Controllability of enzymes

The serine residue of isocitrate dehydrogenase that is phos-phorylated by protein kinase lies within the active site of the enzyme. This situation contrasts with most other examples of covalent modification by protein phosphorylation, where the phosphorylation occurs at a site remote from the active site. What direct effect do you think such active-site phosphorylation might have on the catalytic activity of isocitrate dehydrogenase (See Barford, D., 1991. Molecular mechanisms for the control of enzymic activity by protein phosphorylation. Bioehimiea et Biophysiea Acta 1133 55-62.)... 

ALLOSTERIC HORMONAL MECHANISMS ARE IMPORTANT IN THE METABOLIC CONTROL OF ENZYME-CATALYZED REACTIONS... 

Cohen P Control of Enzyme Activity, 2nd ed. Chapman Hall, 1983. 

A common characteristic of metabolic pathways is that the product of one enzyme in sequence is the substrate for the next enzyme and so forth. In vivo, biocatalysis takes place in compartmentalized cellular structure as highly organized particle and membrane systems. This allows control of enzyme-catalyzed reactions. Several multienzyme systems have been studied by many researchers. They consist essentially of membrane- [104] and matrix- [105,106] bound enzymes or coupled enzymes in low water media [107]. 

The term intermediary metabolism is used to emphasize the fact that metabolic processes occur via a series of individual chemical reactions. Such chemical reactions are usually under the control of enzymes which act upon a substrate molecule (or molecules) and produce a product molecule (or molecules) as shown in Figure 1.1. The substrates and products are referred to collectively as intermediates or metabolites . The product of one reaction becomes the substrate for another reaction and so the concept of a metabolic pathway is created. 

A third means of control of enzyme activity is achieved by altering the conformation of the protein. The actual mechanisms involved are ... 

As indicated in the previous discussion, the control of enzyme activity is understood in terms of kinetic parameters. Differences in Km and or Vmax can also arise when the same chemical (metabolic) reaction is catalysed by two structurally different enzymes. Such is the case with isoenzymes or isoforms of enzymes. 

Control of pymvate dehydrogenase activity is via covalent modification a specific kinase causes inactivation of the PDH by phosphorylation of three serine residues located in the pyruvate decarboxylase/dehydrogenase component whilst a phosphatase activates PDH by removing the phosphates. The kinase and phosphatase enzymes are non-covalently associated with the transacetylase unit of the complex. Here again we have an example of simultaneous but opposite control of enzyme activity, that is, reciprocal regulation. 

There are many examples of phosphorylation/dephosphorylation control of enzymes found in carbohydrate, fat and amino acid metabolism and most are ultimately under the control of a hormone induced second messenger usually, cytosolic cyclic AMP (cAMP). PDH is one of the relatively few mitochondrial enzymes to show covalent modification control, but PDH kinase and PDH phosphatase are controlled primarily by allosteric effects of NADH, acetyl-CoA and calcium ions rather than cAMP (see Table 6.6). 

Zacharis, E., Moore, B.D. and Hailing, P.J., Control of enzyme activity in organic media by solid-state acid-base buffers. J. Am. Chem. Soc., 1997, 119, 12396-12397. 

Stehle, P, Bahsitta, H. P., and Piirst, P. (1986). Analytical control of enzyme-catalyzed peptide-synthesis using capillary isotachophoresis.. Chromatogr. 370, 131—138. 

The third type of inhibition is called allosteric inhibition, and is particularly important in the control of intermediary metabolism This refers to the ability of enzymes to change their shape (tertiary and quaternary structure, see Section 13.3) when exposed to certain molecules. This sometimes leads to inhibition, whereas in other cases it may actually activate the enzyme. The process allows subtle control of enzyme activity according to an organism s demands. Further consideration of this complex phenomenon is outside our immediate needs. 

Control of enzymic activity arising from the modulated access of substrates to a channel leading to the active site. Such a scheme was suggested for aspartate carbamo-yltransferase which has its complement of active sites located on the interior surface of the complex comprised of catalytic and regulatory subunits. Nonetheless, isotope exchange studies of this enzyme suggest that this form of enzyme regulation does not apply in the case of aspartate transcarbamoylase . ... 

Schimke, R.T. and Doyle, D. (1970). Control of enzyme levels in animal tissues. Annu. Rev. Biochem. 39, 929-976. 

Most of the other posttranslational modifications involving the N- or C-terminus (Table 1) as well as the side-chain functionalities (Table 2) of the polypeptide chains occur under the control of enzymes that also dictate the regioselectivity of such chemical transformations. This regioselectivity is difficult to attain by synthetic procedures. Sophisticated protection schemes are required when additional chemistry must be performed on preassembled peptides, unless enzymatic methods can be used to supplement the synthetic strategies. As a consequence, the use of suitably modified amino acids as synthons is generally the preferred approach as will be discussed in the following sections. 

What is the minimum size of synzymes And is there any control effect observed that is analogous to the allosteric control of enzyme activities ... 

The control of enzyme activity by the environment of a polyatomic framework is a vast topic, which I shall not attempt to cover fully in this report. Instead I will concentrate on some selected interactions between and within polypeptide chains that influence enzymatic activity. First, elementary steps involved in ligand-protein, intraprotein, and interprotein interactions are considered. Then enzymes consisting of a single polypeptide chain are discussed, followed by enzymes consisting of multiple polypeptide chains. The concluding sections are concerned with multienzyme complexes and enzymes associated with membranes. 

This review has tried to present an overview of the control of enzymic activity in complex polyatomic frameworks. The examples discussed are intended to be representative obviously many other examples could be cited. The elementary interactions involved in modulating enzymic activity are well understood in terms of thermodynamics, kinetics, and structure. A considerable amount of information is also available for the simplest type of macromolecular framework, enzymes consisting of a single polypeptide chain, although a considerable amount of work remains to be done. 

The catalytic activity of an enzyme in an organic medium often varies by several orders of magnitude depending on the degree of enzyme hydration [9,19]. Control of enzyme hydration or water activity is thus a key issue when optimizing enzymatic conversions in organic solvents. 

Rates of enzyme reactions are often affected by the presence of various chemicals and ions. Enzyme inhibitors combine, either reversibly or irreversibly, with enzymes and cause a decrease in enzyme activity. Effectors control enzyme reactions by combining with the regulatory site(s) of enzymes. There are several mechanisms of reversible inhibition and for the control of enzyme reactions. 

Synthesis of many enzymes is repressed most of the time. The appearance of an enzyme at a particular stage in the life of an organism as well as the differing distributions of isoenzymes within differentiated tissue result from derepression. The control of enzyme synthesis may also be exerted during the splicing of transcripts and at the translational level as well. These control mechanisms are often relatively slow, with response times of hours or even days. However, effects on the synthesis of some hormones, such as insulin (Section G), may be observed within a few minutes. 

Organic chemists have not had much use for prochirality, but it is an important concept for biochemists following the stereochemistry of bio-organic reactions. Almost all biochemical reactions are under the control of enzymes, which function asymmetrically even on symmetrical (but prochiral) molecules. Thus it has been found that only one of the two methylene groups of... 

This section summarizes recent data obtained from biophysical techniques on the nature of the metal ion, catalytic, and modifier sites of E. coli glutamine synthetase. Distance relationships between these sites are given in Fig. 27. This enzyme has many diverse modes of biochemical control. Recent data presented in this review demonstrate that progress is being made toward understanding the structural basis for the control of enzymic catalysis. When x-ray data become available 127), the amino acid groups at the catalytic site will be known and will provide further details concerning the mechanism of the enzymic reaction. 

In allosteric enzymes, the activity of the enzyme is modulated by a non-covalently bound metabolite at a site on a protein other than the catalytic site. Normally, this results in a conformational change, which makes the catalytic site inactive or less active. Covalent modulated enzymes are interconverted between active and inactive forms by the action of other enzymes, some of which are modulated by allosteric-type control. Both of these control mechanisms are responsive to changes in cell conditions and typically the response time in allosteric control is a matter of seconds as compared with minutes in covalent modulation. A third type of control, the control of enzyme synthesis at the transcription stage of protein synthesis (see Appendix 5.6), can take several hours to take effect. 

The concept of control of metabolic activity by allosteric enzymes or the control of enzyme activity by ligand-induced conformational changes arose from the study of metabolic pathways and their regulatory enzymes. A good example is the multi-enzymatic sequence catalysing the conversion of L-threonine to L-isoleucine shown in Fig. 5.32. 

Fig. 5.39. The control of enzyme synthesis by catabolite repression. A control region of the lac operon contains the CAP binding site within the promoter region...

The most common way of regulating metabolic activity is by direct control of enzyme activity. Enzyme activities are usually regulated by noncovalent interaction with small-molecule regulatory factors (see chapter 9) or by a reversible covalent modification, such as phosphorylation or... 

Cohen, P., Control of Enzyme Activity, 2d ed. London and New York Chapman and Hall, 1983. Brief discussion of some types of regulation of activity of metabolic enzymes, emphasizing regulation by covalent modification of the enzymes. 

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