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H Mukasa

Publications and source records attributed to H Mukasa.

At least 55 records · Page 3Linked to original sources
PubMed1982

Purification and properties of Streptococcus mutans extracellular glucosyltransferase.

Extracellular glucosyltransferase (sucrose:1,6-alpha-D-glucan 3-alpha- and 6-alpha-glucosyltransferase) was purified about 10 000-fold from the culture supernatant of Streptococcus mutans 6715. The enzyme preparation was homogeneous on polyacrylamide gel electrophoresis, isoelectric focusing and ultracentrifugation analyses. The specific activity of the enzyme was 34.9 I.U. per mg of protein and the carbohydrate content was less than 1% (w/w). The molecular weight was determined to be 149 000 +/- 5000 by sedimentation equilibrium experiment. The acidic and basic amino acids of the enzyme comprised 29 and 8.4% of total amino acid, respectively, and the isoelectric point was pH 4.1. The enzyme had the optimum pH of 5.5 and the Km value of 2.4 mM for sucrose. The water-soluble glucan, which was de novo-synthesized from sucrose by the purified enzyme, was analyzed by a gas-liquid chromatography-mass spectroscopy and was found to be 1,6-alpha-D-glucan with highly (35%) branched structure of 1,3,6-linked glucose residue.

Amino Acids↗
PubMed1982

Synthesis of glucan on the cell surface of Streptococcus mutans: chemical and scanning electron microscopic studies.

The appearance and continuing growth of extracellular material on Streptococcus mutans HS6 cells in sucrose-containing Merthiolated buffer was observed in a scanning electron microscope and was found to be related to the glucan synthesis on the cell and to adherence of the cell to a smooth surface. Cells grown in broth completely deprived of sucrose by invertase (HS6-IV) had a characteristic, slightly rugged surface structure. On incubation of HS6-IV in the sucrose-containing buffer, a few small globular particles appeared on the surface and grew to an irregular shape (globular to fibrilar) after several hours. The increase in the total glucan content of the cells paralleled the growth of the globular material, to which ferritin-conjugated anti-dextran globulin was found to bind. On the cell surface of cells harvested from conventional broth, both small globular and irregular structures, which possibly formed from sucrose in the broth, existed originally and continued to grow during incubation, along with the material newly appearing on the surface. The accumulation of glucan on the cells resulted in their adherence to a glass surface. The inhibition of growth of the extracellular material on the cells by trypsin, dextranase or anti-glucosyltransferase corresponded to the decrease in glucan synthesis and the loss of adhering ability. These results indicated that the material growing on the cell surface was glucan synthesized by glucosyltransferases.

Adhesiveness↗
PubMed1980

The influence of alcohol on the appearance of frontal midline theta activity.

In 30 healthy male students aged 19 to 28 years of a mean age of 22.6 years, a study was made of the amount of Fm theta by recording EEG patterns from Fz by monopolar lead during the Uchida-Kraepelin serial addition task of a five-minute duration, before and after taking 1.5 ml of 43% whisky per Kg. At the same time, a determination was made of the blood concentration of alcohol, along with the MPI test in all subjects, to examine the relationship between the results obtained and the occurrence of Fm theta. The amount of Fm theta was significantly increased following drinking.

Adult↗
PubMed1979

A recent trend of drug dependence in Kurume district (southern Japan).

With the intention of grasping the trend of drug dependence in recent years, a comparative study was made of the relative prevalence of three different types of drug dependence, i.e., the sociopathic, neurotic and iatrogenic types, in each of the three postwar decades among patients seen at the neuropsychiatric clinic of Kurume University. As a result it was found that the recent trend of drug dependence has the following features: (1) multiplicity of drugs involved, (2) a continuing high incidence of the iatrogenic types, (3) resurgence of the sociopathic type, making the recent pattern of drug dependence similar to that prevailing in postwar years, and (4) prevalence of dependence or abuse among the younger generation. Consideration was also given to the sociopathologic significance of these recent features of drug dependence.

Age Factors↗
PubMed1979

Effect of salts on water-insoluble glucan formation by glucosyltransferase of Streptococcus mutans.

The formation of water-insoluble glucan by extracellular glucosyltransferase from Streptococcus mutans 6715 found to be greatly stimulated by various mono- or divalent cations. An enzyme preparation, obtained by ethanol fractionation, was able to catalyze the formation of water-insoluble glucan from sucrose in the presence of monovalent cations above 100mM or divalent cations above 20 mM at neutral pH. As the concentration of monovalent and divalent cations was reduced to below 10 mM and 1 mM, respectively, the formation of insoluble glucan decreased to a negligible amount. High concentrations of these cations were found to stimulate the formation of insoluble glucan in the following ways: (i) it increased the activity of total glucosyltransferase up to 1.6- and 2.7-fold in the absence and presence of a primer dextran, respectively, and (ii) it changed the formation of soluble glucan to insoluble. It was postulated that one of the essential factors for the formation of insoluble glucan would be to keep more than two water-soluble glucan chains close to enzyme aggregates and that such interaction could be enhanced by the presence of high cation concentrations.

Carbohydrate Metabolism↗
PubMed1975

Serological purification of polysaccharide antigens from Streptococcus mutans serotypes a and d: characterization of multiple antigenic determinants.

The polysaccharide antigen preparations from serotype a and serotype d strains of Streptococcus mutans contained both a serotype-specific antigenic determinant and a common a-d antigenic determinant, as demonstrated by agar gel diffusion studies and a quantitative cross-precipitin assay. The chromatographically purified antigens were isolated by a method which depended on their serological specificity to determine if these two antigenic determinants were located on the same molecule. The a and d polysaccharides were recovered from specific antigen-antibody complexes and characterized with respect to their immunological specificity and chemical composition. Agar gel diffusion tests demonstrated that, in both the a and d preparations, the serotype-specific antigenic determinant and the common a-d antigenic determinant were present in one molecule.

Animals↗
PubMed1974

Mechanism of the Adherence of Streptococcus mutans to Smooth Surfaces III. Purification and Properties of the Enzyme Complex Responsible for Adherence.

Enzymes which possess the ability to cause the adherence of Streptococcus mutans cells to a smooth glass surface were purified 1,100 times by chromatography on agarose gel followed by hydroxylapatite gel. During the purification procedures, the enzymes from strain HS6 (group a) were examined for the synthesis of water-soluble and water-insoluble polysaccharide and the ability to produce adherence. The enzyme preparations producing adherence of the S. mutans cells in the presence of sucrose possessed a molecular size of about 400,000 to 2,000,000 and were composed of approximately equivalent amounts of dextran and levan sucrases and 5 to 30% polysaccharide. The most highly purified preparation contained a negligible amount of contaminating protein as judged by polyacrylamide gel electrophoresis, immunoelectrophoresis, and gel diffusion. In these three tests, the location of the enzyme responsible for the synthesis of insoluble polymer was detected by embedding or covering the enzyme-containing gel with a layer of sucrose-containing agarose gel and observing the formation of insoluble polymer. During purification the ability of all fractions to produce adherence was parallel with the enzyme activity responsible for the synthesis of insoluble polysaccharide from sucrose. About two-thirds of the sucrase enzyme complex in the S. mutans culture fluid synthesized water-soluble polymer. This complex, obtained by filtration through agarose gel, was smaller in molecular size, lower in sugar content, and did not produce adherence, in contrast to the enzyme complex which possessed adherence activity. The inhibition of the enzyme complex synthesizing soluble polymer required more anti-synthetase serum than that required to inhibit the synthesis of water-insoluble polymer. It is not known whether the lack of adherence activity in this enzyme was due to its smaller size and lower sugar content or the absence of unknown factors which are essential for its activity. The carbohydrate in these enzyme preparations, composed of glucose, may represent a primer molecule and/or a remnant of the polymer synthesized by the enzyme. The enzyme activity was not inhibited by anti-dextran globulin.

Journal Article↗
PubMed1974

Mechanism of adherence of Streptococcus mutans to smooth surfaces. II. Nature of the binding site and the adsorption of dextran-levan synthetase enzymes on the cell-wall surface of the streptococcus.

The mechanism of adsorption of the Streptococcus mutans enzymes responsible for the synthesis of insoluble dextran-levan to the S. mutans cell-wall binding sites has been studied. Certain characteristics of these binding sites are presented. The adsorption of these enzymes to the cell surface occurred rapidly without the addition of a source of energy and over a pH range of 3 to 11. The adsorption was inhibited by soluble dextran, probably due to the strong affinity of the polymer to the enzyme. All other polymers and sugars studied showed little or no inhibition. The adsorption was also inhibited by antibody globulin to the a-d immunologically specific group antigen surface polysaccharide of S. mutans and by anti-dextran globulin. The inhibition by anti-a-d globulin is considered to be due to a restriction of access of enzyme to the binding site of the enzyme which may be located in close proximity to the group antigen. On the other hand, anti-dextran globulin appeared to directly inhibit the adsorption by covering the binding site. Dextranase destroyed the binding site and released glucose from the S. mutans cells. These data indicate that S. mutans grown in media containing glucose possesses a small amount of dextran on the cell surface, and that this dextran is, or is a part of, the binding site for enzymes which synthesize the insoluble dextran-levan polymer. Trypsin inhibited the synthesis of insoluble polysaccharide and the adherence of cells. It is not clear in this case that destruction of the binding sites occurred. These data present a partial explanation of the processes which may be concerned in the formation of dental plaque on the smooth surfaces of teeth.

Adsorption↗
PubMed1973

Structure and immunological specificity of the Streptococcus mutans group b cell wall antigen.

The Streptococcus mutans group b antigen of strain FA1 has been defined as to chemical composition and immunological specificity. The antigen in cold trichloroacetic acid extracts was fractionated on diethylaminoethyl-Sephadex A-25 at pH 8.5. Two forms were isolated: a polysaccharide and a mucoprotein. The two polymers reacted as a single substance in agar gel diffusion against specific adsorbed FA1 rabbit antisera but were separated by gel immunoelectrophoresis. No reaction with any other S. mutans or streptococcal group sera occurred. Galactose composed about one-third and galactosamine about 3% of the total weight of each polymer. Rhamnose was a major component of the polysaccharide (47%) but was present only in traces in the mucoprotein. The protein content of the latter was about 40%. No significant quantities of glycerol, phosphorus, or muramic acid were present in either case. Pepsin and trypsin had no effect on the serological specificity of the mucoprotein. d-Galactose and d-galactosamine were strong inhibitors (70%) of the precipitin reaction, whereas d-glucose, d-glucosamine, and N-acetyl-d-glucosamine inhibited between 25 and 35%. The results indicate that the antigen is a major antigenic component of the cell wall and that the specificity of the antigen resides in binding sites which contain both d-galactose and d-galactosamine. Agglutination of whole cells by specific group b antiserum indicates the antibody receptor sites of the polysaccharide antigen are at the surface of the streptococcal cell. The mucoprotein, but not the polysaccharide, was released from the cell by lysozyme. Lysis did not occur. The immunological specificity and other characteristics of the antigen establishes it as the identifying antigen of S. mutans group b.

Agglutination Tests↗
PubMed1973

Extraction, purification, and chemical and immunological properties of the Streptococcus mutans group "a" polysaccharide cell wall antigen.

An antigen of Streptococcus mutans has been extracted from HS6 (group "a") whole cells and repeatedly fractionated by Sephadex chromatography. The antigen is shown to be a polysaccharide and contains the S. mutans group "a" antigenic site and also a second antigenic site which is common to "a" strains and 2 of 3 group "d" strains. Immunological electrophoretic and chromatographic data indicate that the two sites exist in a single molecule. The polysaccharide has a molecular weight of 107,000 and is composed of glucose, galactose, glucosamine, and galactosamine. No significant quantities of lipid, phosphorus, glycerol, or ribitol are present. Immunological specificity of the group "a" polysaccharide site depends primarily on a d-glucose . d-glucose sequence, the "a-d" site on a terminal d-galactose. Water at 100 C and pepsin (pH 2.5) at room temperature are very effective in extracting the polysaccharide from lyophilized S. mutans cells. Trypsin and lysozyme are less effective. The antigen-antibody combining site appears to be located at the cell wall surface. A small quantity of enzyme-resistant protein (5%) is firmly linked to the antigen and is considered to be a remnant of a protein to which the polysaccharide is attached in the cell wall. The composition of the protein does not identify it as a part of the peptidoglycan. No reaction to the purified polysaccharide is obtained with antisera specific for teichoic acid glycerophosphate polymers from streptococci, staphylococci, or lactobacilli.

Alkaline Phosphatase↗
PubMed1973

Mechanism of adherence of Streptococcus mutans to smooth surfaces. I. Roles of insoluble dextran-levan synthetase enzymes and cell wall polysaccharide antigen in plaque formation.

The mechanism of adherence of Streptococcus mutans to smooth glass surfaces has been studied. The results with both viable and heat-killed cells showed that the process required (i) the synthesis of a water-insoluble dextran-levan polymer by cell-bound enzymes and (ii) the participation of a binding site on the surface of the S. mutans cell. Synthesis of the polymer from sucrose in the presence of the cells was required for adherence, and indicates that an "active" form of the polymer was required. Polymer synthesized by cell-free S. mutans enzymes when added to S. mutans cells did not produce adherence. Purified antibody globulin, specific for the a-d site in the polysaccharide S. mutans group a antigen, completely inhibited adherence. Antibody to the second antigen present in the polysaccharide molecule, the a antigen, did not inhibit adherence. The evidence indicates that adherence did not require an antigenic binding site which might be common to all S. mutans strains. The orientation of the synthetase enzyme(s), antigenic binding site, and dextran-levan polymer on the cell surface is under study.

Adsorption↗
PubMed1972

Chemical composition and immunological specificity of the streptococcal group O cell wall polysaccharide antigen.

The group O streptococcal group antigen was shown to be a polysaccharide located in the cell wall of the organism. The antigen could be extracted by one of several methods: (i) 0.5 n NaOH at 37 C, (ii) phenol-water (50:50) at 68 C, (iii) 0.2 n HCl at 100 C, or (iv) 10% trichloroacetic acid at 4 C. The last method yielded more polysaccharide with less protein contamination. The polysaccharide was purified on diethylaminoethyl-Sephadex A-25 and Sephadex G-200. It was composed of two-thirds glucosamine and galactosamine, and the remainder glucose plus galactose. Rhamnose, glycerol, ribitol, and muramic acid were absent. Total phosphorus and amino acids were each less than 0.1%. N-Acetyl-beta-d-glucosamine exerted a strong inhibition of the precipitin reaction and is considered the immunodominant sugar. Glucosamine and glucose possessed a partial inhibitory activity. Galactose and galactosamine were essentially negative. No evidence of cross-reactivity was found between the O polysaccharide and group A and L polysaccharides, and group A and Staphylococcus aureus teichoic acids, which posesss N-acetylglucosamine specificity. The release of limited quantities of N-acetyl-glucosamine from its terminal location by enzyme, and glucose by acid hydrolysis, indicates a limited number of side chains in the O antigen. The glucosamine is in acid-stable linkage in the polysaccharide. Glucose was not released by beta-glucosidase and probably does not occupy a terminal position. The O antigen is the only known streptococcal polysaccharide antigen which does not contain rhamnose. The effect of these factors on the immunological specificity is discussed. O serum, after adsorption with the purified polysaccharide, was used to demonstrate the presence of protein antigens in acid extracts of cells from each of the nine strains examined. These antigens may represent type antigens. Two of these strains, originally described as group O, did not contain the O polysaccharide.

Antibodies, Bacterial↗
PubMed1972

Immunological and chemical studies of the streptococcal group O protein type antigens.

Two antigens present in group O streptococci have been described and are designated as type antigens. No other antigens are present in extracts of group O cells in significant quantity. Antigen I is a cell wall protein, and II is a nucleoprotein of undetermined cell location. The immunological specificity of the latter resides only in the protein. Both antigens have been extracted from the cell with water or 0.2 n HCl (pH 2) at 100 C, and antigen I can be removed by trypsin. After purification, pH 2 was found to destroy the serological activity of each antigen. Antgens I and II and also the O polysaccharide antigen can be distinguished on a single agar-gel diffusion plate with group O whole-cell antiserum. N- and C-terminal amino acid analyses and total amino acid composition prove that I and II are separate molecular species. Each possesses properties which separate them from the streptococcal M, R, and T proteins. Antigen I separated into three fractions, each possessing serological specificity, on a diethylaminoethyl-Sephadex column at pH 8.6 during an increase in buffer molarity. The protein is considered to possess a multiple unit structure. The results indicate that the O streptococci are limited to two serological types. Group O whole-cell antisera, after adsorption with a strain not containing the O polysaccharide, can now be prepared for the positive identification of the types of group O streptococci.

Alkaline Phosphatase↗