Presence and absence of enzymes

Table I. Presence and Absence of Enzymes in Sea Snake Venoms...

Because enzymes are such superb catalysts, it is tempting to ascribe to them powers that they do not have. An enzyme cannot alter the laws of thermo dynamics and consequently cannot alter the equilibrium of a chemical reaction. Consider an enzyme-catalyzed reaction, the conversion of substrate, S, into product, P. Figure 8.2 shows the rate of product formation with time in the presence and absence of enzyme. Note that the amount of product formed... 

Another direct assay method is based on decay kinetics of pulse generated O2 (Takahashi and Asada, 1981). Superoxide was produced within 10 msec by a flash of light through the excitation of flavin mononucleotide in the presence of A,A,A, A -tetramethylethylenedi-amine and oxygen. The kinetics of O2 decay in the presence and absence of enzyme were followed at 240 nm. The catalytic rate constant for bovine erythrocyte copper/zinc superoxide dismutase was found to be 1.75 x... 

CHO cells were exposed to freshly prepared solutions of gallic acid, pH 7.0, in the presence and absence of enzyme (0.5 yg/ml) they were sampled 20 hr later for chromosome analysis. 

The ratio of the equilibrium constants for conversion of substrate from the ground state to the transition state in the presence and absence of enzyme is related to the ratio of the dissociation constants for ES and ES complexes ... 

Portions (50 mU MCA-hydrolsing activity) of purified CinnAE were incubated at 37°C with SBP (10 mg), both in the presence and absence of other carbohydrases, in 100 mM MOPS (pH 6.0) in a final volume of 1 mL. Incubations containing boiled enzyme were performed as controls. Reactions were terminated by boiling (3 min) and the amount of free ferulic acid determined using a method described previously for de-starched wheat bran [18]. The total amount of alkali-extractable ferulic acid present in the SBP was 0.87% [5]. 

Figure 3.1 Equilibrium scheme for enzyme turnover in the presence and absence of reversible inhibitors...

CO3 species was formed and the X-ray structure solved. It is thought that the carbonate species forms on reaction with water, which was problematic in the selected strategy, as water was produced in the formation of the dialkyl carbonates. Other problems included compound solubility and the stability of the monoalkyl carbonate complex. Van Eldik and co-workers also carried out a detailed kinetic study of the hydration of carbon dioxide and the dehydration of bicarbonate both in the presence and absence of the zinc complex of 1,5,9-triazacyclododecane (12[ane]N3). The zinc hydroxo form is shown to catalyze the hydration reaction and only the aquo complex catalyzes the dehydration of bicarbonate. Kinetic data including second order rate constants were discussed in reference to other model systems and the enzyme carbonic anhy-drase.459 The zinc complex of the tetraamine 1,4,7,10-tetraazacyclododecane (cyclen) was also studied as a catalyst for these reactions in aqueous solution and comparison of activity suggests formation of a bidentate bicarbonate intermediate inhibits the catalytic activity. Van Eldik concludes that a unidentate bicarbonate intermediate is most likely to the active species in the enzyme carbonic anhydrase.460... 

The principal tests can be broadly categorized into microbial and mammalian cell assays. In both cases the tests are carried out in the presence and absence of in vitro metabolic activation enzymes, usually derived from rodent liver. 

There are three basic methods for carrying out alternative substrate inhibition studies. In the first, the investigator seeks to observe numerical changes in the coefficients of the double-reciprocal form of the enzyme rate expression in the presence and absence of the alternative substrate. For some mechanisms, only certain coefficients will be altered. This method requires extremely accurate estimates of the magnitudes of the coefficients and should always be supplemented with other kinetic probes . 

In 1976 he was appointed to Associate Professor for Technical Chemistry at the University Hannover. His research group experimentally investigated the interrelation of adsorption, transfer processes and chemical reaction in bubble columns by means of various model reactions a) the formation of tertiary-butanol from isobutene in the presence of sulphuric acid as a catalyst b) the absorption and interphase mass transfer of CO2 in the presence and absence of the enzyme carboanhydrase c) chlorination of toluene d) Fischer-Tropsch synthesis. Based on these data, the processes were mathematically modelled Fluid dynamic properties in Fischer-Tropsch Slurry Reactors were evaluated and mass transfer limitation of the process was proved. In addition, the solubiHties of oxygen and CO2 in various aqueous solutions and those of chlorine in benzene and toluene were determined. Within the framework of development of a process for reconditioning of nuclear fuel wastes the kinetics of the denitration of efQuents with formic acid was investigated. 

Fig. 8.—Effects of Albumin and Zn2+ on the Activity of a-D-Mannosidase.39 [A purified, jack-bean meal preparation was incubated beforehand at 37° and pH 5 in the presence, and absence, of 0.01% of albumin. After 17 hr, ZnS04 (final concentration, 1 m M) was added to a sample of each solution, together with albumin where appropriate to maintain an 0.01% concentration thereof, and incubation was continued. , Enzyme alone O, enzyme + ZnS04 A, enzyme + albumin and A, enzyme + albumin + ZnS04. The results are expressed as a percentage of the activity in the unincubated, enzyme preparation.]...

To elucidate the difference between the enzymatic and nonenzymatic participation of metal ions, it is clearly desirable to be able to compare the effect of a large number of metal ions upon the same reaction both in the presence and absence of the enzyme. For such a study to be feasible it is necessary to work with a metal-activated enzymatic reaction, which will also take place when the metal, but not the enzyme, is omitted. Such a reaction is the decarboxylation of oxaloacetic acid. The mechanism of metal catalysis of this reaction is similar to that assumed for carboxypeptidase, and can be represented as follows (44). 

Another action of phenothiazines is to compete with NADPH for the oxidase, an inference which was based on fulfillment of the criteria for competitive inhibition in double reciprocal plots of 1/[NADPH] vs. 1/rate of formation of O in the presence and absence of inhibitor It seems worth considering that all the effects of phenothiazines might be mediated through this effect and that the process of activation represents the presentation of substrate to the enzyme from which, in the resting state, the substrate is kept separate. 

Citrate synthase from mitochondria has been crystallized and visualized by x-ray diffraction in the presence and absence of its substrates and inhibitors (Fig. 16-8). Each subunit of the homodimeric enzyme is a single polypeptide with two domains, one large and rigid, the other smaller and more flexible, with the active site between them. Oxaloacetate, the first substrate to bind to the enzyme, induces a large conformational... 

Product inhibition (Section A,12) can also provide information about mechanisms. For example, if 1 / v is plotted against 1 / [A] in the presence and absence of the product Q, the product will be found to compete with A and to give a typical family of lines for competitive inhibition. On the other hand, a plot of 1 / v vs 1 / [B] in the presence and absence of Q will indicate noncompetitive inhibition if the binding of substrates is ordered (Eq. 9-43). In other words, only the A-Q pair of substrates are competitive. Product inhibition is also observed with enzymes having ping-pong kinetics (Eq. 9-47) as a result of formation of nonproductive complexes. 

The biochemical basis of CAM-induced stimulation of Ca2+-ATPase activity in carrot cells was studied further by determining the parameters of the Ca2+-translocating reaction of the enzyme in the presence and absence of exogenous CAM, using EGTA-treated plasma membrane [45], The affinity of Ca2+-ATPase for Ca2+ was considerably increased by... 

A Lineweaver-Burk plot of enzyme kinetics in the presence and absence of a noncompetitive inhibitor is shown in Figure E5.5. Umax in the presence of a noncompetitive inhibitor is decreased, but KM is unaffected. The effect of a competitive inhibitor on the direct linear plot is shown in Figure E5.6. 

Mutarotation of 0.3% solutions of the freshly dissolved sugars in 12 ml of 5 mM EDTA, pH 7.4 was followed. Significant differences in mutarotation rates (AK) in the presence and absence of 100 units of bovine kidney enzyme were expressed as the ratio AK/Ksp. Differences of less than 5% in these rate constants were not considered significant. Of the 18 sugars listed, nine have been tested previously as substrates for other mammalian mutarotases with essentially the same pattern as described here. The pattern of specificity indicates that a 3-point attachment of enzyme and substrate is necessary for catalysis of mutarotation. b Data from 72). 

Proteinases and proteinase inhibitors in a sample should be analyzed for their sensitivity to SDS concentration before activity staining is carried out. Theiractivity should be measured in the presence and absence of 0.1 % SDS before proceeding with electrophoresis. The author has characterized many proteinases under such conditions without a loss of activity. Most SDS-sensitive enzymes regain their activity if the gel is washed in the buffer used to dissolve the... 

Solutions of acid phosphatase are particularly sensitive to surface inactivation. Figure 3 (88) shows the inactivation rate of the enzyme in the presence and absence of surface-active detergents. The inactivation process is temperature sensitive and the protection by detergent is total. Most of the enzyme inactivation proceeds with first-order kinetics. A variety of agents—gelatin, bovine serum albumin, egg albumin, and Tween-80—protect the enzyme against inactivation. 

Philip Evans and his coworkers have determined the crystal structures of phosphofructokinase from two species of bacteria, E. coli and Bacillus stearothermophilus. By crystallizing the enzyme in the presence and absence of the substrate and several allosteric effectors, they obtained detailed views of both the T and R conformations. This work has led to an explanation of why phosphofructokinase appears to be constrained largely to all-or-nothing transitions between these two states, rather than adopting a series of intermediate conformations. 

The enzyme contains six Zn2+ per molecule, two per R subunit. The zinc is not required for catalytic activity, but is essential for the maintenance of the quaternary structure. The structure has been determined to a resolution of 2.8 A in the presence and absence of CTP.528 The zinc-binding site is located in the C-terminal region of the R chains, and involves four cysteinyl residues, with tetrahedral geometry. The zinc domain represents the major site of interaction between the R and C chains, explaining the importance of zinc for the association of the subunits and the dissociative effect of mercurial reagents. When E. coli is grown in a zinc-deficient medium, some 70% of the enzyme is found to be dissociated into subunits.529... 

The relative contribution, in intact microsomal preparations, of the two monooxygenases to the formation of N-oxygenated C=N functionalities has been frequently assessed by measurements in the presence and absence of selective enzyme inhibitors or positive effectors. These observations were supplemented by studies using highly purified native or recombinant proteins in reconstituted systems. 

First hints on the nature of regulatory sites in yeast PDC have been obtained from inhibition experiments with the thiol-modifying reagents 4-hydroxy-mer-curi-benzoate and 3-bromo-pyruvamide in the presence and absence of the substrate pyruvate. Upon thiol modification, PDC lost its activation behaviour. Pyruvate was able to protect the enzyme against thiol-modification, suggesting that both modifier and substrate compete for the same sites on the enzyme... 

Figure 9. Loss of ethylene-producing ability in apple slices treated with a mixture of cell-wall-digesting enzymes in presence and absence of methionine (58) (0,9), preclimacteric (A, A), climacteric.

The effect of solvents such as DMSO, ethanol, and methanol, on the MurD coupled enzyme reaction and the ELISA was studied. The MurD enzyme reaction was run in the presence and absence of Mur enzymes to generate minimum and maximum signals. Final solvent concentrations up to 2% had no effect on the reaction when measured by either HPLC or in the ELISA. 

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