α-Galactosidase from sweet chestnut seeds

The major sugars of fresh seeds of Castanea sativa were shown to be raffinose, stachyose and sucrose. Drying seeds at 25° for 14 weeks increased the ratio raffinose: stachyose from 1.1 to 3.5, reduced sucrose content by ca 50 % and decreased total extractable α-galactosidase. The enzyme activity was resolved into two peaks, a high MW form I (apparent MW215 000) and a low MW form II (apparent MW 53 000). The latter form was predominant in the extract of fresh seeds whereas the former was the main form in the 14-week dried seeds. An increase in the amount of enzyme I was also observed when a buffered extract (pH 5.5) of fresh seeds was stored at 4°. Enzymes I and II had pH optima of 4.5 and 6, respectively. Both enzymes hydrolysed p-nitrophenyl α-d-galactoside at a much greater rate than the natural substrates raffinose, stachyose, locust bean gum and carob gum. However, enzyme I showed preference for stachyose as compared to raffinose; the opposite order was observed for enzyme II.     

Travelling seeds? Examples from New Zealand

New Zealand is an archipelago isolated in time and space. Flowering plants have evolved in the almost-complete absence of terrestrial mammals, some of the most prolific seed-dispersers from the rest of the world, and fascinating mechanisms for spreading seeds have developed.     

Sandalwood plantlets from ‘Synthetic seeds’

Somatic embryos of sandalwood (Santalum album) were encapsulated in an alginate matrix to prepare Synthetic seeds. Encapsulated single embryos germinated to form plants with roots and shoots. Embryogenic cell suspensions encapsulated and stored at 4°C for 45 days produced embryos when recultured as suspensions.Abbreviations BAP benzylaminopurine - 2,4-D 2,4-dichlorophenoxyacetic acid - GA gibberellic acid - IAA indole acetic acid - IBA indole butyric acid - MS Murashige and Skoog (1962) - TW tap water - WM White's medium     

Two forms of α-glucosidase from sugar-beet seeds

Two forms of -glucosidase (EC 3.2.1.20), designated as I and II, have been isolated from sugarbeet (Beta vulgaris L.) seeds by a procedure including fractionation with ammonium sulfate and ethanol, carboxymethyl-cellulose column chromatography, and preparative disc gel electrophoresis. The two enzymes were homogeneous by polyacrylamide disc gel electrophoresis. Their molecular weights were 98,000 (I) and 60,000 (II). -Glucosidase I readily hydrolyzed maltose, isomaltose, kojibiose, maltotriose, panose, amylose, soluble starch, amylopectin and glycogen. -Glucosidase II also hydrolyzed maltose, kojibiose and maltotriose but hydrolyzed the other substrates only very weakly or not at all. -Glucosidase I hydrolyzed soluble starch at a faster rate than maltose. It produced isomaltose and panose as the main -glucosyltransfer products from maltose, whereas maltotriose was the main product of -glucosidase II. -Glucosidase I hydrolyzed amylose liberating -glucose. The neutral-sugar content was calculated to be 2.7% for -glucosidase I and 8.8% for -glucosidase II. The main neutral sugar was mannose in -glucosidase I, and glucose in -glucosidase II.     

α-galactosidase from germinating guar (Cyamopsis tetragonolobus) seeds

The changes in α-galactosidase activity in guar (Cyamopsis tetragonolobus) seeds was followed during seven days of germination. The enzyme activity was maximal on the first day of germination and gradually decreased during subsequent days. On the second day of germination the partially purified enzyme upon ion-exchange chromatography on CM-Sephadex C-50 was resolved into α-galactosidase-A (anionic), α-galactosidase-C₁ (cationic) and α-galactosidase-C₂ (cationic) and their relative proportions were 28,12 and 60%, respectively. The combined α-galactosidase C₁ and C₂ activities increased in the first two days of germination followed by significant decrease after the 3rd day onwards, whereas α-galactosidase-A remained fairly constant throughout the germination period, α-Galactosidase-A and C2 had differentK _m and V_max values withp-nitrophenyl α-D-galactopyranoside, raffinose and melibiose as substrates and also differed in their thermal stabilities     

Properties of α-galactosidase II2 from Vicia faba seeds

-Galactosidase II² (MW 43 390) from resting Vicia faba L. seeds had been shown to possess d-glucose/d-mannose-specific lectin activity. Inhibition studies with monosaccharides and an examination of the effects of heat and pH on the catalytic and lectin activities of the enzyme indicate that the enzyme substrate and the lectin haptens bind at different sites on the protein. d-Mannosebinding has been investigated by equilibrium dialysis and spectrophotometrically. Both methods yield K_a values of approx. 3·10³ M^-1 for the interaction and there would appear to be two mannosebinding sites per molecule of enzyme protein. The lectin properties of V. faba -galactosidase II² have been discussed in relation to both V. faba lectin (favin) and other legume -galactosidases.Abbreviations con A concanavalin A - CM-cellulose carboxymethyl cellulose - MW molecular weight - PNPG p-nitrophenyl -d-galactoside - SDS sodium dodecyl sulphate - PAGE polyacrylamide-gel electrophoresis     

Purification and properties of α-galactosidases from Vicia faba seeds

Two forms of alpha-galactosidase, I and II, exist in Vicia faba seeds and these have been purified 3660- and 337-fold respectively. They behaved as homogeneous preparations when examined by ultracentrifugation, disc electrophoresis and gel filtration. The apparent molecular weights of enzymes I and II, as determined by gel filtration, were 209000 and 38000 respectively. The carbohydrate contents of enzymes I and II were 25% and 2.8% respectively, and the enzymes differed in their aromatic amino acid compositions. Enzyme I was split into six inactive subunits in the presence of 6m-urea. alpha-Galactosidases I and II showed different pH optima and K(m) and V(max.) values with p-nitrophenyl alpha-d-galactoside and raffinose as substrates, and also differed in their thermal stabilities.     

Cross-linked enzyme aggregates of β-glucosidase from Prunus domestica seeds

Cross-linked enzyme aggregates (CLEAs) of β-glucosidase were prepared and characterized. Under the optimum conditions, the activity recovery of CLEAs reached 84?%. The reduction by NaBH(4) resulted in slightly lower activities of CLEAs, while their thermostability was enhanced. CLEAs were more thermally stable than free enzyme (half lives, 973 vs. 518?min at 50?°C), while less stable than seed meal (half life, 1,090?min). In 90?% (v/v) t-butanol, the half lives of CLEAs and free enzyme were 53 and 6.7?h, respectively. Besides, the catalytic efficiency (V (max)/K (m)) of CLEAs was comparable to free enzyme (0.42 vs. 0.47?min(-1) mg(-1)). This carrier-free immobilized enzyme had a network structure with multiple layers. The productivity of salidroside using CLEAs reached 150?g/l?g catalyst, while being 6.3?g/l?g with seed meal.     

γ-Glutamylwillardiine and γ-glutamylphenylalanylwillardiine from seeds of Fagus silvatica

γ-l-Glutamyl-l-willardiine [γ-l-glutamyl-3-(1-uracil)-l-l-alanine] and γ-glutamylphenylalanylwillardiine have been isolated from seeds of Fagus silvatica. The structures have been established by spectroscopy, hydrolysis to give the constituent amino acids, and for the tripeptide end-group determination and partial hydrolysis to give the two constituent dipeptides.     

A highly efficient and thermostable α-amylase from soya bean seeds

The α-amylase from soya bean seeds was purified by affinity precipitation, resulting in approx. 20-fold purification with approx. 84% recovery. The purified α-amylase had an optimum pH of 5.5, optimum temperature of 75?°C, Arrhenius energy of activation of 6.03?kcal/mol (1?kcal≈4.184?kJ) and a Km of 2.427?mg/ml (starch substrate). The enzyme had maximum substrate specificity for starch. Among the various metal ions tested, Co2+ and Mn2+ were found to be strong activators. The effect of thiol group modifying agents showed that the thiols of soya bean α-amylase are not directly involved in catalysis. The thermostability of the enzyme makes it suitable for starch liquefaction and the detergent industry respectively.     

The lectin nature of α-galactosidases from Vicia faba seeds

α-Galactosidase from Vicia faba seeds has been resolved into three molecular forms, I, II¹ and II², respectively. Enzyme I is a tetramer (M_r 160 000) consisting of identical sub-units (M_r 44000 ± 2000). All three forms display lectin activity with glucose/mannose specificity. Enzyme I has been further studied with respect to its lectin specificity and various factors affecting this property. The results indicate that the catalytic and the lectin sites reside in the same protein molecule. The results presented are unique in that the enzyme activity is specific for galactose and its lectin activity is specific for glucose/mannose.     

Two novel monoterpene–chalcone conjugates isolated from the seeds of Alpinia katsumadai

Three monoterpene–chalcone conjugates, including two novel compounds isorubraine (2) and sumadain C (3), and a known compound rubraine (1) were isolated from the seeds of Alpinia katsumadai. Their structures and relative configurations were established by NMR spectroscopy and X-ray crystallography. Cytotoxic activities of these compounds were evaluated against cell lines HepG2, MCF-7 and MAD-MB-435, and compound 3 exhibited potent cytotoxic activity.     

Locustoside A — A new purine alkaloid glucoside from seeds of Gleditsia japonica

A new purine alkaloid glucoside designated locustoside A has been isolated from the seeds of Gleditsia japonica, which has been used in oriental traditional medicine. The structure was determined as 7-β-d-glucopyranosyl-3-(3-methyl-2-butenyl)-isoguanine by interpretation of spectroscopic data and was confirmed by X-ray crystallographic analysis.     

Garden variety seeds? Botanical remains from the Petra Garden and Pool Complex

Archaeobotanical samples recovered during the 2004, 2005, 2009 and 2011 excavation seasons at the Petra Garden and Pool Complex (PGPC) in Jordan examined plants that were preserved during different phases of use of the site. The analysis of these remains attempts to determine what plants would have been grown in the garden in antiquity. Results indicated that the material recovered likely represented plants that had been preserved elsewhere in the area and then deposited in the garden to act as fertilizer. The remains were generally charred and were probably derived from fuel and/or waste deposits as it does not appear that the garden was catastrophically burned. Although it is difficult to ascertain garden variety plants under these preservation conditions, an analysis of the archaeobotanical data from the PGPC addresses the questions of what and how plants functioned in the local and regional economy, and how the role of the garden changed through time, in addition to providing insight into the general environmental or ecological conditions present in southern Jordan from the Late Hellenistic through Late Byzantine/Post-Classical periods. Although ornamental species represent a small fraction of materials collected, the presence of these taxa may indicate their occurrence in the garden. Even if garden plants are not readily identifiable, this study aids in addressing the larger issues of trade, economy and environment of this ancient city, as well as the feasibility of identifying garden plants in antiquity.     

A glucose-tolerant β-glucosidase from Prunus domestica seeds: Purification and characterization

A glucose-tolerant β-glucosidase was purified to homogeneity from prune (Prunus domestica) seeds by successive ammonium sulfate precipitation, hydrophobic interaction chromatography and anion-exchange chromatography. The molecular mass of the enzyme was estimated to be 61 kDa by SDS-PAGE and 54 kDa by gel permeation chromatography. The enzyme has a pI of 5.0 by isoelectric focusing and an optimum activity at pH 5.5 and 55 °C. It is stable at temperatures up to 45 °C and in a broad pH range. Its activity was completely inhibited by 5 mM of Ag⁺ and Hg²⁺. The enzyme hydrolyzed both p-nitrophenyl β-d-glucopyranoside with a K_m of 3.09 mM and a V_max of 122.1 μmol/min mg and p-nitrophenyl β-d-fucopyranoside with a K_m of 1.65 mM and a V_max of 217.6 μmol/min mg, while cellobiose was not a substrate. Glucono-δ-lactone and glucose competitively inhibited the enzyme with K_i values of 0.033 and 468 mM, respectively.     

The cytotoxicity of 8-O-4′ neolignans from the seeds of Crataegus pinnatifida

Nine new 8-O-4′ neolignans, named pinnatifidanin B I–IX (1–9), together with 9 known analogs (10–18) were isolated from the seeds of Crataegus pinnatifida. The structures of 1–18 were determined by spectroscopic methods, including 1D, 2D NMR, CD and HRESIMS analysis. Compounds 8–11, 17 and 18 displayed potent cytotoxic activities against human cancer cell lines, and most interestingly, none of the 6 compounds displayed inhibitory activity against human lung cell line (Mrc5). The 6 cytotoxic compounds are considered to be potential as antitumor agents, which could significantly inhibit the cancer cell growth in a dose-dependent manner and are probably safer than positive control drug.     

The partial purification and characterisation of gibberellin 2β-hydroxylases from seeds of Pisum sativum

The gibberellin (GA) 2-hydroxylases in mature and immature seeds of Pisum sativum have been partially purified and characterised. The enzymes are unstable when stored below pH 7.0 or in the absence of a thiol reagent. The optimum assay pH is between 7.4 and 7.8 and activity is dependent upon the presence of -ketoglutarate, Fe²⁺ and ascorbate. The 2-hydroxylase activities for GA₁, GA₄, GA₉ and GA₂₀ are chromatographically inseparable and correspond to a protein of M_r 44000. The rate of GA 2-hydroxylation varies according to substrate and some evidence indicates that the 2-hydroxylase activities for GA₁ and GA₄ and for GA₉ and GA₂₀ may reside in different proteins. During pea seed maturation, the specific activity of the enzyme(s) increases dramatically and reaches a maximum at a time when endogenous GA₉, GA₂₀, GA₂₉ and GA₅₁ are also at their greatest concentration. This correlation is not the result of substrate induction of enzyme activity. Since the GA 2-hydroxylases operate at maximal rate at low substrate concentrations they are incapable of rapidly 2-hydroxylating excessive quantities of (exogenously applied) GA₁ or GA₂₀. On the basis of the kinetic parameters of the GA 2-hydroxylase activities, a generalised model is discussed for the control of the steady-state levels of bioactive hormone under normal physiological conditions.Abbreviations DTE dithioerythritol - EDTA ethylenediaminetetraacetic acid - GA_n gibberellin A_n - HPLC high-performance liquid chromatography - HSS high-speed supernatant - LSS low-speed supernatant - PMSF phenylmethane sulphonyl fluoride     

β-Galactosidase from Ginkgo biloba seeds active against β-galactose-containing N-glycans: purification and characterization

In this study, we purified an acidic β-galactosidase to homogeneity from Ginkgo biloba seeds (β-Gal’ase Gb-1) with approximately 270-fold purification. A molecular mass of the purified β-Gal’ase Gb-1 was estimated about 35 kDa by gel filtration and 32 kDa by SDS-PAGE under non-reducing condition, respectively. On the other hand, β-Gal’ase Gb-1 produced a single band with a molecular mass of 16 kDa by SDS-PAGE under reducing condition. The N-terminal amino acid sequences of 32 kDa and 16 kDa molecules were the same and identified as H-K-A-N-X-V-T-V-A-F-V-M-T-Q-H-, suggesting that β-Gal’ase Gb-1 may function as a homodimeric structure in vivo. When complex-type N-glycans containing β-galactosyl residues were used as substrates, β-Gal’ase Gb-1 showed substantial activity for β1-4 galactosyl residue and modest activity for β1-3 galactosyl residue with an optimum pH near 5.0. Based on these results, the involvement of β-Gal’ase Gb-1 in the degradation of plant complex-type N-glycans is discussed.     

α-isoamylases of rye seeds

Rye seeds contained 5 α-type amylases. Three behaved like typical α-amylases and were called Aα-amylases. Two showed chemical activity like α-amylase, but behaved differently physically and were called Bα-amylases. The latter were partly inactivated at pH 3·3 and at 70°. They were more resistant to EDTA than were the Aα-amylases. Barley and wheat seeds contained amylases behaving like Bα-amylases. Aleurone layers contained relatively large amounts of Aα-amylases. Relative amounts of Aα-and Bα-amylases depended on temperature during germination. Bα-amylases remained active for a longer time after germination than Aα-amylases.