Hydrolytic enzymes catalytic efficiency

Similar explanations, supported by convincing evidence, have been proposed for the high catalytic efficiency and specificity of enzymes (particularly hydrolytic enzymes) relative to ordinary catalysts (Laidler, 23, Wilson, 65, 66). 

Enzymes are proteinaceous catalysts peculiar to living matter. Hundreds have been obtained in purified and crystalline form. Their catalytic efficiency is extremely high—one mole of a pure enzyme may catalyze the transformation of as many as 10,000 to 1,000,000 moles of substrate per minute. While some enzymes are highly specific for only one substrate, others can attack many related substrates. Avery broad classification of enzymes would include hydrolytic enzymes (esterases, proteases), phosphorylases, oxidoreductive enzymes (dehydrogenases, oxidases, peroxidases), transferring enzymes, decarboxylases and others. 

Metal ions are vital to the function of many enzymes that catalyze hydrolytic reactions. Coordination of a water molecule to a metal ion alters its acid-base properties, usually making it easier to deprotonate, which can offer a ready means for catalyzing a hydrolytic reaction. Also, the placement of a metal center in the active site of a hydrolytic enzyme could permit efficient delivery of a catalytic water molecule to the hydrolyzable substrate. In fact, the first enzyme discovered, carbonic an-hydrase, is a metalloenzyme that requires a Zn2+ center for its catalytic activity (32). The function of carbonic anhydrase is to catalyze the hydrolysis of carbon dioxide to bicarbonate ... 

We have noted previously that the catalytic effect of the hydrolytic enzyme chymotrypsin depends critically on the interaction of a hydroxyl and an imidazole group placed in juxtaposition in the enzyme molecule. It was, therefore, particularly tempting to see if an analogous interaction could enhance the catalytic efficiency of PVI, by using copolymers of 4(5)-vinylimidazole with p-vinylphenol (VI/VP) or with vinyl alcohol (VI/VA)... 

Metal ion cofactors have varied roles to enhance the catalytic efficiency of enzymes in hydrolytic reactions, including facilitate substrate binding (water and organic substrate), gathering/template effects, function as an electrostatic catalyst (carbonyl polarization and transition state stabilization), function as a Lewis acid to lower the pA a of metal-water and stabilize the formation of the leaving group. Although their properties make several... 

Enzymes found in hepatocytes,64 neuronal cells,65 and plasma also hydrolyze nerve agents, albeit comparatively weakly. Study of the requirements for hydrolysis at the enzyme active sites could potentially lead to the design of more efficient hydrolytic enzymes that could be used as catalytic scavengers.66... 

To compare the catalytic efficiency of catalysts, it is helpful to compare the enhancement ratios (E.R.). E.R. is calculated by dividing the kcat by the kuncat (the rate constant for the uncatalyzed reaction). Enormous rate enhancements are achieved by enzymes. For example, hydrolytic enzymes often exhibit rate enhancements of 10 -10 2 compared with the spontaneous water-catalyzed or the acid/base-catalyzed reaction at about neutral pH. e For purposes of comparison, the kcat, kuncat, and E.R. values for two hydrolytic abzymes, as well as CCMP fluorohydrolase chromatographed on G-15 Sephadex gel is presented in Table 3. The fluorohydrolase, chromatographed on G-15 Sephadex, has an E.R. four times that of the two abzymes. In addition, the enhancement ratios of natural DFPases from various sources as well as CCMP fluorohydrolase are presented in Table 4. In all cases, the initial E.R. (before any purification) is higher for semisynthetic fluorohydrolases than for any of the natural unpurified DFPases. 

Hydrolytic enzymes, characterized by high specificity and high catalytic reactivity have been the most frequently modeled biopolymers At the active site of the enzyme there are usually several functional groups responsible for the overall catalytic reaction which are covalently bound to remote areas of the enzyme. Collectively these interactions, which are termed as intramolecular multiple catalyst reactions are closely related to an enzyme s specificity and efficiency. 

This model clearly shows that the catalytic machinery involves a dyad of histidine and aspartate together with the oxyanion hole. Hence, it does not involve serine, which is the key amino acid in the hydrolytic activity of lipases, and, together with aspartate and histidine, constitutes the active site catalytic triad. This has been confirmed by constructing a mutant in which serine was replaced with alanine (Serl05Ala), and finding that it catalyzes the Michael additions even more efficiently than the wild-type enzyme (an example of induced catalytic promiscuity ) [105]. 

Hydrolytic catalysis by metal ions is also important in the hydrolysis of nucleic acids, especially RNA (36). Molecules of RNA that catalyze hydrolytic reactions, termed ribozymes, require divalent metal ions to effect hydrolysis efficiently. Thus, all ribozymes are metalloenzymes (6). There is speculation that ribozymes may have been the first enzymes to evolve (37), so the very first enzymes may have been metalloenzymes Recently, substitution of sulfur for the 3 -oxygen atom in a substrate of the tetrahymena ribozyme has been shown to give a 1000-fold reduction in rate of hydrolysis with Mg2+ but no attenuation of the hydrolysis rate with Mn2+ and Zn2+ (38). Because Mn2+ and Zn2+ have stronger affinities for sulfur than Mg2+ has, this feature provides strong evidence for a true catalytic role of the divalent cation in the hydrolytic mechanism, involving coordination of the metal to the 3 -oxygen atom. Other examples of metal-ion catalyzed hydrolysis of RNA involve lanthanide complexes, which are discussed in this volume. 

Hydrolases. Hydrolytic mechanisms are also important in insecticide resistance, despite the apparent low activities in resistant insects when compared to mammalian enzymes (Table III). Some strains of resistant mosquitoes (22), Tribolium beetles (24), and Indianmeal moth (22) have specific resistance for malathion and similar carboxylester insecticides. This is due to increased catalytic hydrolysis, possibly through production of a more efficient enzyme (25.26). Californian tobacco budworms with low level permethrin resistance exhibited twice the normal activity of trans-permethrin carboxylester hydrolase (27). 

Analysis of human CE by Northern blot shows a single band of approximately 2.1 kilobases (kb) (Riddles et al. 1991), and three bands of approximately 2-, 3-, and 4.2-kb occurring with hCE-2 (Schwer et al. 1997). The intensities of the 2.1-kb band were liver 3> heart > stomach > testis > kidney = spleen > colon > other tissues. For hCE-2, the 2-kb band was located in liver > colon > small intestine > heart, the 3-kb band in liver > small intestine > colon > heart, and the 4.2-kb band in brain, testis, and kidney only. Analysis of substrate structure versus efficiency for ester (pyrethroid substrates) revealed that the two CEs recognize different structural features of the substrate (i.e., acid, alcohol, etc.). The catalytic mechanism involves the formation of an acyl-enzyme on an active serine. While earlier studies of pyrethroid metabolism were primarily performed in rodents, knowledge of the substrate structure-activity relationships and the tissue distribution of hCEs are critical for predicting the metabolism and pharmacokinetics of pesticides in humans. Wheelock et al. (2003) used a chiral mixture of the fluorescent substrate cyclopro-panecarboxylic acid, 3-(2,2-dichloroethenyl)-2,2-dimethyl-, cyano(6-methoxy-2-naphthalenyl)methyl ester (CAS No. 395645-12-2) to study the hydrolytic activity of human liver microsomes. Microsomal activity against this substrate was considered to be low (average value of ten samples 2.04 0.68 nmol min mg ), when compared to p-nitrophenyl acetate (CAS No. 830-03-5) at 3,700 2,100 mg ... 

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