Streptomyces castrisilvae sp. nov. and Streptomyces glycanivorans sp. nov., novel soil streptomycetes metabolizing mutan and alternan.

Six bacterial strains, Mut1T, Mut2, Alt1, Alt2, Alt3T, and Alt4, were isolated from soil samples collected in parks in Gothenburg, Sweden, based on their ability to utilize the insoluble polysaccharides α-1,3-glucan (mutan; Mut strains) or the mixed-linkage α-1,3/α-1,6-glucan (alternan; Alt strains). Analysis of 16S rRNA gene sequences identified all strains as members of the genus Streptomyces. The genomes of the strains were sequenced and subsequent phylogenetic analyses identified Mut2 as a strain of Streptomyces laculatispora and Alt1, Alt2 and Alt4 as strains of Streptomyces poriferorum, while Mut1T and Alt3T were most closely related to the type strains Streptomyces drozdowiczii NBRC 101007T and Streptomyces atroolivaceus NRRL ISP-5137T, respectively. Comprehensive genomic and biochemical characterizations were conducted, highlighting typical features of Streptomyces, such as large genomes (8.0-9.6 Mb) with high G+C content (70.5-72.0%). All six strains also encode a wide repertoire of putative carbohydrate-active enzymes, indicating a capability to utilize various complex polysaccharides as carbon sources such as starch, mutan, and cellulose, which was confirmed experimentally. Based on phylogenetic and phenotypic characterization, our study suggests that strains Mut1T and Alt3T represent novel species in the genus Streptomyces for which the names Streptomyces castrisilvae sp. nov. and Streptomyces glycanivorans sp. nov. are proposed, with strains Mut1T (=DSM 117248T=CCUG 77596T) and Alt3T (=DSM 117252T=CCUG 77600T) representing the respective type strains.

α-1,3-Glucanase: present situation and prospect of research.

α-1,3-Glucanases hydrolyze α-1,3-glucan which is an insoluble linear α-1,3-linked homopolymer of glucose and these enzymes are classified into two families of glycoside hydrolases on the basis of amino acid sequence similarity; type-71 α-1,3-glucanases found in fungi and type-87 enzymes in bacteria. α-1,3-Glucan (also called 'mutan') is a major component of dental plaque formed by oral Streptococci and has important physiological roles in various fungal species, including as a component of cell walls, an endogenous carbon source for sexual development, and a virulent factor. Considering these backgrounds, α-1,3-glucanases have been investigated from the perspectives of applications to dental care and development of cell-wall lytic enzymes. Compared with information regarding other glycoside hydrolases such as amylases, cellulases, chitinases, and ß-glucanases, there is limited biochemical and structural information available regarding α-1,3-glucanase. Further research on α-1,3-glucanases on enzyme application to dental care and biological control of pathogenic fungi is expected. In this mini-review, we briefly describe how α-1,3-glucanases are categorized and characterized and present our study findings regarding α-1,3-glucanase from Bacillus circulans KA-304. Furthermore, we briefly discuss potential future applications of α-1,3-glucanases.

Secreted expression of Leuconostoc mesenteroides glucansucrase in Lactococcus lactis for the production of insoluble glucans.

We expressed a glucansucrase, DsrI, from Leuconostoc mesenteroides that catalyzes formation of water-insoluble glucans from sucrose using a nisin-controlled gene expression system in Lactococcus lactis. These polymers have potential for production of biodegradable gels, fibers, and films. We optimized production of DsrI using several different background vectors, signal peptides, strains, induction conditions, and bioreactor parameters to increase extracellular accumulation. Optimal production of the enzyme utilized a high-copy plasmid, pMSP3535H3, which contains a nisin immunity gene, L. lactis LM0230, and bioreactors maintained at pH 6.0 to stabilize the enzyme. We were able to significantly improve growth using the lactic acid inhibitor heme and by continuous removal of lactic acid with anion exchange resins, but enzyme production was less than the controls. The recombinant enzyme under optimized conditions accumulated in the culture medium to approximately 380 mg/L, which was over 150-fold higher compared to the native L. mesenteroides strain. Methods are also included for purification of DsrI utilizing the glucan-binding domain of the enzyme.

Detailed structural characterization of five water-insoluble α-glucans produced by glucansucrases from Streptococcus spp.

Water-insoluble α-glucans synthesized from sucrose by glucansucrases from Streptococcus spp. are essential in dental plaque and caries formation. Because limited information is available on the fine structure of these biopolymers, we analyzed the structures of unmodified glucans produced by five recombinant Streptococcus (S.) mutans DSM 20523 and S. salivarius DSM 20560 glucansucrases in detail. A combination of methylation analysis, endo-dextranase and endo-mutanase hydrolyses, and HPSEC-RI was used. Furthermore, crystal-like regions were analyzed by using XRD and 13C MAS NMR spectroscopy. Our results showed that the glucan structures were highly diverse: Two glucans with 1,3- and 1,6-linkages were characterized in detail besides an almost exclusively 1,3-linked and a linear 1,6-linked glucan. Furthermore, one glucan contained 1,3-, 1,4-, and 1,6-linkages and thus had an unusual, not yet described structure. It was demonstrated that the glucans had a varying structural architecture by using partial enzymatic hydrolyses. Furthermore, crystal-like regions formed by 1,3-glucopyranose units were observed for the two 1,3- and 1,6-linked glucans and the linear 1,3-linked glucan. 1,6-linked regions were mobile and not involved in the crystal-like areas. Altogether, our results broaden the knowledge of the structure of water-insoluble α-glucans from Streptococcus spp.

A comprehensive review on mutan (a mixed linkage of α-1-3 and α-1-6 glucans) from bacterial sources.

Mutan is an extracellular sticky polymer having α-1-3 and α-1-6 glycosidic linkages with a large diversity in molecular weights and structures depending on the source. These compounds are reported to be highly thermostable and also have potential physiochemical and biological applications. The main aim of this review is to provide an overview of glucosyltransferases and their role in mutan synthesis. The production strategies and structural properties of bacterial mutans are discussed with a goal to improve production efficiency. The physicochemical features, chemical modifications, potential industrial applications and future prospects are also discussed. According to data, mutan and its derivatives will play a larger role in medicinal sectors and as thermoplastics in the near future.Abbreviations: ABTS: 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid; BHI: Brain heart infusion broth; 13C (HSQC) NMR: Heteronuclear Single Quantum Coherence NMR; CBMs: Carbohydrate binding modules; DPPH: 2,2-diphenyl-1-picrylhydrazyl; FTIR: Fourier-transform infrared spectroscopy; GC-MS: Gas chromatography-mass spectrometry; GPC: Gel permeation chromatography; Gtfs: Glucosyltransferases; 1H (DQF-COSY): Double-quantum filtered correlation spectroscopy; HPAEC-PAD: High-performance anion exchange chromatography with pulsed amperometric detection; HPLC: High performance liquid chromatography; HPSEC-RI: High-performance size exclusive chromatography coupled with refractive index; HPSEC-MALLS: High-performance size exclusive chromatography with multi-angle laser light scattering detection; MALDI-TOF: Matrix-Assisted Laser Desorption/Ionization-Time of Flight mass spectrometry; Mw: Weight-average molecular weight; MWD: Molecular weight distribution; NMR: Nuclear magnetic resonance spectroscopy; TEM: Transmission electron microscopy; THB: Todd Hewitt Broth; TTY: Tryticase tryptose yeast extract broth.

Detailed Structural Characterization of Glucans Produced by Glucansucrases from Leuconostoc citreum TMW 2.1194.

The water kefir organism Leuconostoc citreum TMW 2.1194 forms highly branched dextrans with O3- and O4-bound side chains. To obtain detailed information on the enzymatic synthesis of these polymers, the four glucansucrases encoded by Leuconostoc citreum TMW 2.1194 were cloned, heterologously expressed, and used for polysaccharide production. Molecular and macromolecular structure of the synthesized glucans were analyzed by methylation analysis, two-dimensional NMR spectroscopy, oligosaccharide analysis after partial hydrolysis, and asymmetric flow field-flow fractionation. It was demonstrated that two glucansucrases form insoluble glucans with variously branched dextran sections and varying portions of consecutive, 1,3-linked glucose units. In contrast, the other two glucansucrases synthesized O3- (Lc6255) and O4-branched (Lc1785) soluble dextrans. Analysis, isolation, and characterization of enzymatically liberated oligosaccharides showed that monomeric and elongated side chains are abundant in both polysaccharides. From the structures and size distributions it was concluded that Lc1785 is mainly responsible for synthesis of fermentatively produced soluble dextrans.

Deletion of uncharacterized domain from α-1,3-glucanase of Bacillus circulans KA-304 enhances heterologous enzyme production in Escherichia coli.

α-1,3-Glucanase (Agl-KA) of Bacillus circulans KA-304 consists of an N-terminal discoidin domain (DS1), a carbohydrate binding module family 6 (CBM6), threonine and proline repeats (TP), a second discoidin domain (DS2), an uncharacterized conserved domain (UCD), and a C-terminal catalytic domain. Previously, we reported that DS1, CBM6, and DS2 have α-1,3-glucan-binding activity and contribute to α-1,3-glucan hydrolysis. In this study, UCD deletion mutant (AglΔUCD) was constructed, and its properties were compared with those of Agl-KA. α-1,3-Glucan hydrolyzing, α-1,3-glucan binding, and protoplast-forming activities of AglΔUCD were almost the same as those of Agl-KA. kcat/Km values of AgΔUCD and Agl-KA were 11.4 and 11.1 s-1 mg-1 mL, respectively. AglΔUCD and Agl-KA exhibited similar characteristics, such as optimal pH, pH stability, optimal temperature, and thermostability. These results suggest that UCD is not α-1,3-glucan-binding and flexible linker domain, and that deletion of UCD does not affect the affinity of N-terminal binding domains and the catalytic action of the C-terminal domain. Subsequently, heterologous UCenzyme productivity of AglΔD in Escherichia coli was compared with that of Agl-KA. The productivity of AglΔUCD was about 4-fold larger than that of Agl-KA after an 8-h induction at 30°C. In the case of induction at 20°C, the productivity of AglΔUCD was also larger than that of Agl-KA. These findings indicate that deletion of only UCD enhances the enzyme productivity in E. coli.

Efecto antibacteriano del extracto etanólico de Camellia sinensis y propóleo, frente a cepas de Streptococcus mutans / Antibacterial effect of the ethanolic extract of Camellia sinensis and propolis, against strains of Streptococcus mutans

Objetivo. Determinar el efecto antibacteriano del extracto etanólico del té verde (Camellia sinensis) y propóleo a una concentración de 10, 20 y 30% a las 24 y 48 horas sobre Streptococcus mutans. Métodos. Estudio experimental, in vitro, comparativo, con muestra no probabilística de 150 discos de papel, distribuidos en 30 placas Petri previamente preparadas con agar sangre e inoculadas con cepas de Streptococcus mutans, se colocaron tres discos embebidos en extracto etanólico al 10, 20 y 30%, un disco en clorhexidina 0,12% (control positivo) y un disco en agua destilada, fueron llevadas a la incubadora y pasadas las 24 horas y 48 horas se midieron los correspondientes halos de inhibición. Los extractos se obtuvieron mediante un proceso de maceración modificado, en aparato de agitación rotatorio. Resultados. El mayor halo inhibitorio del extracto etanólico de Camellia sinensis (C. sinensis) frente a Streptococcus mutans fue en la concentración al 30% a las 24 h y 48 h, mientras que el mayor halo inhibitorio del extracto etanólico de propóleo, fue en la concentración al 30% a las 24 h, por lo tanto, los extractos naturales mostraron ser sensibles en la escala de Duraffourd. Conclusiones. Se evidenció que el propóleo al 30% mostró un efecto antibacteriano similar a la clorhexidina, considerada gold estándar, el tiempo en el que existió mayor efecto antibacteriano del extracto etanólico de C. sinensis y propóleo frente a Streptococcus mutans, fue a las 24 horas, el diámetro de los halos inhibitorios disminuyó, conforme aumentó el tiempo de exposición al microorganismo.

Objective. To determine the antibacterial effect of the ethanolic extract of green tea (Camellia sinensis) and propolis at a concentration of 10, 20 and 30% at 24 and 48 hours on Streptococcus mutans. Methods. Experimental, in vitro, comparative study, with a non-probabilistic sample of 150 paper discs, distributed in 30 Petri dishes previously prepared with blood agar and inoculated with strains of Streptococcus mutans, were placed 3 discs soaked in ethanolic extract at 10, 20 and 30%, 1 disk in 0.12% chlorhexidine (positive control) and 1 disk in distilled water, they were taken to the incubator and after 24 hours and 48 hours the measurements corresponding to the inhibition halos were made. The extracts were gotten by a modified maceration process, in a rotary stirring apparatus. Results. The highest inhibitory halo of the ethanolic extract of C. sinensis against Streptococcus mutans was in the concentration at 30% at 24 h and 48 h, while the highest inhibitory halo of the ethanolic extract of propolis, was in the concentration at 30% at 24 h; therefore, the natural extracts showed to be sensitive on the Duraffourd scale. Conclusions. It was evidenced that 30% propolis showed an antibacterial effect similar to chlorhexidine, considered gold standard, the time in which there was a greater antibacterial effect of the ethanolic extract of Camellia Sinensis and propolis against Streptococcus mutans, was at 24 hours, taking into account that the diameter of the inhibitory halos decreased, as the exposure time to the microorganism increased

Mutan: A mixed linkage α-[(1,3)- and (1,6)]-d-glucan from Streptococcus mutans, that induces osteoclast differentiation and promotes alveolar bone loss.

Mutan is an extracellular polysaccharide of Streptococcus mutans (S. mutans) that consists of α-(1,3)-linked glucose residues in main chains and α-(1,6) bonds in side chains. In the present study, mutan was isolated from S. mutans, and its structural characteristics were determined using Fourier-transform infrared spectroscopy (FT-IR) and (13)C nuclear magnetic resonance (NMR) spectroscopy. The effects of mutan on RANKL-induced osteoclast differentiation in RAW 264.7 cells were examined. Furthermore, microCT and morphometric analyses were used to determine the contribution of mutan to alveolar bone loss in the maxilla of a rat periodontitis model. Mutan increased (more than 2-fold) RANKL-induced osteoclast differentiation in a dose-dependent manner. Mutan also enhanced the alveolar bone loss in the rat maxilla 2.3-fold. In mutan-treated rats, the bone mineral density, bone volume, trabecular number, and trabecular thickness decreased, whereas trabecular separation significantly increased. In addition, mutan and lipopolysaccharide (LPS) induced similar microarray profiles in RAW 264.7 cells. A total of 43 genes related to osteoclastogenesis were differentially expressed after either mutan or LPS treatment. Five-fold increases in the expression of several genes, including IL-1ß, IL-1α, IL-6, and chemokine ligands, were observed in mutan-treated RAW 264.7 cells. These results suggest a molecular mechanism for the inflammation induced by S. mutans during the establishment of periodontal disease.

(1→3)-α-D-Glucan hydrolases in dental biofilm prevention and control: A review.

Dental plaque is a highly diverse biofilm, which has an important function in maintenance of oral and systemic health but in some conditions becomes a cause of oral diseases. In addition to mechanical plaque removal, current methods of dental plaque control involve the use of chemical agents against biofilm pathogens, which however, given the complexity of the oral microbiome, is not sufficiently effective. Hence, there is a need for development of new anti-biofilm approaches. Polysaccharides, especially (1→3),(1→6)-α-D-glucans, which are key structural and functional constituents of the biofilm matrix, seem to be a good target for future therapeutic strategies. In this review, we have focused on (1→3)-α-glucanases, which can limit the cariogenic properties of the dental plaque extracellular polysaccharides. These enzymes are not widely known and have not been exhaustively described in literature.

Effects of Bamboo Salt with Sodium Fluoride on the Prevention of Dental Caries / 치위생과학회지

BACKGROUND: Dental caries is one of several prevalent oral diseases caused by dental plaque biofilms. This study evaluated the anti-cariogenic effects of a bamboo salt (BS) and sodium fluoride (NaF) mixture on oral bacteria.METHODS: The effects of several mixtures of NaF and BS on acid production, growth, and adhesion to glass beads of Streptococcus mutans, and their anti-cariogenic properties were investigated. The growth of S. mutans was measured according to optical density at 3, 6, 9, 12, 15, 18, and 24 hours after treatment using spectrophotometry at a wavelength of 600 nm, while pH was measured using a pH meter. Adhesion of S. mutans was measured according to the weight of glass beads from each group before and after incubation. Gene expression was measured using real-time polymerase chain reaction. Acid production and growth patterns of S. mutans were compared using repeated measures analysis of variance, followed by Scheffe's post-hoc test. The Kruskal–Wallis test was used to compare adhesion, followed by the Mann–Whitney test. Gene expression in the experimental and control samples was compared using the Student's t-test.RESULTS: Growth, acid production, and adhesion of S. mutans were inhibited in all experimental groups. Expression of gft and fructosyltransferase in S. mutans was inhibited in all groups. A mixture of NaF and BS significantly reduced growth, acid production, adhesion, and gene expression of S. mutans compared with the other groups.CONCLUSION: Results of the present study demonstrated that a mixture of NaF and BS was useful as a mouth rinse in preventing dental caries.

Análise mecânica comparativa do destorque de parafusos de minipilar com um novo componente protético, juntamente com avaliação microbiológica do vedamento para implantes de plataforma Cone Morse / Comparative mechanical analysis of the untorsion of miniabutment screws with a new prosthetic component, together with microbiological evaluation of the seal for Cone Morse platform implants

A interface implante pilar (IAI) por se constituir de duas peças inevitavelmente apresentam micro lacuna (GAP), na qual pode ocorrer infiltração bacteriana, permitindo a penetração de microorganismos que colonizam na parte interna do implante levando ao acúmulo de biofilme e, podendo levar ao desenvolvimento da periimplantite. O desgaste da conexão interna do implante é algo que ocorre com frequência, muitas vezes pela fratura do parafuso e/ou, pela perda da rosca interna do implante. A ausência de informações prévias também pode gerar a necessidade da remoção do implante, devido a estas intercorrências, surge a possibilidade da criação de um novo componente para implantes para possibilitar a reabilitação protética, sem ter que passar por uma nova cirurgia de remoção e instalação do implante. O objetivo do trabalho foi mensurar o nível de afrouxamento do parafuso do pilar protético e do minipilar comparando com novo componente protéticos, na tentativa de simular o comportamento do conjunto implante/pilar/prótese. Foram utilizados vinte implantes de plataforma cone morse (CM) da DSP® com seus respectivos mini pilares, na qual foram distribuídos em 2 grupos(n=10): Grupo 1 - implante CM + mini pilar FlexCone® DSP + coroa simplificada pirâmide invertida carga aplicada 3 mm do centro da coroa. E Grupo 2 - implante CM + mini pilar novo + coroa simplificada pirâmide invertida carga aplicada 3 mm do centro da coroa. Foram realizados ciclagem mecânica com carga 133 N, durante 2x106 ciclos, com frequência 2 Hz e temperatura de 37ºC em ambos grupos. Um torquímetro digital foi usado para medir os valores de torque reverso do parafuso protético da coroa e também do pilar protético, antes e após o carregamento. Os resultados do modelo de regressão demonstraram diferenças estatisticamente significativas em função do envelhecimento comparando os grupos da coroa sobre o pilar protético (p = 0.020) e entre os grupos do pilar sobre o implante (p = 0.048), indicando que após o envelhecimento de 2.000.000 de ciclos ao longo do tempo está associado de maneira significativa a essas variáveis no contexto deste estudo. O segundo objetivo deste estudo foi avaliar in vitro a taxa de infiltração bacteriana através da IAI, entre o novo componente protético e a superfície interna do implante, juntamente foi analisado a permeabilidade do IAI para colonização bacteriana. Um total de oitenta implantes foram testados. As estruturas montadas para grupo 1 foi torqueado com 20 N/cm e do G2 foram torqueados com 45 N, ambos imersos em microtubos contendo 200 µl de saliva humana. Após 14 dias de incubação da amostra de bactéria nos implantes, foi realizada uma análise qPCR (reação da cadeia da polimerase em tempo real). O teste revelou que não houve diferenças estatisticamente significativas no crescimento bacteriana entre os grupos em qualquer um dos pontos temporais analisados. Conclui-se que o novo componente testado apresentou um destoque menor do que comparado ao mini pilar FlexCone DSP® e apresentou infiltração bacteriana no GAP da conexão implante-pilar semelhante comparado ao mini pilar original da empresa (AU)

The abutment implant interface (IAI), as it consists of two pieces, inevitably presents a micro gap (GAP), in which bacterial infiltration can occur, allowing the penetration of microorganisms that colonize in the internal part of the implant, leading to the accumulation of biofilm and, which can lead to development of peri-implantitis. Wear of the implant's internal connection is something that occurs frequently, often due to screw fracture and/or loss of the implant's internal thread. The lack of prior information can also generate the need to remove the implant, due to these complications, the possibility arises of creating a new component for implants to enable prosthetic rehabilitation, without having to undergo a new surgery to remove and install the implant. implant. The objective of the work was to measure the level of screw loosening of the prosthetic abutment and the mini-abutment compared with the new prosthetic component, in an attempt to simulate the behavior of the implant/ abutment/prosthesis set. Twenty DSP® morse cone (CM) platform implants were used with their respective mini pillars, which were distributed into 2 groups (n=10): Group 1 - CM implant + FlexCone® DSP mini pillar + simplified crown inverted pyramid load applied 3 mm from the center of the crown. And Group 2 - CM implant + new mini abutment + simplified crown inverted pyramid load applied 3 mm from the center of the crown. Mechanical cycling was carried out with a load of 133 N, for 2x106 cycles, with a frequency of 2 Hz and a temperature of 37ºC in both groups. A digital torque wrench was used to measure the reverse torque values of the prosthetic crown screw and also the prosthetic abutment, before and after loading. The results of the regression model demonstrated statistically significant differences as a function of aging comparing the crown-on-prosthetic abutment groups (p =0.020) and between the abutment-on-implant groups (p = 0.048), indicating that after aging 2,000 ,000 cycles over time is significantly associated with these variables in the context of this study. The second objective of this study was to evaluate in vitro the rate of bacterial infiltration through the IAI, between the new prosthetic component and the internal surface of the implant, together with the permeability of the IAI for bacterial colonization. A total of eighty implants were tested. The assembled structures for group 1 were torqued with 20 N/cm and G2 were torqued with 45 N, both immersed in microtubes containing 200 µl of human saliva. After 14 days of incubation of the bacteria sample in the implants, a qPCR (real-time polymerase chain reaction) analysis was performed. The test revealed that there were no statistically significant differences in bacterial growth between groups at any of the time points analyzed. It is concluded that the new component tested presented a lower impact compared to the FlexCone DSP® mini abutment and presented bacterial infiltration in the GAP of the implant-abutment connection similar to the company's original mini abutment.(AU)

In vitro antibiotic effect of the leaching solution of astragalus on cariogenic bacteria / 实用口腔医学杂志

Astragalus produced in Gansu were chosen as the raw material to leachate. Studied the antibiotic effects of the leaching solution on the cariogenic bacteria and compared with the imported bacteriostatic product MI. Streptococcus mutans and Lactobacilli were cultured in the medium for 24 h. The PH and A600 values were measured. Statistical analysis was conducted by using SPSS 13.0. The leaching solution of astragalus has the same inhibitory effects on the growth and acid production of streptococcus mutans and lactobacilli as MI.

Glucanos extracelulares bacterianos: estructura, biosíntesis y función / Extreacellular bacterial glucans: structure, biosynthesis and function

La caries dental es una de las enfermedades más frecuentes en el ser humano. En su etiología multifactorial, desempeñan un papel importante determinadas bacterias cariogénicas, que en interacción con la superficie del diente promueven su desmineralización. Dentro de los mecanismos mediadores de la adhesión bacteriana, se encuentra la producción de polisacáridos extracelulares bacterianos. En particular los glucanos sintetizados por las glucosiltransferasas, no solo permiten la adherencia, sino que también constituyen una fuente nutricional para las bacterias, por lo tanto, la actividad de dichas enzimas se considera un factor de virulencia bacteriana en la caries dental. Esta revisión bibliográfica tiene el objetivo de esclarecer los aspectos relacionados con la estructura, biosíntesis y función de los glucanos, y enfatizar en la aplicación de estos conocimientos en la prevención de la caries dental(AU)

Dental caries is one of the most common diseases in the human being. Certain cariogenic bacteria play an important role in its multifactorial etiology, since in their interaction with the dental surface they promote its demineralization. The production of extracellular bacterial polyssacharides is among the mechanisms mediating bacterial adhesion. The glucans synthesized by glycosyltransferases not only allow the adherence, but they also are a nutritional source for bacteria and that's why the activity of such enzymes is considered a factor of bacterial virulence in dental caries. This bibliographic review is aimed at making clear the aspects related to the structure, biosynthesis and function of glucans and at giving emphasis to the application of this knowledge in the prevention of dental caries(AU)

Expression of a lysozyme from antheraea pernyi in pastoris and the effects of recombinant Ap-lysozyme on Streptococcus mutans / 实用口腔医学杂志

Objective: To construct the antheraea pernyi lysozyme(ApLZ) expression system using the methylotrophic yeast Pichia pastoris, to assay the antibacterial activity of the recombinant ApLZ against Streptococcus mutans. Methods:The ApLZ expression system was used in the expression and purity of Ap-lysozyme. The antibacterial activity of the recombinant ApLZ against Streptococcus mutans were assayed by using agar diffusion method. Results:Expression system of ApLZ was constructed using the methylotrophic yeast Pichia pastoris as a host. ApLZ was expressed correctly and secreted efficiently when the native signal sequence of ApLZ was used as secretion signal. The level of ApLZ expression in Pichia pastoris peaks at 96 h after the induction of sustaining 5 ml/L methanol. The molecular weight of the recombinant ApLZ is about 20 000. Conclusion:The recombinant ApLZ is active in inhibiting the growth of Streptococcus mutans.

Cloning of mutA and expression of its coding protein / 实用口腔医学杂志

Objective:To obtain mutA gene of Strepto c occus mutans (Ms),and to express it in E.coli DH5?.Methods: mutA gene was amplified by PCR with specific primers from genome of Ms CH43 strain. After sequencing, the gene segment was inserted into vector pProEX and expressed in E.coli DH5?.The protein expression was induced by ITPG an d the protein products were examined by 180 ml/L SDSPAGE electrophorosis. Results:The length of PCR product was 147 bp and was identical to mu tA gene reported by GenBank.The mutA gene product was expressed in E.col i DH5? with Mr of 5.7?10 3.The maximum mutA protein product amount (20% of the total bacterial protein) was obtained when the A 600 value of DH5? was 1.666,IPTG concerntration 1.0 mmol/L and induction time 6 h.Conclus ion:mutA of Ms CH43 can be cloned and expressed in E.coli DH 5?.