Showing papers on "Amylase published in 1979"
TL;DR: In this article, the authors explored completeness of starch hydrolysis in situ in relation to degree of gelatinization, starch content of tissue, evailable enzyme activity, and time allowed for hydrolyisation.
Abstract: This work explored completeness of starch hydrolysis in situ in relation to degree of gelatinization, starch content of tissue, evailable enzyme activity, and time allowed for hydrolysis. Maximum hydrolysis of starch in lyophilized red oak (Quercus rubra L.) root tissue with purified Diazyme (amyloglucosidase) or Clarase (Takadiastase) required high enzyme activity (2.4 U Diazyme or 48 U Clarase per mg starch). Results suggested that at least 70 U Clarase or 5 U Diazyme should be used per mg starch in routine analyses. Neither prolonging gelatinization (more than 15 min) nor hydrolysis (more than 24 to 48 lh) offset inadequate starch hydrolysis caused by insufficient enzyme activity. Starch was completely hydrolyzed in situ after 48 h without gelatinization by 5 U of Diazyme per mg starch. Tissue weight (5 to 100 mg) had no effect on starch hydrolysis by sufficient enzyme. Methanol: chloroform: water (12:5:3 by volume) freed tissues of solubles before starch hydrolysis. No interference with the glucose oxidase analysis of hydrolysates was encountered. In addition, the pigment free methanol–water fractions (soluble sugars, amino acids, organic acids) and chloroform fractions (lipids and pigments) were available or further analysis. Results obtained with red oak were verified with issue from other species such as jack pine (Pinus banksiana lamb.) and white spruce (Picea glauca (Moench) Voss). The resulting technique simply and reliably measured less than 5% starch in 5 mg lyophilized tissue, with a minimum of sample manipulation.
240 citations
TL;DR: The time sequence analysis of the starch digestion pattern of the thin sectioned germinating rice (Oryza sativa L.) seed specimens using the starch film method showed that at the initial stage amylase activity was almost exclusively localized in the epithelium septum between the scutellum and endosperm.
Abstract: The time sequence analysis of the starch digestion pattern of the thin sectioned germinating rice (Oryza sativa L) seed specimens using the starch film method showed that at the initial stage amylase activity was almost exclusively localized in the epithelium septum between the scutellum and endosperm Starch breakdown in the endosperm tissues began afterward; amylase activity in the aleurone layers was detectable only after 2 days Polyacrylamide gel electrofocusing (pH 4 to 6) revealed nearly the same zymogram patterns between endosperm and scutellum extracts, although additional amylase bands appeared in the endosperm extracts at later germination stages (4 to 6 days) These are presumably attributable to the newly synthesized enzyme molecules in the aleurone cells
171 citations
TL;DR: The subcellular localization of the starch biosynthetic and degradative enzymes of spinach leaves was carried out by measuring the distribution of the enzymes in a crude chloroplast pellet and soluble protein fraction and by the separation on sucrose density gradients of intact organelles, chloroplasts, peroxisomes, and mitochondria of a protoplast lysate.
Abstract: The subcellular localization of the starch biosynthetic and degradative enzymes of spinach leaves was carried out by measuring the distribution of the enzymes in a crude chloroplast pellet and soluble protein fraction, and by the separation on sucrose density gradients of intact organelles, chloroplasts, peroxisomes, and mitochondria of a protoplast lysate. ADP-Glucose pyrophosphorylase, starch synthase, and starch-branching enzymes are quantitatively associated with the chloroplasts. The starch degradative enzymes amylase, R-enzyme (debranching activity), phosphorylase, and D-enzyme (transglycosylase) are observed both in the chloroplast and soluble protein fractions, the bulk of the degradative enzyme activities reside in the latter fraction. Chromatography of a chloroplast extract on diethylaminoethyl-cellulose resolves the R- and D-enzymes from amylase and phosphorylase activities although the two latter enzyme activities coeluted. The digestion pattern of amylase with amylopectin as a substrate indicates an endolytic activity but displays properties unlike the typical α-amylase as isolated from endosperm tissue.
129 citations
01 Sep 1979
TL;DR: A strain of the yeast Lipomyces kononenkoae, which converted starch into SCP with a high yield, produced three extracellular amylases which were purified from the culture fluid by Ficoll concentration, dialysis, isopropanol precipitation and DE-cellulose chromatography: an α-amylase, a glucoamylases and a debranching transferase.
Abstract: A strain of the yeast Lipomyces kononenkoae which converted starch into SCP with a high yield, produced three extracellular amylases which were purified from the culture fluid by Ficoll concentration, dialysis, isopropanol precipitation and DE-cellulose chromatography: an α-amylase, a glucoamylase and a debranching transferase. The latter transferred α-1,6-glucosyl units from panose to glucose forming maltose and appeared to have some debranching activity on amylopectin. The α-amylase had the following properties: MW 38000 daltons; no effect of added calcium ions on activity; optimum temperature and pH for activity around 40°C and pH 5.5; ΔH‡ and ΔS‡ of heat inactivation 24360 cal mol−1 and 29.2 cal deg−1 mol−1; range of pH stability pH 4–6.5; pI=7.1; final low molecular weight products of starch hydrolysis, maltose and glucose; Km (40°C, pH 5.5) for starch 2.7 gl−1, for maltotriose 109 gl−1; uncompetitive inhibition by maltose with Ki (40°C, pH 5.5) 29.5 gl−1. The glucoamylase had the following properties: MW 81500 daltons; optimum temperature and pH for activity around 50°C and pH 4.5: ΔH‡ and ΔS‡ of heat inactivation 20400 cal mol−1 and 17.7 cal deg−1 mol−1; range of pH stability pH 4–6.5; pI=6.1; Km (30°C, pH 4.5) for soluble starch 16.2 gl−1, for maltose 0.36 gl−1, for p-nitrophenyl-α-D-glucoside 0.35 gl−1; competitive inhibition by glucose with Ki (30°C, pH 4.5) 4.7 gl−1.
96 citations
94 citations
TL;DR: The attaching ability of a number of bacteria, isolated from bovine rumen and animal intestine, to starch granules was examined in vitro and the attachment of bacteria to starch was inhibited by various carbohydrates.
Abstract: The attaching ability of a number of bacteria, isolated from bovine rumen and animal intestine, to starch granules was examined in vitro. Among bacteria examined in the present investigation, strains possessing the attaching ability to starch were those belonging to an untypable Bacteroides sp., Bacteroides amylophilus, Bacteroides oralis, Bacteroides ruminicola ss. ruminicola, an untypable Bifidobacterium sp., Bifidobacterium thermophilum, Bifidobacterium pseudolongum, and Bifidobacterium longum.The reaction temperature had a profound effect on the attachment of bacteria to starch. All of bacteria listed above attached well to starch at 38°. Strains of the untypable Bacteroides sp., B. oralis, B. thermophilum, B. pseudolongum, and B. longum attached to starch even at the reaction temperature of 4°. On the other hand, those of B. amylophilus did not attach to starch at 4°.The activity of several hydrolases of various kinds of bacteria was examined. The amylase activity of cells of bacteria capable of attaching to starch was remarkably high, compared with that of bacteria incapable of attaching. Bacteria such as B. fibrisolvens and Streptococcus bovis, which were incapable of attaching to starch but possessed intense amylolytic activity, secreted extracellularly large parts of amylase produced by them. Only Peptostreptococcus productus among examined bacteria exhibited remarkably intense urease activity. There was no significant difference in the specific activity of the remaining three hydrolases between bacteria capable and incapable of attaching to starch. The attachment of bacteria to starch was inhibited by various carbohydrates. Carbohydrates like dextrin, amylose, amylopectin, soluble starch, and derivatives of starch except carboxymethyl-starch inhibited the attachment of bacteria to starch. The degree of this inhibition by these carbohydrates depended on their concentrations. Other carbohydrates examined, including derivatives of cellulose and dextran, did not inhibit significantly the attachment of bacteria to starch.An attempt was made to detach or elute bacteria from starch by some means. Cells of B. amylophilus, once attached to starch at 38°, were detached by the cooling treatment at 4°. Although both cells of B. thermophilum and B. pseudolongum were not detached from starch by the cooling treatment at 4°, they were eluted from starch with a solution of dextrin or partially-hydrolysed hydroxypropyl-starch. Cells of Bacteroides sp. 7-4, once attached to starch, were not detached either by the cooling treatment at 4° or by the elution treatment with a solution of above two carbohydrates.
86 citations
TL;DR: A moderate halophile, Micrococcus halobius ATCC 21727, produced an extracellular dextrinogenic amylase when cultivated in media containing 1 to 3 M NaCl, and the products were maltose, maltotriose, and maltotetraose, with lesser amount of glucose.
Abstract: A moderate halophile, Micrococcus halobius ATCC 21727, produced an extracellular dextrinogenic amylase when cultivated in media containing 1 to 3 M NaCl. The amylase was purified from the culture filtrate to an electrophoretically homogenous state by glycogen-complex formation, diethylaminoethyl-cellulose chromatography, and Bio-Gel P-200 gel filtration. The enzyme had maximal activity at pH 6 to 7 in 0.25 M NaCl or 0.75 M KCl at 50 to 55 degrees C. The activity was lost by dialysis against distilled water. Molecular weight was estimated to be 89,000 by sodium dodecyl sulfate-gel electrophoresis. The action pattern on amylose, soluble starch, and glycogen showed that the products were maltose, maltotriose, and maltotetraose, with lesser amount of glucose.
59 citations
TL;DR: Comparison of in vitro translation products of RNAs isolated from embryonic and adult pancreases suggested that the developing exocrine pancreas undergoes at least two differentiative tran- tion events, and the rate of amylase synthe- sis during this developmental period is controlled in large part by the rateof accumulation of amelase mRNA.
58 citations
Journal Article•
TL;DR: It seemed as it trypsin but not amylase was involved in the suppression exerted by pancreatic juice on exocrine pancreatic secretion in the pig.
Abstract: Pigs were surgically prepared with external pancreatic fistulae and duodenal cannulae in order to elucidate whether the proposed intestinal feedback control of pancreatic secretion in the pig--as in rat and man--is exerted by trypsin. Furthermore, the effect of intraluminal amylase on pancreatic secretion was studied. Reintroduction of pancreatic juice into the duodenum or infusion of trypsin into the duodenum depressed the volume of the pancreatic flow and the protein output markedly. Introduction of amylase into the duodenum did not significantly affect the pancreatic secretion. Thus, it seemed as it trypsin but not amylase was involved in the suppression exerted by pancreatic juice on exocrine pancreatic secretion in the pig.
56 citations
TL;DR: Investigation of pure human pancreatic juice obtained by direct cannulation of the main pancreatic duct of 11 healthy volunteer subjects and 10 chronic alcoholics without detectable pancreatic disease revealed the presence of numerous acid hydrolases in this secretion, suggesting they are of lysosomal origin.
Abstract: Investigation of pure human pancreatic juice obtained by direct cannulation of the main pancreatic duct of 11 healthy volunteer subjects and 10 chronic alcoholics without detectable pancreatic disease revealed the presence of numerous acid hydrolases in this secretion. The pH optima and substrate specificities of these enzymes suggest that they are of lysosomal origin. Stimulation of the pancreas by injection of cholecystokinin-pancreozymin (CCK-PZ) (1 Ivy dog unit/kg) resulted in a striking increase in activity of some of these hydrolases (N-acetyl-β-d-glucosaminidase, arylsulfatase, etc.) similar to that observed for trypsin, amylase, and other pancreatic digestive enzymes. In a second group of hydrolases (β-d-glucuronidase, leucine naphthylamidase, etc.) the effect of this hormone was greatly reduced or absent, particularly in normal individuals. In chronic alcoholics enzyme activity in response to CCK-PZ injection was greater than in normal subjects. Although this increase achieved statistical significance (P<0.05) in the case of β-d-glucuronidase only, it was observed for all lysosomal hydrolases tested and suggests either increased synthesis or a more facile release of these enzymes from the pancreas of chronic alcoholics than of normal individuals.
TL;DR: In this article, electron microscopy (SEM) scans were performed on starch granules of banana, lily, and lotus attacked by pancreatin and showed that the granules exhibited a terraced or step-shaped apperance in their inner portions.
TL;DR: Polyacrylamide gel electrophoresis was used to compare the enzymic activities of the bacteria in the intestine of gray mullet, Mugil cephalus, an amphidromous fish with an undeveloped stomach, to find that most of the isolates possessed strong proteolytic activity.
Abstract: Polyacrylamide gel electrophoresis was used to compare the enzymic activities of the bacteria in the intestine of gray mullet, Mugil cephalus, an amphidromous fish with an undeveloped stomach. These bacteria may help the digestion of high molecular weight compounds in the intestine. It was found that most of the isolates possessed strong proteolytic activity, in which the gelatinolytic activity was stronger than the caseinolytic activity. Almost all Vibrio and Enterobacter isolated from the intestine possessed strong protease and amylase isozymal electrophoretic bands. Clear differences between the protease and amylase isozyme zymograms were noticed with Vibrio, Entero-bacter, and the other isolates. Some of the isolates possessed chitinase and lecithinase activities, but none of the isolates tested showed these activity bands by polyacrylamide gel electrophoresis. All the active patterns obtained from Vibrio and Enterobacter showed more and stronger bands than those from the other isolates. On the other hand, the mullet intestine possessed a detectable level of proteolytic activity, but it did not show any chitinase or amylase activity. The enzymic activities in the digestion of high molecular weight compounds were discussed.
TL;DR: The results suggest that substance P may act directly on the parotid cells, and indicate that vasopressin, angiotensin II and neurotensin inhibit the action of substance P on salivation at sites other than the parOTid cells.
Patent•
13 Nov 1979
TL;DR: Heat and acid-stable alpha-amylase enzymes have been shown to retain at least about 70% of their initial activity when held at 90°C and at a pH of 60 for 10 minutes in the absence of calcium ion as mentioned in this paper.
Abstract: Heat and acid-stable alpha-amylase enzymes having the following characteristics: (1) capable of retaining at least about 70% of their initial activity when held at 90° C and at a pH of 60 for 10 minutes in the absence of calcium ion; (2) capable of retaining at least about 50% of their initial activity when held at 90° C at a pH of 60 for 60 minutes in the absence of added calcium ion; and/or (3) capable of retaining at least about 50% of their initial activity at a temperature of 80° C and at a pH of 45 in the presence of 5 mM calcium ion for 10 minutes The preferred alpha-amylases are prepared by culturing a strain of a Bacillus stearothermophilus microorganism in a suitable culture medium The novel alpha-amylases are useful in hydrolyzing and/or liquefying starch and due to their stability at low pH values they can be used in conjunction with other acid stable amylases such as gluco-amylase in either a soluble or an immobilized form
TL;DR: The Bacillus polymyxa strain No. 72 was found to produce 45 units of β-amylase per ml in the medium consisting of 4% soluble starch, 1% meat extract and 1% peptone and 0.3% NaCl (pH 7.0) at 30°C as mentioned in this paper.
Abstract: β-Amylase producing microorganisms were effectively isolated by using an amylase inhibitor (S–AI). Among fortytwo strains isolated, strain No. 72 was found to be the most potent β-amylase producing microorganism, and was identified as Bacillus polymyxa. This strain produced 45 units of β-amylase per ml in the medium consisting of 4% soluble starch, 1% meat extract, 1% peptone and 0.3% NaCl (pH 7.0) at 30°C. Two kinds of β-amylase, that is, 7.3 mg of β-amylase I and 3.9 mg of β-amylase II, were obtained from one liter of culture broth as electrophoretically homogeneous forms. β-Amylases I and II were very similar to each other in their enzymatic properties except the small difference in isoelectric point (I; pH 8.35, II; pH 8.59). β-Amylases I and II were most active at pH 7.5 at 45°C, and stable between pH 4 and 9 for 15 hr at 37°C. Both enzymes were strongly inhibited by PCMB, and reactivated by cysteine. The molecular weights of β-amylase I and II were estimated to be about 44,000. The amino acid compos...
TL;DR: The relatively high conversion rate of starch, and the hydrolysis of cross-linked blue starch by this amylase indicate that the enzyme can cleave its substrates also by an endomechanism.
TL;DR: The mechanism of starch degradation by the yeast Lipomyces starkeyi was studied and it was shown that the amylolytic system is associated with cell walls and is equally well induced by starch or maltose.
Abstract: The mechanism of starch degradation by the yeast Lipomyces starkeyi was studied in strain CBS 1807. It was shown that the amylolytic system is associated with cell walls. It is equally well induced by starch or maltose. The enzyme exhibits considerable α amylase type activity and also liberates a small amount of glucose. Enzyme synthesis occurs during the exponential growth phase. The regulation of the synthesis of the enzyme is discussed.
TL;DR: In cases of effusion with unexplained amylase elevations, the possibility of serous ovarian neoplasia should be considered, even in the absence of demonstrable extra‐ovarian dissemination.
Abstract: Elevated amylase activity was noted in the pleural effusion of a patient who was later found to have a stage I low-grade serous papillary ovarian neoplasm. The effusion resolved spontaneously after resection of the ovarian tumor, which contained large amounts of amylase activity. The ultrastructure of the tumor epithelium resembled that of normal salpinx. Secretory-type cells were present with apical, variably electron-dense secretory granules and cytoplasmic glycogen deposits. Amylase activity in the ovarian neoplasm probably resulted from the presence of functioning endosalpingeal-type epithelium in the tumor. In cases of effusion with unexplained amylase elevations, the possibility of serous ovarian neoplasia should be considered, even in the absence of demonstrable extra-ovarian dissemination.
TL;DR: It has been demonstrated that beta-amylase is uniformly associated with the periphery of starch granules in the starchy endosperm cells.
Abstract: Rabbit antiserum against β-amylase isolated from germinating seeds of rice was produced, and its specific cross-reactivity with β-amylase was confirmed by means of Ouchterlony double immunodiffusion and immunoelectrophoresis procedures. The cellular localization of β-amylase was studied by indirect fluorescence microscopy of thin sectioned germinating rice seed specimens (1-day stage) which had been fixed and treated with purified rabbit anti-β-amylase immunoglobulin G followed by conjugation with fluorescein isothiocyanate-labeled goat antirabbit immunoglobulin G. It has been demonstrated that β-amylase is uniformly associated with the periphery of starch granules in the starchy endosperm cells. The finding is discussed in relation to the general notion concerning the presence of the latent form of β-amylase bound to protein bodies in cereal seeds.
TL;DR: The quantitative effect is the result of an increase in the rate of synthesis of YBR-amylase, and the determinant is cis acting, and studies of other mouse strains suggest that regulatory genetic elements may modulate salivary amylase production.
Abstract: Heterozygotes from matings of the mouse strains YBR/Cv and C3H/As have about 3 times more YBR-amylase than C3H-amylase in the saliva. The determinant for this quantitative effect is located on linkage group XVI close to or within the structural gene for salivary amylase. The quantitative effect is the result of an increase in the rate of synthesis of YBR-amylase, and the determinant is cis acting. Studies of other mouse strains suggest that regulatory genetic elements may modulate salivary amylase production.
TL;DR: The assay was shown to be sensitive, reproducible and specific, but human pancreatic amylase and salivary amylases could not be distinguished by the antiserum used, but the rise in enzymatic activity was not accompanied by increasing amount of amyl enzyme content.
TL;DR: The dependence of the insulin effect upon diet composition demonstrates that the rate of glucose metabolism is the primary factor in the regulation of amylase to lipase ratio.
Abstract: In man and in rat, the diabetic state is associated with diseases of exocrine pancreatic function. In this work, streptozotocin diabetes was shown to lead to a 95% decrease in the amylase to lipase ra
TL;DR: Responsibility to carbachol and CCK-OP is a maturational process in the pancreas that lags behind the development of intracellular processes involved in stimulus-secretion coupling.
Abstract: Mechanisms and development of secretory function were studied in rat pancreas in vitro. Amylase release from term fetal pancreas was refractory to stimulation by carbamylcholine chloride (carbachol) and cholecystokinin-octapeptide (CCK-OP), but was significantly augmented by calcium ionophore (A23187), DBcAMP, 8-Br-cGMP, and theophylline. The latter agent when combined with either cyclic nucleotide analogue further increased secretory responses. At 1 day and 8 days postnatally, responsiveness to carbachol and CCK-OP had been acquired because amylase secretion stimulated by these agents was brisk and at a level comparable to that found in mature tissue. Increasing extracellular calcium concentrations from 1.23 to 5.28 mM had no effect on basal amylase release in either the fetal or 8-day pancreas. No changes in intracellular cAMP concentrations were found at any age under experimental conditions used. Similarily, in fetal tissue, no changes in cGMP concentrations were found in response to carbachol or A23187. However, at 8 days of age, both agents produced two- to four-fold increases in tissue cGMP levels at 1, 2, and 5 min of incubation. These studies confirm that responsiveness to carbachol and CCK-OP is a maturational process in the pancreas that lags behind the development of intracellular processes involved in stimulus-secretion coupling.
TL;DR: In dispersed acini from guinea pig pancreas, adding extracellular manganese increased amylase release and appeared to result from actions ofManganese on one of the later steps in the mechanisms for stimulating the secretory process.
Abstract: In dispersed acini from guinea pig pancreas, adding extracellular manganese increased amylase release. A significant effect could be detected with 0.25 mM manganese, and maximal stimulation occurred with 1 mM manganese. When manganese was added, the rate of amylase release did not change during the first 20 min of incubation and then gradually increased to a new steady state by 80 min, which with 1 mM manganese represented a fourfold increase in the rate of enzyme release. Extracellular manganese inhibited the stimulation of amylase release caused by those secretagogues that mobilize cellular calcium but augmented the stimulation caused by those secretagogues whose actions are mediated by cellular cAMP. The mechanism by which manganese altered stimulated amylase secretion differed from the mechanism by which manganese stimulated basal amylase release because the change in stimulated release was maximal within 10 min, whereas the change in basal release did not occur until after 20 min. The actions of manganese on secretagogue-stimulated amylase release were not attributable to manganese-induced changes in secretagogue-stimulated calcium outflux or cAMP and, instead, appear to result from actions of manganese on one of the later steps in the mechanisms for stimulating the secretory process.
TL;DR: In anaesthetized rabbits amylase secretion from the parotid gland was investigated and evoked by sympathetic nerve stimulation, either alone or superimposed on a parasympathetic background secretion, imitating the resting secretion present in the waking animal.
Abstract: 1. In anaesthetized rabbits amylase secretion from the parotid gland was investigated. Secretion was evoked by sympathetic nerve stimulation, either alone or superimposed on a parasympathetic background secretion, imitating the resting secretion present in the waking animal.
2. Sympathetic nerve stimulation at frequencies below 1 Hz was alone subthreshold for fluid secretion, but could greatly increase the amounts of amylase present in fluid secretion produced by parasympathetic nerve stimulation. The amylase output due to sympathetic nerve stimulation alone at 10 Hz did not exceed that seen in response to a stimulation at 1 Hz superimposed on parasympathetic activity.
3. The amylase output in response to superimposed sympathetic stimulation was not influenced by the rate of fluid secretion, which was altered by stimulating the parasympathetic nerves at different frequencies.
4. Sympathetically-evoked amylase secretion was abolished after β1-block. The amylase secretion remaining on parasympathetic activation was sparse.
5. It is concluded that secretion of amylase in response to sympathetic nerve stimulation requires the presence of a parasympathetic fluid secretion to be washed along the glandular ducts. Parasympathetic activity may also augment the sympathetic effect on amylase secretion.
TL;DR: Breakdown and metabolism of both glucomannan (Salep-Mannan) and starch were studied in tubers of Orchis morio L. during one vegetation period and enzymes responsible for mobilisation of the reserve polysaccharides were shown to be β-mannase, amylase,β-mannosidase, β-glucOSidase and α- glucosid enzyme.
Abstract: Breakdown and metabolism of both glucomannan (Salep-Mannan) and starch were studied in tubers of Orchis morio L. during one vegetation period. Variations in the amount of reducing sugars and sucrose, which resulted from the metabolisation process, were analysed during the same period.
Enzymes responsible for mobilisation of the reserve polysaccharides were shown to be β-mannase, amylase, β-mannosidase, β-glucosidase and α-glucosidase. Variations in the different enzyme activities were measured at the same time intervals.
TL;DR: 1)NA-468アミラーゼ生産菌は胞子,胞zi形成 菌糸の形状,気�o�alityの色,炭素源の同化性等の結果からStreptomyces griseusと同定された.
Abstract: 放線菌アミラーゼの検索を行い,マルトトリオース生成アミラーゼを産生する菌株を見い出したのでその内の1菌株について同定を行い,アミラーゼの精製と酵素的性質について検討を行った.1)NA-468アミラーゼ生産菌は胞子,胞子形成菌糸の形状,気菌糸の色,炭素源の同化性等の結果からStreptomyces griseusと同定された. 2)NA-468アミラーゼは培養炉液より約100倍精製され,ディスク電気泳動的に単一であった.比活性は測定温度40℃ の時5,450u/mgproteinであった.分子量またはサブユニットの分子量はSDS電気泳動法から約55,000と推定された. 3)反応の最適温度,pHはそれぞれ45℃,pH5.6-6.0であった.pH安定性は3.5-6.5の間で最高活性値の80%以上の活性が残存した.また熱安定性では45℃から急激に失活し始め,55℃ で完全に失活した. 4)金属イオンの影響はLi+が活性化に,Hg2+,Cu+,Zn2+が阻害的な効果を示した. 5)NA-468アミラーゼはプルラ.ン,β-シクロデキストリン,ワキシコーンスターチβ-限界デキストリγには作用せず,短鎖アミロースで最高の水解速度を示した.また分岐度の高い基質ほど作用し難い傾向が見られた.水解限度は短鎖アミロース,可溶性澱粉,ワキシコーンスターチの場合それぞれ約100 ,55,51%であった. 6)ペーパークロマトを用いて経時的反応生成物を調べた結果,可溶澱粉からはマルトトリオースのみが生じた.また2次元ペーパーク卩マトの結果からマルトテトラオースが最小基質であり,それ以上のマルトオリゴ糖末端から3番目のグルコシド結合を規則的に水解する作用を有することが観察された. 以上の諸結果からNA-468アミラーゼはいわゆるexo型アミラーゼに共通する特徴を有し,β-アミラーゼ,グルコアミラーゼ等と同様に分類されるのではないかと推測された.
TL;DR: By stepwise introduction of these and other genetic elements into B. subtilis 6160 by transformation and mutation, strains with higher α-amylase producing activity were obtained and the finally obtained strain, T2N26, produced about 1,500-2,000 times more α-Amylase than parental 6160 strain.
Abstract: The production of extracellular α-amylase in Bacillus subtilis is probably regulated by many genetic elements, such as amyR, tmrA7, pap, amyB and sacU. Additional genetic elements, C-108 and A-2 for production of the α-amylase were found in D-cycloserine and ampicillin resistant mutants (C108 and A2) of B. subtilis 6160, respectively. Strain C108 increased the production of α-amylase about 5 times and protease about 80 times compared to parental 6160 strain. Strain A2 showed a nearly 6-fold increased α-amylase production.These genetic elements displayed a synergistic effect with other genetic factors in production of extracellular α-amylase when these elements were transferred by DNA mediated transformation. By stepwise introduction of these and other genetic elements into B. subtilis 6160 by transformation and mutation, strains with higher α-amylase producing activity were obtained. The finally obtained strain, T2N26, produced about 1,500-2,000 times more α-amylase than parental 6160 strain.
TL;DR: Variations in amylase and trypsin activities were studied in Artemia salina L. exposed to concentrations of copper or zinc which impair their growth, and the possibility of growth disturbance by modified development of digestive enzymes is discussed.
Abstract: Variations in amylase and trypsin activities were studied in Artemia salina L. exposed to concentrations of copper or zinc which impair their growth. Disturbances in the levels of these enzymes were compared with decreases in growth rate. Amylase activity was disturbed within 24 h by 2 ppm copper and within 72 h by 5 ppm zinc. Inversely, trypsin activity was disturbed within 24 h by 5 ppm zinc and within 72 h by 2 ppm copper. Disturbances at the enzymatic level occurred more rapidly than decreases in growth rate, except for trypsin in A. salina polluted by copper. In the latter case, both were affected simultaneously. The possibility of growth disturbance by modified development of digestive enzymes is discussed. The possible use of enzymatic variation for rapid detection of sublethal effects of pollutants on aquatic organisms in situ is suggested. However, suitable controls would be necessary. Either polluted and unpolluted areas could be compared if their natural ecology were already well known, or reference areas could be monitored before pollutants are introduced.