Hydrolysis p-nitrophenyl acetate

As one might expect the rate of p-nitrophenyl heptanoate hydrolysis increased at low ethanol concentrations as a result of apolar binding. The rate of p-nitrophenyl acetate hydrolysis also increased markedly at low ethanol concentration. This finding was explained by a conformational effect on the polymer, that is, lower ethanol concentration brings about a shrinkage of the polymer, which increases concerted interactions of the imidazole residues. The hydrolysis of 3-nitro-4-dodecanoyloxybenzoate was found to be 1700 times faster in the presence of poly[4(5)-vinylimidazole] compared to free imidazole (77). A double-displacement mechanism was demonstrated for this system (75). 

Relative reactivity of imidazole and monohydrogen phosphate tfimidazolel/iKHPOj") is order of unity for general base catalysis, but about 10 for nucleophilic catalysis This ratio is 0,25 for ethyl acetate hydrolysis and 4.7 x I0 for p-nitrophenyl acetate hydrolysis 92... 

Kereelec, B., Foglizzo, E., Bonicel, J., Bougis, P.E. and Chapus, C. (1992) Sequence of horse pancreatic lipase as determined by protein and cDNA sequencing. Implications for p-nitrophenyl acetate hydrolysis by pancreatic lipases. Eur. 

In order to bypass the problem of designing a pocket from scratch, Bolon and Mayo [27] introduced a catalytically active His residue in thioredoxin, a well-defined 108-residue protein for which much structural and functional information was available. The design was based on the well-known reaction mechanism of p-nitrophenyl acetate hydrolysis and thioredoxin was redesigned by computation to accommodate a histidine with an acylated side chain to mimic transition state stabilization. The thioredoxin mutant was catalytically active and the reaction followed saturation kinetics with a k at of 4.6 x 10 s and a Km of 170 xM. The catalytic efficiency, after correction for differential protonation and nucleophilicity, can be estimated to be a factor of 50 greater than that of 4-methylimidazole, due to nucleophilic catalysis and proximity effects, see Section 5.2.3. 

The Bronsted coefficient p for imidazole catalysis of p-nitrophenyl acetate hydrolysis is 0.8 [29], and the second-order rate constants of all His residues can therefore be related to that of 4-methylimidazole to determine whether there are effects on reactivity beyond those of differential nudeophilicity and levels of protonation. The reactivity of His residues in the pH independent region may be estimated from rate constants, pH and values. The second-order rate constant of the 4-methylimidazole catalyzed hydrolysis of mono-p-nitrophenyl fumarate at pH 5.85 and 290 K is 1.02 x 10 s. From this value and the pfC of 7.9, the second-... 

Inhibition of bovine CA by 5-methyl-l,10-phenanthroline and aniline involves direct tertiary co-ordination to the zinc ion. With the latter reagent, however, H n.m.r. relaxation of metal-bound water is unchanged on aniline co-ordination suggesting either that a histidine ligand is replaced or that a fifth co-ordination site is available. By contrast, methanol binds at a hydrophobic region ca. 6 A from the metal site and shows non-competitive inhibition of p-nitrophenyl acetate hydrolysis activity. ... 

Figure 2 shows variation of rates of hydrolysis of p-nitrophenyl acetate in the presence of various catalysts. It shows that B-CD does not accelerate p-nitrophenyl acetate hydrolysis, but 6-CD-histamine enhances the rate significantly. A mixture of B-CD and histamine acceler-rates the hydrolysis, but it is nearly equal to the case of histamine only, and lower than the case of B-CD-histamine. This result reveals that the ability of B CD-histamine in hydrolysis is caused jointly by a histamine group and a hydrophobic cavity of B-CD. 

Others polymers were also engaged in p-nitrophenyl acetate hydrolysis. CD-poly (vinylamine) polymers showed a higher catalytic activity than P-CD alone and a cooperative action between CD residnes and NH2 gronps [19]. Seven CD-poly (vinylamine) polymers were synthesized. The snbstitntion degree (SD) of amino fnnctions substituted by p-CD ranged from 0.3 to 5 mol% (Scheme 2.5). 

Figure 6-8 is a pH-rate profile for the hydrolysis of p-nitrophenyl acetate. The slopes of the straight-line portions are —1,0, and -L 1, reading in the acid to base direction, and this system can be described by... 

These data are for the nucleophilic catalysis of the hydrolysis of p-nitrophenyl acetate by imidazoles and benzimidazoles at pH 8.0. Tbe apparent second-order catalytic rate constants are defined by... 

An artificial metalloenzyme (26) was designed by Breslow et al. 24). It was the first example of a complete artificial enzyme, having a substrate binding cyclodextrin cavity and a Ni2+ ion-chelated nucleophilic group for catalysis. Metalloenzyme (26) behaves a real catalyst, exhibiting turnover, and enhances the rate of hydrolysis of p-nitrophenyl acetate more than 103 fold. The catalytic group of 26 is a -Ni2+ complex which itself is active toward the substrate 1, but not toward such a substrate having no metal ion affinity at a low catalyst concentration. It is appearent that the metal ion in 26 activates the oximate anion by chelation, but not the substrate directly as believed in carboxypeptidase. 

The hydrolysis of p-nitrophenyl acetate and bis(p-nitrophenyl phosphate) are frequently used to probe hydrolytic activity. A problem with some other dinuclear systems is that the Zn units are held together by bridging ligands which can be cleaved on reaction with the substrate.440 This is not the case in a ditopic ligand such as those designed by Lippard and co-workers based on Kemp s triacid imide with a xylyl spacer.441,442 Both zinc dimers and mixed metal dimers were formed and a structure characterized with a bridging phosphodiester (Figure 6). 

Hua I, Hochemer RH, Hoffmann MR (1995) Sonolytic hydrolysis of p-nitrophenyl acetate The role of supercritical water. J Phys Chem 99 2335-2342... 

The most effective catalyst for the hydrolysis of p-nitrophenyl acetate was reported to be a cycloheptaamylose derivative containing approximately two imidazole groups per cycloheptaamylose molecule (Cramer and Mackensen, 1970). At pH 7.5 and 23°, this material accelerates the rate of release of phenol from p-nitrophenyl acetate by a factor of 300 when compared with the hydrolysis of this substrate in the absence of catalyst. However, when compared with an equivalent concentration of imidazole, which is an effective catalyst for ester hydrolysis at neutral pHs, the rate accelerations imposed by this cycloheptaamylose derivative are only two- to threefold. Cramer and Mackensen attributed this rate enhancement to nucleophilic displacement of phenol from the included ester by a cycloheptaamylose hydroxyl group, assisted internally by the attached imidazole group... 

The hydrolysis of esters by the nickel derivative (271) provided an early example of the use of a metal-capped cyclodextrin as a catalyst (shown here as its p-nitrophenyl acetate inclusion complex) (Breslow Overman, 1970 Breslow, 1971). The synthesis of this host involves the following steps (i) covalent binding of the pyridine dicarboxylic acid moiety to cyclodextrin, (ii) coordination of Ni(n) to this species, and (iii)... 

Effect of Various Histidine Peptides on the Rate of Hydrolysis of p-Nitrophenyl Acetate at pH 7.73 ... 

For the hydrolysis of p-nitrophenyl acetate at pH 7.7 the most effective catalyst was Gly-His-Gly-Gly-His-Gly. However, this peptide had only 50% of the catalytic activity of imidazole. For the seven peptides the range of catalytic effectiveness was found to be 30-50% that of imidazole. 

Manecke and his collaborators have synthesized polymers of vinylimid-azole hydroxamic acid (75) (22, 23, 24). Hydrolysis of p-nitrophenyl acetate... 

The oxamate group has been incorporated into a vinyl polymer by Kirsh and Kabanov 41). These workers prepared a copolymer of 4-vinyl-A -(phenacyloxime)pyridinium bromide and vinylpyridine. For the hydrolysis of p-nitrophenyl acetate the oxamate polymer produced a significant rate enhancement over the monomeric analogs. 

Bruice and Sturtevant, (1959) and Bruice, (1959) found extremely facile intramolecular nucleophilic attack by neighbouring imidazole in the hydrolysis of p-nitrophenyl 7-(4-imidazoyl)butyrate [19]. The rate constant for imidazole participation (release of p-nitro-phenolate) in this reaction is nearly identical with the rate constant for a-chymotrypsin catalysed release of p-nitrophenolate ion [190 min in equation (11) at pH 7 and 25°] from p-nitrophenyl acetate. Comparison of the rate constant for intramolecular imidazole participation to that for the analogous bimolecular reaction (imidazole attack on p-nitrophenyl acetate) (s" /m s )... 

Reaction of the m-nitrophenyl ester of pyridine-2,5-dicarboxylic acid with cyclodextrin (see Section 3) gives a picolinate ester [52] of a cyclodextrin secondary hydroxyl group (Breslow, 1971 Breslow and Overman, 1970) which will bind metal ions or a metal ion-pyridine carboxaldoxime complex. Such a complex will catalyse hydrolysis of p-nitrophenyl acetate bound within the cyclodextrin cavity leading to a rate constant approximately 2000-fold greater at... 

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