Biomedical subjects

S K Harlander

Publications and source records attributed to S K Harlander.

22 records · Page 2Linked to original sources

PubMed1977

Streptococcus mutans dextransucrase: functioning of primer dextran and endogenous dextranase in water-soluble and water-insoluble glucan synthesis.

The extracellular enzyme activities of Streptococcus mutans 6715 that synthesize glucans from sucrose were concentrated and partially purified by ammonium sulfate precipitation and gel permeation column chromatography. Polyacrylamide gel analysis demonstrated that all of the major proteins precipitated by ammonium sulfate were quantitatively recovered in the high-molecular-weight, enzyme-containing aggregates found in the void volume of the gel column. Anion-exchange column chromatography was used to fractionate the aggregates into preparations, alpha and beta, which produced water-insoluble and water-soluble glucans, respectively. Polyacrylamide gel analysis showed that alpha and beta contained unique proteins and dextransucrase (EC 2.4.1.5) activities. Studies on the time course of glucan synthesis by alpha demonstrated that this enzyme preparation contained dextranase activity, which partially degraded nascent alcohol-insoluble glucan into alcohol-soluble products that were subsequently reincorporated into insoluble product. The beta enzyme preparation contained no detectable dextranase activity. Mixing experiments in the absence of primer dextran demonstrated that the dextranase activity present in alpha could modify glucan production by beta. CsCl density gradient analysis of product glucans demonstrated that exogenous primer dextrans were used as acceptor molecules by both the alpha and beta enzyme preparations, and that water-soluble glucans synthesized by beta could be converted into water-insoluble glucans by alpha. It is proposed that the structural heterogeneity of the native glucans produced from sucrose by S. mutans is a result of the concerted action of glucan-forming dextransucrases and endohydrolytic dextranase activity.

Chromatography, Gel↗

PubMed1976

Streptococcus mutans dextransucrase: availability of disaggregated enzyme after growth in a chemically defined medium.

The soluble dextransucrase (EC 2.4.1.5) activity produced by Streptococcus mutans strain 6715 during growth on a chemically defined synthetic medium (FMS) was compared to enzyme from glucose broth cultures (TSB). Growth on the two media was similar. The specific activity of ammonium sulfate-precipitated FMC enzyme was 17 times greater than similar TSB enzyme preparations. The FMC enzyme was stimulated 11-fold, whereas the TSB enzyme was stimulated 1.2-fold by the addition of exogenous primer dextran. In contrast to the TSB enzyme, the FMC activity could be disaggregated to a low-molecular-weight form by 1 M salt. Thus, low-molecular-weight S. mutans dextransucrase activity free of contaminating primer glucan may be readily obtained after growth of the bacterium in a chemically defined sucrose-free medium.

Culture Media↗

PubMed1975

Dextranases from oral bacteria: inhibition of water-insoluble glucan production and adherence to smooth surfaces by Streptococcus mutans.

The effect of dextranases (EC 3.2.1.11) from the oral isolates Actinomyces israelii and Bacteroides ochraceus on water-insoluble glucan production by the Streptococcus mutans dextransucrase (EC 2.4.1.5) and sucrose-dependent adherence to smooth glass surfaces by S. mutans was studied. Collection on membrane filters of water-insoluble polysaccharides synthesized from radioactive sucrose was used to demonstrate the marked sensitivity of insoluble glucan formation to the presence of dextranase. Concentrations of A. israelii dextranase as low as 0.002 U/ml inhibited insoluble glucan formation by 60%. Similar results were obtained the the B. ochraceus enzyme. An assay for sucrose-stimulated adherence of S. mutans to smooth surfaces involved attachment of radioactively labeled nongrowing cells to the bottom of glass scintillation vials. This facile and sensitive assay was utilized to demonstrate that sucrose-dependent adherence was affected by low levels of dextranase from either A. israelii or B. ochraceus. Enzyme at 0.005 U/ml reduced adherence of S. mutans by 80%. Treatment of S. mutans cells previously attached to glass with low concentrations of the dextranases resulted in removal of 50% to 60% of the bacteria. The results indicate that dextranase-producing oral bacteria may affect sucrose-dependent colonization of S. mutans on the tooth surface and offer a possible explanation for both the difficulties involved in implanting this bacterium into the human mouth and the limited intraoral transmission of S. mutans from one tooth surface to another.

Actinomyces↗

PubMed1975

Production of elevated levels of dextransucrase by a mutant of Streptococcus mutans.

A mutant (S19) of Sreptococcus mutans strain 6715 which produces elevated levels of dextransucrase (EC 2.4.1.5) was isolated. Soluble enzyme in culture supernatant solutions from S19 polymerized the glucosyl moiety of sucrose into alcohol-insoluble and water-insoluble glucans at a rate three to six times greater than that of the parent strain. Washed-cell suspensions of S19 also contained increased amounts of cell-associated enzyme. Adherence of S19 to glass in the presence of sucrose occurred at twice the rate of strain 6715. The Km values for sucrose and primer dextran were similar for the mutant and parent enzymes. Mutant S19 should facilitate studies on the mechanism of adherence of S. Mutans and the control of dextransucrase production by this bacterium.

Glucosyltransferases↗