THE METABOLISM OF THE ERYTHROCYTE: IX. DIPHOSPHOPYRIDINE NUCLEOTIDASE OF ERYTHROCYTES

1956 ◽  
Vol 34 (1) ◽  
pp. 46-60 ◽  
Author(s):  
S. G. A. Alivisatos ◽  
S. Kashket ◽  
O. F. Denstedt
Keyword(s):  
Ion Transport ◽  
External Medium ◽  
Nicotinamide Adenine ◽  
Rabbit Erythrocyte ◽  
Purine Derivatives ◽  
Quaternary Nitrogen ◽  
Enzyme Substrate ◽  
Adenylic Acid ◽  
Ph Range ◽  
Hydrolysis Of

The rabbit erythrocyte possesses an active DPN-ase which is firmly attached to the membrane of the cell. Evidently, the enzyme is oriented in the membrane in such a way as to be able to act upon DPN added to the external medium. The enzyme splits DPN at the bond linking the quaternary nitrogen of the nicotinamide moiety with the ribose component. Despite the release of an H+ ion during hydrolysis of DPN, the activity of the enzyme remains practically constant over the pH range from 4.5 to 10.0. DPN-ase also splits nicotinamide from TPN, but its affinity for the latter is only about one third of that for DPN. Nicotinamide, adenine, and the substituted purine derivatives theobromine, theophylline, and xanthine inhibit DPN-ase, while compounds such as ribose, adenylic acid, caffeine, and nembutal are without effect in this respect. Of all the substances tested theobromine proved to be the most powerful inhibitor. The mode of enzyme–substrate attachment and the possible involvement of DPN-ase in ion transport are discussed in the light of these findings.

THE METABOLISM OF THE ERYTHROCYTE: IX. DIPHOSPHOPYRIDINE NUCLEOTIDASE OF ERYTHROCYTES

1956 ◽  
Vol 34 (1) ◽  
pp. 46-60 ◽  
Author(s):  
S. G. A. Alivisatos ◽  
S. Kashket ◽  
O. F. Denstedt
Keyword(s):  
Ion Transport ◽  
External Medium ◽  
Nicotinamide Adenine ◽  
Rabbit Erythrocyte ◽  
Purine Derivatives ◽  
Quaternary Nitrogen ◽  
Enzyme Substrate ◽  
Adenylic Acid ◽  
Ph Range ◽  
Hydrolysis Of

The rabbit erythrocyte possesses an active DPN-ase which is firmly attached to the membrane of the cell. Evidently, the enzyme is oriented in the membrane in such a way as to be able to act upon DPN added to the external medium. The enzyme splits DPN at the bond linking the quaternary nitrogen of the nicotinamide moiety with the ribose component. Despite the release of an H+ ion during hydrolysis of DPN, the activity of the enzyme remains practically constant over the pH range from 4.5 to 10.0. DPN-ase also splits nicotinamide from TPN, but its affinity for the latter is only about one third of that for DPN. Nicotinamide, adenine, and the substituted purine derivatives theobromine, theophylline, and xanthine inhibit DPN-ase, while compounds such as ribose, adenylic acid, caffeine, and nembutal are without effect in this respect. Of all the substances tested theobromine proved to be the most powerful inhibitor. The mode of enzyme–substrate attachment and the possible involvement of DPN-ase in ion transport are discussed in the light of these findings.

THE METABOLISM OF THE ERYTHROCYTE: XII. DIPHOSPHOPYRIDINE NUCLEOTIDE NUCLEOSIDASE OF THE RABBIT ERYTHROCYTE

1956 ◽  
Vol 34 (1) ◽  
pp. 141-145 ◽  
Author(s):  
A. Malkin ◽  
O. F. Denstedt
Keyword(s):  
Cleavage Product ◽  
Rabbit Erythrocyte ◽  
Quaternary Nitrogen ◽  
Rate Of Hydrolysis ◽  
Additional Support ◽  
Hydrolysis Of

Evidence is presented in support of the hypothesis that in the hydrolysis of DPN by the DPN nucleosidase of rabbit erythrocyte stroma, the substrate is attached to the enzyme at more than one site. Cleavage of DPN at the nicotinamide–ribose bond is inhibited by nicotinamide, ADP, and adenine. Considering the nature of the inhibition and the extent of the inhibition, it is suggested that DPN attaches to the enzyme surface at the quaternary nitrogen of the nicotinamide and at the pyrophosphate group of the DPN molecule. Additional support for this supposition accrues from the observation that NMN, a cleavage product of DPN, can serve as a substrate for DPN-ase but the rate of hydrolysis is much slower than that with DPN.

THE METABOLISM OF THE ERYTHROCYTE: XII. DIPHOSPHOPYRIDINE NUCLEOTIDE NUCLEOSIDASE OF THE RABBIT ERYTHROCYTE

1956 ◽  
Vol 34 (2) ◽  
pp. 141-145 ◽  
Author(s):  
A. Malkin ◽  
O. F. Denstedt
Keyword(s):  
Cleavage Product ◽  
Rabbit Erythrocyte ◽  
Quaternary Nitrogen ◽  
Rate Of Hydrolysis ◽  
Additional Support ◽  
Hydrolysis Of

Evidence is presented in support of the hypothesis that in the hydrolysis of DPN by the DPN nucleosidase of rabbit erythrocyte stroma, the substrate is attached to the enzyme at more than one site. Cleavage of DPN at the nicotinamide–ribose bond is inhibited by nicotinamide, ADP, and adenine. Considering the nature of the inhibition and the extent of the inhibition, it is suggested that DPN attaches to the enzyme surface at the quaternary nitrogen of the nicotinamide and at the pyrophosphate group of the DPN molecule. Additional support for this supposition accrues from the observation that NMN, a cleavage product of DPN, can serve as a substrate for DPN-ase but the rate of hydrolysis is much slower than that with DPN.

Optimization of Enzymatic Hydrolysis of Cucurbitin Using Response Surface Methodology: Improvement of the Functional Properties

2011 ◽  
Vol 7 (5) ◽  
Author(s):  
Ljiljana Popovic ◽  
Draginja Peričin ◽  
Žužana Vaštag ◽  
Senka Popovic
Keyword(s):  
Response Surface Methodology ◽  
Enzymatic Hydrolysis ◽  
Response Surface ◽  
Functional Properties ◽  
Food Systems ◽  
Degree Of Hydrolysis ◽  
Foaming Properties ◽  
Enzyme Substrate ◽  
Ph Range ◽  
Hydrolysis Of

Optimization of enzymatic hydrolysis of cucurbitin, extracted from pumpkin (Cucurbita pepo) oil cake with bromelain, was carried out by response surface methodology (RSM). Second-order polynomial model (R2=0.791) has been proposed for the effect of time (t), and enzyme/substrate ratio (E/S) on degree of hydrolysis (DH). Conditions for obtaining maximal value of DH were determined (E/S= 0.0132 (w/w), t= 42 min). Furthermore, according to the regression equation, conditions for production of hydrolysates with target DH values were chosen. Comparison of the functional properties of cucurbitin and its hydrolysates (DH= 10%, 20% and 30%) were carried out. The solubility of the hydrolysates gradually increased with the increase of DH, in pH range of 6–8. The hydrolysate with DH=10% had the best emulsifying properties (EA=0.632 ± 0.02 A500nm, ES= 44.2min), and extent of hydrolysis decreased both emulsifying activity and emulsifying stability. All studied hydrolysates exhibited higher oil-holding capacity and have improved foaming properties compared to the original protein. DH has influence on foam capacity. The highest foam capacity has hydrolysate with DH=20% (150.3±3.66 %). Obtained results suggest that limited protease hydrolysis of cucurbitin with bromelain produces the hydrolysates with improved functional properties and indicate their possible use in different food systems.

Schiff base equilibria. II. Beryllium complexes of N-n-butylsalicylideneimine and the hydrolysis of the Be2+ ion

1965 ◽  
Vol 18 (5) ◽  
pp. 651 ◽  
Author(s):  
RW Green ◽  
PW Alexander
Keyword(s):  
Aqueous Solution ◽  
Schiff Base ◽  
Complex Formation ◽  
Distribution Coefficient ◽  
Dilute Aqueous Solution ◽  
The Stability ◽  
Ph Range ◽  
Hydrolysis Of ◽  
Beryllium Complexes ◽  
Equilibria In Aqueous Solution

The Schiff base, N-n-butylsalicylideneimine, extracts more than 99.8% beryllium into toluene from dilute aqueous solution. The distribution of beryllium has been studied in the pH range 5-13 and is discussed in terms of the several complex equilibria in aqueous solution. The stability constants of the complexes formed between beryllium and the Schiff base are log β1 11.1 and log β2 20.4, and the distribution coefficient of the bis complex is 550. Over most of the pH range, hydrolysis of the Be2+ ion competes with complex formation and provides a means of measuring the hydrolysis constants. They are for the reactions: Be(H2O)42+ ↔ 2H+ + Be(H2O)2(OH)2, log*β2 - 13.65; Be(H2O)42+ ↔ 3H+ + Be(H2O)(OH)3-, log*β3 -24.11.

Micellar catalysis of organic reactions. VIII. Kinetic studies of the hydrolysis of 2-acetyloxybenzoic acid (aspirin) in the presence of micelles

1982 ◽  
Vol 35 (7) ◽  
pp. 1357 ◽  
Author(s):  
TJ Broxton
Keyword(s):  
Aqueous Solution ◽  
Cetyltrimethylammonium Bromide ◽  
Cetylpyridinium Chloride ◽  
Kinetic Studies ◽  
Base Catalysis ◽  
Plateau Region ◽  
General Base ◽  
Mechanism Of Hydrolysis ◽  
Ph Range ◽  
Hydrolysis Of

The hydrolysis of 2-acetyloxybenzoic acid in the pH range 6-12 has been studied in the presence of micelles of cetyltrimethylammonium bromide (ctab) and cetylpyridinium chloride (cpc). In the plateau region (pH 6-8) the hydrolysis is inhibited by the presence of micelles, while in the region where the normal BAC2 hydrolysis (pH > 9) occurs the reaction is catalysed by micelles of ctab and cpc. The mechanism of hydrolysis in the plateau region is shown to involve general base catalysis by the adjacent ionized carboxy group both in the presence and absence of micelles. This reaction is inhibited in the presence of micelles because the substrate molecules are solubilized into the micelle and water is less available in this environment than in normal aqueous solution.

Effect of Modifiers on the Hydrolysis of Basic and Neutral Peptides by Carboxypeptidase B

1975 ◽  
Vol 53 (7) ◽  
pp. 747-757 ◽  
Author(s):  
Graham J. Moore ◽  
N. Leo Benoiton
Keyword(s):  
Active Site ◽  
Active Sites ◽  
Kinetic Behavior ◽  
Carboxypeptidase A ◽  
Phenyllactic Acid ◽  
Carboxypeptidase B ◽  
Substrate Complex ◽  
Enzyme Substrate ◽  
Basic Peptides ◽  
Hydrolysis Of

The initial rates of hydrolysis of Bz-Gly-Lys and Bz-Gly-Phe by carboxypeptidase B (CPB) are increased in the presence of the modifiers β-phenylpropionic acid, cyclohexanol, Bz-Gly, and Bz-Gly-Gly. The hydrolysis of the tripeptide Bz-Gly-Gly-Phe is also activated by Bz-Gly and Bz-Gly-Gly, but none of these modifiers activate the hydrolysis of Bz-Gly-Gly-Lys, Z-Leu-Ala-Phe, or Bz-Gly-phenyllactic acid by CPB. All modifiers except cyclohexanol display inhibitory modes of binding when present in high concentration.Examination of Lineweaver–Burk plots in the presence of fixed concentrations of Bz-Gly has shown that activation of the hydrolysis of neutral and basic peptides by CPB, as reflected in the values of the extrapolated parameters, Km(app) and keat, occurs by different mechanisms. For Bz-Gly-Gly-Phe, activation occurs because the enzyme–modifier complex has a higher affinity than the free enzyme for the substrate, whereas activation of the hydrolysis of Bz-Gly-Lys derives from an increase in the rate of breakdown of the enzyme–substrate complex to give products.Cyclohexanol differs from Bz-Gly and Bz-Gly-Gly in that it displays no inhibitory mode of binding with any of the substrates examined, activates only the hydrolysis of dipeptides by CPB, and has a greater effect on the hydrolysis of the basic dipeptide than on the neutral dipeptide. Moreover, when Bz-Gly-Lys is the substrate, cyclohexanol activates its hydrolysis by CPB by increasing both the enzyme–substrate binding affinity and the rate of the catalytic step, an effect different from that observed when Bz-Gly is the modifier.The anomalous kinetic behavior of CPB is remarkably similar to that of carboxypeptidase A, and is a good indication that both enzymes have very similar structures in and around their respective active sites. A binding site for activator molecules down the cleft of the active site is proposed for CPB to explain the observed kinetic behavior.

scholarly journals Purification and properties of three (1→3)-β-d-glucanase isoenzymes from young leaves of barley (Hordeum vulgare)

1993 ◽  
Vol 289 (2) ◽  
pp. 453-461 ◽  
Author(s):  
M Hrmova ◽  
G B Fincher
Keyword(s):  
Hordeum Vulgare ◽  
Elevated Temperatures ◽  
Gel Filtration ◽  
Degree Of Polymerization ◽  
Exchange Chromatography ◽  
Purification And Properties ◽  
Young Leaves ◽  
Monomeric Proteins ◽  
Ph Range ◽  
Hydrolysis Of

Three (1->3)-beta-D-glucan glucanohydrolase (EC 3.2.1.39) isoenzymes GI, GII and GIII were purified from young leaves of barley (Hordeum vulgare) using (NH4)2SO4 fractional precipitation, ion-exchange chromatography, chromatofocusing and gel-filtration chromatography. The three (1->3)-beta-D-glucanases are monomeric proteins of apparent M(r)32,000 with pI values in the range 8.8-10.3. N-terminal amino-acid-sequence analyses confirmed that the three isoenzymes represent the products of separate genes. Isoenzymes GI and GII are less stable at elevated temperatures and are active over a narrower pH range than is isoenzyme GIII, which is a glycoprotein containing 20-30 mol of hexose equivalents/mol of enzyme. The preferred substrate for the enzymes is laminarin from the brown alga Laminaria digitata, an essentially linear (1->3)-beta-D-glucan with a low degree of glucosyl substitution at 0-6 and a degree of polymerization of approx. 25. The three enzymes are classified as endohydrolases, because they yield (1->3)-beta-D-oligoglucosides with degrees of polymerization of 3-8 in the initial stages of hydrolysis of laminarin. Kinetic analyses indicate apparent Km values in the range 172-208 microM, kcat. constants of 36-155 s-1 and pH optima of 4.8. Substrate specificity studies show that the three isoenzymes hydrolyse substituted (1->3)-beta-D-glucans with degrees of polymerization of 25-31 and various high-M(r), substituted and side-branched fungal (1->3;1->6)-beta-D-glucans. However, the isoenzymes differ in their rates of hydrolysis of a (1->3;1->6)-beta-D-glucan from baker's yeast and their specific activities against laminarin vary significantly. The enzymes do not hydrolyse (1->3;1->4)-beta-D-glucans, (1->6)-beta-D-glucan, CM-cellulose, insoluble (1->3)-beta-D-glucans or aryl beta-D-glycosides.

scholarly journals Purification and Characterization of 2,6-β-d-Fructan 6-Levanbiohydrolase fromStreptomyces exfoliatus F3-2

2000 ◽  
Vol 66 (1) ◽  
pp. 252-256 ◽  
Author(s):  
Katsuichi Saito ◽  
Kazuya Kondo ◽  
Ichiro Kojima ◽  
Atsushi Yokota ◽  
Fusao Tomita
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Extracellular Enzyme ◽  
Molecular Weights ◽  
Sds Page ◽  
Monomeric Structure ◽  
Ph Range ◽  
Hydrolysis Of

ABSTRACT Streptomyces exfoliatus F3-2 produced an extracellular enzyme that converted levan, a β-2,6-linked fructan, into levanbiose. The enzyme was purified 50-fold from culture supernatant to give a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weights of this enzyme were 54,000 by SDS-PAGE and 60,000 by gel filtration, suggesting the monomeric structure of the enzyme. The isoelectric point of the enzyme was determined to be 4.7. The optimal pH and temperature of the enzyme for levan degradation were pH 5.5 and 60°C, respectively. The enzyme was stable in the pH range 3.5 to 8.0 and also up to 50°C. The enzyme gave levanbiose as a major degradation product from levan in an exo-acting manner. It was also found that this enzyme catalyzed hydrolysis of such fructooligosaccharides as 1-kestose, nystose, and 1-fructosylnystose by liberating fructose. Thus, this enzyme appeared to hydrolyze not only β-2,6-linkage of levan, but also β-2,1-linkage of fructooligosaccharides. From these data, the enzyme from S. exfoliatus F3-2 was identified as a novel 2,6-β-d-fructan 6-levanbiohydrolase (EC 3.2.1.64 ).

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