Replacement of Loops at the Entrance of the Active Pocket of Streptococcus thermophilus 4,6-α-Glucanotransferase Changes Its Catalytic Activity and Product Specificity.

To explore the roles of loops around active pocket in the reuteran type 4,6-α-glucanotransferase (StGtfB) from S. thermophilus, they were individually or simultaneously replaced with those of an isomalto/maltopolysaccharides type 4,6-α-glucanotransferase from L. reuteri. StGtfB with the replaced loops A1, A2 (A1A2) and A1, A2, B (A1A2B), respectively, showed 1.41- and 0.83-fold activities of StGtfB. Two mutants reduced crystallinity and increased starch disorder at 2, 4, and 8 U/g more than StGtfB and increased DP ≤ 5 short branches of starch by 38.01% at 2 U/g, much more than StGtfB by 4.24%. A1A2B modified starches had the lowest retrogradation over 14 days. A1A2 modified starches had the highest percentage of slowly digestible fractions, ranging from 40.32% to 43.34%. StGtfB and its mutants bind substrates by hydrogen bonding and van der Waals forces at their nonidentical amino acid residues, suggesting that loop replacement leads to a different conformation and changes activity and product structure.

Fermentation enrichment, structural characterization and immunostimulatory effects of ß-glucan from Quinoa.

We developed an efficient mixed-strain co-fermentation method to increase the yield of quinoa ß-glucan (Q+). Using a 1:1 mass ratio of highly active dry yeast and Streptococcus thermophilus, solid-to-liquid ratio of 1:12 (g/mL), inoculum size of 3.8 % (mass fraction), fermentation at 32 °C for 27 h, we achieved the highest ß-glucan yield of (11.13 ± 0.80)%, representing remarkable 100.18 % increase in yield compared to quinoa ß-glucan(Q-) extracted using hot water. The structure of Q+ and Q- were confirmed through Fourier Transform Infrared (FTIR) and Nuclear Magnetic Resonance (NMR) spectroscopies. Q+ contained 41.66 % ß-glucan, 3.93 % protein, 2.12 % uronic acid; Q- contained 37.21 % ß-glucan, 11.49 % protein, and 1.73 % uronic acid. The average molecular weight of Q+(75.37 kDa) was lower than that of Q- (94.47 kDa). Both Q+ and Q- promote RAW264.7 cell proliferation without displaying toxicity. They stimulate RAW264.7 cells through the NF-κB and MAPK signaling pathways, primarily inducing NO and pro-inflammatory cytokines by upregulating CD40 expression. Notably, Q+ exhibited stronger immunostimulatory activity compared to Q-. In summary, the fermentation enrichment method yields higher content of quinoa ß-glucan with increased purity and stronger immunostimulatory properties. Further study of its bioimmunological activity and structure-activity relationship may contribute to the development of new immunostimulants.

Activation of Csm6 ribonuclease by cyclic nucleotide binding: in an emergency, twist to open.

Type III CRISPR systems synthesize cyclic oligoadenylate (cOA) second messengers as part of a multi-faceted immune response against invading mobile genetic elements (MGEs). cOA activates non-specific CRISPR ancillary defence nucleases to create a hostile environment for MGE replication. Csm6 ribonucleases bind cOA using a CARF (CRISPR-associated Rossmann Fold) domain, resulting in activation of a fused HEPN (Higher Eukaryotes and Prokaryotes Nucleotide binding) ribonuclease domain. Csm6 enzymes are widely used in a new generation of diagnostic assays for the detection of specific nucleic acid species. However, the activation mechanism is not fully understood. Here we characterised the cyclic hexa-adenylate (cA6) activated Csm6' ribonuclease from the industrially important bacterium Streptococcus thermophilus. Crystal structures of Csm6' in the inactive and cA6 bound active states illuminate the conformational changes which trigger mRNA destruction. Upon binding of cA6, there is a close to 60° rotation between the CARF and HEPN domains, which causes the 'jaws' of the HEPN domain to open and reposition active site residues. Key to this transition is the 6H domain, a right-handed solenoid domain connecting the CARF and HEPN domains, which transmits the conformational changes for activation.

Structural elucidation of the exopolysaccharide from Streptococcus thermophilus XJ53 and the effect of its molecular weight on immune activity.

EPS53, a homogeneous exopolysaccharide (EPS) was isolated from Streptococcus thermophiles XJ53 fermented in skimmed milk via anion exchange column chromatography. The relative molecular weight of EPS53 was above 6.7 × 105 g/mol; its repeating structural unit of EPS53 consisted of ß-T-Galp, ß-1,3-Galf, α-1,3-Glcp and ß-1,3,6-Glcp residues in a molar ratio of 1:1:1:1, with ß-T-Galp attached to the O-6 position of ß-1,3,6-Glcp，identical to the EPS produced from S. thermophilus SFi39. EPS53-D, purified under similar conditions as EPS53 except for the deproteinization of trichloroacetic acid (TCA), had a lower molecular weight but the same repeating structural unit. The effects of EPS53 and EPS53-D on proliferation, phagocytosis and nitric oxide (NO) release of macrophage RAW264.7 were compared. EPS53 exhibited stronger immune activity than EPS53-D, suggesting that the molecular weight might have an important effect on the activity of EPS molecules. Treatment with TCA might affect the activities of native EPSs produced by fermentation.

Structure, physicochemical characterization, and antioxidant activity of the highly arabinose-branched exopolysaccharide EPS-M2 from Streptococcus thermophilus CS6.

Streptococcus thermophilus CS6 could produce the high exopolysaccharide (EPS) level in optimized skimmed milk medium. However, physicochemical properties and structure of these polymers have not been fully characterized. In this study, two purified fractions (EPS-M1 and EPS-M2) exhibited good rheology, thermostability and antioxidant activity. Further monosaccharide composition, molecular weight and NMR analysis indicated EPS-M2 was composed of galactose, arabinose and glucose (5:2.5:1) with an average molecular weight of 2.22 × 104 Da and its suggested repeating unit was →6)-[α-L-Araf-(1 â†’ 3)]-ß-D-Galp-(1 â†’ 4)-ß-D-Galp-(1 â†’ 6)-[α-L-Araf-(1 â†’ 5)-{α-L-Araf-(1 â†’ 3)}-α-L-Araf-(1 â†’ 3)]-ß-D-Galp-(1 â†’ 4)-ß-D-Galp-(1 â†’ 6)-[ß-D-Galp-(1 â†’ 5)-α-L-Araf-(1 â†’ 5)-α-L-Araf-(1 â†’ 3)]-ß-D-Galp-(1 â†’ 6)-[ß-D-Galp-(1 â†’ 5)-α-L-Araf-(1 â†’ 5)-{α-L-Araf-(1 â†’ 3)}-α-L-Araf-(1 â†’ 3)]-ß-D-Galp-(1→. High EPS production relied on the expression of eps gene cluster and key enzymes of nucleotide sugar metabolism. Overall, EPS-M2 from a potential functional starter S. thermophilus CS6 provided opportunities for natural thickener, stabilizer, and antioxidant agent exploration in the food industry.

Coevolution of the bacterial pheromone ComS and sensor ComR fine-tunes natural transformation in streptococci.

Competence for natural transformation extensively contributes to genome evolution and the rapid adaptability of bacteria dwelling in challenging environments. In most streptococci, this process is tightly controlled by the ComRS signaling system, which is activated through the direct interaction between the (R)RNPP-type ComR sensor and XIP pheromone (mature ComS). The overall mechanism of activation and the basis of pheromone selectivity have been previously reported in Gram-positive salivarius streptococci; however, detailed 3D-remodeling of ComR leading up to its activation remains only partially understood. Here, we identified using a semirational mutagenesis approach two residues in the pheromone XIP that bolster ComR sensor activation by interacting with two aromatic residues of its XIP-binding pocket. Random and targeted mutagenesis of ComR revealed that the interplay between these four residues remodels a network of aromatic-aromatic interactions involved in relaxing the sequestration of the DNA-binding domain. Based on these data, we propose a comprehensive model for ComR activation based on two major conformational changes of the XIP-binding domain. Notably, the stimulation of this newly identified trigger point by a single XIP substitution resulted in higher competence and enhanced transformability, suggesting that pheromone-sensor coevolution counter-selects for hyperactive systems in order to maintain a trade-off between competence and bacterial fitness. Overall, this study sheds new light on the ComRS activation mechanism and how it could be exploited for biotechnological and biomedical purposes.

Structural characterisation of EPS of Streptococcus thermophilus S-3 and its application in milk fermentation.

The application of Streptococcus thermophilus S-3 into yogurt production was studied and the structural properties of the generated exopolysaccharides (EPS-S3) were characterized. The proposed structure of EPS-S3 was obtained. EPS-S3 contained a high ratio of N-Acetyl-galactosamine with the Mw of 574 kDa, which was higher than that of AR333 (314 kD) leading to higher apparent viscosity. Streptococcus thermophilus strain S-3 was co-cultured with Lactobacillus delbrueckii for yogut production which highly increased the acidifying rate and post-acidification rate. The quality of the co-cultured yogurts in terms of apparent viscosity, syneresis capacity, water holding capacity and rheological properties were much better than that by using Lactobacillus bulgaricus only. The production mechanism of EPS-S3 from gene regulated level was also discussed which is helpful to facilitate the application of Streptococcus thermophilus strain into milk production.

Structure-function studies of Rgg binding to pheromones and target promoters reveal a model of transcription factor interplay.

Regulator gene of glucosyltransferase (Rgg) family proteins, such as Rgg2 and Rgg3, have emerged as primary quorum-sensing regulated transcription factors in Streptococcus species, controlling virulence, antimicrobial resistance, and biofilm formation. Rgg2 and Rgg3 function is regulated by their interaction with oligopeptide quorum-sensing signals called short hydrophobic peptides (SHPs). The molecular basis of Rgg-SHP and Rgg-target DNA promoter specificity was unknown. To close this gap, we determined the cryoelectron microscopy (cryo-EM) structure of Streptococcus thermophilus Rgg3 bound to its quorum-sensing signal, SHP3, and the X-ray crystal structure of Rgg3 alone. Comparison of these structures with that of an Rgg in complex with cyclosporin A (CsA), an inhibitor of SHP-induced Rgg activity, reveals the molecular basis of CsA function. Furthermore, to determine how Rgg proteins recognize DNA promoters, we determined X-ray crystal structures of both Streptococcus dysgalactiae Rgg2 and S. thermophilus Rgg3 in complex with their target DNA promoters. The physiological importance of observed Rgg-DNA interactions was dissected using in vivo genetic experiments and in vitro biochemical assays. Based on these structure-function studies, we present a revised unifying model of Rgg regulatory interplay. In contrast to existing models, where Rgg2 proteins are transcriptional activators and Rgg3 proteins are transcriptional repressors, we propose that both are capable of transcriptional activation. However, when Rgg proteins with different activation requirements compete for the same DNA promoters, those with more stringent activation requirements function as repressors by blocking promoter access of SHP-bound conformationally active Rgg proteins. While a similar gene expression regulatory scenario has not been previously described, in all likelihood it is not unique to streptococci.

Discovery and Biosynthesis of Streptosactin, a Sactipeptide with an Alternative Topology Encoded by Commensal Bacteria in the Human Microbiome.

Mammalian microbiomes encode thousands of biosynthetic gene clusters (BGCs) and represent a new frontier in natural product research. We recently found an abundance of quorum sensing-regulated BGCs in mammalian microbiome streptococci that code for ribosomally synthesized and post-translationally modified peptides (RiPPs) and contain one or more radical S-adenosylmethionine (RaS) enzymes, a versatile superfamily known to catalyze some of the most unusual reactions in biology. In the current work, we target a widespread group of streptococcal RiPP BGCs and elucidate both the reaction carried out by its encoded RaS enzyme and identify its peptide natural product, which we name streptosactin. Streptosactin is the first sactipeptide identified from Streptococcus spp.; it contains two sequential four amino acid sactionine macrocycles, an unusual topology for this compound family. Bioactivity assays reveal potent but narrow-spectrum activity against the producing strain and its closest relatives that carry the same BGC, suggesting streptosactin may be a long-suspected fratricidal agent of Streptococcus thermophilus. Our results highlight mammalian streptococci as a rich source of unusual enzymatic chemistries and bioactive natural products.

Structural characterization of exopolysaccharide from Streptococcus thermophilus ASCC 1275.

In this study, we purified and characterized exopolysaccharide (EPS) produced by a high-EPS-producing dairy starter bacterium, Streptococcus thermophilus ASCC 1275. Crude EPS was extracted from S. thermophilus ASCC 1275 and partially purified using dialysis. Further purification and fractionation of exopolysaccharide was conducted using HPLC on a Superose 6 column (Cytiva/Global Life Sciences Solutions, Marlborough, MA). Glycosyl composition analysis, linkage analysis along with 1-dimensional and 2-dimensional nuclear magnetic resonance spectroscopy were performed to deduce the structure of EPS. Three fractions (F) obtained from gel permeation chromatography were termed F1 (2.6%), F2 (45.8%), and F3 (51.6%) with average molecular weights of approximately 511, 40, and 5 kDa, respectively. Monosaccharide composition analysis revealed the dominance of glucose, galactose, and mannose in all 3 fractions. Major linkages observed in F3 were terminal galactopyranosyl (t-Gal), 3-linked glucopyranosyl (3-Glc), 3-linked galactofuranosyl (3-Galf), and 3,6-linked glucopyranosyl (3,6-Glc) and major linkages present in F2 were 4-Glc (48 mol%), followed by terminal mannopyranosyl (t-Man), 2- + 3-linked mannopyranosyl (2-Man+3-Man), and 2,6-linked mannopyranosyl (2,6-Man; total ∼28 mol%). The 1-dimensional and 2-dimensional nuclear magnetic resonance spectroscopy revealed that F2 comprised mannans linked by (1→2) linkages and F3 consisted of linear chains of α-d-glucopyranosyl (α-d-Glcp), ß-d-glucopyranosyl (ß-d-Glcp), and ß-d-galactofuranosyl (ß-d-Galf) connected by (1→3) linkages; branching was through (1→6) linkage in F3. A possible structure of EPS in F2 and F3 was proposed.

Effect of different biopolymer-based structured systems on the survival of probiotic strains during storage and in vitro digestion.

BACKGROUND: This study aimed to evaluate the protective effect of different biopolymer systems on the viability of two probiotics (Lactobacillus rhamnosus and Streptococcus thermophilus) during storage and in vitro digestion. Methylcellulose (MC), sodium alginate (SA), and whey protein (WP)-based structures were designed and characterized in terms of pH, rheological properties, and visual appearance. RESULTS: The results highlighted that the WP-system ensured probiotic protection during both storage and in vitro digestion. This result was attributed to a combined effect of the physical barrier offered by the protein gel network and whey proteins as a nutrient for microbes. On the other hand, surprisingly, the viscous methylcellulose-based system was able to guarantee good microbial viability during storage. However, this was not confirmed during in vitro digestion. The opposite results were obtained for sodium alginate beads. CONCLUSION: The results suggest that the capacity of a polymeric structure to protect probiotic bacteria is a combination of structural organization and system formulation. © 2020 Society of Chemical Industry.

Development of nutritious probiotic sportsman drink.

Whey plays an important role in the sports nutrition because of high quality proteins and essential amino acid profile. Nine formulations of sportsman drinks were made using Cheddar, Mozzarella and Paneer whey with normal as well as additional fermentation. The developed sportsman drinks were evaluated for physico-chemical analyses, amino acid profile, viscosity and total plate count along with sensory response during two month storage. Drink having Cheddar whey (T4) with additional fermentation was better in terms of quality and nutrition. Furthermore, amino acid profile considered it a complete and balanced source of essential and non-essential amino acids. Amongst essential amino acids, highest values was recorded for branched chain amino acids like leucine (73.16±3.09) followed by lysine (61.56±0.61) and valine (44.13±1.86)mg/g protein. The dietary significance of sportsman drink can be enhanced through additional fermentation using Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophillus.

Production and molecular structure of heteropolysaccharides from two lactic acid bacteria.

In the dairy industry, exopolysaccharides (EPS) produced in situ from lactic acid bacteria are of great interest because of their contribution to product texture. Some EPS cause ropiness which might be linked to specific physical and chemical EPS properties. EPS show a broad variety of chemical structures and, because analysis is rather complex, it is still a major challenge to establish structure-function relationships. The aim of this study was to produce EPS with different degree of ropiness, perform in-depth structural elucidations and relate this information to their behaviour in aqueous solutions. After cultivation of Streptococcus thermophilus DGCC7919 and Lactococcus lactis LL-2A and subsequent EPS isolation, both EPS showed similar macromolecular properties, but pronounced differences in monosaccharide composition and glycosidic linkages. Our data suggests that mainly the side chains in the EPS from LL-2A might be responsible for a higher ropiness than that observed for EPS from DGCC7919.

Formation and Stoichiometry of CRISPR-Cascade Complexes with Varying Spacer Lengths Revealed by Native Mass Spectrometry.

The adaptive immune system of bacteria and archaea against viral DNA is based on clustered, regularly interspaced, short palindromic repeats (CRISPRs) which are encoded in the host genome and translated into CRISPR RNAs (crRNAs) containing single spacer sequences complementary to foreign DNA. crRNAs assemble with CRISPR-associated (Cas) proteins forming surveillance complexes that base-pair with viral DNA and mediate its degradation. As specificity of degradation is provided by the crRNA spacer sequence, genetic engineering of the CRISPR system has emerged as a popular molecular tool, for instance, in gene silencing and programmed DNA degradation. Elongating or shortening the crRNA spacer sequence are therefore promising ventures to modify specificity toward the target DNA. However, even though the stoichiometry of wild-type complexes is well established, it is unknown how variations in crRNA spacer length affect their stoichiometry. The CRISPR-associated antiviral defense surveillance complexes of Streptococcus thermophilus (StCascade complexes) contain crRNA and five protein subunits. Using native mass spectrometry, we studied the formation and stoichiometry of StCascade complexes assembled on a set of crRNAs with different spacer lengths. We assigned all relevant complexes and gained insights into the stoichiometry of the complexes as well as their preferred assembly. We found that stable complexes, which incorporate or lose a (Cas7)2(Cse2)1-module, assemble on crRNA varied in length by 12-nucleotide units, while varying crRNA length in six-nucleotide units results in heterogeneous mixtures of complexes. Combining our results from the various variants, we generated an assembly pathway revealing general features of I-E type Cascade complex formation.

A Combined Proteomics, Metabolomics and In Vivo Analysis Approach for the Characterization of Probiotics in Large-Scale Production.

The manufacturing processes of commercial probiotic strains may be affected in different ways in the attempt to optimize yield, costs, functionality, or stability, influencing gene expression, protein patterns, or metabolic output. Aim of this work is to compare different samples of a high concentration (450 billion bacteria) multispecies (8 strains) formulation produced at two different manufacturing sites, United States of America (US) and Italy (IT), by applying a combination of functional proteomics, metabolomics, and in vivo analyses. Several protein-profile differences were detected between IT- and US-made products, with Lactobacillus paracasei, Streptococcus thermophilus, and Bifidobacteria being the main affected probiotics/microorganisms. Performing proton nuclear magnetic spectroscopy (1H-NMR), some discrepancies in amino acid, lactate, betaine and sucrose concentrations were also reported between the two products. Finally, we investigated the health-promoting and antiaging effects of both products in the model organism Caenorhabditis elegans. The integration of omics platforms with in vivo analysis has emerged as a powerful tool to assess manufacturing procedures.

Exopolysaccharide from Streptococcus thermophilus as stabilizer in fermented dairy: Binding kinetics and interactions with casein of milk.

Exopolysaccharides (EPSs) from lactic acid bacteria have great effect on the quality of fermented milk products. However, the mechanism for the quality improvement has not been well described. This study aimed to investigate the molecular binding kinetics and interactions between EPS obtained from Streptococcus thermophilus AR333 (EPS333) and casein of milk (CM) in a simulated acidifying process. The results indicated that EPS333 had a significant effect on the stability of casein micelles at acidic pH (6.0-4.5) according to the turbidity, ζ-potential, particle size and distribution analysis. The adsorption-desorption study by bio-layer interferometry identified the direct affinity binding between EPS333 and CM, the interactive moiety of casein was α-casein, rather than ß- or κ-casein. Fluorescence quenching analysis revealed that the force types of interaction between EPS333 and CM were dynamically changeable during the acidifying process, mainly from electrostatic interaction at pH 7.0-6.5, to hydrophobic or hydrogen bonding at pH 6.5-5.5, and then transferred to electrostatic interaction again at pH 5.5-5.0. Conclusively, EPS333 could bind with CM directly via different binding forces during acidifying process to stabilize the properties of casein micelles.

Profiles of Streptococcus thermophilus MN-ZLW-002 nutrient requirements in controlled pH batch fermentations.

This study aimed to evaluate the profiles of Streptococcus thermophilus nutrient requirements to guide the design of media for high cell density culturing. The growth kinetics, physiological state, and nutrient requirement profiles of S. thermophilus were analyzed in chemically defined media. The results showed that the intracellular ATP concentration, H+ -ATPase activity, NADH/NAD+ , and NH3 concentrations varied with intracellular pH. The nutrient components with the highest amounts required were Leu and Asp; ascorbic acid and p-amino benzoic acid; K+ and PO43- ; and guanine and uracil. The nutrient components with the largest required ratios were Arg, His, and Met; folic acid, cyanocobalamine, biotin, and nicotinic acid; Ca2+ and Mg2+ ; and guanine and uracil. In this study, different nutrient components were primarily used at different phase. Trp, Tyr, calcium pantothenate, thiamine, guanine, and Mg2+ were mainly used from late-lag to midexponential phase. Met, Pro, Phe, Ala, Gly, nicotinic acid, and riboflavin were mainly used from midexponential to late-exponential phase. The highest bioavailabilities of nutrient components were also found at diverse phase. Met, Leu, Ile, Asn, Glu, Lys, Pro, Gly, riboflavin, nicotinic acid, adenine, uracil, inosine, and Ca2+ had the highest bioavailability from late-lag to midexponential phase. Lactose, Glu, Asp, His, Trp, Cys, Val, Arg, Phe, Ala, Ser, Thr, Tyr, folate and cobalamin, calcium pantothenate, ascorbic acid, thiamine, biotin, p-amino benzoic acid, vitamin B6 , K+ , Mg2+ , guanine, xanthine, and PO43- had the highest bioavailability from midexponential to late-exponential phase. This study elucidated the nutrient requirement profiles with culture time and biomass at various average growth rates during the growth of S. thermophilus. The present results will help to formulate complex media for high cell density cultivation and provide the theoretical basis for S. thermophilus feeding strategies.

Prophylactic use of probiotic chocolate modulates intestinal physiological functions in constipated rats.

BACKGROUND: This study investigated the in vivo prophylactic effect of probiotic chocolate on constipation. Rats were administered chocolate containing 2.5 × 1010 CFU g-1 of probiotics daily for 4 weeks and treated with loperamide (5 mg kg-1 ) daily at the fourth week of treatment. RESULTS: Probiotic chocolate treatment significantly (P < 0.05) increased the intestinal motility, colon length, fecal moisture content and number of excreted fecal pellets in constipated rats. Moreover, quantitative real-time polymerase chain reaction data and histological images also revealed that both probiotic chocolate LYC and BB12 treatments were capable of upregulating the mRNA expression levels of colonic ZO-1, occludin and AQP8, leading to the maintenance of the defensive barrier function in the constipated rats compared with the negative controls. Interestingly, these treatments also modulated gut bacterial populations by increasing the abundance levels of Lactobacillus and Bifidobacterium, as well as reducing the abundance level of Enterobacteriaceae. CONCLUSION: The present study demonstrated that probiotic chocolate LYC and BB12 could potentially be used as alternative agents for prophylactic constipation. © 2018 Society of Chemical Industry.

Intracellular GSH of Streptococcus thermophilus shows anti-oxidative activity against low-density lipoprotein oxidation in vitro and in a hyperlipidaemic hamster model.

Streptococcus thermophilus YIT 2001 (ST-1), a lactic acid bacterial strain, was shown to have inhibitory effects on the oxidation of low-density lipoprotein (LDL) and the development of aortic fatty lesions in an animal model, and lower the serum levels of malondialdehyde-modified LDL, an oxidative modification product of LDL, in a clinical trial. This study aimed to identify the intracellular active component of ST-1 associated with anti-oxidative activity against LDL oxidation. High-performance liquid chromatography-electrospray ionisation mass spectrometry analysis after fractionation of the cellular extract by reversed-phase chromatography demonstrated that the active fraction contained reduced glutathione (GSH). GSH showed anti-oxidative activity in a dose-dependent manner, while this activity disappeared following thiol derivatisation. ST-1 had the strongest anti-oxidative activity against LDL oxidation and the highest level of intracellular GSH among five strains of S. thermophilus. In addition, the anti-oxidative activity of ST-1 after thiol derivatisation decreased by about half, which was similar to that of three other strains containing poor or no intracellular GSH or thiol components. Moreover, anti-oxidative activity against LDL oxidation was observed in hyperlipidaemic hamsters fed with high GSH ST-1 cells but not in those given low GSH cells. These findings suggest that intracellular GSH in ST-1 may provide beneficial effects via anti-oxidative activity against LDL oxidation and excess oxidative stress in the blood.

Characterization of Exopolysaccharide Produced by Streptococcus thermophilus CC30.

An exopolysaccharide (EPS) producing strain CC30 was isolated from raw milk and identified as Streptococcus thermophilus with morphological and 16S sequencing analysis. The strain was shown to produce 1.95 g/L of EPS when grown in skim milk lactose medium at 30°C by increasing the viscosity of the medium. The EPS was isolated and purified, and it was shown to consist of glucose and galactose in 1 : 1 ratio, with molecular weights ranging from 58 to 180 kDa. FTIR spectroscopy indicated the EPS to have amide, hydroxyl, and carboxyl groups. Under Atomic Force Microscopy, EPS showed spike-like lumps of EPS. Scanning Electron Microscopy (SEM) studies showed that it had irregular lumps with a coarse surface. The EPS displayed pseudoplastic nature. Thermogravimetric analysis (TGA) reported a degradation temperature of 110.84°C. The purified EPS exhibited reducing activity, hydrogen peroxide radical scavenging activity, and emulsification activity. The results of the present study indicated that EPS producing Streptococcus thermophilus could serve as a promising candidate for further exploitation in food industry.