Purification and properties of a phenol carboxylic acid acyl esterase from Aspergillus flavus

1971 ◽  
Vol 17 (11) ◽  
pp. 1455-1463 ◽  
Author(s):  
J. J. Child ◽  
T. Oka ◽  
F. J. Simpson ◽  
H. G. Krishnamurty
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Carboxylic Acid ◽  
Aspergillus Flavus ◽  
Protocatechuic Acid ◽  
Hydroxybenzoic Acid ◽  
Purification And Properties ◽  
Wide Range ◽  
Hydrolysis Of ◽  
Active Preparation

Aspergillus flavus produces an extracellular esterase that hydrolyses phenolic carboxylic acid acyl esters. An assay based upon the measurement of the rate of release of phloroglucinol on hydrolysis of the ester of phloroglucinol and protocatechuic acid is described. The most active preparation hydrolyzed 30.8 μmoles of substrate per minute per milligram of protein and was active against a wide range of esters of meta and para hydroxybenzoic acid derivatives.The enzyme was isolated as a homogeneous protein, as judged by ultracentrifugation and by electrophoresis and had an isoelectric point of pH 4.45. The molecular weight was determined as 166 000. The enzyme is a glycoprotein containing 42.8% carbohydrate and the amino acid composition is described.

Degradation of rutin by Aspergillus flavus. Purification of the dioxygenase, quercetinase

1971 ◽  
Vol 17 (1) ◽  
pp. 111-118 ◽  
Author(s):  
T. Oka ◽  
F. J. Simpson ◽  
J. J. Child ◽  
Sister Cecily Mills
Keyword(s):  
Molecular Weight ◽  
Carbon Monoxide ◽  
Amino Acid ◽  
Carboxylic Acid ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Aspergillus Flavus ◽  
Single Enzyme

Evidence is presented that a single enzyme, quercetinase, is responsible for the degradation of quercetin by Aspergillus flavus to yield carbon monoxide and a depside, 2-protocatechuoylphloroglucinol carboxylic acid. A procedure for the isolation of the dioxygenase as a homogeneous protein is described. The most purified preparation degraded 10 800 μmoles of quercetin/h mg protein and was homogeneous as judged by ultracentrifugation and by electrophoresis. The molecular weight was determined as 111 000 + 4000. Km values for quercetin and oxygen as substrates were 5.2 × 10−6 M and 1.2 × 10−4 M respectively. The enzyme is a glycoprotein containing 27.5% carbohydrate and the amino acid composition is presented.

Purification and properties of glutamate-phenylpyruvate aminotransferase from the ruminal protozoan Entodinium caudatum

2004 ◽  
Vol 55 (9) ◽  
pp. 991
Author(s):  
Md. Ruhul Amin ◽  
Ryoji Onodera ◽  
R. Islam Khan ◽  
R. John Wallace ◽  
C. Jamie Newbold
Keyword(s):  
Amino Acid ◽  
Amino Acid Metabolism ◽  
Acid Metabolism ◽  
Maximum Activity ◽  
Purification And Properties ◽  
Sds Page ◽  
Wide Range ◽  
S 100 ◽  
A Cell

Entodinium species are important in catabolic protein metabolism by the mixed ruminal microbial population. This study was conducted to purify, and investigate properties of one of the enzymes involved in amino acid metabolism by Entodinium caudatum, glutamate-phenylpyruvate aminotransferase (GPA; EC 2.6.1.64). GPA was purified 74-fold from a cell-free extract by ammonium sulfate precipitation and column chromatography with phenyl-superose, DEAE-Toyopearl 650M, Sephacryl S-100 HR, and Sephadex G-100. The molecular mass of GPA was estimated by SDS–PAGE to be 65.0 kDa. The optimum pH was 6.0 and it was found to be reactive over a wide range of pH from 5.0 to 10.5. Maximum activity of GPA occurred at 45°C and the activity declined at temperatures over 55°C. GPA was stable below 60°C. Aminooxyacetate and phenylhydrazine were highly inhibitory, and SDS, EDTA, and some heavy metal ions also inhibited activity. The purification and characterisation of the enzyme will help to isolate the gene and ultimately to understand the role of GPA in both anabolic and catabolic amino acid metabolism by Entodinium caudatum.

scholarly journals Purification and Properties of Staphylocoagulase

1975 ◽  
Author(s):  
A.D. Muller ◽  
B. M. Bas ◽  
H. C. Hemker
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Amino Acid Composition ◽  
Aspartic Acid ◽  
Acid Composition ◽  
Isoelectric Point ◽  
Terminal Amino Acid ◽  
Purification And Properties ◽  
Terminal Amino

Staphylocoagulase, an exoprotein of coagulase positive staphylocoagulase, has been purified to a state in which only trace amounts of contaminating proteins are detectable.Purification was more than 35,000 fold, which is 7 times more than the highest value reported in the literature. The yield was about 15%.Aspartic acid was found as a single N-terminal amino acid in this preparation. The molecular weight is 61,000 and the isoelectric point lies at pH 4.53.The amino acid composition was determined.

scholarly journals Electrophoretic and zymographic techniques for production monitoring of two lipase forms from Candida antarctica DSM 70725

2012 ◽  
Vol 66 (2) ◽  
pp. 201-206
Author(s):  
Aleksandra Dimitrijevic ◽  
Dusan Velickovic ◽  
Ratko Jankov ◽  
Nenad Milosavic
Keyword(s):  
Molecular Weight ◽  
Large Scale ◽  
Fermentation Broth ◽  
Candida Antarctica ◽  
Simple Method ◽  
Wide Range ◽  
Work Production ◽  
Lipase A ◽  
Dimeric Form ◽  
Hydrolysis Of

Yeast Candida antarctica produces two lipase forms, which are widely used as catalysts in variety of organic reactions, many of which are applied on a large scale. In this work, production of two forms of lipase from C. antarctica DSM 70725 (CAL A and CAL B) was monitored during seven days of cultivation in the optimal medium using different electrophoretic and zymographic techniques. According to electrophoresis after silver staining, C. antarctica lipase A (molecular mass 45 kDa) was produced starting from the second day of cultivation. C. antarctica lipase B (CAL B) was also produced starting from the second day, but protein was present in the fermentation broth predominantly as dimer (molecular weight 66 kDa), while presence of monomeric form of CAL B (molecular weight of 33 kDa) was observed starting from the fourth day of cultivation. Both types of zymograms (based on hydrolysis and synthesis reactions) were used for detection of lipase activity in the fermentation broth. C. antarctica lipase A showed activity only in hydrolytic zymogram, when ?-naphtyl butyrate was used as substrate. In the same zymogram, with ?-naphtyl acetate as substrate no CAL A activity was detected. Similarly, CAL A showed no activity in synthesis based zymograms towards oleic acid and octanol as substrates, indicating that CAL A is not active towards very short or long-chain substrates. As opposite of CAL A, both monomeric and dimeric form of CAL B were detected in the all zymograms, suggesting that CAL B is active towards wide range of substrates, regardless to the chain length. Thus, zymogram based on hydrolysis of ?-naphtyl butyrate represents a simple method for monitoring the production of two forms of lipase from C. antarctica, that greatly differ in their characteristics.

scholarly journals The purification and properties of butyryl-coenzyme A dehydrogenase from Peptostreptococcus elsdenii

1971 ◽  
Vol 125 (3) ◽  
pp. 879-887 ◽  
Author(s):  
Paul C. Engel ◽  
V. Massey
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Simple Procedure ◽  
Potassium Bromide ◽  
Prosthetic Group ◽  
Pig Liver ◽  
Peak Absorption ◽  
Purification And Properties ◽  
Green Form ◽  
Spectrophotometric Titrations

Butyryl-CoA dehydrogenase prepared by a simple procedure from Peptostreptococcus elsdenii has a molecular weight of approx. 150000. The enzyme has FAD as its prosthetic group. The amino acid analysis is reported. This enzyme, like most of the corresponding mammalian ones, is green. The absorption band at 710nm can be abolished irreversibly by dithionite reduction and air reoxidation; it can be abolished reversibly by phenylmercuric acetate or potassium bromide. The enzyme as isolated appears to be a mixture of a green and a yellow form, both of which are active. This view is supported by the variable ‘greenness’ of different preparations and the biphasic curve obtained in anaerobic spectrophotometric titrations with dithionite. It can be calculated from the titration results that fully green enzyme would have a peak-to-peak absorption ratio (E710/E430) as great as 0.54. The green form is much less rapidly reduced by dithionite than the yellow form, but is nevertheless much more readily reduced by dithionite than the enzyme from pig liver. It is also more readily reoxidized by air and shows less tendency to form a semiquinone. Treatment with sodium borohydride produces an unusual reduced species that is probably the 3,4-dihydroflavin.

scholarly journals Antioxidant and Functional Activities of MRPs Derived from Different Sugar–Amino Acid Combinations and Reaction Conditions

Antioxidants ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 1840
Author(s):  
David D. Kitts
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Antioxidant Capacity ◽  
Maillard Reaction ◽  
Reducing Sugars ◽  
Food System ◽  
Radical Scavenging ◽  
Trolox Equivalent Antioxidant Capacity ◽  
Maillard Reaction Products ◽  
Wide Range

The Maillard reaction (MR), or non-enzymatic browning, involves reducing sugars reacting with amino acids, peptides, or proteins when heated to produce an abundance of products that contribute to sensory, nutritional, and functional qualities of the food system. One example of an important functional quality of MR relates to antioxidant capacity, which has relevance to preserve food quality and also to extend a potential role that may promote gastrointestinal health. The addition of Alphacel (10%), a non-reactive polysaccharide, to MR reactants produced small significant (p < 0.05) reductions in yield of soluble Maillard reaction products (MRPs), sugar loss, and color change of products formed respectively, for reducing sugars. A similar effect was also noticed for different free-radical scavenging capacity (p < 0.05), using chemical (e.g., 2,2-diphenyl-1-picrylhydrazyl (DPPH)), Trolox equivalent antioxidant capacity (TEAC), and oxygen radical absorbance capacity (ORAC) assays. An inflamed Caco-2 cell model revealed nitric oxide (NO) inhibitory activity for Glu-amino acid MRPs, which contrasted the NO stimulatory activity obtained with Fru-amino acid MRPs, especially when glycine was used as the amino acid. Pre-treating Caco-2 cells with Fru-glycine MRPs protected against loss in trans-epithelial resistance (TEER) (p < 0.05) and reduced (p < 0.05) disruption of Caco-2 intestinal epithelial tight-junction (TJ) protein cells when exposed to 7.5% ethanol. A low molecular weight Fru-glycine (e.g., <1 kDa) fraction contributed to the protective effect, not observed with the corresponding high molecular weight MRP fraction. The presence of Alphacel had minimal effect on generating MRPs with relative modified protection against intestinal dysfunction in cultured Caco-2 cells. Rather, different types of sugar–amino acid combinations used to generate MRPs contributed more to mitigate injury in stress-induced Caco-2 cells. With the growing evidence that MRPs have a wide range of bioactive activities, this study concludes that specificity of substrate precursors that produce MRPs in heated foods is a critical factor for antioxidant and related cellular functions that represent a healthy gut.

Properties of Purified Quinonoid Dihydropterin Reductase

1973 ◽  
Vol 51 (9) ◽  
pp. 1229-1239 ◽  
Author(s):  
Surinder Cheema ◽  
S. J. Soldin ◽  
Antoinetta Knapp ◽  
T. Hofmann ◽  
K. G. Scrimgeour
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Adrenal Medulla ◽  
Kinetic Properties ◽  
Terminal Amino Acid ◽  
Sheep Liver ◽  
Wide Range ◽  
Sheep Brain ◽  
Terminal Amino ◽  
Covalently Linked

Quinonoid dihydropterin reductase has been purified to homogeneity from sheep liver, sheep brain, and beef adrenal medulla. Each of these enzymes has a molecular weight of about 45 000–55 000, and is composed of two subunits of half that weight. The subunits of the sheep liver reductase have identical charge, size, and N-terminal amino acid residue. The reductase exists in solution over a wide range of concentrations as the dimer. A dimer covalently linked by dimethylsuberimidate retains full activity. A number of kinetic properties of quinonoid dihydropterin reductase, including inhibition by thiol reagents and by pterin analogues, are reported.

Astringency in fruits. I. Microbial degradation of catechin

1969 ◽  
Vol 15 (3) ◽  
pp. 303-306 ◽  
Author(s):  
T. Chandra ◽  
W. Madhavakrishna ◽  
Y. Nayudamma
Keyword(s):  
Carboxylic Acid ◽  
Aspergillus Fumigatus ◽  
Aspergillus Flavus ◽  
Microbial Degradation ◽  
Protocatechuic Acid ◽  
Penicillium Species ◽  
Streptomyces Species ◽  
Enrichment Techniques

Four molds (Aspergillus fumigatus, A. terreus, A. niger, Penicillium species) and a Streptomyces species capable of degrading catechin were isolated from soil by enrichment techniques. These organisms and Aspergillus flavus (dicat), isolated from a dicatechin medium, were compared for their ability to degrade catechin. The studies indicated that A. flavus (dicat) was the most active of the organisms tested. The degradation of catechin by A. flavus (dicat) was optimal at pH 5.0 and 0.3% catechin. Phloroglucinol carboxylic acid and protocatechuic acid were identified as products of the degradation.

Acid Hydrolysis of Proteins for Chromatographic Analysis of Amino Acids

1987 ◽  
Vol 70 (1) ◽  
pp. 147-151 ◽  
Author(s):  
Robert W Zumwalt ◽  
Joseph S Absheer ◽  
Floyd E Kaiser ◽  
Charles W Gehrke
Keyword(s):  
Amino Acid ◽  
Matrix Effects ◽  
Bond Cleavage ◽  
Peptide Bond ◽  
Extensive Study ◽  
Performic Acid ◽  
Acid Oxidation ◽  
Peptide Bond Cleavage ◽  
Wide Range ◽  
Hydrolysis Of

Abstract The conditions used to hydrolyze proteins are vital in determining amino acid compositions because they necessarily represent a compromise aimed at yielding the best estimate of amino acid composition. Variations in ease of peptide bond cleavage, differences in amino acid stabilities, and matrix effects from nonproteinaceous components all militate against a single set of hydrolysis conditions that quantitatively hydrolyze every peptide bond and concurrently cause no destruction of any amino acid. This presentation summarizes and reviews an extensive study which evaluated a number of variations in the techniques and procedures of the classical 6N HC1, 110°C, 24 h hydrolysis of protein. The objectives of the recent investigation were: (/) to compare hydrolysis at 145°C, 4 h with 110°C, 24 h for proteins in a wide range of different sample matrixes; (2) to compare protein hydrolysis at 110°C, 24 h conducted in sealed glass ampoules after vacuum removal of air with hydrolysis in glass tubes with Teflon-lined screw caps after removal of air by vacuum, nitrogen purge, and sonication; (3) to evaluate a performic acid oxidation procedure before hydrolysis for the analysis of cystine and methionine in the different sample matrixes; (4) to evaluate multiple hydrolysis times at 145°C; (5) to evaluate the variation of interlaboratory hydrolysates prepared at 145°C, 4 h in 2 different laboratories on the amino acid analysis of an array of protein-containing matrixes. The major sources of inaccuracy and lack of precision arising from the application of ion-exchange or gas chromatography, both of which provide excellent accuracy and precision, are prechromatographic sample handling and the method used for hydrolysis of the protein sample itself. Hydrolysate preparation is the area that requires the most attention to solve problems of variability of amino acid analyses.

scholarly journals Colouring Matters of the Aphidoidea XLII. Purification and Properties of the Cyclising Enzyme [Protoaphin Dehydratase (Cyclising)] Concerned with Pigment Transformations in the Woolly Aphid Eriosoma lanigerum Hausmann (Hemiptera: Insecta)

1977 ◽  
Vol 30 (3) ◽  
pp. 173 ◽  
Author(s):  
DW Cameron ◽  
WH Sawyer ◽  
VM Trikojus
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Amino Acid Analysis ◽  
Eriosoma Lanigerum ◽  
Purification And Properties ◽  
Woolly Aphid

Dried extracts of woolly aphid were treated with n-butanol, then by chromatography on DEAESephadex A50 and finally by filtration on Sephadex G150 to yield a substantially homogeneous protein catalysing the conversion of the aglucone of protoaphin into xanthoaphin. Traces of lowmolecular- weight contaminants were removed by chromatography on Sephadex G 100. The enzyme, which has a molecular weight of 120000�2000 and a high content of p-structure, was inhibited by naphthoresorcinol. Its glycoprotein nature was indicated by amino acid analysis.

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