[Heterologous expression of a novel ß-glucosidase BglD2 and its application in polydatin-hydrolyzing].

A novel ß-glucosidase BglD2 with glucose and ethanol tolerant properties was screened and cloned from the deep-sea bacterium Bacillus sp. D1. The application potential of BglD2 toward polydatin-hydrolyzing was also evaluated. BglD2 exhibited the maximal ß-glucosidase activity at 45 °C and pH 6.5. BglD2 maintained approximately 50% of its origin activity after incubation at 30 °C and pH 6.5 for 20 h. BglD2 could hydrolyze a variety of substrates containing ß (1→3), ß (1→4), and ß (1→6) bonds. The activity of ß-glucosidase was enhanced to 2.0 fold and 2.3 fold by 100 mmol/L glucose and 150 mmol/L xylose, respectively. BglD2 possessed ethanol-stimulated and -tolerant properties. At 30 °C, the activity of BglD2 enhanced to 1.2 fold in the presence of 10% ethanol and even remained 60% in 25% ethanol. BglD2 could hydrolyze polydatin to produce resveratrol. At 35 °C, BglD2 hydrolyzed 86% polydatin after incubation for 2 h. Thus, BglD2 possessed glucose and ethanol tolerant properties and can be used as the potential candidate of catalyst for the production of resveratrol from polydatin.

An unusual GH1 ß-glucosidase from marine sediment with ß-galactosidase and transglycosidation activities for superior galacto-oligosaccharide synthesis.

A novel ß-glucosidase, BglD1 with high ß-galactosidase and transglycosidation activities, was screened and cloned from the deep-sea bacterium Bacillus sp. D1. BglD1 exhibited the maximal ß-glucosidase and ß-galactosidase activities at 55-60 °C and pH 5.5-6.0. The enzyme maintained approximately 50% of its original activity at 35 °C and pH 6.0 after 120-h incubation. When applied to synthesize galacto-oligosaccharides (GOS), BglD1 generated 118.3 g/L GOS (33.8% (w/w)) from 350 g/L lactose, with trisaccharide Gal-ß(1 → 3)-Lac and disaccharide Gal-ß(1 → 4)-Gal as the main components. Furthermore, BglD1 could hydrolyze lactose in milk and produce GOS simultaneously. Using milk as the substrate, BglD1 hydrolyzed 88.5% lactose and produced 3.3 g/L GOS after incubation at 30 °C for 1 h. To improve the transglycosidation activity, a mutant BglD1:E224T was generated based on the semi-rational design. The GOS yield of BglD1:E224T was 11.5% higher than that of BglD1 when using lactose solution as the substrate. Thus, BglD1 and the mutant could be used as beneficial alternatives of the existing ß-galactosidases for the production of GOS.

Leucothrix sargassi sp. nov., isolated from a marine alga [Sargassum natans (L.) Gaillon].

A novel bacterial strain, C3212T, was isolated from a marine alga collected from the sea shore of Yantai, China. The strain was Gram-stain-negative, rod-shaped, aerobic, non-motile, and oxidase- and catalase-positive. Growth was observed at 8-37 °C (optimum, 28 °C), at pH 6.0-9.0 (optimum, pH 7.0) and in the presence of 1.0-7.0 % (w/v) NaCl (optimum, 4.0 %). The major respiratory quinone was ubiquinone-8 (Q-8). The polar lipids of strain C3212T consisted of diphosphatidylglycerol (cardiolipin), phosphatidylglycerol, phosphatidylethanolamine, an unidentified aminophospholipid, an unidentified phospholipid and an unidentified polar lipid. The major fatty acids were C16 : 1ω6c and/or C16 : 1ω7c, and C18 : 1ω6c and/or C18 : 1ω7c. The DNA G+C content of strain C3212T was 44.3 mol%. Phylogenetic analysis based on 16S rRNA gene sequences showed that the novel strain was related most closely to Leucothrix pacifica XH122T, Leucothrix arctica IMCC 9719T and Leucothrix mucor DSM 2157T with similarities of 98.0, 97.5 and 94.3 %, respectively. Estimated DNA-DNA hybridization values were 14.2, 20.7 and 13.9 % between strain C3212T and L. pacifica XH122T, L. arctica IMCC 9719T and L. mucor DSM 2157T, respectively. Phenotypic, phylogenetic and genomic analyses revealed that strain C3212T represents a novel species of the genus Leucothrix, for which the name Leucothrix sargassi sp. nov. is proposed. The type strain is C3212T (=MCCC 1K03600T=KCTC 72121T).

Fabrication of cellulose-based halogen-free flame retardant and its synergistic effect with expandable graphite in polypropylene.

In this paper, a mono-component, cellulose-based and intumescent halogen-free flame retardant (HECPM) was prepared by introducing phosphate groups and melamine groups onto the structure of cellulose. And the chemical structure and surface morphology of HECPM was confirmed by FTIR, EDS and SEM, respectively. The thermal degradation tests demonstrate HECPM possesses great expansion property during the inducement of heat and the char residue of HECPM is higher than 43% at 600 °C. Moreover, the synergistic effect of HECPM and EG in flame-retarding polypropylene (PP) was also verified. When 30% HECPM/EG in ratio of 1/3 were introduced into PP, the LOI value reaches 31.5% and it can obtain UL-94 V-0 rating. The conjunct action of HECPM and EG in promoting the char-forming process was also achieved in comparison with the calculated and experimental TGA curves. Along with the investigation on the residue, a potential condensed-phase flame retardant mechanism was primarily proposed.

Methionine Promotes Milk Protein and Fat Synthesis and Cell Proliferation via the SNAT2-PI3K Signaling Pathway in Bovine Mammary Epithelial Cells.

Methionine (Met) plays a critical regulatory role in milk production, however, the molecular mechanism of action of Met is largely unknown. This study therefore aimed to investigate the influence of Met on milk synthesis in and proliferation of bovine mammary epithelial cells (BMECs) and explore the underlying mechanism. BMECs cultured in fetal bovine serum (FBS) free Dulbecco's modified eagle's medium (DMEM)/F-12 medium were treated with Met (0, 0.3, 0.6, 0.9, and 1.2 mM). Results showed that Met (0.6 mM) significantly increased milk protein and fat synthesis and cell proliferation. Met stimulation also increased mTOR phosphorylation and protein expression of SREBP-1c and Cyclin D1. Gene function study approaches further revealed that SNAT2 is a key regulator of these signaling pathways. PI3K inhibition experiments demonstrated that SNAT2 stimulates these pathways through regulating PI3K activity, and SNAT2 inhibition experiments further revealed that SNAT2 is required for Met to activate PI3K. Furthermore, immunofluorescence observation detected that Met stimulates SNAT2 cytoplasmic expression. Collectively, these findings demonstrate that Met positively regulates milk protein and fat synthesis and cell proliferation via the SNAT2-PI3K signaling pathway in BMECs.