Heterotropic allosteric enzymes

For heterotropic allosteric enzymes, those whose modulators are metabolites other than the normal substrate, it is difficult to generalize about the shape of the substrate-saturation curve. An activator may cause the curve to become more nearly hyperbolic, with a decrease in Z0.5 but no change in Fmax, resulting in an increased reaction velocity at a fixed substrate concentration (V0 is higher for any value of [S] Fig. 6-29b, upper curve). 

Other heterotropic allosteric enzymes respond to an activator by an increase in Fmax with little change in if0i5 (Fig. 6-29c). A negative modulator (an inhibitor) may produce a more sigmoid substrate-saturation curve,... 

When binding of a substrate molecule at an enzyme active site promotes substrate binding at other sites, this is called positive homotropic behavior (one of the allosteric interactions). When this co-operative phenomenon is caused by a compound other than the substrate, the behavior is designated as a positive heterotropic response. Equation (6) explains some of the profile of rate constant vs. detergent concentration. Thus, Piszkiewicz claims that micelle-catalyzed reactions can be conceived as models of allosteric enzymes. A major factor which causes the different kinetic behavior [i.e. (4) vs. (5)] will be the hydrophobic nature of substrate. If a substrate molecule does not perturb the micellar structure extensively, the classical formulation of (4) is derived. On the other hand, the allosteric kinetics of (5) will be found if a hydrophobic substrate molecule can induce micellization. 

One of the first known examples of allosteric feedback inhibition was the bacterial enzyme system that catalyzes the conversion of L-threonine to L-isoleucine in five steps (Fig. 6-28). In this system, the first enzyme, threonine dehydratase, is inhibited by isoleucine, the product of the last reaction of the series. This is an example of heterotropic allosteric inhibition. Isoleucine is quite specific as an inhibitor. No other intermediate in this sequence inhibits threonine dehydratase, nor is any other enzyme in the sequence inhibited by isoleucine. Isoleucine binds not to the active site but to another specific site on the enzyme molecule, the regulatory site. This binding is noncovalent and readily reversible if the isoleucine concentration decreases, the rate of threonine dehydration increases. Thus threonine dehydratase activity responds rapidly and reversibly to fluctuations in the cellular concentration of isoleucine. 

Fig. 9-10 Behavior of an MWC allosteric enzyme in the presence of positive and negative heterotropic effectors. The activator term, y, in Eq. (9.62) causes the curve to become more hyperbolic, whereas the inhibitor term (j3) renders it more sigmoidal. The curves were constructed using Eq. (9.62) with L = 1,000 and n - 4.

The product of this reaction, oxaloacetate, can either enter the gluconeogenic pathway (Chap. 11) by way of malate or condense with acetyl-CoA to yield citrate. Pyruvate carboxylase is an allosteric enzyme, and it is activated by the heterotropic effector, acetyl-CoA. Thus, pyruvate in the mitochondria is the substrate for either pyruvate dehydrogenase or pyruvate carboxylase, the activities of which, in turn, are controlled by reactants associated with the citric acid cycle. The interplay among pyruvate dehydrogenase, pyruvate carboxylase, pyruvate, and the citric acid cycle is shown in Fig. 12-9. 

Relationship between the initial velocity (v) and the substrate concentration [S] for an allosteric enzyme that shows a heterotropic effect with constant Vmax but with varying ATq.s- Curve a is obtained in the absence of any modulators, curve b in the presence of a positive modulator, and curve c in the presence of a negative modulator. Regulation is achieved by modulation of ATq.s without change in F ax. 

The tryptophan synthase bienzyme complex from enteric bacteria provides an important example wherein RSSF has been used to good advantage for the study of both enzyme mechanism and protein structure-function relationships. This enzyme complex is composed of heterologous a- and P2-subunits arranged in a nearly linear a-(3-(l-a array (81). The a-subunit catalyzes the aldolytic cleavage of IGP to indole and G3P, while the P-subunit catalyzes the PLP-dependent condensation of i-Ser and indole to yield i-Trp. The aP-reaction is essentially the sum of the individual a- and P-reactions (scheme I). Indole, the common intermediate produced at the a-site, is direcdy channeled to the P-active site via a tunnel located in the interior of the protein complex which directly interconnects the a- and P-catalytic centers (81-84). Although the individual subunits may be isolated and are functional, formation of the bienzyme complex not only increases the catalytic activities of the separate subunits by nearly 100-fold, but also alters the thermodynamic stability of P-site reaction intermediates and introduces heterotropic allosteric interactions between sites. 

Enzyme activity can also be affected by binding of substrate and nonsubstrate Mgands, which can act as activators or inhibitors, at a site other than the active site. These enzymes are called allosteric. These responses can be homotropic or heterotropic. Homotropic responses refer to the allosteric modulation of enzyme activity strictly by substrate molecules heterotropic responses refer to the allosteric modulation of enzyme activity by nonsubstrate molecules or combinations of substrate and nonsubstrate molecules. The allosteric modulation can be positive (activation) or negative (inhibition). Many allosteric enzymes also display cooperativity, making a clear differentiation between allosterism and cooperativity somewhat difficult. 

Heterotropic effectors The effector may be different from the substrate, in which case the effect is said to be heterotropic. For example, consider the feedback inhibition shown in Figure 5.17. The enzyme that converts A to B has an allosteric site that binds the end-product, E. If the concentration of E increases (for example, because it is not used as rapidly as it is synthesized), the initial enzyme in the pathway is inhibited. Feedback inhibition provides the cell with a product it needs by regulating the flow of substrate molecules through the pathway that synthesizes that product. [Note Heterotropic effectors are commonly encountered, for example, the glycolytic enzyme phosphofructokinases allosterically inhibited by citrate, which is not a substrate for the enzyme (see p. 97).]... 

Let us first define two terms. Homotropic effects are allosteric interactions that occur when several identical molecules are bound to a protein. The binding of substrate molecules to different sites on an enzyme, such as the binding of aspartate to ATGase, is an example of a homotropic effect. Heterotropic effects are allosteric interactions that occur when different substances (such as inhibitor and substrate) are bound to the protein. In the ATGase reaction, inhibition by GTP and activation by ATP are both heterotropic effects. 

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