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5-Aminolevunic acid, a natural plant growth regulator, can thin pear fruits by inhibiting pollen tube growth through Ca 2+ -ATPase-mediated Ca 2+ efflux. Here, using a non-invasive micromeasurement technology, we detected the occurrence of an ALA-induced Ca 2+ efflux. Transmission electron microscope observations showed that a disruption of the intracellular Ca pool, including endoplasmic reticulum and mitochondria, was also linked to ALA-decreased Ca 2+ in pollen tubes. Then we further explored the mechanisms how ALA-decreased Ca 2+ inhibits pollen tube growth. We observed that ALA-inhibited pollen tube growth was accompanied by tube swelling and disruption in an ALA concentration-dependent manner. Ultrastructural studies further showed that ALA increased the number of vesicles, and the vesicle aggregation resulted in the morphological abnormality of pollen tubes. Confocal laser scanning microscopy using the fluorescent marker FM 4-64 confirmed the increasing number of vesicles induced by ALA in the pollen tubes. Ultrastructural analysis of pollen tubes treated by ALA with exogenous CaCl 2 or/ and EGTA not only indicated that a Ca 2+ signal plays a crucial role in ALA-mediated increase of vesicles, but also suggested that destruction of endoplasmic reticulum and Golgi bodies is one of the vesicle sources. Confocal microscopy of FM 4-64 further demonstrated that Ca 2+ signal involves in ALA-mediated increase of vesicles also through promoting endocytosis and inhibiting exocytosis. In addition, treatment of taxol, a microtubule stabilizer, partially rescued the effects of ALA on pollen tube growth and FM 4-64 staining, indicating that microtubules are involved in ALA-regulated pollen tube growth. Taken together, we conclude that ALA-decreased Ca 2+ disrupts endoplasmic reticulum and Golgi apparatus, enhances endocytosis, suppresses exocytosis, impairs vesicle transport, and finally leads to vesicle accumulation and abnormality of pollen tubes. Our findings provide new information on how ALA regulates pollen tube growth. [ABSTRACT FROM AUTHOR]

CuI-catalyzed decarboxylative phosphonylation of terminal alkyne-substituted cyclic carbonates/carbamates with phosphine oxides has been developed relying on the highly regioselective addition at the α -position of the in situ generated copper-allenylidene intermediates. A series of 4-phosphonyl 2,3-allenols and 4-phosphonyl 2,3-allenamines were smoothly obtained in moderate to excellent yields under mild conditions. The synthetic utility of this methodology was demonstrated by large-scale synthesis and versatile transformations of product. [Display omitted] [ABSTRACT FROM AUTHOR]

Abstract: Glycosidase-mediated glycosylation of alkynyl alcohols and azide-containing alcohols was followed by a click reaction, affording various types of triazole glycosides. The activities of triazole glycosides detected in subsequent bioassays show that this procedure is a feasible approach to the development of anti-fungal drugs. [Copyright &y& Elsevier]

Abstract: A new resource of biocatalyst for asymmetric reduction of aromatic ketones has been discovered for the first time from a common plant seed, adzuki bean, i.e. Phaseolus angularis (Willd.) W.F. Wight. The study investigated the best methods to prepare the biocatalyst and its ability to reduce ketones. Our results indicated that the biocatalyst from adzuki bean could reduce various aromatic ketones at relatively high concentrations (e.g. 100mM), exhibiting excellent stereoselectivity (>98% e.e.). In addition, it was found that NADPH acts as the reducing cofactor, which can be regenerated by the crude enzyme system itself using glucose as an auxiliary substrate. [Copyright &y& Elsevier]

Abstract: Among several fruit seeds, apple seed was identified as a new promising β-glucosidase source for alkyl O-glucoside synthesis by reverse hydrolysis, since it showed high hydrolytic activities on a broad spectrum of β-glucosides. From the crude extract of apple seed meal, a major glucosidase isoenzyme was purified to homogeneity, with a purification factor of 47-fold and an overall yield of 12.8%, by ammonium sulfate fractionation and chromatographic separation through DEAE-Cellulose, Butyl-Toyopearl and Sephadex G150 columns. The purified enzyme is a homodimer; each subunit has a molecular mass of about 60kDa as determined by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and the pI of this β-glucosidase is about 5.7. These properties indicate that the apple seed β-glucosidase is totally different from any of the almond β-glucosidase isozymes reported so far. Furthermore, the purified enzyme of apple seed displays higher thermal stability than the commercially supplied β-glucosidase from almond, with half-lives of 42.9h and 14.2h, respectively, as preserved at 50°C in an aqueous environment. The optimum pH of the apple seed β-glucosidase was 6.0 and the stable pH range was 5.0–9.0, both similar to those of the commercial almond β-glucosidase. [Copyright &y& Elsevier]

Abstract: A new soil isolate, Fusarium proliferatum ECU2042, was found capable of producing a ginsenoside Rg3 hydrolase (a special β-glucosidase) which is useful for production of a very potent antitumor agent Rh2. By investigating its fermentation progress, the optimal time for β-glucosidase biosynthesis was determined as 72h. The crude β-glucosidase was partially purified by 49.4-fold from the cell-free extract with an activity yield of 13%. The catalytic performance of the partially purified β-glucosidase was examined, giving optima of pH and temperature at pH 5.0–6.0 and 50°C. It was found that the F. proliferatum β-glucosidase can also hydrolyze cellobiose, but not active towards a frequently used artificial substrate p-nitrophenyl β-d-glucopyranoside (pNPG). In a lab-scale preparation, the bio-hydrolytic reaction was carried out at 50°C for 24h in 50ml NaAc/HAc buffer (100mM, pH 5.0) containing 50mg Rg3 and 12mg lyophilized crude enzyme extract, giving the ginsenoside Rh2 in 60.3% conversion. [Copyright &y& Elsevier]

Abstract: A novel β-glucosidase from Fusarium proliferatum ECU2042 (FPG) was successfully purified to homogeneity with a 506-fold increase in specific activity. The molecular mass of the native purified enzyme (FPG) was estimated to be approximately 78.7kDa, with two homogeneous subunits of 39.1kDa, and the pI of this enzyme was 4.4, as measured by two-dimensional electrophoresis. The optimal activities of FPG occurred at pH 5.0 and 50°C, respectively. The enzyme was stable at pH 4.0–6.5 and temperatures below 60°C, and the deactivation energy (E d) for FPG was 88.6kJmo1−1. Moreover, it was interesting to find that although the purified enzyme exhibited a very low activity towards p-nitrophenyl β-d-glucoside (pNPG), and almost no activity towards cellobiose, a relatively high activity was observed on ginsenoside Rg3. The enzyme hydrolyzed the 3-C, β-(1→2)-glucoside of ginsenoside Rg3 to produce ginsenoside Rh2, but did not sequentially hydrolyze the β-d-glucosidic bond of Rh2. The K m and V max values of FPG for ginsenoside Rg3 were 2.37mM and 0.568μmol (hmg protein)−1, respectively. In addition, this enzyme also exhibited significant activities towards various alkyl glucosides, aryl glucosides and several natural glycosides. [Copyright &y& Elsevier]