Cloning and Bioinformatics Analysis of Rosa rugosa β-1,3-Glucanase Gene (RrGlu)

In order to reveal which role the callose played in R. rugosa pollination incompatibility, the full-length cDNA sequence of β-1,3-glucanase gene was cloned for the first time from the stylus of Rosa rugosa “Tanghong” with RT-PCR and RACE methods and named as RrGlu. The full-length cDNA is 1380 bp with an open reading frame of 1041 bp, encoding 346 amino acids. The derived protein has a molecular weight of 37.85 kD, a calculated pI of 9.12, a pfam00332 conserved domain at position 36 - 345, and belongs to glycosyl hydrolase family 17. The derived protein is a hydrophilic protein secreted into the vacuole. There is a signal peptide cleavage site at position 34 - 35, a transmembrane domain at position 13 - 32, six Ser phosphorylation sites, three Thr phosphorylation sites, three Tyr phosphorylation sites, one N-glycosylation site, and five O-glycosylation sites. There are 31.50% α-helixes, 30.92% random coil, 25.14% extended peptide chain, and 12.43% β-corner structure. This protein and the Glu protein from eight other species, including Prunus persica, share a sequence homology of greater than 72%;all of the proteins contain a pfam00332 conserved domain and a β-1,3-glucanase active center sequence (LIVM)-X-(LIVMFYW)3-(STAG)-E-(ST)-G-W-P-(ST)-X-G. Furthermore, their phylogenetic relationships are consistent with their traditional classifications. These results were meaningful to reveal the molecular mechanism of R. rugosa pollination incompatibility and improve the theory and techniques of breeding ornamental R. rugosa.

Production of β-1,3-glucanase and chitinase of two biocon-trol agents and their possible modes of action

Pichia membranefaciens Hansen and Candida guilliermondii (Cast) Langeronet Guerra are two antagonists of R. stolonifer on harvested nectarine and peach fruits. In this study, β-1,3-glucanase and chitinase activities of the antagonists were induced in vitro and in vivo. The highest β-1, 3-glucanase activity was detected in Lilly-Barnett minimal salt medium supplemented with glucose in combination with CWP of R. stolonifer as a carbon source. The β-1,3-glucanase activity of P. membranefaciens reached the maximum level, being 114.0 SU (specific activity unit), and that of C. guilliermondii reached 103.2 SU. The lowest β-1,3-glucanase activity was observed in the medium containing glucose as sole car-bon source. P. membranefaciens was able to produce signifi-cantly higher levels of chitinase (exochitinase and endochiti-nase) in vitro than C. guilliermondii grown in Czapeck mini-mal medium. An increase in β-1,3-glucanase and chitinase activity was also triggered by wounding, adding of carbon sources

ZmGns,a maize class I β-1,3-glucanase,is induced by biotic stresses and possesses strong antimicrobial activity

Plant b-1,3-glucanases are members of the pathogenesis-related protein 2（PR-2） family,which is one of the 17 PR protein families and plays important roles in biotic and abiotic stress responses.One of the differentially expressed proteins（spot 842） identified in a recent proteomic comparison between five pairs of closely related maize（Zea mays L.） lines differing in aflatoxin resistance was further investigated in the present study.Here,the corresponding cDNA was cloned from maize and designated as ZmGns.ZmGns encodes a protein of338 amino acids containing a potential signal peptide.The expression of Zm Gns was detectible in all tissues studied with the highest level in silks.ZmGns was significantly induced by biotic stresses including three bacteria and the fungus Aspergillus flavus.ZmGns was also induced by most abiotic stresses tested and growth hormones including salicylic acid.In vivo,ZmGns showed a significant inhibitory activity against thebacterial pathogen Pseudomonas syringae pv.tomato DC3000 and fungal pathogen Botrytis cinerea when it overexpressed in Arabidopsis.Its high level of expression in the silk tissue and its induced expression by phytohormone treatment,as well as by bacterial and fungal infections,suggest it plays a complex role in maize growth,development,and defense.

Induction of Volatile Organic Compounds in Leaves of Lycopersicon Esculentum by Nd^(3+)

The effects of Nd^3＋ on the quality and quantity of volatile organic compounds （VOCs） in the leaves of Lycopersicon esculentum were studied. The results demonstrate that Nd^3＋ can increase the total amount of VOC by 75% after treatment for 120 h, as compared with the control. Phyto-oxylipins, terpenoids and aromatic compounds were increased by 73%, 38% and 21%, respectively. （E）-2-hexenal, the most abundant constituent is increased by 74%, β- phellandrene and α-caryophyllene in terpenoids,

Continuous Saccharification of Laminarin by Immobilized Laminarinase ULam111 Followed by Ethanol Fermentation with a Marine-Derived Yeast

We isolated a novel laminarinase ULam111 from Flavobacterium sp. strain UMI-01. Purified ULam111 showed degradation activity against laminarin with the specific activity of 224 ± 18 U/mg at 30°C and pH 6.0. Its optimum temperature was 50°C, and degradation activities against laminarin were observed at 4°C - 80°C. With a laminarin degradation system, we investigated the preparation and properties of immobilized ULam111 with the use of the 11 types of carriers. The high activity recoveries of immobilized ULam111 were as follows: 19.4% for IB-S60P carrier beads (the non-ionic type), 15.6% for IB-S60S carrier beads (the non-ionic type), 11.9% for IB-150P carrier beads (the covalent type), and 7.1% for IB-C435 carrier beads (the cationic type). With the repeated use of immobilized ULam111, the enzyme activities immobilized on IB-S60S and those on IB-S60P remained at 40% and 30% respectively after the sixth trial. We selected IB-S60S as suitable beads for enzyme immobilization, and we attempted to construct a reactor system with ULam111 immobilized on IB-S60S beads. In this system, 1.2 - 1.9 g/L glucose was repeatedly produced from 30 mg/mL laminarin solutions after 20 hr when the reactor operation was repeated 10 times. We examined ethanol fermentation from the saccharified solutions with a marine-derived yeast (Saccharomyces cerevisiae C-19), and 0.51 - 0.58 g/L bioethanol was produced from the saccharified solution that contained 1.71 - 1.86 g/L of glucose.

Glu R2-3Y Inhibits the Acquisition and Reinstatement of Morphine-Induced Conditioned Place Preference in Rats

Accumulating evidence indicates that a-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptors（AMPARs） are involved in the relapse to abused drugs.However, the role of AMPARs containing the Glu R2 subunit in opiate addiction is still unclear. Glu R2-3Y,an interfering peptide, prevents the endocytosis of AMPARs containing the Glu R2 subunit. In this study, we explored the effect of intravenous injection of Glu R2-3Y on the acquisition, expression, and reinstatement of morphine-induced conditioned place preference（m CPP） in rats. We found that infusion of Glu R2-3Y（1.5 nmol/g） one hour before morphine during the conditioning phase inhibited the acquisition of m CPP, while an identical injection one hour before the post-conditioning test had no influence on the expression of m CPP. Injection of Glu R2-3Y（1.5 nmol/g） after m CPP extinction blocked the morphine-induced reinstatement of m CPP. Our results strongly support the hypothesis that inhibition of AMPAR endocytosis provides a new target for the treatment of opiate addiction.

Steroidal Plant Growth Promoters vs. Phytopathogens, via Enzymatic Regulation;An in <i>Silico</i>Approach

Steroidal plant growth promoters (SPGP) have been continuously studied due to their high activity increasing biomass and resistance to diverse stress factors. In our hands, a new SPGP family of 22-oxocholestanic compounds stands out at a comparative level to brassinosteroids (BSs). The potential activity of new SPGP against phytopathogens was studied through in silico molecular docking, these assays were performed with relevant ensymes of phytopatogens Chitinase B and 1,3-β-Glucanase. Nine Chitinase B inhibitors and two 1,3-β-Glucanase inhibitors were proposed. The launched study analyzed the interactional and spatial level, determining the presence of interactions with key amino acids in receptors in comparison to reference inhibitors. Even more, the AVR4 and ECP6 effectors were also examined. No compound that blocks ECP6 was found;due to, probably, the influence of its highly hydrophilic environment. In the case of AVR4, two SPGP showed a better docking score (DS) than a chitin fragment (endogenous ligand);this fact demonstrates the latent potential of the 22-oxocholestanic derivatives against phytopathogens, with a specific regulation via proliferation inhibition. Moreover, this SPGP does not affect the symbiotic fungi that are beneficial for the natural plant system.