Cinnamate-4-Hydroxyiase in Polyporus hispidus

1973 ◽  
Vol 51 (6) ◽  
pp. 731-734 ◽  
Author(s):  
C. P. Vance ◽  
A. M. D. Nambudiri ◽  
G. H. N. Towers
Keyword(s):  
Enzyme Activity ◽  
Enzymatic Activity ◽  
Microsomal Fraction ◽  
Mitochondrial Fraction ◽  
Maximum Activity ◽  
Coumaric Acid ◽  
Free System ◽  
Optimum Ph ◽  
A Cell ◽  
Plant Enzyme

A cell-free system capable of hydroxylating cinnamic acid to p-coumaric acid has been isolated from 10-day-oid cultures of Polyporus hispidus. The enzyme requires NADPH and FAD for maximum activity. Enzyme activity appears to be localized in the microsomal fraction with slight activity occurring in the mitochondrial fraction. The optimum pH for enzymatic activity is 7.5. This is the first report on any properties of this enzyme in a fungus. The enzyme differs from the known plant enzyme in its FAD requirement.

Synthesis of β-glucan by cell-free extracts of Aureobasidium pullulans

1987 ◽  
Vol 33 (2) ◽  
pp. 123-127 ◽  
Author(s):  
Malcolm A. J. Finkelman ◽  
Alex Vardanis
Keyword(s):  
Enzyme Activity ◽  
Acetic Acid ◽  
Bovine Serum Albumin ◽  
Aureobasidium Pullulans ◽  
Enzyme Preparation ◽  
Glucanase Activity ◽  
Free System ◽  
Enzyme Preparations ◽  
Partial Activation ◽  
A Cell

A cell-free system catalysing the synthesis of β-glucan from uridine-diphosphoglucose was prepared from Aureobasidium pullulans. The activity was stable in the presence of 1 M sucrose and 10 mM MgSO4. The polymer produced was insoluble in H2O or acetic acid (0.5 M) and soluble in NaOH (0.5 M). Several enzyme preparations containing β-glucanase activity degraded the polymer to various extents. Synthesis of the polymer was enhanced by the presence of cellobiose and bovine serum albumin, but not by NaF or ATP. A Lineweaver–Burke plot of enzyme activity versus substrate concentration revealed biphasic kinetics. The enzyme preparation was subject to partial activation by trypsin and chymotrypsin.

scholarly journals The secretion of tropoelastin by chick-embryo artery cells

1985 ◽  
Vol 230 (1) ◽  
pp. 217-225 ◽  
Author(s):  
N A Saunders ◽  
M E Grant
Keyword(s):  
Plasma Membrane ◽  
Chick Embryo ◽  
Microsomal Fraction ◽  
Single Gene ◽  
Subcellular Fractions ◽  
Secretory Vesicles ◽  
Half Time ◽  
Free System ◽  
Close Relationship ◽  
A Cell

Chymotryptic fingerprint analyses of tropoelastin a and tropoelastin b demonstrated a very close relationship between these two polypeptides synthesized in a cell-free system under the direction of chick-embryo polyribosomal mRNA. A similar study on tropoelastin polypeptides extracted in their hydroxylated and under-hydroxylated forms from artery cells incubated with [3H]valine in the absence and presence of alpha alpha'-bipyridine or 3,4-dehydroproline confirmed this close relationship and suggested that tropoelastins a and b are likely to be the products of a single gene. Pulse-chase experiments in which the synthesis and secretion of tropoelastin by artery cells were monitored demonstrated that, after a pulse with [3H]proline, the polypeptides rapidly appeared in the medium and the half-time of tropoelastin secretion was approx. 30 min. Further pulse-chase studies, in which [3H]tropoelastin contents of subcellular fractions were determined, showed that rough and smooth microsomal fractions contained maximal amounts of tropoelastin at different times. The quantity of tropoelastin in the smooth-microsomal fraction was always only a small proportion of that in the rough-microsomal fraction, suggesting rapid translocation of the polypeptides to the plasma membrane. Incubation of the cells with 0.1 mM-colchicine did not markedly alter the rate of secretion or the distribution of tropoelastin between the subcellular fractions, whereas when 1 microM-monensin was included in the incubations the polypeptides were retained in the rough microsomal fraction. The results are consistent with the proposal that tropoelastin may follow a pathway of secretion from rough endoplasmic reticulum to the plasma membrane via secretory vesicles.

scholarly journals Purification and characterization of amylase from local isolate Pseudomonas sp.SPH4

2012 ◽  
Vol 6 (1) ◽  
pp. 69-79
Author(s):  
Hala M. Ali ◽  
Ghazi M. Aziz
Keyword(s):  
Enzyme Activity ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Silver Ions ◽  
Enzyme Characterization ◽  
Maximum Activity ◽  
Mercury Ions ◽  
Local Isolate ◽  
Optimum Ph

The amylase produced from local isolate Pseudomonas sp. SPH4 was purified by precipitation with 30% saturation ammonium sulphate, followed by ion-exchange chromotography using DEAE-cellulose column, and Gel filtration using Sephacryl S-300 column.The two iso-enzymes (a, b) were purified to (2.83, 3.47) times in the last step with an enzymes yields of (32.36, 76.34)% respectively. Enzyme characterization of the two iso-enzymes indicated that the optimum pH for the two iso-enzymes a and b were (7, 7.5) respectively, while the optimum pH for the iso-enzymes stability were (6.5, 7) respectively. The maximum activity for iso-enzymes (a, b) appeared at 45ºC and stable for 15 min at 30-50ºC and lost approximately 50% of it's activity at rang above 75ºC. Enzyme characterization results showed that the chlorides of silver and mercury had inhibitory effect on enzyme activity, the remaining enzyme activity for the iso-enzymes (a, b) were (46.66, 36.36)% for silver ions and (41.33, 33.63)% for mercury ions at 5 mM respectively, and (28, 28.18)% for silver ions and (25.33, 19.09)% for mercury ions at 10 mM respectively. The iso-enzymes a and b were affected by chelating agent ethylene diamine tetra acetic acid (EDTA) at concentration 2mM the remaining activity (45.33, 43.63)% respectively, and 5mM the remaining activity (28, 28.18)% respectivily, and these iso-enzymes (a, b) refered to metalloenzymes. The iso-enzymes (a, b) were kept their activity when treated by reducing agent (2-mercaptoethanol) at 2 mM the remaining activity (92, 92.72)% respectively, and 5 mM the remaining activity (85.3, 89.09)% respectivily. The iso-enzymes (a, b) were kept their activity when treated by phenyl methyl sulphonyl fluoride (PMSF) at concentration 1mM the remaining activity (93.33, 90.90)% respectivily,and 5 mM the remaining activity (90.66, 87.27)% respectivily, and these indicated that these iso-enzymes didnot referred to serineamylases group.

Methylamine dehydrogenase from the obligate methylotroph Methylomonas methylovora

1977 ◽  
Vol 23 (4) ◽  
pp. 402-406 ◽  
Author(s):  
R. J. Mehta
Keyword(s):  
Enzyme Activity ◽  
Mercuric Chloride ◽  
Isonicotinic Acid ◽  
Acid Hydrazide ◽  
Sodium Borate ◽  
Phenazine Methosulfate ◽  
Methylamine Dehydrogenase ◽  
Obligate Methylotroph ◽  
Free System ◽  
A Cell

An obligate methyltroph Methylomonas methylovora oxidized methylamine, formaldehyde, and formate. Enzymes oxidizing these substrates were detected in a cell-free system. Phenazine methosulfate-linked methylamine dehydrogenese was purified 21-fold. The enzyme had optimum activity at pH 7.5 and was stable at 60 °C for 5 min. The enzyme activity was inhibited by parachloromercuric benzoate, isonicotinic acid hydrazide, mercuric chloride, and sodium borate.

scholarly journals Extraction, precipitation and characterization of urease from Vicia faba L.

2020 ◽  
Vol 31 (1) ◽  
pp. 9
Author(s):  
Dalal Subree Bedan
Keyword(s):  
Enzyme Activity ◽  
Ammonium Sulfate ◽  
Vicia Faba ◽  
Urease Activity ◽  
Enzyme Characterization ◽  
Maximum Activity ◽  
Mineral Salts ◽  
Urease Enzyme ◽  
Vicia Faba L ◽  
Optimum Ph

This study aimed to extract urease enzyme from available plant source which was broad beans (Vicia faba L.) using aqueous solution in a ratio1:3(w:v) .The crude extract appeared enzyme activity 33.3 U/ml. Results of this study revealed the possibility to precipitate urease enzyme after extraction using different precipitation methods consisted of Acetone precipitation; Alcohol precipitation and Ammonium sulfate precipitation, these ways of urease precipitation gave enzyme activities 74.8; 43.6 and 82.2 U/ml respectively .The enzyme precipitation using 60% saturation ratio of Ammonium sulfate gave the maximum activity of urease precipitation among other precipitation methods. Enzyme characterization appeared the optimum pH of urease activity was 8.0 and gave enzyme activity 93.8U/ml, while the optimum pH of urease stability was 6.0; the enzyme maintains its activity. On the other side the optimum temperature of precipitated urease activity was 50°C and the enzyme activity reached 99.3U/ml while the optimal temperature of urease stability was 40°C ,the precipitated urease maintain 100% of activity. The effect of other factors on urease action were studied also, these are consisted of the storage time of enzyme ,urease maintain its activity for 8days and the activity begun to decline after that time; the effect of time reusability was also revealed that the enzyme could be used for six times and maintain its activity. The influence of different mineral salts on precipitated urease were also recorded ;these salts were CuSO4 , MgSO4, MgCl2,KCl,CaCl2and FeSo4, results appeared that magnesium and calcium salts were activator for the precipitated urease while copper ,potassium and ferrous salts were inhibitor of the precipitated urease. Urea concentration as an enzyme substrate were also examined and results appeared the optimal substrate concentration was 30mM; enzyme activity reached to125.2U/ml.

scholarly journals Characterization and Effects of β–Galactosidase in the Crude Extracts of Cuminum cyminum and Curcuma longa in Preparation of Delactosed Milk and Whey

2016 ◽  
Vol 5 (1) ◽  
pp. 1
Author(s):  
Omar M. Atrooz
Keyword(s):  
Enzyme Activity ◽  
Curcuma Longa ◽  
Maximum Activity ◽  
Cow’S Milk ◽  
Enzyme Kinetic ◽  
Optimum Temperature ◽  
Cuminum Cyminum ◽  
Crude Extracts ◽  
Optimum Ph ◽  
Hydrolysis Of

<p>β-galactosidase (EC 3.2.1.23) was extracted from <em>Cuminum cyminum </em>and<em> Curcuma longa</em>. The crude extracts of these plants were then characterized in term of pH, temperature, and enzyme kinetic. The crude extracts were also used in hydrolysis of lactose in milk and whey. The enzyme activity was measured by its ability to hydrolyze the substrate o-nitrophenyl β -D-galactopyranoside (ONPG).</p><p>It was found that β-galactosidase in the crude extracts of <em>Cuminum cyminum </em>exhibited maximum activity at pH 8.0 and optimum temperature at 60 °C. While, β-galactosidase in the crude extracts of <em>Curcuma longa</em> have optimum pH at 5.0 and 7.0 and optimum temperature at 50 °C.The K<sub>m</sub> and V<sub>max</sub> values of the β-galactosidase in the crude extracts of <em>Cuminum cyminum</em> and <em>Curcuma longa </em>were 4.16 mM and 0.087 μmol/min, and 2.63 mM and 0.333μmol/min, respectively.</p><p>The results showed that 96.84-97.08% of lactose was hydrolyzed in cow’s milk and whey when treated with crude extracts of <em>Cuminum cyminum</em> and 90-98.6% when treated with crude extracts of <em>Curcuma longa</em>.</p>

Lipase activity of pigeon heart muscle particulate fractions and its metabolic significance

1963 ◽  
Vol 204 (1) ◽  
pp. 165-167 ◽  
Author(s):  
J. C. George ◽  
P. Thomas Iype
Keyword(s):  
Fatty Acids ◽  
Enzyme Activity ◽  
Lipase Activity ◽  
Heart Muscle ◽  
Microsomal Fraction ◽  
Mitochondrial Fraction ◽  
Hydrolysis Of ◽  
Metabolic Significance

The lipase activity in the different particulate fractions of the pigeon heart muscle was determined manometrically. The microsomal fraction was found to have more of the enzyme activity than the mitochondrial one. The mitochondrial fraction was found to be incapable of oxidizing tributyrin and triolein in vitro. The possible role of the microsomes in the esterification of fatty acids and hydrolysis of fat is discussed.

scholarly journals Asoprisnil, a Selective Progesterone Receptor Modulator (SPRM), Inhibits Melanosome Export in B16F10 Cells and HEMn-DP Melanocytes

Molecules ◽  
2020 ◽  
Vol 25 (16) ◽  
pp. 3581
Author(s):  
Shilpi Goenka ◽  
Sanford R. Simon
Keyword(s):  
Enzyme Activity ◽  
Progesterone Receptor ◽  
Human Keratinocytes ◽  
Tyrosinase Activity ◽  
Free System ◽  
Cell Free System ◽  
Receptor Modulator ◽  
B16f10 Cells ◽  
A Cell ◽  
Tyrosinase Enzyme

Previous studies have reported that estrogen hormone promotes melanogenesis while progesterone inhibits it. A selective estrogen receptor modulator (SERM), tamoxifen, has been shown to promote melanogenesis; however, to date, there have been no reports on the effects of a selective progesterone receptor modulator (SPRM) on melanogenesis. In the present study, we hypothesized that asoprisnil (AP), a SPRM, inhibits melanogenesis. AP was tested for cytotoxicity to B16F10 mouse melanoma cells for screening the nontoxic concentrations using MTS cytotoxicity assay. Extracellular and intracellular melanin levels were estimated at nontoxic concentrations of AP. To evaluate the direct effect of AP on tyrosinase enzyme, tyrosinase activity and copper chelating activities were measured. Next, the effects of AP on melanogenesis were tested in normal human melanocytes, neonatal, darkly pigmented (HEMn-DP). Our results demonstrate that AP was nontoxic at a concentration range of 10–50 μM in B16F10 cells; AP at 50 μM significantly suppressed extracellular melanin levels comparable to kojic acid at 500 μM, with no significant effect on intracellular melanin levels. The mechanism of melanogenesis inhibition was studied to assess if AP downregulated tyrosinase activity in cell lysates or in a cell-free system. However, AP was found to increase intracellular tyrosinase activity without any effect on tyrosinase enzyme activity or copper chelating activity in a cell-free system, indicating that AP inhibits melanogenesis by mechanisms other than direct effects on tyrosinase enzyme activity. The capacity of AP to inhibit melanosome export was further validated in HEMn-DP cells; AP significantly suppressed dendricity at concentrations of 20 and 30 μM in the absence of effects on melanin synthesis or intracellular tyrosinase activity. In addition, AP was nontoxic to human keratinocytes (HaCaT) at these concentrations, validating its safety for topical use. Taken together, our preliminary results demonstrate that AP might be repurposed as a candidate therapeutic for treatment of hyperpigmentation disorders via a unique mechanism, which encompasses a selective inhibition of melanosome export.

scholarly journals A novel Thermothelomyces heterothallicus PA2S4T fungus isolated from the soil induces chitinase production using orange peel flour

2021 ◽  
Vol 17 (9) ◽  
Author(s):  
Victoria Pommer ◽  
Paula Daniela Helfenstein Rother ◽  
Letícia Mara Rasbold ◽  
José Luis Da Conceição Silva ◽  
Alexandre Maller ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Enzymatic Activity ◽  
Carbon Sources ◽  
Orange Peel ◽  
Biotechnological Application ◽  
Optimum Ph ◽  
Chitinase Production ◽  
Stationary Conditions ◽  
Agricultural Industries ◽  
Optimum Ph And Temperature

Chitinases are enzymes capable of hydrolysing the β-1,4 bonds of chitin releasing chitooligosaccharides and N-acetylglucosamine and are widely used in food, pharmaceutical, and agricultural industries. Microorganisms are potential producers of this enzyme; however, there are no reports in the literature on the production of chitinase by fungi of the genus Thermothelomyces. Thus, this work aimed to investigate the production of extracellular chitinase using alternative carbon sources by the fungus isolated from soil, Thermothelomyces heterothallicus PA2S4T. The fungus was cultivated in a liquid medium supplemented with carbon sources and incubated at 40°C under stationary conditions for seven days. Orange peel flour was the best inducer for extracellular chitinase, with 82.3 U/mL of enzymatic activity. The highest production of chitinase was obtained on the tenth day, and the optimum pH and temperature for enzyme activity were 4.5 and 50ºC, respectively. Therefore, the fungus T. heterothallicus PA2S4T proved to be promising in the production of extracellular chitinase, which presents pH and temperature characteristics favourable to biotechnological application.

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