Human Biochemistry, Volume 1 - Murray R. 1993

Structure and Functions of Proteins and Enzymes
Enzymes: Regulation of Activity
Regulation of Catalytic Activity of Enzymes

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When The activity of an enzyme changes As a result of physiological processes, we often cannot determine whether this is due to A change in The amount of the enzyme or in its catalytic efficiency. We will refer to all changes in enzyme activity that occur while the amount of the enzyme remains constant as changes in its catalytic efficiency.

Class="center">Table 10.1. Selected rat Liver Enzymes whose rates of synthesis vary depending on environmental conditions¹⁾

¹⁾ All data, except for HMG-CoA reductase, are taken from Annu. Rev. Biochem. 1970: 39: 929.

Reactant Concentrations

Studying the kinetic and regulatory Properties of Enzymes provides deeper insight into The Nature of physiological processes occurring in intact Cells, Tissues, and the Organism as a whole. However, most information has been obtained by studying enzymes in vitro under conditions that differ significantly from those in living cells. Therefore, extrapolating these data to in vivo conditions requires caution. For example, substrate concentrations in vitro may differ markedly from those found in vivo.

Enzyme Compartmentation

The Role of compartmentation (spatial segregation) of metabolic processes in Eukaryotic cells, including mammalian cells, can hardly be overstated. The localization of specific metabolic processes in the Cytosol or within cellular Organelles facilitates the independent regulation of these pathways. Well-developed metabolic compartmentation is particularly characteristic of higher life forms, enabling the most fine-tuned REGULATION OF METABOLISM. At the same time, this creates a new challenge: The transport of metabolites across separating barriers. This problem is solved through "shuttle mechanisms" that convert a metabolite into a form capable of crossing the barrier. Then, on the other side of the barrier, the metabolite is converted back into its original form. The presence of barriers necessitates the functioning of, for example, cytosolic and mitochondrial isoforms of certain enzymes. Because these enzyme forms are physically separated, their independent regulation is facilitated. The role of shuttle mechanisms in maintaining equilibrium among metabolic pools of reducing equivalents, Citric Acid Cycle intermediates, and various other intermediates is discussed in Chapter 17.

Macromolecular Complexes

The assembly of a set of enzymes that catalyze a multi-step sequence of metabolic reactions into a macromolecular complex allows for the coordination of enzymatic activity and channels the movement of intermediates along the metabolic pathway. Adequate spatial arrangement of the enzymes facilitates The transfer of a product from one enzyme to another without prior equilibration with the bulk metabolic pool. This provides more efficient Metabolic control than when the complex components are isolated from one another. Furthermore, Conformational Changes in one component can be transmitted to Other components of the complex via Protein-Protein Interactions, thereby amplifying regulatory effects.

Effective Concentrations of Substrates, Coenzymes, and Cations

Average intracellular concentrations of a substrate, coenzyme, or metal ion tell us little about enzyme behavior in vivo. It is necessary to know the concentrations of the relevant metabolites in the immediate microenvironment of the enzyme in question. However, even measuring metabolite concentrations in individual cellular compartments fails to account for local concentration gradients within a compartment caused, for example, by proximity to a site of metabolite entry or synthesis. Finally, insufficient attention is often paid to the distinction between total and free metabolite concentrations. For instance, the total concentration of 2,3-bisphosphoglycerate in erythrocytes is very high, whereas the concentration of free bisphosphoglycerate in these cells is comparable to that in other tissues. Erythrocytes contain approximately 5 mM Hemoglobin, which binds 1 mol of bisphosphoglycerate per mole of deoxygenated tetramer. Therefore, with a total bisphosphoglycerate content of 4 mM, the concentration of free bisphosphoglycerate in venous Blood erythrocytes is significantly lower. Similar phenomena are observed for other metabolites in the presence of Proteins that bind them tightly, substantially lowering the concentration of free metabolites.

Michaelis–Menten kinetic analysis is based on the assumption that the total Substrate Concentration equals the free substrate concentration. As we can see, this assumption may not hold true in vivo, where the free substrate concentration is frequently of the same order of magnitude as the Enzyme Concentration.

Metal Ions, which play catalytic and structural roles in the action of many enzymes (accounting for over a quarter of all known enzymes, see Chapter 9), can also exert regulatory Functions, especially when ATP serves as the substrate. In reactions where the ATP–metal ion complex acts as the substrate, maximum activity is most frequently observed at an ATP-to-metal molar ratio of ≃ 1. An excess of either metal or ATP exerts an inhibitory effect. Because nucleoside di- and triphosphates form stable complexes with divalent cations, intracellular nucleotide concentrations also affect intracellular concentrations of free metal ions and, consequently, the activity of certain enzymes. For example, in the absence of metal ions, Glutamine Synthetase in E. coli assumes a "relaxed," catalytically inactive conformation. Mg²⁺ or Mn²⁺ ions convert the synthetase into an active, "tensed" form. Moreover, adenylylation of the synthetase alters its Specificity—the enzyme preferentially utilizes Mn²⁺ rather than Mg²⁺. Finally, the activity of the adenylylated enzyme becomes sensitive to the ATP/Mg²⁺ ratio, whereas the unadenylylated form lacks this property.