Allosteric activation, of enzymes

Compounds that act as allosteric activators of enzymes are often precursors of the pathway, so this is a mechanism for feed-forward activation, increasing the activity of a controlling enzyme in anticipation of increased availability of substrate. 

Pyruvate carboxylase is the most important of the anaplerotie reactions. It exists in the mitochondria of animal cells but not in plants, and it provides a direct link between glycolysis and the TCA cycle. The enzyme is tetrameric and contains covalently bound biotin and an Mg site on each subunit. (It is examined in greater detail in our discussion of gluconeogenesis in Chapter 23.) Pyruvate carboxylase has an absolute allosteric requirement for acetyl-CoA. Thus, when acetyl-CoA levels exceed the oxaloacetate supply, allosteric activation of pyruvate carboxylase by acetyl-CoA raises oxaloacetate levels, so that the excess acetyl-CoA can enter the TCA cycle. 

Two particularly interesting aspects of the pyruvate carboxylase reaction are (a) allosteric activation of the enzyme by acyl-coenzyme A derivatives and (b) compartmentation of the reaction in the mitochondrial matrix. The carboxy-lation of biotin requires the presence (at an allosteric site) of acetyl-coenzyme A or other acylated coenzyme A derivatives. The second half of the carboxylase reaction—the attack by pyruvate to form oxaloacetate—is not affected by CoA derivatives. 

Because this enzyme catalyzes the committed step in fatty acid biosynthesis, it is carefully regulated. Palmitoyl-CoA, the final product of fatty acid biosynthesis, shifts the equilibrium toward the inactive protomers, whereas citrate, an important allosteric activator of this enzyme, shifts the equilibrium toward the active polymeric form of the enzyme. Acetyl-CoA carboxylase shows the kinetic behavior of a Monod-Wyman-Changeux V-system allosteric enzyme (Chapter 15). 

Ghanges in the availability of substrates are responsible for most changes in metabolism either directly or indirectly acting via changes in hormone secretion. Three mechanisms are responsible for regulating the activity of enzymes in carbohydrate metabolism (1) changes in the rate of enzyme synthesis, (2) covalent modification by reversible phosphorylation, and (3) allosteric effects. 

Binding of a reversible inhibitor to an enzyme is rapidly reversible and thus bound and unbound enzymes are in equilibrium. Binding of the inhibitor can be to the active site, or to a cofactor, or to some other site on the protein leading to allosteric inhibition of enzyme activity. The degree of inhibition caused by a reversible inhibitor is not time-dependent the final level of inhibition is reached almost instantaneously, on addition of inhibitor to an enzyme or enzyme-substrate mixture. 

The enzyme is also inhibited by fructose 2,6-bi pho phate, which also functions as an allosteric activator of glycolysis. 

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

Phosphofructokinase-1 is a regulatory enzyme (Chapter 6), one of the most complex known. It is the major point of regulation in glycolysis. The activity of PFK-1 is increased whenever the cell s ATP supply is depleted or when the ATP breakdown products, ADP and AMP (particularly the latter), are in excess. The enzyme is inhibited whenever the cell has ample ATP and is well supplied by other fuels such as fatty acids. In some organisms, fructose 2,6-bisphosphate (not to be confused with the PFK-1 reaction product, fructose 1,6-bisphosphate) is a potent allosteric activator of PFK-1. The regulation of this step in glycolysis is discussed in greater detail in Chapter 15. 

Feedback can also be positive. Since AMP is a product of the hydrolysis of ATP, its accumulation is a signal to activate key enzymes in metabolic pathways that generate ATP. For example, AMP causes allosteric activation of glycogen phosphorylase, which catalyzes the first step in the catabolism of glycogen. As is shown in Fig. 11-5, the allosteric site for AMP or IMP binding is more than 3 nm away from the active site. Only a... 

The effects of ATP, AMP, and fructose 2,6-bisphos-phate on phosphofructokinase have been discussed in Chapter 11, Section C. Fructose 2,6-P2 is a potent allosteric activator of phosphofructokinase and a strong competitive inhibitor of fructose 1,6-bisphosphatase (Fig. 11-2). It is formed from fructose 6-P and ATP by the 90-kDa bifunctional phosphofructo-2-kinase/ fructose 2,6-bisphosphatase. Thus, the same protein forms and destroys this allosteric effector. Since the bifunctional enzyme is present in very small amounts, the rate of ATP destruction from the substrate cycling is small. 

Muscle pyruvate kinase (PK) responds hyperbolically to its substrate, PEP, but the liver form of the enzyme responds sigmoidally. Fructose-1,6-bisphos-phate is an allosteric activator of liver pyruvate kinase, but it apparently has no effect on the muscle enzyme. 

Stadtman, E. R. (1966). Allosteric Regulation of Enzymic Activity. Adv Enzymol 28 41. 

The activity of enzymes (and indeed of the functionality of proteins in general) can be regulated by reversibly binding ligands (allosteric effectors) and by covalent modification (that can be either reversible or irreversible). 

The most comprehensive set of kinetic studies on AdoCbl homolysis have been performed with AdoCbl-dependent ribonucleotide reductase by Stubbe and coworkers (Licht et al., 1999a Licht et al., 1999b). In this enzyme, AdoCbl is used to generate a thiyl radical on a cysteine residue it is this thiyl radical that abstracts hydrogen from the 3-position of the ribonucleotide to faeilitate reduction at C-2. In the presence of dGTP, an allosteric activator of the enzyme, the enzyme catalyzes the reversible cleavage of AdoCbl and formation of thiyl radical in the absence of substrate. This partial reaction proceeds rapidly enough for it to be mechanistically relevant and provides a system to study AdoCbl homolysis which is both simple and amenable to detailed kinetic analysis. 

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