scholarly journals Improving the Lethal Effect of Cpl-7, a Pneumococcal Phage Lysozyme with Broad Bactericidal Activity, by Inverting the Net Charge of Its Cell Wall-Binding Module

2014 ◽  
Vol 58 (5) ◽  
pp. 2996-2996
Author(s):  
R. Diez-Martinez ◽  
H. D. de Paz ◽  
N. Bustamante ◽  
E. Garcia ◽  
M. Menendez ◽  
...  
Keyword(s):  
Cell Wall ◽  
Bactericidal Activity ◽  
Lethal Effect ◽  
Net Charge ◽  
Cell Wall Binding

scholarly journals Improving the Lethal Effect of Cpl-7, a Pneumococcal Phage Lysozyme with Broad Bactericidal Activity, by Inverting the Net Charge of Its Cell Wall-Binding Module

2013 ◽  
Vol 57 (11) ◽  
pp. 5355-5365 ◽  
Author(s):  
Roberto Díez-Martínez ◽  
Héctor de Paz ◽  
Noemí Bustamante ◽  
Ernesto García ◽  
Margarita Menéndez ◽  
...  
Keyword(s):  
Cell Wall ◽  
Lethal Effect ◽  
Lytic Activity ◽  
Net Charge ◽  
Killing Effect ◽  
Content Type ◽  
Cell Wall Binding ◽  
Pass Through ◽  
Multiresistant Strains

ABSTRACTPhage endolysins are murein hydrolases that break the bacterial cell wall to provoke lysis and release of phage progeny. Recently, these enzymes have also been recognized as powerful and specific antibacterial agents when added exogenously. In the pneumococcal system, most cell wall associated murein hydrolases reported so far depend on choline for activity, and Cpl-7 lysozyme constitutes a remarkable exception. Here, we report the improvement of the killing activity of the Cpl-7 endolysin by inversion of the sign of the charge of the cell wall-binding module (from −14.93 to +3.0 at neutral pH). The engineered variant, Cpl-7S, has 15 amino acid substitutions and an improved lytic activity againstStreptococcus pneumoniae(including multiresistant strains),Streptococcus pyogenes, and other pathogens. Moreover, we have demonstrated that a single 25-μg dose of Cpl-7S significantly increased the survival rate of zebrafish embryos infected withS. pneumoniaeorS. pyogenes, confirming the killing effect of Cpl-7Sin vivo. Interestingly, Cpl-7S, in combination with 0.01% carvacrol (an essential oil), was also found to efficiently kill Gram-negative bacteria such asEscherichia coliandPseudomonas putida, an effect not described previously. Our findings provide a strategy to improve the lytic activity of phage endolysins based on facilitating their pass through the negatively charged bacterial envelope, and thereby their interaction with the cell wall target, by modulating the net charge of the cell wall-binding modules.

scholarly journals A Putative Amidase Endolysin Encoded by Clostridium perfringens St13 Exhibits Specific Lytic Activity and Synergizes with the Muramidase Endolysin Psm

Antibiotics ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 245
Author(s):  
Hiroshi Sekiya ◽  
Maho Okada ◽  
Eiji Tamai ◽  
Toshi Shimamoto ◽  
Tadashi Shimamoto ◽  
...  
Keyword(s):  
Cell Wall ◽  
Clostridium Perfringens ◽  
Antimicrobial Agents ◽  
Catalytic Domain ◽  
Food Poisoning ◽  
Binding Activity ◽  
Lytic Activity ◽  
Intestinal Bacterium ◽  
Terminal Domain ◽  
Cell Wall Binding

Clostridium perfringens is an often-harmful intestinal bacterium that causes various diseases ranging from food poisoning to life-threatening fulminant disease. Potential treatments include phage-derived endolysins, a promising family of alternative antimicrobial agents. We surveyed the genome of the C. perfringens st13 strain and identified an endolysin gene, psa, in the phage remnant region. Psa has an N-terminal catalytic domain that is homologous to the amidase_2 domain, and a C-terminal domain of unknown function. psa and gene derivatives encoding various Psa subdomains were cloned and expressed in Escherichia coli as N-terminal histidine-tagged proteins. Purified His-tagged full-length Psa protein (Psa-his) showed C. perfringens-specific lytic activity in turbidity reduction assays. In addition, we demonstrated that the uncharacterized C-terminal domain has cell wall-binding activity. Furthermore, cell wall-binding measurements showed that Psa binding was highly specific to C. perfringens. These results indicated that Psa is an amidase endolysin that specifically lyses C. perfringens; the enzyme’s specificity is highly dependent on the binding of the C-terminal domain. Moreover, Psa was shown to have a synergistic effect with another C. perfringens-specific endolysin, Psm, which is a muramidase that cleaves peptidoglycan at a site distinct from that targeted by Psa. The combination of Psa and Psm may be effective in the treatment and prevention of C. perfringens infections.

Effect of a yeast-cell-wall glucan on the bactericidal activity of rainbow trout macrophages

1993 ◽  
Vol 3 (4) ◽  
pp. 267-277 ◽  
Author(s):  
Jorunn B. Jørgensen ◽  
Gregory J.E. Sharp ◽  
Christopher J. Secombes ◽  
Børre Robertsen
Keyword(s):  
Cell Wall ◽  
Rainbow Trout ◽  
Yeast Cell ◽  
Bactericidal Activity ◽  
Yeast Cell Wall

scholarly journals The structure of Rv3717 reveals a novel amidase fromMycobacterium tuberculosis

2013 ◽  
Vol 69 (12) ◽  
pp. 2543-2554 ◽  
Author(s):  
Atul Kumar ◽  
Sanjiv Kumar ◽  
Dilip Kumar ◽  
Arpit Mishra ◽  
Rikeshwer P. Dewangan ◽  
...  
Keyword(s):  
Cell Wall ◽  
Bartonella Henselae ◽  
General Mechanism ◽  
Isolation And Identification ◽  
Cell Wall Binding ◽  
New Family ◽  
A Cell ◽  
Sad Phasing ◽  
Cell Wall Binding Domain ◽  
Net Positive Charge

BacterialN-acetylmuramoyl-L-alanine amidases are cell-wall hydrolases that hydrolyze the bond betweenN-acetylmuramic acid and L-alanine in cell-wall glycopeptides. Rv3717 ofMycobacterium tuberculosishas been identified as a unique autolysin that lacks a cell-wall-binding domain (CBD) and its structure has been determined to 1.7 Å resolution by the Pt-SAD phasing method. Rv3717 possesses an α/β-fold and is a zinc-dependent hydrolase. The structure reveals a short flexible hairpin turn that partially occludes the active site and may be involved in autoregulation. This type of autoregulation of activity of PG hydrolases has been observed inBartonella henselaeamidase (AmiB) and may be a general mechanism used by some of the redundant amidases to regulate cell-wall hydrolase activity in bacteria. Rv3717 utilizes its net positive charge for substrate binding and exhibits activity towards a broad spectrum of substrate cell walls. The enzymatic activity of Rv3717 was confirmed by isolation and identification of its enzymatic products by LC/MS. These studies indicate that Rv3717, anN-acetylmuramoyl-L-alanine amidase fromM. tuberculosis, represents a new family of lytic amidases that do not have a separate CBD and are regulated conformationally.

scholarly journals Telavancin, a Multifunctional Lipoglycopeptide, Disrupts both Cell Wall Synthesis and Cell Membrane Integrity in Methicillin-Resistant Staphylococcus aureus

2005 ◽  
Vol 49 (3) ◽  
pp. 1127-1134 ◽  
Author(s):  
Deborah L. Higgins ◽  
Ray Chang ◽  
Dmitri V. Debabov ◽  
Joey Leung ◽  
Terry Wu ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Membrane Potential ◽  
Cell Membrane ◽  
Bactericidal Activity ◽  
Bacterial Cell ◽  
Cell Membrane Potential ◽  
Gram Positive ◽  
Methicillin Resistant ◽  
Bacterial Cell Membrane

ABSTRACTThe emergence and spread of multidrug-resistant gram-positive bacteria represent a serious clinical problem. Telavancin is a novel lipoglycopeptide antibiotic that possesses rapid in vitro bactericidal activity against a broad spectrum of clinically relevant gram-positive pathogens. Here we demonstrate that telavancin's antibacterial activity derives from at least two mechanisms. As observed with vancomycin, telavancin inhibited late-stage peptidoglycan biosynthesis in a substrate-dependent fashion and bound the cell wall, as it did the lipid II surrogate tripeptideN,N′-diacetyl-l-lysinyl-d-alanyl-d-alanine, with high affinity. Telavancin also perturbed bacterial cell membrane potential and permeability. In methicillin-resistantStaphylococcus aureus, telavancin caused rapid, concentration-dependent depolarization of the plasma membrane, increases in permeability, and leakage of cellular ATP and K+. The timing of these changes correlated with rapid , concentration-dependent loss of bacterial viability, suggesting that the early bactericidal activity of telavancin results from dissipation of cell membrane potential and an increase in membrane permeability. Binding and cell fractionation studies provided direct evidence for an interaction of telavancin with the bacterial cell membrane; stronger binding interactions were observed with the bacterial cell wall and cell membrane relative to vancomycin. We suggest that this multifunctional mechanism of action confers advantageous antibacterial properties.

Lateral flow assay-based bacterial detection using engineered cell wall binding domains of a phage endolysin

2017 ◽  
Vol 96 ◽  
pp. 173-177 ◽  
Author(s):  
Minsuk Kong ◽  
Joong Ho Shin ◽  
Sunggi Heu ◽  
Je-Kyun Park ◽  
Sangryeol Ryu
Keyword(s):  
Cell Wall ◽  
Lateral Flow ◽  
Lateral Flow Assay ◽  
Bacterial Detection ◽  
Binding Domains ◽  
Cell Wall Binding

scholarly journals Rapid Multiplex Detection and Differentiation of Listeria Cells by Use of Fluorescent Phage Endolysin Cell Wall Binding Domains

2010 ◽  
Vol 76 (17) ◽  
pp. 5745-5756 ◽  
Author(s):  
Mathias Schmelcher ◽  
Tatiana Shabarova ◽  
Marcel R. Eugster ◽  
Fritz Eichenseher ◽  
Vincent S. Tchang ◽  
...  
Keyword(s):  
Cell Wall ◽  
Modular Organization ◽  
Binding Assays ◽  
Surface Plasmon Resonance Analysis ◽  
Fluorescent Marker ◽  
Binding Domains ◽  
Cell Wall Binding ◽  
Fluorescent Markers ◽  
Severe Infections ◽  
And Control

ABSTRACT The genus Listeria comprises food-borne pathogens associated with severe infections and a high mortality rate. Endolysins from bacteriophages infecting Listeria are promising tools for both their detection and control. These proteins feature a modular organization, consisting of an N-terminal enzymatically active domain (EAD), which contributes lytic activity, and a C-terminal cell wall binding domain (CBD), which targets the lysin to its substrate. Sequence comparison among 12 different endolysins revealed high diversity among the enzyme's functional domains and allowed classification of their CBDs into two major groups and five subclasses. This diversity is reflected in various binding properties, as determined by cell wall binding assays using CBDs fused to fluorescent marker proteins. Although some proteins exhibited a broad binding range and recognize Listeria strains representing all serovars, others target specific serovars only. The CBDs also differed with respect to the number and distribution of ligands recognized on the cells, as well as their binding affinities. Surface plasmon resonance analysis revealed equilibrium affinities in the pico- to nanomolar ranges for all proteins except CBD006, which is due to an internal truncation. Rapid multiplexed detection and differentiation of Listeria strains in mixed bacterial cultures was possible by combining CBDs of different binding specificities with fluorescent markers of various colors. In addition, cells of different Listeria strains could be recovered from artificially contaminated milk or cheese by CBD-based magnetic separation by using broad-range CBDP40 and subsequently identified after incubation with two differently colored CBD fusion proteins of higher specificity.

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