scholarly journals Ursolic Acid Targets Glucosyltransferase and Inhibits Its Activity to Prevent Streptococcus mutans Biofilm Formation

2021 ◽  
Vol 12 ◽  
Author(s):  
Yucui Liu ◽  
Yanxin Huang ◽  
Cong Fan ◽  
Zhongmei Chi ◽  
Miao Bai ◽  
...  
Keyword(s):  
Streptococcus Mutans ◽  
Ursolic Acid ◽  
Biofilm Formation ◽  
Natural Compound ◽  
Structural Integrity ◽  
Site Directed Mutagenesis ◽  
Extracellular Polysaccharides ◽  
Virulence Determinants ◽  
Pentacyclic Triterpene ◽  
Catalytic Center

Streptococcus mutans (S. mutans), the prime pathogen of dental caries, can secrete glucosyltransferases (GTFs) to synthesize extracellular polysaccharides (EPSs), which are the virulence determinants of cariogenic biofilms. Ursolic acid, a type of pentacyclic triterpene natural compound, has shown potential antibiofilm effects on S. mutans. To investigate the mechanisms of ursolic acid-mediated inhibition of S. mutans biofilm formation, we first demonstrated that ursolic acid could decrease the viability and structural integrity of biofilms, as evidenced by XTT, crystal violet, and live/dead staining assays. Then, we further revealed that ursolic acid could compete with the inherent substrate to occupy the catalytic center of GTFs to inhibit EPS formation, and this was confirmed by GTF activity assays, computer simulations, site-directed mutagenesis, and capillary electrophoresis (CE). In conclusion, ursolic acid can decrease bacterial viability and prevent S. mutans biofilm formation by binding and inhibiting the activity of GTFs.

scholarly journals Molecular mechanisms of inhibiting glucosyltransferases for biofilm formation in Streptococcus mutans

2021 ◽  
Vol 13 (1) ◽  
Author(s):  
Qiong Zhang ◽  
Qizhao Ma ◽  
Yan Wang ◽  
Hui Wu ◽  
Jing Zou
Keyword(s):  
Streptococcus Mutans ◽  
Biofilm Formation ◽  
Molecular Mechanisms ◽  
Therapeutic Interventions ◽  
Extracellular Polysaccharides ◽  
Combination Strategies ◽  
Wide Range ◽  
Challenges And Opportunities ◽  
Virulence Property ◽  
Inhibitory Molecules

AbstractGlucosyltransferases (Gtfs) play critical roles in the etiology and pathogenesis of Streptococcus mutans (S. mutans)- mediated dental caries including early childhood caries. Gtfs enhance the biofilm formation and promotes colonization of cariogenic bacteria by generating biofilm extracellular polysaccharides (EPSs), the key virulence property in the cariogenic process. Therefore, Gtfs have become an appealing target for effective therapeutic interventions that inhibit cariogenic biofilms. Importantly, targeting Gtfs selectively impairs the S. mutans virulence without affecting S. mutans existence or the existence of other species in the oral cavity. Over the past decade, numerous Gtfs inhibitory molecules have been identified, mainly including natural and synthetic compounds and their derivatives, antibodies, and metal ions. These therapeutic agents exert their inhibitory role in inhibiting the expression gtf genes and the activities and secretion of Gtfs enzymes with a wide range of sensitivity and effectiveness. Understanding molecular mechanisms of inhibiting Gtfs will contribute to instructing drug combination strategies, which is more effective for inhibiting Gtfs than one drug or class of drugs. This review highlights our current understanding of Gtfs activities and their potential utility, and discusses challenges and opportunities for future exploration of Gtfs as a therapeutic target.

scholarly journals Effects of Norspermidine on Dual‐Species Biofilms Composed of Streptococcus mutans and Streptococcus sanguinis

2019 ◽  
Vol 2019 ◽  
pp. 1-9
Author(s):  
Yan Sun ◽  
Yihuai Pan ◽  
Yu Sun ◽  
Mingyun Li ◽  
Shengbin Huang ◽  
...  
Keyword(s):  
Streptococcus Mutans ◽  
Metabolic Activity ◽  
Biofilm Formation ◽  
Acid Production ◽  
Single Species ◽  
Extracellular Polysaccharides ◽  
Streptococcus Sanguinis ◽  
Crystal Violet Assay ◽  
Sem Imaging

The present study aimed at investigating the influence of norspermidine on the formation of dual-species biofilms composed of Streptococcus mutans (S. mutans) and Streptococcus sanguinis (S. sanguinis). Crystal violet assay was conducted to assess the formation of single-species biofilms of S. mutans and S. sanguinis, and the growth curve was carefully observed to monitor the growth of these two species of bacteria. Fluorescence in situ hybridization (FISH) and MTT array were used to analyze the composition and metabolic activity of the dual-species biofilms, respectively. Extracellular polysaccharides (EPS)/bacteria staining, anthrone method, and scanning electron microscopy (SEM) imaging were conducted to study the synthesis of EPS by dual-species biofilms. Lactic acid assay and pH were measured to detect dual-species biofilm acid production. We found that norspermidine had different effects on S. mutans and S. sanguinis including their growth and biofilm formation. Norspermidine regulated the composition of the dual-species biofilms, decreased the ratio of S. mutans in dual-species biofilms, and reduced the metabolic activity, EPS synthesis, and acid production of dual-species biofilms. Norspermidine regulated dual-species biofilms in an ecological way, suggesting that it may be a potent reagent for controlling dental biofilms and managing dental caries.

scholarly journals Molecule Targeting Glucosyltransferase Inhibits Streptococcus mutans Biofilm Formation and Virulence

2015 ◽  
Vol 60 (1) ◽  
pp. 126-135 ◽  
Author(s):  
Zhi Ren ◽  
Tao Cui ◽  
Jumei Zeng ◽  
Lulu Chen ◽  
Wenling Zhang ◽  
...  
Keyword(s):  
Streptococcus Mutans ◽  
Dental Plaque ◽  
Biofilm Formation ◽  
Extracellular Polysaccharides ◽  
Content Type ◽  
Oral Infections ◽  
Concomitant Reduction ◽  
New Strategies

ABSTRACTDental plaque biofilms are responsible for numerous chronic oral infections and cause a severe health burden. Many of these infections cannot be eliminated, as the bacteria in the biofilms are resistant to the host's immune defenses and antibiotics. There is a critical need to develop new strategies to control biofilm-based infections. Biofilm formation inStreptococcus mutansis promoted by major virulence factors known as glucosyltransferases (Gtfs), which synthesize adhesive extracellular polysaccharides (EPS). The current study was designed to identify novel molecules that target Gtfs, thereby inhibitingS. mutansbiofilm formation and having the potential to prevent dental caries. Structure-based virtual screening of approximately 150,000 commercially available compounds against the crystal structure of the glucosyltransferase domain of the GtfC protein fromS. mutansresulted in the identification of a quinoxaline derivative, 2-(4-methoxyphenyl)-N-(3-{[2-(4-methoxyphenyl)ethyl]imino}-1,4-dihydro-2-quinoxalinylidene)ethanamine, as a potential Gtf inhibitor.In vitroassays showed that the compound was capable of inhibiting EPS synthesis and biofilm formation inS. mutansby selectively antagonizing Gtfs instead of by killing the bacteria directly. Moreover, thein vivoanti-caries efficacy of the compound was evaluated in a rat model. We found that the compound significantly reduced the incidence and severity of smooth and sulcal-surface cariesin vivowith a concomitant reduction in the percentage ofS. mutansin the animals' dental plaque (P< 0.05). Taken together, these results represent the first description of a compound that targets Gtfs and that has the capacity to inhibit biofilm formation and the cariogenicity ofS. mutans.

scholarly journals Mechanistic analysis of a synthetic inhibitor of the Pseudomonas aeruginosa LasI quorum-sensing signal synthase

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
O. Lidor ◽  
A. Al-Quntar ◽  
E. C. Pesci ◽  
D. Steinberg
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Quorum Sensing ◽  
In Silico ◽  
Biofilm Formation ◽  
Homoserine Lactone ◽  
Site Directed Mutagenesis ◽  
Great Promise ◽  
Acyl Homoserine Lactone ◽  
Virulence Determinants

Abstract Pseudomonas aeruginosa is an opportunistic Gram-negative pathogen responsible for many human infections. LasI is an acyl-homoserine lactone synthase that produces a quorum-sensing (QS) signal that positively regulates numerous P. aeruginosa virulence determinants. The inhibition of the LasI protein is therefore an attractive drug target. In this study, a novel in silico to in vitro complementation was applied to screen thiazolidinedione-type compounds for their ability to inhibit biofilm formation at concentrations not affecting bacterial growth. The compound (z)-5-octylidenethiazolidine-2, 4-dione (TZD-C8) was a strong inhibitor of biofilm formation and chosen for further study. Structural exploration of in silico docking predicted that the compound had high affinity for the LasI activity pocket. The TZD-C8 compound was also predicted to create hydrogen bonds with residues Arg30 and Ile107. Site-directed mutagenesis (SDM) of these two sites demonstrated that TZD-C8 inhibition was abolished in the lasI double mutant PAO-R30D, I107S. In addition, in vitro swarming motility and quorum sensing signal production were affected by TZD-C 8, confirming this compound alters the cell to cell signalling circuitry. Overall, this novel inhibitor of P. aeruginosa quorum sensing shows great promise and validates our mechanistic approach to discovering inhibitors of LuxI-type acyl-homoserine lactone synthases.

scholarly journals Raffinose Induces Biofilm Formation by Streptococcus mutans in Low Concentrations of Sucrose by Increasing Production of Extracellular DNA and Fructan

2017 ◽  
Vol 83 (15) ◽  
Author(s):  
Ryo Nagasawa ◽  
Tsutomu Sato ◽  
Hidenobu Senpuku
Keyword(s):  
Streptococcus Mutans ◽  
Biofilm Formation ◽  
Cell Surface Hydrophobicity ◽  
Extracellular Dna ◽  
Regulatory Function ◽  
Extracellular Polysaccharides ◽  
Oral Flora ◽  
Content Type ◽  
Positive Effects ◽  
Low Concentrations

ABSTRACT Streptococcus mutans is the primary etiological agent of dental caries and causes tooth decay by forming a firmly attached biofilm on tooth surfaces. Biofilm formation is induced by the presence of sucrose, which is a substrate for the synthesis of extracellular polysaccharides but not in the presence of oligosaccharides. Nonetheless, in this study, we found that raffinose, which is an oligosaccharide with an intestinal regulatory function and antiallergic effect, induced biofilm formation by S. mutans in a mixed culture with sucrose, which was at concentrations less than those required to induce biofilm formation directly. We analyzed the possible mechanism behind the small requirement for sucrose for biofilm formation in the presence of raffinose. Our results suggested that sucrose contributed to an increase in bacterial cell surface hydrophobicity and biofilm formation. Next, we examined how the effects of raffinose interacted with the effects of sucrose for biofilm formation. We showed that the presence of raffinose induced fructan synthesis by fructosyltransferase and aggregated extracellular DNA (eDNA, which is probably genomic DNA released from dead cells) into the biofilm. eDNA seemed to be important for biofilm formation, because the degradation of DNA by DNase I resulted in a significant reduction in biofilm formation. When assessing the role of fructan in biofilm formation, we found that fructan enhanced eDNA-dependent cell aggregation. Therefore, our results show that raffinose and sucrose have cooperative effects and that this induction of biofilm formation depends on supportive elements that mainly consist of eDNA and fructan. IMPORTANCE The sucrose-dependent mechanism of biofilm formation in Streptococcus mutans has been studied extensively. Nonetheless, the effects of carbohydrates other than sucrose are inadequately understood. Our findings concerning raffinose advance the understanding of the mechanism underlying the joint effects of sucrose and other carbohydrates on biofilm formation. Since raffinose has been reported to have positive effects on enterobacterial flora, research on the effects of raffinose on the oral flora are required prior to its use as a beneficial sugar for human health. Here, we showed that raffinose induced biofilm formation by S. mutans in low concentrations of sucrose. The induction of biofilm formation generally generates negative effects on the oral flora. Therefore, we believe that this finding will aid in the development of more effective oral care techniques to maintain oral flora health.

scholarly journals The vicK gene of Streptococcus mutans mediates its cariogenicity via exopolysaccharides metabolism

2021 ◽  
Vol 13 (1) ◽  
Author(s):  
Yalan Deng ◽  
Yingming Yang ◽  
Bin Zhang ◽  
Hong Chen ◽  
Yangyu Lu ◽  
...  
Keyword(s):  
Dental Caries ◽  
Streptococcus Mutans ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Single Species ◽  
Laser Scanning Microscopy ◽  
Gas Chromatography Mass Spectrometry ◽  
Extracellular Polysaccharides ◽  
Confocal Laser

AbstractStreptococcus mutans (S. mutans) is generally regarded as a major contributor to dental caries because of its ability to synthesize extracellular polysaccharides (EPS) that aid in the formation of plaque biofilm. The VicRKX system of S. mutans plays an important role in biofilm formation. The aim of this study was to investigate the effects of vicK gene on specific characteristics of EPS in S. mutans biofilm. We constructed single-species biofilms formed by different mutants of vicK gene. Production and distribution of EPS were detected through atomic force microscopy, scanning electron microscopy and confocal laser scanning microscopy. Microcosmic structures of EPS were analyzed by gel permeation chromatography and gas chromatography-mass spectrometry. Cariogenicity of the vicK mutant was assessed in a specific pathogen-free rat model. Transcriptional levels of cariogenicity-associated genes were confirmed by quantitative real-time polymerase chain reaction. The results showed that deletion of vicK gene suppressed biofilm formation as well as EPS production, and EPS were synthesized mostly around the cells. Molecular weight and monosaccharide components underwent evident alterations. Biofilms formed in vivo were sparse and contributed a decreased degree of caries. Moreover, expressional levels of genes related to EPS synthesis were down-regulated, except for gtfB. Our report demonstrates that vicK gene enhances biofilm formation and subsequent caries development. And this may due to its regulations on EPS metabolism, like synthesis or microcosmic features of EPS. This study suggests that vicK gene and EPS can be considered as promising targets to modulate dental caries.

scholarly journals Transcriptional Profiling Reveals the Importance of RcrR in the Regulation of Multiple Sugar Transportation and Biofilm Formation in Streptococcus mutans

mSystems ◽  
2021 ◽  
Author(s):  
Tao Gong ◽  
Xiaoya He ◽  
Jiamin Chen ◽  
Boyu Tang ◽  
Ting Zheng ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Chronic Disease ◽  
Oral Cavity ◽  
Streptococcus Mutans ◽  
Biofilm Formation ◽  
Transcriptional Profiling ◽  
Extracellular Polysaccharides ◽  
Tooth Decay ◽  
Content Type ◽  
Cariogenic Bacterium

The human oral cavity is a constantly changing environment. Tooth decay is a commonly prevalent chronic disease mainly caused by the cariogenic bacterium Streptococcus mutans . S. mutans is an oral pathogen that metabolizes various carbohydrates into extracellular polysaccharides (EPSs), biofilm, and tooth-destroying lactic acid.

scholarly journals Analyses of the Effects of Arginine, Nicotine, Serotype and Collagen-Binding Proteins on Biofilm Development by 33 Strains of Streptococcus mutans

2021 ◽  
Vol 2 ◽  
Author(s):  
Dawn R. Wagenknecht ◽  
Richard L. Gregory
Keyword(s):  
Endothelial Cell ◽  
Streptococcus Mutans ◽  
Cell Invasion ◽  
Biofilm Formation ◽  
Extracellular Polysaccharides ◽  
Collagen Binding ◽  
Bacterial Proliferation ◽  
Biofilm Production ◽  
Biofilm Development ◽  
Bacterial Mass

Streptococcus mutans serotype k strains comprise &lt;3% of oral isolates of S. mutans but are prominent in diseased cardiovascular (CV) tissue. Collagen binding protein (CBP) genes, cbm and cnm, are prevalent in serotype k strains and are associated with endothelial cell invasion. Nicotine increases biofilm formation by serotype c strains of S. mutans, but its effects on serotype k strains and strains with CBP are unknown. Saliva contains arginine which alters certain properties of the extracellular polysaccharides (EPS) in S. mutans biofilm. We examined whether nicotine and arginine affect sucrose-induced biofilm of S. mutans serotypes k (n = 23) and c (n = 10) strains with and without CBP genes. Biofilm mass, metabolism, bacterial proliferation, and EPS production were assessed. Nicotine increased biomass and metabolic activity (p &lt; 0.0001); arginine alone had no effect. The presence of a CBP gene (either cbm or cnm) had a significant effect on biofilm production, but serotype did not. Nicotine increased bacterial proliferation and the effect was greater in CBP + strains compared to strains lacking CBP genes. Addition of arginine with nicotine decreased both bacterial mass and EPS compared to biofilm grown in nicotine alone. EPS production was greater in cnm + than cbm + strains (p &lt; 0.0001). Given the findings of S. mutans in diseased CV tissue, a nicotine induced increase in biofilm production by CBP + strains may be a key link between tobacco use and CV diseases.

scholarly journals Acetylation of glucosyltransferases regulates Streptococcus mutans biofilm formation and virulence

2021 ◽  
Vol 17 (12) ◽  
pp. e1010134
Author(s):  
Qizhao Ma ◽  
Yangyang Pan ◽  
Yang Chen ◽  
Shuxing Yu ◽  
Jun Huang ◽  
...  
Keyword(s):  
Streptococcus Mutans ◽  
Biofilm Formation ◽  
Extracellular Polysaccharides ◽  
Lysine Acetylation ◽  
Post Translational Modification ◽  
Caries Model ◽  
Acetyl Group ◽  
Important Virulence Factor ◽  
Protein Acetyltransferase

Lysine acetylation is a frequently occurring post-translational modification (PTM), emerging as an important metabolic regulatory mechanism in prokaryotes. This process is achieved enzymatically by the protein acetyltransferase (KAT) to specifically transfer the acetyl group, or non-enzymatically by direct intermediates (acetyl phosphate or acetyl-CoA). Although lysine acetylation modification of glucosyltransferases (Gtfs), the important virulence factor in Streptococcus mutans, was reported in our previous study, the KAT has not been identified. Here, we believe that the KAT ActG can acetylate Gtfs in the enzymatic mechanism. By overexpressing 15 KATs in S. mutans, the synthesized water-insoluble extracellular polysaccharides (EPS) and biofilm biomass were measured, and KAT (actG) was identified. The in-frame deletion mutant of actG was constructed to validate the function of actG. The results showed that actG could negatively regulate the water-insoluble EPS synthesis and biofilm formation. We used mass spectrometry (MS) to identify GtfB and GtfC as the possible substrates of ActG. This was also demonstrated by in vitro acetylation assays, indicating that ActG could increase the acetylation levels of GtfB and GtfC enzymatically and decrease their activities. We further found that the expression level of actG in part explained the virulence differences in clinically isolated strains. Moreover, overexpression of actG in S. mutans attenuated its cariogenicity in the rat caries model. Taken together, our study demonstrated that the KAT ActG could induce the acetylation of GtfB and GtfC enzymatically in S. mutans, providing insights into the function of lysine acetylation in bacterial virulence and pathogenicity.

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