Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Antimicrobial activity of phage GR1 and its endolysin LysGR1 and LysGR1_EAD.

Article Snippet: To propagate the isolated phage, the lysate of a single bacteriophage GR1 plaque was inoculated with the G. stearothermophilus ATCC 10149 culture (OD 600 = 0.5) and incubated at 50°C for 4 h. After centrifugation (15,000 × g at 4°C for 5 min), the supernatant was filtered using a 0.22 μm pore size filter.

Techniques: Activity Assay

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Modular structure of a novel endolysin LysGR1. (A) Schematic representation of LysGR1 and its EAD and CBD. LysGR1 has a conserved amidase domain as an EAD at the N-terminus and tandem repeats of LysM motifs as a putative CBD at the C-terminus. LysGR1_EAD contains only an amidase-3 domain, and the CBD construct contains two LysM domains. (B) Amino acid sequence alignment of LysGR1 with other endolysins from Bacillus and Sporosarcina infecting phages. Phage_G: N-acetylmuramoyl-L-alanine amidase from Bacillus megaterium phage G, Lietuvens: N-acetylmuramoyl-L-alanine amidase from Sporosarcina phage Lietuvens, vB_BcM_Sam: N-acetylmuramoyl-L-alanine amidase from Bacillus phage vB_BcM_Sam112. Each domain of LysGR1 indicated with colored square bracket: red: Amidase_3; light blue: first LysM; dark blue: second LysM. Conserved residues are shaded in black and gray (dark gray: > 70% conserved; light grey: > 40% conserved). (C) Dose-dependent lytic activity of recombinant LysGR1 against G. stearothermophilus ATCC 10149 cells.

Article Snippet: To propagate the isolated phage, the lysate of a single bacteriophage GR1 plaque was inoculated with the G. stearothermophilus ATCC 10149 culture (OD 600 = 0.5) and incubated at 50°C for 4 h. After centrifugation (15,000 × g at 4°C for 5 min), the supernatant was filtered using a 0.22 μm pore size filter.

Techniques: Construct, Sequencing, Activity Assay, Recombinant

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Antimicrobial and binding activity of LysGR1 and its domains. (A) Comparisons of the lytic activity of LysGR1 and its EAD against G. stearothermophilus ATCC 10149 cells. (B) Time-killing activity of LysGR1 and EAD against G. stearothermophilus ATCC 10149 cells was determined in PBS buffer. The error bars indicate the standard deviation and data were replicated for three independent times and analyzed based on the PBS results by t -test, * p < 0.05, ** p < 0.01, *** p < 0.005. (C) Binding capacity of LysGR1_CBD toward G. stearothermophilus KCTC 3775 cells. (D) Binding spectrum of LysGR1_CBD.

Article Snippet: To propagate the isolated phage, the lysate of a single bacteriophage GR1 plaque was inoculated with the G. stearothermophilus ATCC 10149 culture (OD 600 = 0.5) and incubated at 50°C for 4 h. After centrifugation (15,000 × g at 4°C for 5 min), the supernatant was filtered using a 0.22 μm pore size filter.

Techniques: Binding Assay, Activity Assay, Standard Deviation

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Antimicrobial activity of phage GR1 and its endolysin LysGR1 and LysGR1_EAD.

Article Snippet: The exponentially growing G. stearothermophilus ATCC 10149 cells (OD 600 = 0.6) were washed once and resuspended to OD 600 = 1.0 by PBS buffer and treated with 1.6 μM of LysGR1 and EAD, respectively.

Techniques: Activity Assay

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Modular structure of a novel endolysin LysGR1. (A) Schematic representation of LysGR1 and its EAD and CBD. LysGR1 has a conserved amidase domain as an EAD at the N-terminus and tandem repeats of LysM motifs as a putative CBD at the C-terminus. LysGR1_EAD contains only an amidase-3 domain, and the CBD construct contains two LysM domains. (B) Amino acid sequence alignment of LysGR1 with other endolysins from Bacillus and Sporosarcina infecting phages. Phage_G: N-acetylmuramoyl-L-alanine amidase from Bacillus megaterium phage G, Lietuvens: N-acetylmuramoyl-L-alanine amidase from Sporosarcina phage Lietuvens, vB_BcM_Sam: N-acetylmuramoyl-L-alanine amidase from Bacillus phage vB_BcM_Sam112. Each domain of LysGR1 indicated with colored square bracket: red: Amidase_3; light blue: first LysM; dark blue: second LysM. Conserved residues are shaded in black and gray (dark gray: > 70% conserved; light grey: > 40% conserved). (C) Dose-dependent lytic activity of recombinant LysGR1 against G. stearothermophilus ATCC 10149 cells.

Article Snippet: The exponentially growing G. stearothermophilus ATCC 10149 cells (OD 600 = 0.6) were washed once and resuspended to OD 600 = 1.0 by PBS buffer and treated with 1.6 μM of LysGR1 and EAD, respectively.

Techniques: Construct, Sequencing, Activity Assay, Recombinant

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Antimicrobial and binding activity of LysGR1 and its domains. (A) Comparisons of the lytic activity of LysGR1 and its EAD against G. stearothermophilus ATCC 10149 cells. (B) Time-killing activity of LysGR1 and EAD against G. stearothermophilus ATCC 10149 cells was determined in PBS buffer. The error bars indicate the standard deviation and data were replicated for three independent times and analyzed based on the PBS results by t -test, * p < 0.05, ** p < 0.01, *** p < 0.005. (C) Binding capacity of LysGR1_CBD toward G. stearothermophilus KCTC 3775 cells. (D) Binding spectrum of LysGR1_CBD.

Article Snippet: The exponentially growing G. stearothermophilus ATCC 10149 cells (OD 600 = 0.6) were washed once and resuspended to OD 600 = 1.0 by PBS buffer and treated with 1.6 μM of LysGR1 and EAD, respectively.

Techniques: Binding Assay, Activity Assay, Standard Deviation

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Antimicrobial activity of phage GR1 and its endolysin LysGR1 and LysGR1_EAD.

Article Snippet: Geobacillus stearothermophilus phage GR1 was isolated from roadside soil, in Seoul, using G. stearothermophilus ATCC 10149 as a host.

Techniques: Activity Assay

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Modular structure of a novel endolysin LysGR1. (A) Schematic representation of LysGR1 and its EAD and CBD. LysGR1 has a conserved amidase domain as an EAD at the N-terminus and tandem repeats of LysM motifs as a putative CBD at the C-terminus. LysGR1_EAD contains only an amidase-3 domain, and the CBD construct contains two LysM domains. (B) Amino acid sequence alignment of LysGR1 with other endolysins from Bacillus and Sporosarcina infecting phages. Phage_G: N-acetylmuramoyl-L-alanine amidase from Bacillus megaterium phage G, Lietuvens: N-acetylmuramoyl-L-alanine amidase from Sporosarcina phage Lietuvens, vB_BcM_Sam: N-acetylmuramoyl-L-alanine amidase from Bacillus phage vB_BcM_Sam112. Each domain of LysGR1 indicated with colored square bracket: red: Amidase_3; light blue: first LysM; dark blue: second LysM. Conserved residues are shaded in black and gray (dark gray: > 70% conserved; light grey: > 40% conserved). (C) Dose-dependent lytic activity of recombinant LysGR1 against G. stearothermophilus ATCC 10149 cells.

Article Snippet: Geobacillus stearothermophilus phage GR1 was isolated from roadside soil, in Seoul, using G. stearothermophilus ATCC 10149 as a host.

Techniques: Construct, Sequencing, Activity Assay, Recombinant

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Antimicrobial and binding activity of LysGR1 and its domains. (A) Comparisons of the lytic activity of LysGR1 and its EAD against G. stearothermophilus ATCC 10149 cells. (B) Time-killing activity of LysGR1 and EAD against G. stearothermophilus ATCC 10149 cells was determined in PBS buffer. The error bars indicate the standard deviation and data were replicated for three independent times and analyzed based on the PBS results by t -test, * p < 0.05, ** p < 0.01, *** p < 0.005. (C) Binding capacity of LysGR1_CBD toward G. stearothermophilus KCTC 3775 cells. (D) Binding spectrum of LysGR1_CBD.

Article Snippet: Geobacillus stearothermophilus phage GR1 was isolated from roadside soil, in Seoul, using G. stearothermophilus ATCC 10149 as a host.

Techniques: Binding Assay, Activity Assay, Standard Deviation

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Antimicrobial activity of phage GR1 and its endolysin LysGR1 and LysGR1_EAD.

Article Snippet: A turbidity reduction assay showed that the lytic activity of LysGR1 against G. stearothermophilus ATCC 10149 cells is dose-dependent , while it also exhibited a broad lytic spectrum against all the tested G. stearothermophilus strains ( ).

Techniques: Activity Assay

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Modular structure of a novel endolysin LysGR1. (A) Schematic representation of LysGR1 and its EAD and CBD. LysGR1 has a conserved amidase domain as an EAD at the N-terminus and tandem repeats of LysM motifs as a putative CBD at the C-terminus. LysGR1_EAD contains only an amidase-3 domain, and the CBD construct contains two LysM domains. (B) Amino acid sequence alignment of LysGR1 with other endolysins from Bacillus and Sporosarcina infecting phages. Phage_G: N-acetylmuramoyl-L-alanine amidase from Bacillus megaterium phage G, Lietuvens: N-acetylmuramoyl-L-alanine amidase from Sporosarcina phage Lietuvens, vB_BcM_Sam: N-acetylmuramoyl-L-alanine amidase from Bacillus phage vB_BcM_Sam112. Each domain of LysGR1 indicated with colored square bracket: red: Amidase_3; light blue: first LysM; dark blue: second LysM. Conserved residues are shaded in black and gray (dark gray: > 70% conserved; light grey: > 40% conserved). (C) Dose-dependent lytic activity of recombinant LysGR1 against G. stearothermophilus ATCC 10149 cells.

Article Snippet: A turbidity reduction assay showed that the lytic activity of LysGR1 against G. stearothermophilus ATCC 10149 cells is dose-dependent , while it also exhibited a broad lytic spectrum against all the tested G. stearothermophilus strains ( ).

Techniques: Construct, Sequencing, Activity Assay, Recombinant

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Antimicrobial and binding activity of LysGR1 and its domains. (A) Comparisons of the lytic activity of LysGR1 and its EAD against G. stearothermophilus ATCC 10149 cells. (B) Time-killing activity of LysGR1 and EAD against G. stearothermophilus ATCC 10149 cells was determined in PBS buffer. The error bars indicate the standard deviation and data were replicated for three independent times and analyzed based on the PBS results by t -test, * p < 0.05, ** p < 0.01, *** p < 0.005. (C) Binding capacity of LysGR1_CBD toward G. stearothermophilus KCTC 3775 cells. (D) Binding spectrum of LysGR1_CBD.

Article Snippet: A turbidity reduction assay showed that the lytic activity of LysGR1 against G. stearothermophilus ATCC 10149 cells is dose-dependent , while it also exhibited a broad lytic spectrum against all the tested G. stearothermophilus strains ( ).

Techniques: Binding Assay, Activity Assay, Standard Deviation

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Biochemical properties of LysGR1 and its EAD. The effects of temperature (A) , NaCl (B) , and pH (C) on the lytic activity of LysGR1 and the effects of temperature (D) , NaCl (E) , and pH (F) on the lytic activity of LysGR1_EAD against B. amyloliquefaciens KACC 15877 cells. Each column represents the mean of triplicate experiments, and the error bars indicate the standard deviation.

Article Snippet: A turbidity reduction assay showed that the lytic activity of LysGR1 against G. stearothermophilus ATCC 10149 cells is dose-dependent , while it also exhibited a broad lytic spectrum against all the tested G. stearothermophilus strains ( ).

Techniques: Activity Assay, Standard Deviation

Journal: Frontiers in Microbiology

Article Title: LysGR1, a novel thermostable endolysin from Geobacillus stearothermophilus bacteriophage GR1

doi: 10.3389/fmicb.2023.1178748

Figure Lengend Snippet: Biofilm reduction activity of LysGR1 (A) and its EAD (B) in 24-well polystyrene microplates. G. stearothermophilus ATCC 12980 biofilms on the surface of polystyrene were treated with various concentrations of LysGR1 and its EAD for 1 h. Each column represents the mean of triplicate experiments, and the error bars indicate the standard deviation. Data were analyzed based on the buffer-treated negative control by t -test, * p < 0.05, ** p < 0.01, *** p < 0.005.

Article Snippet: A turbidity reduction assay showed that the lytic activity of LysGR1 against G. stearothermophilus ATCC 10149 cells is dose-dependent , while it also exhibited a broad lytic spectrum against all the tested G. stearothermophilus strains ( ).

Techniques: Activity Assay, Standard Deviation, Negative Control