Specificity, enzymatic reaction

The previous chapters taught us how to ask questions about specific enzymatic reactions. In this chapter we will attempt to look for general trends in enzyme catalysis. In doing so we will examine various working hypotheses that attribute the catalytic power of enzymes to different factors. We will try to demonstrate that computer simulation approaches are extremely useful in such examinations, as they offer a way to dissect the total catalytic effect into its individual contributions. 

Protein phosphorylation is a specific enzymatic reaction in which one protein serves as a substrate for a protein kinase. Protein kinases are phophotransferases. They catalyze the transfer of a phosphate group from ATP to an acceptor amino acid in the substrate protein (fig. 2.10). A detailed discussion of protein kinases can be found in chapter 7. 

The ubiquitinylation of proteins is a complex process which involves several specific enzymatic reactions. Three sequential steps can be distinguished (fig. 2.15A) ... 

Both in nature and in test tubes many complicated reactions take place with apparent ease. Highly specific enzymatic reactions involving complementarity of substrate and receptor both with respect to their spatial structure and electrostatic fields lie outside the scope of this book. Here we would like only to mention a few... 

Although the Michaelis-Menten equation places emphasis on V max, published kinetic studies of enzymes generally do not disclose the V max for specific enzymatic reactions. Instead papers report kcaV Deep in the experimental section, a paper will mention the total concentration of the enzyme, [Et], used in the kinetic study. Since k2 = kcat, with both kcat and [Et], one may use Equation 4.10 to estimate Vmax for the reaction of interest. 

An alternative set of non-specific enzymatic reactions can reveal masked polar functional groups which might be present in a drug. For example, there are enzymes which can demethylate a methyl ether to reveal a more polar hydroxyl group. Once again, the more polar product (metabolite) is excreted more efficiently. 

By preserving thermo-sensitive compounds, the new process is also particularly attractive for vegetable oUs, which represent very important part of the food and health industry. In addition to reducing the viscosity of the oil for purification purposes, we can also foresee its composition being modified by specific enzymatic reactions at the pore level this is the concept of the active membrane, which follows the immobilization of enzymes at membrane walls. The goal would be to obtain high-added-value products from cheap and plentiful raw substrates. 

Cells are able to cany out all of the functions of life. During cell duplication (reproduction), the DNA is replicated (one double-stranded molecule is converted to two double-stranded molecules) by specific enzymatic reactions, and the newly formed DNA is separated into two separate cells produced by growth and division of the former single cell. In bacteria, this asexual process of reproduction is the primary means of multiplication to produce new individuals. In contrast, protists, plants, fungi, and... 

Besides the design of synthetic molecules, several papers reported the presence of specific enzymatic reactions able to produce also in the cells compounds able to act as reversible FAAH inhibitors (Maccarrone and Finazzi-Agro, 2004a). In particular, it has been found that oxidative metabolites of AEA generated by various lipoxygenases, i.e. the hydroxyanandamides (HAEAs see 12-OH-AEA in Table 4.1), are powerful inhibitors of FAAH (van der Stelt et al., 2002). Instead, derivatives of AEA generated by cyclooxygenase-2, and termed prosta-mides, have been recently shown to be inelfective on FAAH activity (Matias et al., 2004). 

The modern tests employ specific enzymatic reactions and provide a fairly accurate measurement over the broad range of blood glucose concentrations. The concentration of gluco.se can be read visually (Fig. 1) or with the aid of portable reflectance meters. It is important to follow the measuring procedure exactly. Improper storage of reagent strips can affect the results. Any grossly abnormal or unexpected results should be confimied in the laboratory. 

Triamcinolone may be prepared by a specific enzymatic reaction with fluorocortisone whereby the a-hydroxy group is strategically introduced at C-16. The resulting product is subsequently ineubated with the microbe Corynehacterium simplex which causes the dehydrogenation between C-1 and C-2 to produce the official compound triamcinolone. 

Water-soluble vitamins participate in a vast number of enzymatic reactions, in most cases in the form of co-enzymes. Many of these enzymatic reactions are well characterized. Many biochemical pathways in the body necessitate several different and specific enzymatic reactions, each having its own vitamin-containing co-factors. 

HS-C protein is poly(ADP-ribose) polymerase. To test this possibility further, the protein was subjected to ADP-ribosylation with [i C]NAD and nicked DNA. The reaction product was subjected to SDS-PAGE. Most of the radioactivity was located near the origin or at a position that approximately corresponded to the Coomassie blue-stained protein band in a parallel track (data not shown). Since poly(ADP-ribose) polymerase is capable of auto-ADP-ribosylation and this modification can alter electrophoretic mobility (6), the two radioactive peaks observed after ADP-ribosylation of HS-C protein probably correspond to the different levels of ADP-ribosylation occurring in the enzyme molecule. That the radioactive peaks were indeed generated by a specific enzymatic reaction was shown by the lack of incorporation of [i C]NAD at 4°C or in the absence of nicked DNA (data not shown). 

Ehrbar, M., Rizzi, S. C., Schoenmakers, R. G., et al. (2007) Biomolecular hydrogels formed and degraded via site-specific enzymatic reactions. Biomacromolecules, 8, 3000-7. 

PD Fraser, M Albrecht, G Sandmann. Development of high-performance liquid chromatographic systems for the separation of radiolabelled carotenes and precursors formed in specific enzymatic reactions. J Chromatogr 645 265-272, 1993. 

As shown in Table 1.2, metalloenzymes contain specific numbers of metal ions at the active sites in specific proteins. The presence of metal ions allows metalloenzymes to perform functions as biocatalysts for specific enzymatic reactions including gene (DNA, RNA) synthesis, metabolism, antioxidation and so forth. 

Disaccharides such as sucrose can have their structures established by methylation followed by hydrolysis as well as various specific enzymatic reactions. 

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