Genetic Analysis of Streptococcus mutans Virulence

Curtiss, rd R

doi:10.1007/978-3-642-70586-1_14

Cited by 61 publications

(70 citation statements)

References 117 publications

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“…It has been postulated, based on mutational analyses, that S. sobrinus carries a single dextranase gene (4). To see if the same held true for S. salivarius PC-1, chromosomal DNA was analyzed by Southern hybridization with both biotinlabeled pPD13 and the remaining 3.7-kb XhoI fragment (Fig.…”

Section: Methodsmentioning

confidence: 99%

Molecular cloning of the extracellular endodextranase of Streptococcus salivarius

Lawman

Bleiweis

1991

J Bacteriol

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We report the cloning in Escherichia coli of the gene encoding an extracellular endodextranase (a-1,6-glucanhydrolase, EC 3.2.1.11) from Streptococcus salivarius PC-1. Recombinants from a S. salivarius PC-l-Lambda ZAP II genomic library specifying dextranase activity were identified as plaques surrounded by zones of clearing on blue dextran agar. One such clone, PD1, had a 6.3-kb EcoRI fragment insert which encoded a 190-kDa protein with dextranase activity. The recombinant strain also produced two lowermolecular-mass polypeptides (90 and 70 kDa) that had dextranase activity. Native dextranase was recovered from concentrated culture fluids of S. salivarius as a single ll0-kDa polypeptide. PD1 phage lysate and PC-1 culture supernatant fluid extract were used to measure substrate specificity of the recombinant and native forms of dextranase, respectively. Analysis of these reaction products by thin-layer chromatography revealed the expected isomaltosaccharide products yielded by the recombinant-specified enzyme but was unable to resolve the larger polysaccharide products of the native enzyme. Furthermore. S. salivarius utilized neither the substrates nor the products of dextran hydrolysis for growth.Sucrose metabolism in the oral streptococci has been the subject of numerous studies, primarily because it is believed to be the major contributory factor in plaque formation and the subsequent development of dental caries (6,7,11). The metabolism of sucrose includes the synthesis of extracellular polysaccharides, both glucans and fructans, catalyzed by glucosyltransferase (GTF) and fructosyltransferase enzymes, respectively. These polymers are, in turn, degraded by dextranase (a-1,6-glucanhydrolase, EC 3.2.1.11), which has been shown to behave as an endoenzyme breaking glucans down to isomaltosaccharides (26), and fructanase (,-D-fructan fructohydrolase, EC 3.2.1.80), which hydrolyzes fructans to fructose (3).While sucrose metabolism has been the topic of much discussion as it relates to the potential virulence of oral streptococci, little attention has been focused on its potential to serve as a model for studying the complex regulatory network of enzymes and cofactors necessary for cellular response to irregular signals from the environment. Streptococcus salivarius is an adept colonizer of the human mouth and constitutes up to 60% of the streptococci in saliva and on the dorsal side of the tongue (11). It possesses the entire array of enzymes necessary for the production and degradation of the extracellular polysaccharides derived from sucrose and has been shown to regulate these enzymes in an extremely complex fashion (25). For these reasons, S. salivarius is an ideal organism on which to base a model for the control of extracellular enzymes in gram-positive organisms. Evidence suggesting that the availability of jextranase and certain glycosyltransferases is controlled to a great extent by posttranslational events, rather than genetic regulation, has been presented (25). To lay the foundation for future genetic a...

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Section: Methodsmentioning

confidence: 99%

Molecular cloning of the extracellular endodextranase of Streptococcus salivarius

Lawman

Bleiweis

1991

J Bacteriol

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“…GbpC shares homology with the AgI/II family of proteins that at one time were reported to possess glucan-binding properties. However, this claim could be traced only to a personal communication, and experimental verification could not be found (Curtiss, 1985).…”

Section: Mechanisms Of Glucan Bindingmentioning

confidence: 99%

Glucan-binding Proteins of the Oral Streptococci

Banas

Vickerman

2003

Critical Reviews in Oral Biology & Medicine

291

262

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ABSTRACT:The synthesis of extracellular glucan is an integral component of the sucrose-dependent colonization of tooth surfaces by species of the mutans streptococci. In investigators' attempts to understand the mechanisms of plaque biofilm development, several glucan-binding proteins (GBPs) have been discovered. Some of these, the glucosyltransferases, catalyze the synthesis of glucan, whereas others, designated only as glucan-binding proteins, have affinities for different forms of glucan and contribute to aspects of the biology of their host organisms. The functions of these latter glucan-binding proteins include dextran-dependent aggregation, dextranase inhibition, plaque cohesion, and perhaps cell wall synthesis. In some instances, their glucan-binding domains share common features, whereas in others the mechanism for glucan binding remains unknown. Recent studies indicate that at least some of the glucan-binding proteins modulate virulence and some can act as protective immunogens within animal models. Overall, the multiplicity of GBPs and their aforementioned properties are testimonies to their importance. Future studies will greatly advance the understanding of the distribution, function, and regulation of the GBPs and place into perspective the facets of their contributions to the biology of the oral streptococci.

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“…Whether lectin-like proteins are involved in intercellular adhesion of S. epidermidis, as is the case in the aggregation of Streptococcus mutans (17,59), is now of interest. This problem can now be attacked by affinity chromatography with the purified specific polysaccharide as a ligand and extracts of cell wall proteins of the isogenic biofilm-negative transposon mutant M10 or M11.…”

Section: Vol 178 1996 S Epidermidis Polysaccharide Intercellular Amentioning

confidence: 99%

The intercellular adhesin involved in biofilm accumulation of Staphylococcus epidermidis is a linear beta-1,6-linked glucosaminoglycan: purification and structural analysis

et al. 1996

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The primary attachment to polymer surfaces followed by accumulation in multilayered cell clusters leads to biofilm production of Staphylococcus epidermidis, which is thought to contribute to virulence in biomaterialrelated infections. We purified a specific polysaccharide antigen of biofilm-producing S. epidermidis 1457 and RP62A, which was recently shown to have a function in the accumulative phase of biofilm production by mediating intercellular adhesion (D. Mack, M. Nedelmann, A. Krokotsch, A. Schwarzkopf, J. Heesemann, and R. Laufs, Infect. Immun. 62:3244-3253, 1994). Following Sephadex G-200 gel filtration, this antigen was separated by Q-Sepharose chromatography into a major polysaccharide, polysaccharide I (>80%), which did not bind to Q-Sepharose, and a minor polysaccharide, polysaccharide II (<20%), which was moderately anionic. As shown by chemical analyses and nuclear magnetic resonance spectroscopy, polysaccharide I is a linear homoglycan of at least 130 ␤-1,6-linked 2-deoxy-2-amino-D-glucopyranosyl residues. On average, 80 to 85% of them are N acetylated; the rest are non-N-acetylating and positively charged. Chain cleavage by deamination with HNO 2 revealed a more or less random distribution of the non-N-acetylated glucosaminyl residues, with some prevalence of glucosaminyl-rich sequences. Cation-exchange chromatography separated molecular species whose content of non-N-acetylated glucosaminyl residues varied between 2 and 26%. Polysaccharide II is structurally related to polysaccharide I but has a lower content of non-N-acetylated Dglucosaminyl residues and contains phosphate and ester-linked succinate, rendering it anionic. Enzyme-linked immunosorbent assay inhibition with various monosaccharides revealed the ␤-anomeric form and the acetylated amino group of the D-glucosaminyl residues as important for reactivity with the specific antiserum. The unbranched polysaccharide structure favors long-range contacts and interactions between polysaccharide strands and the cell wall and/or lectin-like proteins, leading to intercellular adhesion and biofilm accumulation. The structure of the polysaccharide is, so far, considered to be unique and, according to its function, is referred to as S. epidermidis polysaccharide intercellular adhesin (PIA).At present, coagulase-negative staphylococci, mostly Staphylococcus epidermidis, represent the most frequent cause by far of nosocomial sepsis and are the most prominent organisms responsible for infections associated with implanted biomaterials like intravascular catheters, peritoneal dialysis catheters, cerebrospinal fluid shunts, prosthetic heart valves, and prosthetic joints, resulting in substantial morbidity and mortality (25,37,54,56).By scanning electron microscopy, coagulase-negative staphylococci were shown to colonize intravascular catheters in large adherent biofilms composed of multilayered cell clusters embedded in an amorphous extracellular material, which is composed of exopolysaccharides referred to as slime or glycocalyx (13,16,28,45,51). In vitro...

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Genetic Analysis of Streptococcus mutans Virulence

Cited by 61 publications

References 117 publications

Molecular cloning of the extracellular endodextranase of Streptococcus salivarius

Molecular cloning of the extracellular endodextranase of Streptococcus salivarius

Glucan-binding Proteins of the Oral Streptococci

The intercellular adhesin involved in biofilm accumulation of Staphylococcus epidermidis is a linear beta-1,6-linked glucosaminoglycan: purification and structural analysis

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