Catalytic activity, enzymes reaction

The catalytically active enzyme substrate complex is an interactive structure in which the enzyme causes the substrate to adopt a form that mimics the transition-state intermediate of the reaction. Thus, a poor substrate would be one that was less effective in directing the formation of an optimally active enzyme transition-state intermediate conformation. This active conformation of the enzyme molecule is thought to be relatively unstable in the absence of substrate, and free enzyme thus reverts to a conformationally different state. 

However, there are disadvantages to using immobilised cells. The cell may contain numerous catalytically active enzymes, which may catalyse unwanted side reactions. Also, the cell membrane itself may serve as a diffusion barrier, and may reduce productivity. The matrix may sharply reduce productivity if the microorganism is sensitive to product inhibition. One of the disadvantages of immobilised cell reactors is that the physiological state of the microorganism cannot be controlled. 

Notably, natural variation in the type III PKS active site cavity, like that observed in Ipomoea and Petunia, does not result in functionally impaired enzymes, but in fact, generates catalytically active enzymes that display both altered substrate and product specificities. Sequential increases in the side chain volume of position 256 in alfalfa CHS2 result in decreases in polyketide chain length and predictable shifts in the ratio of tetraketide to triketide reaction products.32 These results functionally link the volume of the elongation/cyclization lobe in type III PKS to chain length determination. 

The production of abzymes on a substantial scale is tedious [577] and preparative-scale reactions are still in the micromolar range [578,579]. Furthermore, to avoid the occurrence of catalytically active enzyme impurities, the purity of the abzymes must be very high. 

An edited volume entitled Molecular Movements and Chemical Reactivity as Conditioned by Membranes, Enzymes, and Other Macromolecules includes sections on synzymes (synthetic polymers with enzyme-like catalytic activities), diffusion-reaction in structured media and membranes bearing enzymes, and structures and energetics of proteins and their active sites. ... 

Chemical reactions between biochemical compounds are enhanced by biological catalysts called enzymes, which consist mostly or entirely of globular proteins. In many cases a cofactor is needed to combine with an otherwise inactive protein to produce the catalytically active enzyme complex. The two distinct varieties of cofactors are coenzymes, which are complex organic molecules, and metal ions. Enzymes catalyze six major classes of reactions 1) Oxidoreductases (oxidation-reduction reactions), 2) Transferases (transfer of functional groups), 3) Hydrolases (hydrolysis reactions), 4) Lyases (addition to double bonds, 5) Isomerases (isomerization reactions) and 6) Ligases (formation of bonds with ATP (adenosine triphosphate) cleavage) [1]. 

Most reactions in cells are carried out by enzymes [1], In many instances the rates of enzyme-catalysed reactions are enhanced by a factor of a million. A significantly large fraction of all known enzymes are proteins which are made from twenty naturally occurring amino acids. The amino acids are linked by peptide bonds to fonn polypeptide chains. The primary sequence of a protein specifies the linear order in which the amino acids are linked. To carry out the catalytic activity the linear sequence has to fold to a well defined tliree-dimensional (3D) stmcture. In cells only a relatively small fraction of proteins require assistance from chaperones (helper proteins) [2]. Even in the complicated cellular environment most proteins fold spontaneously upon synthesis. The detennination of the 3D folded stmcture from the one-dimensional primary sequence is the most popular protein folding problem. 

In contrast to the situation in the absence of catalytically active Lewis acids, micelles of Cu(DS)2 induce rate enhancements up to a factor 1.8710 compared to the uncatalysed reaction in acetonitrile. These enzyme-like accelerations result from a very efficient complexation of the dienophile to the catalytically active copper ions, both species being concentrated at the micellar surface. Moreover, the higher affinity of 5.2 for Cu(DS)2 compared to SDS and CTAB (Psj = 96 versus 61 and 68, respectively) will diminish the inhibitory effect due to spatial separation of 5.1 and 5.2 as observed for SDS and CTAB. 

Many globular proteins are enzymes They accelerate the rates of chemical reactions m biological systems but the kinds of reactions that take place are the fundamental reactions of organic chemistry One way m which enzymes accelerate these reactions is by bringing reactive func tions together m the presence of catalytically active functions of the protein... 

The earliest examples of analytical methods based on chemical kinetics, which date from the late nineteenth century, took advantage of the catalytic activity of enzymes. Typically, the enzyme was added to a solution containing a suitable substrate, and the reaction between the two was monitored for a fixed time. The enzyme s activity was determined by measuring the amount of substrate that had reacted. Enzymes also were used in procedures for the quantitative analysis of hydrogen peroxide and carbohydrates. The application of catalytic reactions continued in the first half of the twentieth century, and developments included the use of nonenzymatic catalysts, noncatalytic reactions, and differences in reaction rates when analyzing samples with several analytes. 

Potentiometry is another useful method for determining enzyme activity in cases where the reaction Hberates or consumes protons. This is the so-called pH-stat method. pH is kept constant by countertitration, and the amount of acid or base requited is measured. An example of the use of this method is the determination of Hpase activity. The enzyme hydroly2es triglycerides and the fatty acids formed are neutralized with NaOH. The rate of consumption of NaOH is a measure of the catalytic activity. 

Usually, a rapid binding step of the inhibitor I to the enzyme E leads to the formation of the initial noncovalent enzyme-inhibitor complex E-I. This is usually followed by a rate determining catalytic step, leading to the formation of a highly reactive species [E—I ]. This species can either undergo reaction with an active site amino acid residue of the enzyme to form the covalent enzyme-inhibitor adduct E—I", or be released into the medium to form product P and free active enzyme E. 

Enzymes are excellent catalysts for two reasons great specificity and high turnover rates. With but few exceptions, all reac tions in biological systems are catalyzed by enzymes, and each enzyme usually catalyzes only one reaction. For most of the important enzymes and other proteins, the amino-acid sequences and three-dimensional structures have been determined. When the molecular struc ture of an enzyme is known, a precise molecular weight could be used to state concentration in molar units. However, the amount is usually expressed in terms of catalytic activity because some of the enzyme may be denatured or otherwise inactive. An international unit (lU) of an enzyme is defined as the amount capable of producing one micromole of its reaction product in one minute under its optimal (or some defined) reaction conditions. Specific activity, the activity per unit mass, is an index of enzyme purity. 

Oxidation of P-nicotinamide adenine dinucleotide (NADH) to NAD+ has attracted much interest from the viewpoint of its role in biosensors reactions. It has been reported that several quinone derivatives and polymerized redox dyes, such as phenoxazine and phenothiazine derivatives, possess catalytic activities for the oxidation of NADH and have been used for dehydrogenase biosensors development [1, 2]. Flavins (contain in chemical structure isoalloxazine ring) are the prosthetic groups responsible for NAD+/NADH conversion in the active sites of some dehydrogenase enzymes. Upon the electropolymerization of flavin derivatives, the effective catalysts of NAD+/NADH regeneration, which mimic the NADH-dehydrogenase activity, would be synthesized [3]. 

The turnover number of an enzyme, is a measure of its maximal catalytic activity, is defined as the number of substrate molecules converted into product per enzyme molecule per unit time when the enzyme is saturated with substrate. The turnover number is also referred to as the molecular activity of the enzyme. For the simple Michaelis-Menten reaction (14.9) under conditions of initial velocity measurements, Provided the concentration of... 

But k must always be greater than or equal to k h / (A i + kf). That is, the reaction can go no faster than the rate at which E and S come together. Thus, k sets the upper limit for A ,. In other words, the catalytic effieiency of an enzyme cannot exceed the diffusion-eontroUed rate of combination of E and S to form ES. In HgO, the rate constant for such diffusion is approximately (P/M - sec. Those enzymes that are most efficient in their catalysis have A , ratios approaching this value. Their catalytic velocity is limited only by the rate at which they encounter S enzymes this efficient have achieved so-called catalytic perfection. All E and S encounters lead to reaction because such catalytically perfect enzymes can channel S to the active site, regardless of where S hits E. Table 14.5 lists the kinetic parameters of several enzymes in this category. Note that and A , both show a substantial range of variation in this table, even though their ratio falls around 10 /M sec. 

Many enzymes require metal ions for maximal activity. If the enzyme binds the metal very tightly or requires the metal ion to maintain its stable, native state, it is referred to as a metalloenzyme. Enzymes that bind metal ions more weakly, perhaps only during the catalytic cycle, are referred to as metal activated. One role for metals in metal-activated enzymes and metalloenzymes is to act as electrophilic catalysts, stabilizing the increased electron density or negative charge that can develop during reactions. Among the enzymes that function in this... 

The specificity of enzyme reactions can be altered by varying the solvent system. For example, the addition of water-miscible organic co-solvents may improve the selectivity of hydrolase enzymes. Medium engineering is also important for synthetic reactions performed in pure organic solvents. In such cases, the selectivity of the reaction may depend on the organic solvent used. In non-aqueous solvents, hydrolytic enzymes catalyse the reverse reaction, ie the synthesis of esters and amides. The problem here is the low activity (catalytic power) of many hydrolases in organic solvents, and the unpredictable effects of the amount of water and type of solvent on the rate and selectivity. 

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