Isolation and characterisation of sialidase from a strain of Streptococcus oralis

Byers, Helen L.; Tarelli, Edward; Homer, Karen A.; Beighton, David

doi:10.1099/0022-1317-49-3-235

Cited by 42 publications

(28 citation statements)

References 44 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, sialidase genes have previously been found in S. oralis, a close relation of the pneumococcus (7,25). The sequences of the protein alleles reported in our work were very similar to that of pneumococcal NanA, and this may indicate that tight control of the NanA structure is more important for biological activity than that of Ply and the Mly homologue.…”

Section: Figsupporting

confidence: 54%

Detection of Large Numbers of Pneumococcal Virulence Genes in Streptococci of the Mitis Group

Johnston

Hinds

Smith³

et al. 2010

J Clin Microbiol

View full text Add to dashboard Cite

Seven streptococcal isolates from the mitis group were analyzed for the presence of pneumococcal gene homologues by comparative genomic hybridization studies with microarrays based on open reading frames from the genomes of Streptococcus pneumoniae TIGR4 and R6. The diversity of pneumolysin (ply) and neuraminidase A (nanA) gene sequences was explored in more detail in a collection of 14 S. pseudopneumoniae and 29 mitis group isolates, respectively. The mitis group isolates used in the microarray experiments included a type strain (NCTC 12261), two S. mitis isolates from the nasopharynxes of children, one S. mitis isolate from a case of infective endocarditis, one S. mitis isolate from a dental abscess, and one S. oralis isolate and one S. pseudopneumoniae isolate from the nasopharynxes of children. The results of the microarray study showed that the 5 S. mitis isolates had homologues to between 67 and 82% of pneumococcal virulence genes, S. oralis hybridized to 83% of pneumococcal virulence genes, and S. pseudopneumoniae hybridized to 92% of identified pneumococcal virulence genes. Comparison of the pneumolysin, mitilysin (mly), and newly identified pseudopneumolysin (pply) gene sequences revealed that mly and pply genes are more closely related to each other than either is to ply. In contrast, the nanA gene sequences in the pneumococcus and streptococci from the mitis group are closely clustered together, sharing 99.4 to 99.7% sequence identity with pneumococcal nanA alleles.Streptococcus pneumoniae, also known as the pneumococcus, is an important human pathogen causing high levels of mortality worldwide. Diseases caused include meningitis, pneumonia, and otitis media. S. pneumoniae is a member of the viridans group streptococci (VGS) and is closely related to Streptococcus mitis and Streptococcus oralis, sharing Ͼ99% homology by 16S rRNA gene analysis (13,23). Recently, a subset of "atypical" pneumococci were identified that were distinct from both S. mitis and S. pneumoniae by housekeeping gene analysis (48). It is likely that these isolates belonged to the species Streptococcus pseudopneumoniae, which has since been identified, with isolates appearing phenotypically similar to S. mitis but undistinguishable from S. pneumoniae in genotypic tests (2). As a result of the close relationships between these species, clear identification can be problematic. A recent study showed that definitive species identification can be achieved by phylogenetic analysis of 4 housekeeping genes. Furthermore, although it has been reported that comparison of superoxide dismutase genes (sodA) was sufficient to identify most species, this did not distinguish between S. pneumoniae and S. pseudopneumoniae isolates (19).Despite the close genetic relationships between these species, while S. pneumoniae can be a severe pathogen, S. oralis and S. mitis are usually commensal organisms of mucous membranes. However, on rare occasions, they are able to cause opportunistic disease in immunocompromised patients, particularly those with replacem...

show abstract

Section: Figsupporting

confidence: 54%

Detection of Large Numbers of Pneumococcal Virulence Genes in Streptococci of the Mitis Group

Johnston

Hinds

Smith³

et al. 2010

J Clin Microbiol

View full text Add to dashboard Cite

show abstract

“…Quantitative sialidase assays involved measuring 4-methylumbelliferone (4-MU) fluorescence, released from 4-MU-NeuNAc by sialidase activity, in a Fluoroskan Ascent FL fluorimeter (Labsystems Thermo, United Kingdom) with an excitation wavelength of 380 nm and an emission wavelength of 460 nm (6,7). To determine the optimal pH for sialidase activity, assays were carried out with 50 M 4-MU-NeuNAc in 70 mM sodium citrate buffer (pH 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, or 6.0), 70 mM sodium phosphate buffer (pH 6.0, 6.5, 7.0, 7.5, or 8.0), 70 mM potassium phosphate buffer (pH 6.5, 7.0, or 7.5), or 70 mM Tris-HCl (pH 7.5, 8.0, 8.5, or 8.9) and appropriately diluted cell lysates, such that the rate of release of 4-MU from 4-MU-NeuNAc was linear with respect to time.…”

Section: Methodsmentioning

confidence: 99%

An Orthologue of Bacteroides fragilis NanH Is the Principal Sialidase in Tannerella forsythia

Thompson¹,

Homer²,

Rao³

et al. 2009

J Bacteriol

View full text Add to dashboard Cite

Sialidase activity is a putative virulence factor of the anaerobic periodontal pathogen Tannerella forsythia, but it is uncertain which genes encode this activity. Characterization of a putative sialidase, SiaHI, by others, indicated that this protein alone may not be responsible for all of the sialidase activity. We describe a second sialidase in T. forsythia (TF0035), an orthologue of Bacteroides fragilis NanH, and its expression in Escherichia coli. Sialidase activity of the expressed NanH was confirmed by using 2-(4-methylumbelliferyl)-␣-D-N-acetylneuraminic acid as a substrate. Biochemical characterization of the recombinant T. forsythia NanH indicated that it was active over a broad pH range, with optimum activity at pH 5.5. This enzyme has high affinity for 2-(4-methylumbelliferyl)-␣-D-N-acetylneuraminic acid (K m of 32.9 ؎ 10.3 M) and rapidly releases 4-methylumbelliferone (V max of 170.8 ؎ 11.8 nmol of 4-methylumbelliferone min ؊1 mg of protein ؊1 ). E. coli lysates containing recombinant T. forsythia NanH cleave sialic acid from a range of substrates, with a preference for ␣2-3 glycosidic linkages. The genes adjacent to nanH encode proteins apparently involved in the metabolism of sialic acid, indicating that the NanH sialidase is likely to be involved in nutrient acquisition.

show abstract

“…Oral bacteria have been shown to express a wide variety of glycosidases that permits the degradation of complex substrates, such as salivary glycoproteins (e.g. Beighton & Whiley, 1990;Byers et al, 2000). This metabolic activity was clearly shown by Bradshaw et al (1994) in a study employing hog gastric mucin as the major carbon and energy source in a chemostat inoculated with a mixture of 10 organisms selected for their hydrolytic enzyme activity.…”

Section: Introductionmentioning

confidence: 92%

Differential metabolic activity by dental plaque bacteria in association with two preparations of MUC5B mucins in solution and in biofilms

2009

View full text Add to dashboard Cite

Salivary mucin, MUC5B, is an oligomeric glycoprotein, heterogeneous in size and with a diverse repertoire of oligosaccharides, which differ in composition and charge. Since complex salivary glycoproteins are considered to be the major source of nutrients for the oral supragingival microbiota, the major aim of the current study was to determine whether different preparations of non-denatured MUC5B could be isolated exhibiting different biological properties in relation to the microflora associated with the surfaces of the oral cavity. Two preparations, solMUC5B and gelMUC5B, were isolated by density-gradient centrifugation and were shown to have different buoyant densities, carbohydrate content and surface-adsorbing characteristics. To ascertain differences in biological activity, the two mucin preparations, both in solution and adsorbed to a model surface, were incubated with freshly isolated dental plaque and assayed for metabolic (dehydrogenase) activity with the fluoresecent substrate CTC (5-cyano-2,3-ditolyl tetrazolium chloride). The plaque bacteria exhibited higher metabolism with the solMUC5B preparation in solution, with 79.4 % active plaque cells compared to the controls without mucin (9.6 %), while gelMUC5B showed 48.2 % active cells with the same plaque population. In contrast, the same mucins adhered to a surface elicited a significantly lower metabolic response, with surfaceassociated plaque cells showing only 12.1 % active cells with solMUC5B and 29.2 % with gelMUC5B. These results suggested that the metabolism by the plaque cells adsorbed to surface-associated mucins was downregulated compared to the same cells suspended in mucin solution. This was confirmed in an experiment where active dispersed plaque/solMUC5B suspensions were shown to lose significant metabolic activity (e.g. 74.9 to 19.3 %) when allowed to interact with gelMUC5B adsorbed to a surface. Clearly, the solMUC5B and gelMUC5B preparations exhibited different biological activity when assayed with freshly plaque bacteria in suspension and in a biofilm.

show abstract

Isolation and characterisation of sialidase from a strain of Streptococcus oralis

Cited by 42 publications

References 44 publications

Detection of Large Numbers of Pneumococcal Virulence Genes in Streptococci of the Mitis Group

Detection of Large Numbers of Pneumococcal Virulence Genes in Streptococci of the Mitis Group

An Orthologue of Bacteroides fragilis NanH Is the Principal Sialidase in Tannerella forsythia

Differential metabolic activity by dental plaque bacteria in association with two preparations of MUC5B mucins in solution and in biofilms

Contact Info

Product

Resources

About