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The popularity of light/energy devices for cosmetic purposes (e.g., skin care) is increasing. However, the effects and underlying mechanisms remain poorly understood. Commencing in the 1960s, various studies have evaluated the beneficial effects of a light source on cells and tissues. The techniques evaluated include low-level light (laser) therapy and photobiomodulation (PBM). Most studies on PBM used red light sources, but, recently, many studies have employed near-infrared light sources including those of wavelength 800 nm. Here, we used a light-emitting diode (LED) array with a wavelength of 863 nm to treat DMBA/TPA-induced mouse skin tumors; treatment with the array delayed tumor development and reduced the levels of systemic inflammatory cytokines. These results suggest that light therapy could be beneficial. However, the effects were small. Further studies on different skin tumors using an optimized LED setup are required. Combination therapies (conventional methods and an LED array) may be useful.

Soybean pods, separated and enclosed from the outside environment, are considered a suitable place to find new microbes. A Gram-stain-negative, aerobic bacterium, bacterial strain (YB22T) was isolated from the pod of Glycine max (soybean) collected from a rural area in Republic of Korea and characterized by using polyphasic taxonomy. Cells of the strain were rod-shaped (approximately 0.4–0.6 µm wide and 4.0–5.0 µm long), non-flagellated and formed silver-yellow colonies. Cells grew at 25–35 °C (optimum, 28–30 °C), at pH 5.0–9.0 (optimum, pH 7.0) and with 0–2.0% NaCl (optimum, 0 % NaCl). 16S rRNA gene sequencing showed that strain YB22T was phylogenetically closest to the genus Elizabethkingia , and showed highest similarities to Elizabethkingia occulta G4070T (96.7 %), Elizabethkingia meningoseptica ATCC 13253T (96.7 %), Elizabethkingia miricola DSM 14571T (96.6 %), Elizabethkingia bruuniana G0146T (96.5 %), Elizabethkingia ursingii G4122T (96.4 %) and Elizabethkingia anophelis R26T (96.2 %). Average amino acid identity values between strain YB22T and other taxa in the genus Elizabethkingia were all above the threshold range of genus determination. Average nucleotide identity and digital DNA–DNA hybridization values between strain YB22T and other phylogenetic relatives were all found to be below the threshold range for species determination. The respiratory quinone of strain YB22T was menaquinone 6 (MK-6) and the predominant cellular fatty acids were iso-C15 : 0 (47.8 %) and iso-C17 : 0 3-OH (18.5 %). The major polar lipids were phosphatidylethanolamine, four unidentified aminolipids and three unidentified polar lipids. The phylogenetic analysis and physiological and biochemical data showed that strain YB22T should represent a novel species in the genus Elizabethkingia , for which the name Elizabethkingia argenteiflava sp. nov. is proposed. The type strain for this novel species is YB22T (=KCCM 43263T=JCM 32097T).

Photobiomodulation (PBM) is attracting increased attention in the fields of dermatology and cosmetics. PBM with a variety of light parameters has been used widely in skin care, but can cause certain types of unwanted cells to proliferate in the skin; this can lead to skin tumors, such as papillomas and cancers. We constructed a mouse model of human skin tumors using DMBA as an initiator and TPA as a promoter, and confirmed that LEDs with a wavelength of 642 nm (red light) increased tumor size, epidermal thickness, and systemic proinflammatory cytokine levels. These results indicated that skin tumor cell proliferation may result from the use of 642 nm LEDs, suggesting the need for regulation of skin care based on LED light therapy.

Purpose: To analyze the effects of an aqueous extract of Sericea lespedeza (SL) on rat male menopause.Methods: Levels of nitric oxide (NO), endothelial nitric oxide synthase (NOS), cGMP, and prostaglandin E2 (PGE2) in the penile corpus cavernosum of the rats were evaluated using appropriate kits. Serum levels of dihydrotestosterone (DHT), testosterone, sex hormone-binding globulin (SHBG), and 17-beta hydroxysteroid dehydrogenases (17β-HSD) were measured with enzyme-linked immunosorbent assay kits. Total and motile sperms were counted on a hemocytometer. Histological changes in rat testis and epididymis were analyzed with hematoxylin and eosin staining.Results: The levels of NO, NOS, and cGMP (but not PGE2) increased in a dose-dependent manner (p< 0.05) upon administration of an aqueous extract of SL (AESL), while levels of DHT, 17β-HSD, and testosterone increased in the group administered with 300 mg/kg of AESL. Epididymal sperm count increased by 24 % in such rats compared to controls (p < 0.05).Conclusion: The aqueous extract of SL improves sperm count and muscle mass in rats by increasing the levels of NO, NOS, cGMP and testosterone. Thus, SL extract can potentially be developed as an alternative therapeutic agent for clinical management of TDS. Keywords: NO-cGMP, Testosterone, Hormones, Sperm count, Muscle mass, Sericea lespedeza, Lespedeza cuneata

Background/aim Recurrence and metastasis of cancer caused by cancer stem cells (CSCs) is a challenge to overcome. Low level laser therapy is a new treatment strategy to suppress their invasiveness. We have assessed the inhibitory effects of 470 nm blue LED on the invasiveness of them to determine the molecular mechanisms of anti-invasiveness. Materials and methods The effects of blue LEDs on their viability, proliferation and invasion were analyzed using MTT and transwell methods. In addition, the anti-invasiveness effect of blue LED on them was evaluated by zymography, semi-quantitative RT-PCR and western blot analysis. Results Irradiation with blue LED at 3 J/cm2 resulted in inhibition of their viability, proliferation and invasiveness. Their matrix metalloproteinase 2 (MMP-2) and MMP-9 activities were reduced by blue LED irradiation. Semi-quantitative RT-PCR also showed similar results. In addition, western blotting analyses showed that cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2) synthesis were significantly inhibited by LED irradiation in CD133+ colorectal CSCs. Conclusion Down-regulation of the COX-2/PGE2 signaling pathway by blue LED irradiation led to reduce expression of MMP-2 and MMP-9, inhibiting the invasiveness of CD133+ colorectal CSC.

We evaluated changes in cell viability and morphology in response to low-level light irradiation and underlying variations in the levels of heat shock proteins (HSPs). Human fibroblasts were irradiated with a light-emitting diode (LED) array at 660 nm (50 mW for 15, 30, and 60 minutes). Cell viability and morphological changes were evaluated via epifluorescence analysis; we also assessed cell viability and length changes. The expression levels of adenosine triphosphate (ATP) and various HSPs (HSP27, 60, 70, and 90) were analyzed by immunohistochemical staining, Western blotting and microarray analysis. After LED irradiation, cellular viability and morphology changed. Of the several HSPs analyzed, the HSP90 level increased significantly, suggesting that this protein played roles in the morphological and cellular changes. Thus, low-level irradiation triggered cellular changes mediated by increased HSP90 expression; this may explain why skin irradiation enhances wound-healing.

PikD is a widely known pathway-specific regulator for controlling pikromycin production in Streptomyces venezuelae ATCC 15439, which is a representative of the large ATP-binding regulator of the LuxR family (LAL) in Streptomyces sp. RapH and FkbN also belong to the LAL family of transcriptional regulators, which show greatest homology with the ATP-binding motif and helix-turn-helix DNA-binding motif of PikD. Overexpression of pikD and heterologous expression of rapH and fkbN led to enhanced production of pikromycin by approximately 1.8-, 1.6-, and 1.6-fold in S. venezuelae, respectively. Cross-complementation of rapH and fkbN in the pikD deletion mutant (ΔpikD) restored pikromycin and derived macrolactone production. Overall, these results show that heterologous expression of rapH and fkbN leads to the overproduction of pikromycin and its congeners from the pikromycin biosynthetic pathway in S. venezuelae, and they have the same functionality as the pathwayspecific transcriptional activator for the pikromycin biosynthetic pathway in the ΔpikD strain. These results also show extensive "cross-communication" between pathway-specific regulators of streptomycetes and suggest revision of the current paradigm for pathwayspecific versus global regulation of secondary metabolism in Streptomyces species.

We identified another tcsABCD genes encoding biosynthesis of the allylmalonyl-CoA in new FK506 producing strain, Streptomyces sp. KCCM 11116P. The functionality of tcsABCD is confirmed by gene deletion. To validate the functionality of these genes, tcsABCD genes were heterologously expressed in Streptomyces hygroscopicus ATCC 14891 using pSE34. The resulting strain produced the FK506 which showed a molecular ion [M+NH4]+ at m/z 821 as verified by LC/ESI–MS. To address the function of TcsB in the allylmalonyl-CoA pathway, we pursued the construction of two different combinations: the heterologous FabH-like β-ketoacyl-acyl carrier protein synthase III gene expression and chemical complementation. Plasmid-based expression of the fabH homolog dpsC in ΔTcsB using pSE34 and pSET152(ermE*) led to restoration of approximately 32.2 and 28.7 % of the levels of FK506 relative to Streptomyces sp. KCCM 11116P, respectively. Furthermore, the addition of propionic acid to the culture of ΔTcsB restored FK506 production up to 35.6 % of overall FK506 levels in Streptomyces sp. KCCM 11116P. We have engineered production of 36-methyl-FK506 in ΔTcsB. The heterologous expression of Syteptomyces glaucescens FabH in ΔTcsB gave rise to 5.9-fold increase in the 36-methyl-FK506 level as compared with mutasynthetic production by 4-methylpentanoic acid. Supplementing the culture broth of ΔTcsB expressing S. glaucescens FabH with valine resulted in an additional 2.5-fold increase in the 36-methyl-FK506 titer. Overall, this study shows that TcsB was found to maintain a very high-specific activity for propionyl-CoA in allylmalonyl-CoA biosynthetic work and S. glaucescens FabH which use isobutyryl-CoA as substrate can initiate methyl side chain biosynthesis in 36-methyl-FK506 in ΔTcsB.

A sco3956-deletion mutant (ΔSCO3956) of Streptomyces coelicolor was generated to characterize the S-adenosylmethionine (SAM)-induced, ATP-binding cassette transporter (ABC transporter) ATP-binding protein, SCO3956. It produced actinorhodin (ACT) and undecylprodigiosin (RED) decreased by approx. 82 and 64 %, respectively. In addition, the effect of exogenous SAM was lost in the ΔSCO3956. Plasmid-based complementation of sco3956 in ΔSCO3956 restored ACT and RED levels of ΔSCO3956 to wild-type levels (ACT: 20 ± 1.4 mg g(-1) DCW and RED: 5.3 ± 0.6 mg g(-1) DCW) and the exogenous effect significantly increased ACT and RED by approx. 129 and 135 %, respectively, when compared to the exogenous SAM non-treated sco3956 complementation strain. Thus, the ABC transporter ATP-binding protein, SCO3956, plays a critical role in ACT and RED production serving as a transducer of SAM signaling.

RedP is proposed to initiate undecylprodiginine biosynthesis in Streptomyces coelicolor by condensing an acyl-CoA with malonyl-ACP and is homologous to FabH that catalyzes the same reaction for initiation of fatty acid biosynthesis. Herein, we report the substrate specificities of RedP and FabH from assays using pairings of two acyl-CoA substrates (acetyl-CoA and isobutyryl-CoA) and two malonyl-ACP substrates (malonyl-RedQ and malonyl-FabC). RedP activity was observed only with a pairing of acetyl-CoA and malonyl-RedQ, consistent with its proposed role in initiating the formation of acetyl-CoA-derived prodiginines. Malonyl-FabC is not a substrate for RedP, indicating that ACP specificity is one of the factors that permit a separation between prodiginine and fatty acid biosynthetic processes. FabH demonstrated greater catalytic efficiency for isobutyryl-CoA in comparison with acetyl-CoA using malonyl-FabC, consistent with the observation that in streptomycetes, a broad mixture of fatty acids is synthesized, with those derived from branched-chain acyl-CoA starter units predominating. Diminished FabH activity was also observed using malonyl-RedQ with the same preference for isobutyryl-CoA, completing biochemical and genetic evidence that in the absence of RedP this enzyme can produce branched-chain alkyl prodiginines.

The allyl moiety of the immunosuppressive agent FK506 is structurally unique among polyketides and critical for its potent biological activity. Here, we detail the biosynthetic pathway to allylmalonyl-coenzyme A (CoA), from which the FK506 allyl group is derived, based on a comprehensive chemical, biochemical, and genetic interrogation of three FK506 gene clusters. A discrete polyketide synthase (PKS) with noncanonical domain architecture presumably in coordination with the fatty acid synthase pathway of the host catalyzes a multistep enzymatic reaction to allylmalonyl-CoA via trans-2-pentenyl-acyl carrier protein. Characterization of this discrete pathway facilitated the engineered biosynthesis of novel allyl group-modified FK506 analogues, 36-fluoro-FK520 and 36-methyl-FK506, the latter of which exhibits improved neurite outgrowth activity. This unique feature of FK506 biosynthesis, in which a dedicated PKS provides an atypical extender unit for the main modular PKS, illuminates a new strategy for the c...

FK506 is a 23-membered polyketide macrolide with immunosuppressant activity produced by Streptomyces species. The production of FK506 in S. clavuligerus CKD1119 (KCTC 10561BP) was improved by enhancing the supply of biosynthetic precursors. This improvement was approximately 2.5-fold (15 mg/l) with the supplementation of 10 mM methyl oleate, which is the probable source of acyl-CoAs, to R2YE medium. When the level of FK506 production reached its maximum, the intracellular concentration of methylmalonyl-CoA in S. clavuligerus CKD1119 supplemented with methyl oleate was 12.5-fold higher than that of the unsupplemented strain, suggesting that an increased methylmalonyl-CoA level caused the high-level production of FK506. The following three pathways for the production of (2S)-methylmalonyl-CoA were evaluated to identify the effective precursor supply pathway that can support the high production of FK506 in S. clavuligerus CKD1119: propionyl-CoA carboxylase, methylmalonyl-CoA mutase (MCM), and malonyl/methylmalonyl-CoA ligase. Of the three pathways examined, the MCM pathway supported the highest levels of FK506 production. The expression of MCM in S. clavuligerus CKD1119 led to a threefold and 1.5-fold increase in the methylmalonyl-CoA pool and FK506 production, respectively. Supplementing the culture broth of S. clavuligerus CKD1119 expressing MCM with methyl oleate resulted in an additional twofold increase in the FK506 titer (17.8 mg/l). Overall, these results show that the methylmalonyl-CoA supply is a limiting factor for FK506 biosynthesis and that among the three pathways analyzed, the MCM pathway is the most effective precursor supply pathway supporting the highest titer of FK506 in S. clavuligerus CKD1119.

The role of the putative extracytoplasmic function sigma (σ) factor FujE, which has not been characterized as a member of the FK506 biosynthetic gene cluster, on FK506 production was identified by gene deletion, overexpression, and transcription analysis experiments in Streptomyces sp. strain KCCM 11116P. Inactivation of fujE had no effect on FK506 production, growth, or morphological differentiation. Overexpression of fujE with integrative vectors increased FK506 production by 2.87-fold (24.5 ± 1.4 mg·L–1) compared with the wild type (8.5 ± 0.5 mg·L–1). Semiquantitative reverse transcription – polymerase chain reaction analysis indicated that the overexpression of fujE stimulates the transcription of the FK506 biosynthetic genes. These results demonstrated that fujE is a new member of the FK506 biosynthetic gene cluster.

A FK506 analogue containing a non-natural starter unit was obtained through mutasynthesis by feeding cultures of Streptomyces sp. KCTC 11604BP fkbO deletion mutant with 3-cyclohexene-1-carboxylic acid. The structure of the new compound, 32-dehydroxy-FK506, and its biological activities were determined.

FK506 production by a mutant strain (Streptomyces sp. RM7011) induced by N-methyl-N′-nitro-N-nitrosoguanidine and ultraviolet mutagenesis was improved by 11.63-fold (94.24 mg/l) compared to that of the wild-type strain. Among three different metabolic pathways involved in the biosynthesis of methylmalonyl-CoA, only expression of propionyl-CoA carboxylase (PCC) pathway led to a 1.75-fold and 2.5-fold increase in FK506 production and the methylmalonyl-CoA pool, respectively, compared to those of the RM7011 strain. Lipase activity of the high FK506 producer mutant increased in direct proportion to the increase in FK506 yield, from low detection level up to 43.1 U/ml (12.6-fold). The level of specific FK506 production and lipase activity was improved by enhancing the supply of lipase inducers. This improvement was approximately 1.88-fold (71.5 mg/g) with the supplementation of 5 mM Tween 80, which is the probable effective stimulator in lipase production, to the R2YE medium. When 5 mM vinyl propionate was added as a precursor for PCC pathway to R2YE medium, the specific production of FK506 increased approximately 1.9-fold (71.61 mg/g) compared to that under the non-supplemented condition. Moreover, in the presence of 5 mM Tween 80, the specific FK506 production was approximately 2.2-fold (157.44 mg/g) higher than that when only vinyl propionate was added to the R2YE medium. In particular, PCC expression in Streptomyces sp. RM7011 (RM7011/pSJ1003) together with vinyl propionate feeding resulted in an increase in the FK506 titer to as much as 1.6-fold (251.9 mg/g) compared with that in RM7011/pSE34 in R2YE medium with 5 mM Tween 80 supplementation, indicating that the vinyl propionate is more catabolized to propionate by stimulated lipase activity on Tween 80, that propionyl-CoA yielded from propionate generates methylmalonyl-CoA, and that the PCC pathway plays a key role in increasing the methylmalonyl-CoA pool for FK506 biosynthesis in RM7011 strain. Overall, these results show that a combined approach involving classical random mutation and metabolic engineering can be applied to supply the limiting factor for FK506 biosynthesis, and vinyl propionate could be successfully used as a precursor of important methylmalonyl-CoA building blocks.

FK506 is an important 23-member polyketide macrolide with immunosuppressant activity. Its entire biosynthetic gene cluster was previously cloned from Streptomyces sp. strain KCTC 11604BP, and sequence analysis identified three putative regulatory genes, tcs2 , tcs7 , and fkbN , which encode proteins with high similarity to the AsnC family transcriptional regulators, LysR-type transcriptional regulators, and LAL family transcriptional regulators, respectively. Overexpression and in-frame deletion of tcs2 did not affect the production of FK506 or co-occurring FK520 compared to results for the wild-type strain, suggesting that tcs2 is not involved in their biosynthesis. fkbN overexpression improved the levels of FK506 and FK520 production by approximately 2.0-fold, and a deletion of fkbN caused the complete loss of FK506 and FK520 production. Although the overexpression of tcs7 decreased the levels of FK506 and FK520 production slightly, a deletion of tcs7 caused 1.9-fold and 1.5-fold increases in FK506 and FK520 production, respectively. Finally, fkbN overexpression in the tcs7 deletion strain resulted in a 4.0-fold (21 mg liter −1 ) increase in FK506 production compared to that by the wild-type strain. This suggests that fkbN encodes a positive regulatory protein essential for FK506/FK520 biosynthesis and that the gene product of tcs7 negatively regulates their biosynthesis, demonstrating the potential of exploiting this information for strain improvement. Semiquantitative reverse transcription-PCR (RT-PCR) analyses of the transcription levels of the FK506 biosynthetic genes in the wild-type and mutant strains proved that most of the FK506 biosynthetic genes are regulated by fkbN in a positive manner and negatively by tcs7 .

A novel extracellular phospholipase C (PLC) was purified from a marine streptomycete. It had a molecular mass of 28 kDa as estimated by SDS-polyacrylamide gel electrophoresis. Its enzyme activity was optimal at pH 8.0 at 45 degrees Celsius. The PLC hydrolyzed only phosphatidylcholine. Its activity was enhanced 300% by Na(+) (200 mM), suggesting that the purified PLC is a typical marine-type enzyme.

Marine bacterial isolates were screened for phospholipase C (PLC) activity on PCY agar plates containing phosphatidylcholine (PC) as substrate. The strain that showed the highest activity on a PCY screening agar plate and a thin-layer chromatography was identified as a strain of Pseudoalteromonas and subsequently designated Pseudoalteromonas sp. J937. The extracellular PLC of the strain J937 was purified to a specific activity of 33 U mg(-1) protein by serial ion exchange and gel filtration column chromatography. It had a molecular mass of 32 kDa estimated by SDS-PAGE. The optimal pH and temperature of the enzyme were about pH 8 and 45 degrees C, respectively. The PLC hydrolyzed phosphatidylethanolamine as well as PC but not other glycerophospholipids. Its activity was enhanced by 150% with Ca2+ (200 mM) and by 180% with Na+ (500 mM), suggesting that the purified PLC is a marine-type enzyme.

SummaryThe enzyme RedP is thought to initiate the biosynthesis of the undecylpyrolle component of the antibiotic undecylprodiginine produced by Streptomyces coelicolor. RedP has homology to FabH, which initiates fatty acid biosynthesis by condensing the appropriate acyl-CoA starter unit with malonyl ACP. We have generated a redP-deletion mutant of S. coelicolor M511 (SJM1) and shown that it produces reduced levels of prodiginines and two new analogs, methylundecylprodiginine and methyldodecylprodiginine. Incorporation studies with perdeuterated valine were consistent with these being generated using methylbutyryl-CoA and isobutyryl-CoA as starter units, respectively. Plasmid-based expression of a streptomycete fabH in the SJM1 mutant led to restoration of overall prodiginine titers but the same overall ratio of undecylprodiginines and novel prodiginines. Thus, the redP FabH can be replaced by FabH enzymes with different substrate specificities and provides a method for generating novel prodiginines.

To obtain eicosapentaenoic acid (EPA)-producing bacteria, some 600 strains of bacteria were isolated from Antarctic sediment and marine organisms during the summer expedition of 1999-2000 and 7 EPA-producing bacteria were obtained through screening with TLC and GC. A strain designated as L93 showed the highest EPA production, which was gram-negative, rod-shaped bacterium. L93 strain was identified as Shewanella sp., from the sequence analysis of 16S rDNA. Optimal conditions temperature and pH for the growth and EPA production were about and pH 7. In addition, its production was optimized by 50%(w/v) sea salt. We establish the optimal production system to produce about 320 mg per liter by using this optimal EPA production conditions. EPA-methyl ester was purified from cultured L93 strain to a purity of higher than 97% and typical purification yield is greater than 72% of the input amount via urea complexation and HPLC.

Background Clostridium butyricum is a butyric acid-producing anaerobic bacteriuma, and commonly present as gut microbiota in humans. This species has been used as a probiotic for the prevention of diarrhea in humans. In this study, we report the draft genome of C. butyricum DKU-01, which was isolated from infant feces, to better understand the characteristics of this strain so that it can later be used in the development of probiotic products. Results A total of 79 contigs generated by hybrid assembly of sequences obtained from Roche 454 and Illumina Miseq sequencing systems were investigated. The assembled genome of strain DKU-01 consisted of 4,519,722 bp (28.62% G + C content) with a N50 contig length of 108,221 bp and 4,037 predicted CDSs. The extracted 16S rRNA gene from genome sequences of DKU-01 was similar to Clostridium butyricum with 99.63% pairwise similarity. The sequence of strain DKU-01 was compared with previously reported genome sequences of C. butyricum. The value of average nucleotide identity between strains DKU-01 and C. butyricum 60E3 was 98.74%, making it the most similar strain to DKU-01. Conclusions We sequenced the DKU-01 strain isolated from infant feces, and compared it with the available genomes of C. butyricum on a public database. Genes related to Fructooligosaccharide utilization were detected in the genome of strain DKU-01 and compared with other genera, such as Bifidobacterium and Streptococcus. We found that strain DKU-01 can metabolize a wide range of carbohydrates in comparative genome result. Further analyses of the comparative genome and fermentation study can provide the information necessary for the development of strain DKU-01 for probiotics. Electronic supplementary material The online version of this article (doi:10.1186/s13099-015-0055-3) contains supplementary material, which is available to authorized users.