D Wunder

University of Rochester, Rochester, NY, United States

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Publications (7)13.98 Total impact

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    ABSTRACT: This study examines the possible effect of the antimicrobial peroxidase system on the activity of streptococcal glucosyltransferases B, C and D (GtfB, GtfC and GtfD), either in solution (GtfB and GtfC) or when adsorbed to hydroxyapatite (GtfC and GtfD) at pH 6.5. The lactoperoxidase (LP) system (LP, H(2)O(2), SCN(-)) had no effect on the activity of dissolved GtfC, but the activity of dissolved GtfB was enhanced. The LP system, however, strongly inhibited the activities of both GtfC and GtfD in their adsorbed form. LP enzyme, without its substrates, inhibited all three Gtf enzymes: GtfB and GtfC in concentrations between 10 and 100 microg/ml in liquid phase and adsorbed GtfC and GtfD in concentrations between 25 and 50 microg/ml. This inhibition was in part abolished in liquid phase, but not in solid phase, if the substrates of LP were added. This study shows that the lactoperoxidase system can exert inhibitory activity against streptococcal Gtfs without generating oxidizing agents.
    Caries Research 01/2002; 36(2):116-21. · 2.51 Impact Factor
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    ABSTRACT: Glucosyltransferase (GTF) plays an essential role in the formation of the biofilm known as dental plaque and in the pathogenesis of dental caries. Mutans streptococci produce at least three distinct GTFs (GtfB, C and D), each of which forms a glucan polymer from sucrose. Glucan is a major constituent of plaque biofilm. GTF adsorbed to a surface forms glucans that differ in structure from those formed by the same enzyme in solution. In the present study, activities of GtfB and GtfC in solution or adsorbed on a surface were inhibited in the presence of a polyclonal antiserum (DS-1) to a mixture of GTFs and by immunoglobulin G (IgG) prepared from DS-1; in contrast, enzyme activity was enhanced by normal rabbit serum (NRS) and IgG from NRS. GtfD activity on a surface was enhanced by both antiserum DS-1 and NRS, and IgG prepared from either serum; GtfD activity in solution was slightly inhibited by each of the sera. The structure of GtfB and GtfC glucans formed in the presence of antiserum differed from that of controls based on linkage analyses, and on their susceptibilities to the glucanohydrolases mutanase (alpha-1,3 hydrolase) and dextranase (alpha-1,6 hydrolase); soluble products from the enzymatic digestion also differed. The results show that the effects of antibody on enzyme activity are more complex than simple inhibition or enhancement and that the presence of antibody may influence glucan structure, which clearly could impact plaque formation. The results have implications for the formation and properties of biofilms formed in other environments.
    Caries Research 01/2002; 36(2):108-15. · 2.51 Impact Factor
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    ABSTRACT: The aim of our study was to determine whether the structure of glucans formed by glucosyltransferase from Streptococcus sanguinis (GtfSs) on a surface differ from those formed in solution and to explore the effects of antiserum to Gtfs, control normal rabbit serum, starch hydrolysates (STH) and dextran on S. sanguinis (GtfSs) glucan. Linkage analyses showed that solution-formed glucans are predominantly alpha-1,6-linked and have a small amount of alpha-1,3-linked glucose. Surface-formed glucans have enhanced susceptibility to mutanase. Solution- and surface-formed glucans made in the presence or absence of sera, STH, and dextran contain linkages which differ in both amount and type from control glucans. The GtfSs enzyme in solution exposed to antiserum behaves as if it is adsorbed to a surface. Binding of Streptococcus mutans GS-5 and Actinomyces viscosus OMZ105E (Ny1) to S. sanguinis glucan differs if the glucan is formed in the presence of antiserum. The information could help to define the role of glucans in the formation of pellicle, colonization of tooth surfaces and the accumulation of dental plaque.
    Caries Research 01/2001; 35(1):67-74. · 2.51 Impact Factor
  • D Wunder, W H Bowen
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    ABSTRACT: Previous studies have shown that glucosyltransferase enzymes (Gtfs) of Streptococcus mutans adsorbed to saliva coated hydroxyapatite (sHA) have distinct properties from the same enzymes in solution. The purpose of the present study was to determine the effects on enzyme activity of polyclonal antibodies raised to Gtfs in a soluble form and bound to sHA. Antiserum was raised in six New Zealand White rabbits using the purified glucosyltransferase enzymes (Gtfs) of S. mutans, GtfB, GtfC, or GtfD as soluble antigens or adsorbed to hydroxyapatite (HA, insolubilized). The antisera were examined for their ability to react to these Gtfs and Gtf from Streptococcus sanguis (GtfSs) in Western blot formats as well as inhibit enzyme activity in solution and insolubilized. Antibodies raised against GtfB or GtfC detected all Gtf enzymes examined in Western blots; antibodies raised to GtfD reacted strongly to GtfD and GtfSs, poorly to GtfC, and was non-reactive with GtfB. Antibodies to GtfB or GtfC inhibited activity of GtfB and GtfC in solution by 90% or more. Enzyme activity adsorbed to sHA was inhibited from 70% to 80% by the same antisera. These same antibodies possessed no specific effect on the activities of either GtfD or GtfSs. Antibodies raised to the GtfD enzyme inhibited activity of GtfD (80% to 90% inhibition) and GtfSs activity (50% to 80%) in solution. In contrast the GtfD antibodies had no effect on the activity of either GtfB or GtfC enzymes in solution. Modest inhibitory effects were noted on GtfC and GtfSs enzymes bound to sHA, but no inhibition was observed for sHA-bound GtfB or GtfD. These data show that some antibodies effective against enzymes in solution may have significantly lesser inhibitory effects against the same enzymes insolubilized. Further, presentation of Gtf antigen immobilized to HA has only a minor influence on the production of antibodies inhibitory to Gtf activity.
    Oral Diseases 10/2000; 6(5):289-96. · 2.38 Impact Factor
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    ABSTRACT: To investigate the effects of the peroxidase system (LP, H2O2, SCN) and its components on the activity of glucosyltransferase D (GtfD) originating from Streptococcus mutans (S. mutans). GtfD was incubated in buffered-KCl assay mixture that contained 20 microM dextran and 100 mM sucrose including 14C-glucose. The activity of GtfD was measured with a scintillation counter. The effects of lactoperoxidase system and its components on GtfD activity were examined by incubating different components separately and together with GtfD. The lactoperoxidase system-generated hypothiocyanite (OSCN) had no effect on the activity of streptococcal GtfD, while the lactoperoxidase enzyme inhibited GtfD. The ratio between the 2 enzymes, LP and GtfD, was important for the inhibition. However, if LP was combined with its substrates, SCN and/or a high concentration of H2O2, it enhanced the activity of GtfD. Peroxidase-generated antimicrobial agent HOSCN/OSCN is not inhibitory against GtfD, whereas low concentrations of the LP-enzyme inhibit GtfD.
    The Chinese journal of dental research: the official journal of the Scientific Section of the Chinese Stomatological Association (CSA) 09/2000; 3(2):61-4.
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    ABSTRACT: We have shown in previous studies that the glucosyltransferase (Gtf) enzymes of Streptococcus mutans have distinct properties when adsorbed to a surface. In the present study, we compared the activity of Gtf from Streptococcus sanguis, designated GtfSs, in solution and on the surface of saliva-coated hydroxyapatite (sHA) beads, and determined the ability of its product glucan to support the adherence of oral microorganisms. Gtf from S. sanguis 804 NCTC 10904 was purified from culture supernatant fluids by means of hydroxyapatite chromatography. Enzyme and the substrate were prepared in buffers at pH values from 3.5 to 7.5. Maximum activity of GtfSs occurred between pH 5.5 and pH 6.5, whether in solution or adsorbed onto a surface. The solubilized and insolubilized enzymes showed highest activity at 40 degrees C; activity was reduced by 50(+/-2)% at 20 and 30 degrees C. The enzyme did not form glucans in either phase at 10 or 60 degrees C. The K(m), determined from Lineweaver-Burk plots, for the enzyme in solution was 4.3(+/-0.4) mmol/l sucrose, and the K(m) for the enzyme on sHA beads was 5.0(+/-1.0) mmol/l sucrose. The ability of the GtfSs glucan synthesized on the surface of sHA beads to support the adherence of oral bacteria was investigated. (3)H-thymidine-labeled bacteria (S. mutans GS-5, S. sobrinus 6715, S. sobrinus 6716, S. sanguis 10904, Actinomyces viscosus OMZ105E, A. viscosus 2085, and A. viscosus 2086) were incubated with sHA beads coated with GtfSs glucan. S. mutans GS-5 displayed the highest level of binding numerically. These results show that the GtfSs of S. sanguis is active on sHA beads, that the pH optimum for activity on a surface differs slightly from that in solution, and that its product glucan can support the adherence of oral microorganisms.
    Caries Research 01/2000; 34(4):295-302. · 2.51 Impact Factor
  • D Wunder, W H Bowen
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    ABSTRACT: Glucosyltransferase (Gtf) activity mediates sucrose-dependent adherence of mutans streptococci to the tooth surface, is essential for the cariogenicity of these micro-organisms, and contributes significantly to the exopolysaccharide component of the dental-plaque matrix. Clearly, agents that inhibit Gtfs could have therapeutic benefit. Here the effects of agents that inhibit Gtfs in solution and adsorbed to a surface were explored. Various classes of chemical reagents were tested for their ability to inhibit the enzymes responsible for insoluble-glucan synthesis (GtfB), insoluble/soluble glucan synthesis (GtfC), and soluble-glucan (GtfD) from Streptococcus mutans. Standard inhibition assays were done with Gtf enzyme in solution or with Gtf adsorbed to parotid saliva-coated hydroxylapatite (surface phase). Reagents tested included the metallic cations Li+, Zn2+, Cu2+, Fe2+ and Fe3+; the oxidizing compounds hypochlorite, Rose Bengal, perborate, and sodium-meta-periodate; and a panel of sugars and sugar analogues including sorbitol, xylitol, 1',4',6' trideoxy-trichloro-galactosucrose (TGS), and 1-deoxynojirimycin (dNJ). In solution, Gtf activity was inhibited significantly, at the highest concentrations tested: by the metal ions Zn2+, Cu2+, Fe2+ and Fe3+ (approx. 40-80% inhibition); by Rose Bengal and hypochlorite (approx. 80-90% inhibition); and by TGS and dNJ (approx. 50-80%). However, surface-adsorbed Gtfs displayed increased resistance to inhibition by the same metal cations and oxidizing compounds that inhibited them in solution. In contrast, both TGS and dNJ possessed similar inhibition profiles for both surface-bound Gtf and enzyme in solution. These data indicate that the nature of the inhibitor is important, and also whether the Gtf enzyme is in solution or adsorbed to saliva-coated hydroxylapatite.
    Archives of Oral Biology 04/1999; 44(3):203-14. · 1.55 Impact Factor