Cross-Linked Peptidoglycan Mediates Lysostaphin Binding to the Cell Wall Envelope of<i>Staphylococcus aureus</i>

Gründling, Angelika; Schneewind, Olaf

doi:10.1128/jb.188.7.2463-2472.2006

Cited by 147 publications

(146 citation statements)

References 72 publications

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“…3). These findings are similar to the results obtained with lysostaphin and ALE-1, a glycyl-glycine endopeptidase homologous to lysostaphin, which showed a (1,22), which bind to cross-linked peptidoglycan and recognize the Gly 5 interpeptide cross bridge (12,22). Our results suggest that a similar targeting mechanism exists for the 11 endolysin and that, in this respect, the phage enzyme closely resembles its staphylococcal counterparts.…”

supporting

confidence: 79%

Lytic Activity of Recombinant Bacteriophage φ11 and φ12 Endolysins on Whole Cells and Biofilms of Staphylococcus aureus

Saß

Bierbaum

2007

Appl Environ Microbiol

205

156

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The recombinant 11 endolysin hydrolyzed heat-killed staphylococci as well as staphylococcal biofilms. Cell wall targeting appeared to be a prerequisite for lysis of whole cells, and the combined action of the endopeptidase and amidase domains was necessary for maximum activity. In contrast, the 12 endolysin was inactive and caused aggregation of the cells.Infections caused by Staphylococcus aureus and Staphylococcus epidermidis still play a major role in human and animal disease. In particular staphylococcal biofilms on indwelling devices are difficult to treat due to their inherent antibiotic resistance (5,7,26). In this context, it is important to develop alternative treatment strategies to combat staphylococcal infections, particularly in view of the ability of staphylococci to acquire resistance to commonly used antibiotics (10,23,29). Phage lysins, or endolysins, have received considerable attention as possible antimicrobial agents against gram-positive bacteria and have been applied to a variety of pathogens, such as Bacillus anthracis (21), Streptococcus pneumoniae (9), and S. aureus (25). Endolysins are active against dead cells and even living, planktonic cells (9, 25), but their ability to lyse the complex structure of staphylococcal biofilms has not yet been investigated. In this approach, we cloned and heterologously overexpressed the lysis genes of the bacteriophages 11 and 12 of S. aureus NCTC8325 in Escherichia coli for subsequent analysis of the lytic activity of the enzymes and their single subdomains on cell walls, whole cells, and biofilms. Knowledge of the lytic activity of both endolysins is limited. Their nucleotide sequences have been published (16), and the 11 endolysin has been shown to possess a D-alanyl-glycyl endopeptidase and an N-acetylmuramyl-L-alanine amidase activity on crude cell walls of S. aureus OS2 (24).Sequence comparison, cloning, and overexpression of 11 and 12 endolysins. The 11 and 12 endolysin sequences were BLAST searched against the NCBI protein database. Both endolysins are modular enzymes which consist of three distinct domains coding for an N-terminal CHAP (cysteine, histidine-dependent amidohydrolases/peptidases) domain with hydrolytic function, a central amidase domain (N-acetylmuramyl-L-alanine amidase), and a C-terminal SH3b domain, which is involved in cell wall recognition (1). In spite of their similar domain architecture, these endolysins show low sequence identity (26.9%).The endolysin genes of the bacteriophages 11 (open reading frame [ORF] 53; 1,473 bp; accession number NC_004615) and 12 (ORF 49; 1,455 bp; accession number NC_004616) were amplified by PCR from genomic DNA of S. aureus NCTC8325, using the primers listed in Table 1. In order to test the activity of the 11 endolysin subunits, the endopeptidase unit (11endo, amino acids [aa] 1 to 180) and the amidase unit (11ami, aa 180 to 371) as well as each unit plus the cell wall binding domain (11endo/CBD, aa 1 to 180/371 to 490, and 11ami/CBD, aa 180 to 490) were constructed separately (Fig. 1). In ad...

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supporting

confidence: 79%

Lytic Activity of Recombinant Bacteriophage φ11 and φ12 Endolysins on Whole Cells and Biofilms of Staphylococcus aureus

Saß

Bierbaum

2007

Appl Environ Microbiol

205

156

View full text Add to dashboard Cite

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“…This CBD, shared by lysostaphin (a bacteriocin from Staphylococcus simulans bv. staphylolyticus) and its homologue, hydrolase ALE-1, has been uncovered to recognize pentaglycine cross bridges in PG (68,69). Therefore, the fact that most of the SH3 domains are found in Staphylococcus-like endolysins is not surprising.…”

Section: Resultsmentioning

confidence: 99%

Molecular Aspects and Comparative Genomics of Bacteriophage Endolysins

Oliveira

Melo

Santos

et al. 2013

J Virol

251

247

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Phages are recognized as the most abundant and diverse entities on the planet. Their diversity is determined predominantly by their dynamic adaptation capacities when confronted with different selective pressures in an endless cycle of coevolution with a widespread group of bacterial hosts. At the end of the infection cycle, progeny virions are confronted with a rigid cell wall that hinders their release into the environment and the opportunity to start a new infection cycle. Consequently, phages encode hydrolytic enzymes, called endolysins, to digest the peptidoglycan. In this work, we bring to light all phage endolysins found in completely sequenced double-stranded nucleic acid phage genomes and uncover clues that explain the phage-endolysin-host ecology that led phages to recruit unique and specialized endolysins.

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“…Since the killing required contact of the cells, it seemed unlikely that soluble substances, such as bacteriocin (21,60) or killer toxin (53), were involved. We also showed that soluble capsular polysaccharides from C. neoformans inhibited killing.…”

mentioning

confidence: 99%

Contribution of the Mannan Backbone of Cryptococcal Glucuronoxylomannan and a Glycolytic Enzyme ofStaphylococcus aureusto Contact-Mediated Killing ofCryptococcus neoformans

et al. 2007

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The fungal pathogen Cryptococcus neoformans is killed by the bacterium Staphylococcus aureus, and the killing is inhibited by soluble capsular polysaccharides. To investigate the mechanism of killing, cells in coculture were examined by scanning and transmission electron microscopy. S. aureus attached to the capsule of C. neoformans, and the ultrastructure of the attached C. neoformans cells was characteristic of dead cells. To identify the molecules that contributed to the fungal-bacterial interaction, we treated each with NaIO 4 or protease. Treatment of C. neoformans with NaIO 4 promoted adherence. It was inferred that cleavage of xylose and glucuronic acid side chains of glucuronoxylomannan (GXM) allowed S. aureus to recognize mannose residues in the backbone, which resisted periodate oxidation. On the other hand, treatment of S. aureus with protease decreased adherence, suggesting that protein contributed to attachment in S. aureus. In confirmation, side chain-cleaved polysaccharide or defined ␣-(133)-mannan inhibited the killing at lower concentrations than native GXM did. Also, these polysaccharides reduced the adherence of the two species and induced clumping of pure S. aureus cells. ␣-(133)-Mannooligosaccharides with a degree of polymerization (DP) of >3 induced cluster formation of S. aureus in a dose-dependent manner. Surface plasmon resonance analyses showed interaction of GXM and surface protein from S. aureus; the interaction was inhibited by oligosaccharides with a DP of >3. Conformations of ␣-(133) oligosaccharides were predicted. The three-dimensional structures of mannooligosaccharides larger than triose appeared curved and could be imagined to be recognized by a hypothetical staphylococcal lectin. Native polyacrylamide gel electrophoresis of staphylococcal protein followed by electroblotting, enzyme-linked immunolectin assay, protein staining, and N-terminal amino acid sequencing suggested that the candidate protein was triosephosphate isomerase (TPI). The enzymatic activities were confirmed by using whole cells of S. aureus. TPI point mutants of S. aureus decreased the ability to interact with C. neoformans. Thus, TPI on S. aureus adheres to the capsule of C. neoformans by recognizing the structure of mannotriose units in the backbone of GXM; we suggest that this contact is required for killing of C. neoformans.

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Cross-Linked Peptidoglycan Mediates Lysostaphin Binding to the Cell Wall Envelope ofStaphylococcus aureus

Cited by 147 publications

References 72 publications

Lytic Activity of Recombinant Bacteriophage φ11 and φ12 Endolysins on Whole Cells and Biofilms of Staphylococcus aureus

Lytic Activity of Recombinant Bacteriophage φ11 and φ12 Endolysins on Whole Cells and Biofilms of Staphylococcus aureus

Molecular Aspects and Comparative Genomics of Bacteriophage Endolysins

Contribution of the Mannan Backbone of Cryptococcal Glucuronoxylomannan and a Glycolytic Enzyme ofStaphylococcus aureusto Contact-Mediated Killing ofCryptococcus neoformans

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