The control of enzyme activity

The first involves various feedback and feed forward control mechanisms associated with metabolic pathways. Here a chemical present in the cell, usually an end product of a metabolic sequence, will influence the activity of an enzyme at the beginning of the pathway. This is usually achieved by the presence of an allosteric enzyme whose properties are significantly changed by effector molecules (Fig. 5.31). 

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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.)... 

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. 

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 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. 

The control of enzyme activity by allosteric regulation of key enzymes is of immense benefit to an organism because it allows the concentration of cellular products to be maintained within very narrow limits. 

List three mechanisms for the control of enzyme activity. 

A further compHcation remains in the control of enzyme activities which to a large extent is dependent upon expression during the fermentation. One potential solution is to add supplementary enzymes to the cell-free extract. Such a precedent has already been set by a few reported cases where whole cells were mixed with the isolated enzyme for ex vivo cofactor recycle. E>espite these problems, there is no doubt that as genetic engineering for expression of the desired enzyme is improved, more systems wiU be tested in the cell-free environment [26]. At the very least it is clear that cell-free extracts, combined with network topology analysis can provide an excellent basis for effective analysis and targeting of the network so as to insulate the desired pathway from undesired enzymatic reactions [27]. 

Since the control of enzyme activity can be analyzed quantitatively by using power approximations, it is possible to apply this to the control of an activity by the adenine nucleotides. Let us assume that an activity ( ) is influenced by the concentrations of the three nucleotides, then... 

Reagents. The measurement of enzyme activities requires rigid control of the analytical conditions, including accurate measurement of reagent and sample volumes, and careful control of temperature, pH and reagent stability. 

The above conclusion is supported by the results shown in figure 4. Just as inhibitors of the 5-HT uptake carrier can antagonize MDMA-induced [ H]5-HT release in vitro, coadministration of MDMA with an uptake inhibitor such as citalopram can completely block the acute depletion of 5-HT. Although citalopram also antagonized the MDMA-induced decrease in TPH activity, there was still a significant loss of enzyme activity when compared to control. This implies that if MDMA requires access to the interior of the nerve terminals to affect TPH activity, it does not require the activity of the uptake carrier to gain entrance. Hence, these results are consistent with the outcome of synaptosomal uptake experiments with [ HJMDMA (Schmidt et al. 1987), which show that MDMA is not actively concentrated by a carrier system. Furthermore, it is apparent that the loss of enzyme activity alone is not sufficient to reduce 5-HT concentrations, but that release via the carrier must occur simultaneously, to deplete the terminal once synthetic capacity is reduced. 

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

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. 

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 . ... 

Loss of the Activity and the Change in Primary Structure, Ozone causes the loss of enzymic activity in lysozyme as shown in Figure 1, The enzyme is inactivated linearly until the activity is reduced to 5%, Prolonging exposure of lysozyme to ozone causes further decrease in enzymic activity. Control experiments, using gas streams without ozone, were carried out in the same conditions and over the same periods. Figure 2 shows that the activity of both control and ozonized lysozyme varies with pH change. Although the ozonized product shows more extensive dependence on pH than the native one, the optimum is the same. 

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

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. 

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 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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