Showing papers on "Amylase published in 1999"

Bioavailability of starch in bread rich in amylose: metabolic responses in healthy subjects and starch structure

Abstract: Objective: This study investigated whether postprandial metabolic responses to bread could be lowered by substituting high amylose maize starch for a part of the flour. Design and subjects: Eight healthy subjects consumed test meals of equivalent nutritional composition based on white wheat bread, bread rich in amylose (HAWB) and spaghetti as a breakfast meal. Blood samples were collected to measure insulin and glucose concentration during two hours after consumption. The degree of starch crystallinity was investigated by X-ray diffraction and DSC analysis. Results: HAWB produced low glycaemic (60±18) and insulinaemic (57±20) indexes similar to those of spaghetti (83±46, 61±16). In vitro amylase hydrolysis of the three foods showed that high amylose content in HAWB significantly lowered starch degradation in bread without affecting hydrolysis kinetics. Addition of amylose in dough increased the resistant starch content of HAWB (14% of dry matter). The resistant starch fraction was mainly composed of crystalline amylose (B-type X-ray diffraction pattern, melting temperature 105°C) attributable to native high amylose maize starch incompletely gelatinised during bread-cooking. Conclusions: Bread produced by the substitution of high amylose maize starch for a part of wheat flour showed a low glycaemic index. Resistant starch in HAWB corresponded to native crystalline amylose not gelatinised during normal bread-processing conditions. Sponsorship: Barilla, via Montova 166, 43100, Parma, Italy.

Expression of amylase and glucose oxidase in the hypopharyngeal gland with an age-dependent role change of the worker honeybee (Apis mellifera L.).

Abstract: Worker honeybees change their behaviour from the role of nurse to that of forager with age. We have isolated cDNA clones for two honeybee (Apis mellifera L.) genes, encoding alpha-amylase and glucose oxidase homologues, that are expressed in the hypopharyngeal gland of forager bees. The predicted amino acid sequence of the putative Apis amylase showed 60.5% identity with Drosophila melanogaster alpha-amylase, whereas that of Apis glucose oxidase showed 23.8% identity with Aspergillus niger glucose oxidase. To determine whether the isolated cDNAs actually encode these enzymes, we purified amylase and glucose oxidase from homogenized forager-bee hypopharyngeal glands. We sequenced the N-terminal regions of the purified enzymes and found that they matched the corresponding cDNAs. mRNAs for both enzymes were detected by Northern blotting in the hypopharyngeal gland of the forager bee but not in the nurse-bee gland. These results clearly indicate that expression of the genes for these carbohydrate-metabolizing enzymes, which are needed to process nectar into honey, in the hypopharyngeal gland is associated with the age-dependent role change of the worker.

Nucleic acid molecule coding for beta-amylase, plants synthesizing a modified starch, method of production and applications

Abstract: The present invention relates to nucleic acid molecules coding for a protein with beta-amylase activity from potato and to a method for the production of transgenic plant cells and plants synthesizing a modified starch. The invention also relates to vectors and host cells containing the nucleic acid molecules according to the invention, the plant cells and plants produced using the method according to the invention and the starch synthesized by the plant cells and plants according to the invention. The invention also relates to a method of production of said starch.

Modes of action of acarbose hydrolysis and transglycosylation catalyzed by a thermostable maltogenic amylase, the gene for which was cloned from a Thermus strain.

Abstract: Several maltogenic amylases (EC 3.2.1.-) and closely related enzymes were cloned from gram-positive bacteria, including Bacillus species (4, 13). The enzymes were different from typical amylases in that they (i) were not secreted outside the cell, (ii) preferred cyclodextrins to starch or pullulan as a substrate, and (iii) exhibited both transglycosylation and hydrolysis activities on various substrates. They hydrolyzed starch and β-cyclodextrin mainly to maltose and pullulan to panose. Many of these properties, if not all, were shared by some amylolytic enzymes, including neopullulanases (EC 3.2.1.135) and cyclomaltodextrinases (EC 3.2.1.54; CDases) (7, 10, 17, 20, 26). The action modes of two maltogenic amylases (4, 13) and a CDase (17) isolated from three different Bacillus species were investigated by time course experiments with soluble starch or maltotriose as a substrate. The enzymes transferred a sugar molecule (donor) released after the hydrolysis of an α-(1,4)-glycosidic linkage to a reducing end of another sugar molecule (acceptor) by forming an α-(1,6)-glycosidic linkage. The coupled transglycosylation and hydrolysis activities of these enzymes were used for the production of branched oligosaccharides (BOS) from liquefied starch (15, 23), giving a more efficient process than the traditional one (31). The maltogenic amylases from Bacillus licheniformis (BLMA [13]), Bacillus stearothermophilus (BSMA, [4]), and B. subtilis (unpublished data) could hydrolyze acarbose, an amylase inhibitor, at different levels of efficiency. Acarbose is a pseudotetrasaccharide that has a pseudosugar ring at the nonreducing end [4,5,6-trihydroxy-3-(hydroxymethyl)-2-cyclohexene-1-yl] linked to the nitrogen of 4-amino-4,6-dideoxy-d-glucopyranose (4-amino-4-deoxy-d-quinovo-pyranose), which is linked via an α-(1,4)-glycosidic linkage to maltose. The pseudotrisaccharide (PTS) resulting from the hydrolysis of acarbose by these enzymes was transferred to the C-6 of glucose forming isoacarbose. This indicated that the catalytic properties unique to maltogenic amylases are probably due to differences in the tertiary structures of the proteins. The primary structures of maltogenic amylases in four regions were well conserved, and their secondary structure was likely to constitute a (β/α)8-barrel domain as with other amylolytic enzymes (11, 12). The characterization of amylolytic enzymes that exhibit transglycosylation and/or cyclodextrin hydrolyzing activity at the level of protein structure and enzymatic properties would be quite useful for understanding catalytic activities and substrate binding patterns more precisely. In this paper, we report on the cloning and physicochemical properties of another maltogenic amylase of a Thermus strain (ThMA) that was capable of hydrolyzing acarbose and transferring PTS to various acceptors. The enzyme was isolated from a thermophilic gram-negative bacterium, Thermus strain IM6501, and was more stable at high temperatures than other maltogenic amylases. Studies of the transferring activity of the thermostable enzyme by using acarbose and methylation of the resulting transfer products revealed additional modes of transglycosylation. Transglycosylation of a donor sugar molecule (PTS) to an acceptor molecule by forming an α-(1,3)-glycosidic linkage was demonstrated for the first time by using acarbose and ThMA.

A Corn Trypsin Inhibitor with Antifungal Activity Inhibits Aspergillus flavus α-Amylase

Abstract: In this study, we found that the inhibition of fungal growth in potato dextrose broth (PDB) medium by the 14-kDa corn trypsin inhibitor (TI) protein, previously found to be associated with host resistance to aflatoxin production and active against various fungi, was relieved when exogenous α-amylase was added along with TI. No inhibitory effect of TI on fungal growth was observed when Aspergillus flavus was grown on a medium containing either 5% glucose or 1% gelatin as a carbon source. Further investigation found that TI not only inhibited fungal production of extracellular α-amylase when A. flavus was grown in PDB medium containing TI at 100 μg ml-1 but also reduced the enzymatic activity of A. flavus α-amylase by 27%. At a higher concentration, however, TI stimulated the production of α-amylase. The effect of TI on the production of amyloglucosidase, another enzyme involved in starch metabolism by the fungus, was quite different. It stimulated the production of this enzyme during the first 10 ...

Genetic and Biochemical Evidence for the Involvement of α-1,4 Glucanotransferases in Amylopectin Synthesis

Abstract: We describe a novel mutation in the Chlamydomonas reinhardtii STA11 gene, which results in significantly reduced granular starch deposition and major modifications in amylopectin structure and granule shape. This defect simultaneously leads to the accumulation of linear malto-oligosaccharides. The sta11-1 mutation causes the absence of an alpha-1,4 glucanotransferase known as disproportionating enzyme (D-enzyme). D-enzyme activity was found to be correlated with the amount of wild-type allele doses in gene dosage experiments. All other enzymes involved in starch biosynthesis, including ADP-glucose pyrophosphorylase, debranching enzymes, soluble and granule-bound starch synthases, branching enzymes, phosphorylases, alpha-glucosidases (maltases), and amylases, were unaffected by the mutation. These data indicate that the D-enzyme is required for normal starch granule biogenesis in the monocellular alga C. reinhardtii.

Purification and properties of the raw starch-degrading α- amylase of Bacillus sp. IMD 434

Abstract: The raw starch-degrading a-amylase of Bacillus sp. IMD 434 was purified to homogeneity by acetone precipitation, ion- exchange chromatography and hydrophobic interaction chromatography. The enzyme had a relative molecular mass of 69,200, displayed maximum activity at pH 6.0 and 65 °C and released large amounts of glucose and maltose on hydrolysis of starch.

A highly thermostable amylase from a newly isolated thermophilic Bacillus sp. WN11

Abstract: A thermostable amylase-producing Bacillus sp. WN11 was isolated from Wondo Genet hot spring in Ethiopia. The enzyme had a temperature optimum of 75–80 °C. Over 80% of its peak activity was in the pH range of 5–8, with an optimum at 5·5. Thermal stability of the enzyme at 105 °C was higher with the addition of starch. The stabilizing effect of starch was concentration-dependent, showing better stability with increasing concentration of starch. At liquefying temperature (105 °C), addition of Ca2+ did not result in further improvement of the stabilizing effect of starch. This indicates that in the presence of starch, WN11 amylase does not require Ca2+ as a stabilizer at liquefying temperatures as high as 105 °C.

Digestive capacity of the rabbit during the post-weaning period, according to the milk/solid feed intake pattern before weaning.

Abstract: The milk/solid feed intake pattern and digestive capacity were compared in two groups of young rabbits reared between 16 and 32 days in litters of four (L4 group, n = 18 litters) or ten pups (L10, n = 20) in order to modulate milk availability. Compared to the L4 group, rabbits in the L10 group presented a lower milk intake (22.3 versus 28.8 g x day(-1)) and a higher solid feed intake (12.9 versus 8.0 g x day(-1)) between 16 and 32 days of age. Live weight and weight gain were significantly lower in the L10 (-15%) than in the L4 group from 22 until 42 days. No significant difference was observed among the two groups for post-weaning digestibility and for the intraluminal specific or total activities of amylase and maltase. The total intraluminal activity (pool of enzyme) of amylase and maltase doubled between weaning (32 days) and 42 days. At weaning, the specific activity of amylase was similar in the jejunal and ileal segments (8.5 AU x g(-1)). From 32 to 42 days, the specific amylase activity doubled in the jejunum, while it remained steady in the ileum. The intraluminal specific maltase activity did not significantly change between 32 and 42 days, while it was twice as high in the jejunum compared to the ileum (3.5 versus 1.8 micromoles hydrolysed maltose x g(-1), respectively).

Expression of multiple complex polysaccharide-degrading enzyme systems by marine bacterium strain 2-40

Abstract: Saprophytic marine bacterium strain 2-40 (2-40) can degrade numerous complex polysaccharides (CP) including agar, alginic acid, carrageenan, carboxymethylcellulose, chitin, beta-glucan, laminarin, pectin, pullulan, starch, and xylan. The growth of 2-40 was assessed in minimal media containing one of 16 CP or simple carbohydrates, with the result that all supported growth. Each of the carbohydrase systems was elicited at highest levels by the homologous substrate. Each, excluding amylase, was repressed when 2-40 was cultured in glucose minimal synthetic media. Cyclic adenosine monophosphate alleviated the repression. Agarose as sole carbon source supported the synthesis of the most heterologous complex carbohydrase systems, although, generally, at a lower level of activity than the homologous CP.

Enzymatic Degradation of Epichlorohydrin Crosslinked Starch Microspheres by α-Amylase

Abstract: Purpose. The influence of chemical parameters on the sensitivity to enzymatic degradation by α-amylase of starch microspheres cross-linked by epichlorohydrin was studied.

Morphological Change, Sugar Content, and ɑ-amylase Activity of Rice Seeds under Various Priming Conditions

Abstract: An experiment was carried out to find out the changes in morphology, sugars, and -amylase activity during the priming of rice seeds (Oryza sativa L. cv. ＇Ilpumbyeo＇). For priming, seeds were soaked in -0.6 MPa PEG solution at 15 for 4 days (properly primed) and at for 4 and 10 days (over-primed) and dried at room temperature. The size of coleoptile and differentiated leaves of properly primed seeds were bigger and coleoptile was separated from the other part of embryo compared with non-primed and over-primed seeds. As priming of seeds advanced, compound starch grains in the endosperm disintegrated into tiny starch granules, and small holes were found in the tiny starch granules and a cavities developed between embryo and endosperm. The radicle and plumule of properly primed germinating seeds developed faster than non-primed and overprimed germinating seeds. Sucrose, maltose, and raffinose contents of properly primed seeds decreased, while content of glucose and fructose and -amylase activity increased. However, sugar content and -amylase activity of over-primed seeds were lower compared with non-primed seeds or properly primed seeds.

Studies on catabolite repression in solid state fermentation for biosynthesis of fungal amylases

Abstract: Catabolite repression by glucose of the biosynthesis of alpha amylase and amyloglucosidase by Aspergillus niger CFTRI 1105 was studied in a solid state fermentation (SSF) and in submerged fermentation (SMF) systems and the results were compared. The addition of glucose did not enhance the production of alpha-amylase and amyloglucosidase in an earlier fermentation system. However, a drastic reduction in alphaamylase production was observed in submerged fermentation by the addition of 5·0 mg ml-1 glucose and of amyloglucosidase production by 10 mg ml-1 glucose. Glucose concentrations above 50 mg ml-1 completely suppressed the production of both enzymes in the initial hours. In contrast, in the SSF system the repression was negligible, even when the glucose level was raised to 150 mg g�1 wheat bran for both alpha and amyloglucosidase synthesis.