectrum Bactericidal Activity against Bacillus cereus Sensu Lato Isolates

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