The Experts below are selected from a list of 108 Experts worldwide ranked by ideXlab platform
Christopher M. Overall - One of the best experts on this subject based on the ideXlab platform.
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Discovery of a proteolytic flagellin family in diverse bacterial phyla that assembles enzymatically active flagella
Nature Communications, 2017Co-Authors: Ulrich Eckhard, Hina Bandukwala, Michael J. Mansfield, Giada Marino, Jiujun Cheng, Iain Wallace, Todd Holyoak, Trevor C. Charles, John Austin, Christopher M. OverallAbstract:Bacterial flagella are cell locomotion and occasional adhesion organelles composed primarily of the polymeric protein flagellin, but to date have not been associated with any enzymatic function. Here, we report the bioinformatics-driven discovery of a class of enzymatic flagellins that assemble to form proteolytically active flagella. Originating by a metallopeptidase insertion into the central flagellin hypervariable region, this flagellin family has expanded to at least 74 bacterial species. In the pathogen, Clostridium haemolyticum , metallopeptidase-containing flagellin (which we termed flagellinolysin) is the second most abundant protein in the flagella and is localized to the extracellular flagellar surface. Purified flagellar filaments and recombinant flagellin exhibit proteolytic activity, cleaving nearly 1000 different peptides. With ~ 20,000 flagellin copies per ~ 10-μm flagella this assembles the largest proteolytic complex known. Flagellum-mediated extracellular proteolysis expands our understanding of the functional plasticity of bacterial flagella, revealing this family as enzymatic biopolymers that mediate interactions with diverse peptide substrates. So far no enzymatic activity has been attributed to flagellin, the major component of bacterial flagella. Here the authors use bioinformatic analysis and identify a metallopeptidase insertion in flagellins from 74 bacterial species and show that recombinant flagellin and flagellar filaments have proteolytic activity.
Ulrich Eckhard - One of the best experts on this subject based on the ideXlab platform.
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Identification and characterization of the proteolytic flagellin from the common freshwater bacterium Hylemonella gracilis.
Scientific Reports, 2020Co-Authors: Ulrich Eckhard, Michael J. Mansfield, Constantin Blöchl, Benjamin Jenkins, Christian G. Huber, Andrew C. Doxey, Hans BrandstetterAbstract:Flagellins are the protein components of bacterial flagella and assemble in up to 20,000 copies to form extracellular flagellar filaments. An unusual family of flagellins was recently discovered that contains a unique metalloprotease domain within its surface-exposed hypervariable region. To date, these proteolytic flagellins (also termed flagellinolysins) have only been characterized in the Gram-positive organism Clostridium haemolyticum, where flagellinolysin was shown to be proteolytically active and capable of cleaving extracellular protein substrates. The biological function of flagellinolysin and its activity in other organisms, however, remain unclear. Here, using molecular biochemistry and proteomics, we have performed an initial characterization of a novel flagellinolysin identified from Hylemonella gracilis, a Gram-negative organism originally isolated from pond water. We demonstrate that H. gracilis flagellinolysin (HgrFlaMP) is an active calcium-dependent zinc metallopeptidase and characterize its cleavage specificity profile using both trypsin and GluC-derived peptide libraries and protein substrates. Based on high-throughput degradomic assays, HgrFlaMP cleaved 784 unique peptides and displayed a cleavage site specificity similar to flagellinolysin from C. haemolyticum. Additionally, by using a set of six protein substrates, we identified 206 protein-embedded cleavage sites, further refining the substrate preference of HgrFlaMP, which is dominated by large hydrophobic amino acids in P1', and small hydrophobic or medium-sized polar residues on the amino-terminal side of the scissile bond. Intriguingly, recombinant HgrFlaMP was also capable of cleaving full-length flagellins from another species, suggesting its potential involvement in interbacterial interactions. Our study reports the first experimentally characterized proteolytic flagellin in a Gram-negative organism, and provides new insights into flagellum-mediated enzymatic activity.
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Discovery of a proteolytic flagellin family in diverse bacterial phyla that assembles enzymatically active flagella
Nature Communications, 2017Co-Authors: Ulrich Eckhard, Hina Bandukwala, Michael J. Mansfield, Giada Marino, Jiujun Cheng, Iain Wallace, Todd Holyoak, Trevor C. Charles, John Austin, Christopher M. OverallAbstract:Bacterial flagella are cell locomotion and occasional adhesion organelles composed primarily of the polymeric protein flagellin, but to date have not been associated with any enzymatic function. Here, we report the bioinformatics-driven discovery of a class of enzymatic flagellins that assemble to form proteolytically active flagella. Originating by a metallopeptidase insertion into the central flagellin hypervariable region, this flagellin family has expanded to at least 74 bacterial species. In the pathogen, Clostridium haemolyticum , metallopeptidase-containing flagellin (which we termed flagellinolysin) is the second most abundant protein in the flagella and is localized to the extracellular flagellar surface. Purified flagellar filaments and recombinant flagellin exhibit proteolytic activity, cleaving nearly 1000 different peptides. With ~ 20,000 flagellin copies per ~ 10-μm flagella this assembles the largest proteolytic complex known. Flagellum-mediated extracellular proteolysis expands our understanding of the functional plasticity of bacterial flagella, revealing this family as enzymatic biopolymers that mediate interactions with diverse peptide substrates. So far no enzymatic activity has been attributed to flagellin, the major component of bacterial flagella. Here the authors use bioinformatic analysis and identify a metallopeptidase insertion in flagellins from 74 bacterial species and show that recombinant flagellin and flagellar filaments have proteolytic activity.
Yutaka Tamura - One of the best experts on this subject based on the ideXlab platform.
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phylogenetic analysis and pcr detection of Clostridium chauvoei Clostridium haemolyticum Clostridium novyi types a and b and Clostridium septicum based on the flagellin gene
Veterinary Microbiology, 2002Co-Authors: Yoshimasa Sasaki, Noriyasu Takikawa, Akemi Kojima, Hiroshi Aoki, Yasuaki Ogikubo, Yutaka TamuraAbstract:Abstract The flagellin genes ( fliC ) of Clostridium chauvoei , Clostridium haemolyticum , Clostridium novyi types A and B, and Clostridium septicum were analysed by PCR amplification and DNA sequencing. The five Clostridium species have at least two copies of the flagellin gene ( fliC ) arranged in tandem on the chromosome. The deduced N- and C-terminal aminoacid sequences of the flagellin proteins (FliCs) of these clostridia are well conserved but their central region aminoacid sequences are not. Phylogenic analysis based on the N-terminal aminoacid sequence of the FliC protein revealed that these clostridia, which belong to Clostridium 16S rDNA phylogenic cluster I ( Collins et al., 1994 ), are more closely related to Bacillus subtilis than to Clostridium difficile , which belongs to the cluster XI. Moreover, a multiplex polymerase reaction (PCR) system based on the fliC sequence was developed to rapidly identify C. chauvoei , C. haemolyticum , C. novyi types A and B, and C. septicum . PCR of each Clostridium amplified a species-specific band. The multiplex PCR system may be useful for rapid identification of pathogenic clostridia.
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Phylogenetic positions of Clostridium novyi and Clostridium haemolyticum based on 16S rDNA sequences.
International Journal of Systematic and Evolutionary Microbiology, 2001Co-Authors: Yoshimasa Sasaki, Noriyasu Takikawa, Akemi Kojima, Mari Norimatsu, Shoko Suzuki, Yutaka TamuraAbstract:The partial sequences (1465 bp) of the 16S rDNA of Clostridium novyi types A, B and C and Clostridium haemolyticum were determined. C. novyi types A, B and C and C. haemolyticum clustered with Clostridium botulinum types C and D. Moreover, the 16S rDNA sequences of C. novyi type B strains and C. haemolyticum strains were completely identical; they differed by 1 bp (level of similarity > 99.9%) from that of C. novyi type C, they were 98.7% homologous to that of C. novyi type A (relative positions 28-1520 of the Escherichia coli 16S rDNA sequence) and they exhibited a higher similarity to the 16S rDNA sequence of C. botulinum types D and C than to that of C. novyi type A. These results suggest that C. novyi types B and C and C. haemolyticum may be one independent species generated from the same phylogenetic origin.
Michael J. Mansfield - One of the best experts on this subject based on the ideXlab platform.
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Identification and characterization of the proteolytic flagellin from the common freshwater bacterium Hylemonella gracilis.
Scientific Reports, 2020Co-Authors: Ulrich Eckhard, Michael J. Mansfield, Constantin Blöchl, Benjamin Jenkins, Christian G. Huber, Andrew C. Doxey, Hans BrandstetterAbstract:Flagellins are the protein components of bacterial flagella and assemble in up to 20,000 copies to form extracellular flagellar filaments. An unusual family of flagellins was recently discovered that contains a unique metalloprotease domain within its surface-exposed hypervariable region. To date, these proteolytic flagellins (also termed flagellinolysins) have only been characterized in the Gram-positive organism Clostridium haemolyticum, where flagellinolysin was shown to be proteolytically active and capable of cleaving extracellular protein substrates. The biological function of flagellinolysin and its activity in other organisms, however, remain unclear. Here, using molecular biochemistry and proteomics, we have performed an initial characterization of a novel flagellinolysin identified from Hylemonella gracilis, a Gram-negative organism originally isolated from pond water. We demonstrate that H. gracilis flagellinolysin (HgrFlaMP) is an active calcium-dependent zinc metallopeptidase and characterize its cleavage specificity profile using both trypsin and GluC-derived peptide libraries and protein substrates. Based on high-throughput degradomic assays, HgrFlaMP cleaved 784 unique peptides and displayed a cleavage site specificity similar to flagellinolysin from C. haemolyticum. Additionally, by using a set of six protein substrates, we identified 206 protein-embedded cleavage sites, further refining the substrate preference of HgrFlaMP, which is dominated by large hydrophobic amino acids in P1', and small hydrophobic or medium-sized polar residues on the amino-terminal side of the scissile bond. Intriguingly, recombinant HgrFlaMP was also capable of cleaving full-length flagellins from another species, suggesting its potential involvement in interbacterial interactions. Our study reports the first experimentally characterized proteolytic flagellin in a Gram-negative organism, and provides new insights into flagellum-mediated enzymatic activity.
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Discovery of a proteolytic flagellin family in diverse bacterial phyla that assembles enzymatically active flagella
Nature Communications, 2017Co-Authors: Ulrich Eckhard, Hina Bandukwala, Michael J. Mansfield, Giada Marino, Jiujun Cheng, Iain Wallace, Todd Holyoak, Trevor C. Charles, John Austin, Christopher M. OverallAbstract:Bacterial flagella are cell locomotion and occasional adhesion organelles composed primarily of the polymeric protein flagellin, but to date have not been associated with any enzymatic function. Here, we report the bioinformatics-driven discovery of a class of enzymatic flagellins that assemble to form proteolytically active flagella. Originating by a metallopeptidase insertion into the central flagellin hypervariable region, this flagellin family has expanded to at least 74 bacterial species. In the pathogen, Clostridium haemolyticum , metallopeptidase-containing flagellin (which we termed flagellinolysin) is the second most abundant protein in the flagella and is localized to the extracellular flagellar surface. Purified flagellar filaments and recombinant flagellin exhibit proteolytic activity, cleaving nearly 1000 different peptides. With ~ 20,000 flagellin copies per ~ 10-μm flagella this assembles the largest proteolytic complex known. Flagellum-mediated extracellular proteolysis expands our understanding of the functional plasticity of bacterial flagella, revealing this family as enzymatic biopolymers that mediate interactions with diverse peptide substrates. So far no enzymatic activity has been attributed to flagellin, the major component of bacterial flagella. Here the authors use bioinformatic analysis and identify a metallopeptidase insertion in flagellins from 74 bacterial species and show that recombinant flagellin and flagellar filaments have proteolytic activity.
Trevor C. Charles - One of the best experts on this subject based on the ideXlab platform.
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Discovery of a proteolytic flagellin family in diverse bacterial phyla that assembles enzymatically active flagella
Nature Communications, 2017Co-Authors: Ulrich Eckhard, Hina Bandukwala, Michael J. Mansfield, Giada Marino, Jiujun Cheng, Iain Wallace, Todd Holyoak, Trevor C. Charles, John Austin, Christopher M. OverallAbstract:Bacterial flagella are cell locomotion and occasional adhesion organelles composed primarily of the polymeric protein flagellin, but to date have not been associated with any enzymatic function. Here, we report the bioinformatics-driven discovery of a class of enzymatic flagellins that assemble to form proteolytically active flagella. Originating by a metallopeptidase insertion into the central flagellin hypervariable region, this flagellin family has expanded to at least 74 bacterial species. In the pathogen, Clostridium haemolyticum , metallopeptidase-containing flagellin (which we termed flagellinolysin) is the second most abundant protein in the flagella and is localized to the extracellular flagellar surface. Purified flagellar filaments and recombinant flagellin exhibit proteolytic activity, cleaving nearly 1000 different peptides. With ~ 20,000 flagellin copies per ~ 10-μm flagella this assembles the largest proteolytic complex known. Flagellum-mediated extracellular proteolysis expands our understanding of the functional plasticity of bacterial flagella, revealing this family as enzymatic biopolymers that mediate interactions with diverse peptide substrates. So far no enzymatic activity has been attributed to flagellin, the major component of bacterial flagella. Here the authors use bioinformatic analysis and identify a metallopeptidase insertion in flagellins from 74 bacterial species and show that recombinant flagellin and flagellar filaments have proteolytic activity.