Regulating Metabolic Pathways

An inhibitor is any agent that interferes with the activity of an enzyme. Inhibitors may affect the binding of enzyme to substrate, or catalysis (via modification of the enzyme s active site), or both. Researchers use enzyme inhibitors to define metabolic pathways and to understand enzyme reaction mechanisms. Many drugs are designed as inhibitors of target enzymes. Inhibition is also a natural phenomenon. Cells regulate metabolic pathways by specific inhibition of key enzymes. 

The quantitative study of enzyme catalysis, referred to as enzyme kinetics, provides information about reaction rates. Kinetic studies also measure the affinity of enzymes for substrates and inhibitors and provide insight into reaction mechanisms. Enzyme kinetics has several practical applications. These include a greater comprehension of the forces that regulate metabolic pathways and the design of improved therapies. 

Development of differentiated cells of bacilli or strep-tomycetes is accompanied by accumulation of adenosine and guanosine highly phosphorylated nucleotides. Guano-sine-5 -diphosphate-3 -diphosphate (ppGpp) is known to act as a pleiotropic effector regulating metabolic pathways and transcription of different operons during nu-... 

The number of substances that can be measured by monoenzymatic approaches in electrochemical biosensors is limited, because in the majority of biocatalytic reactions electrochemically active compounds are not involved. To form readily detectable species, different enzymatic reactions have to be coupled, as is already routine in wet biochemical analysis [13]. This coupling can be accomplished in ways analogous to those present in a living cell. Here, nature provides us a variety of ways of regulating metabolic pathways. Thus, like in nature, catalytic activities of different enzymes can be combined in biosensors either in sequence, competing pathways, or in cycles (Table 4). 

Phosphorylation of the hydroxyl group of a serine, threonine or tyrosine residue in the enzyme (Figure 3-3), thus altering the chemical nature of its catalytic site. As discussed in section 10.3, phosphorylation of the enzyme is also important in regulating metabolic pathways, especially in response to hormone action. 

There is an obvious need to regulate metabolic pathways within individual cells, so as to ensure that catabolic and biosynthetic pathways are not attempting to operate at the same time. There is also a need to integrate and coordinate metabolism throughout the body, so as to ensure a steady provision of metabolic fuels and to control the extent to which different fuels are used by different tissues. 

In a broad sense, interconvertible forms of the same enzyme are compartmented in being rendered temporarily inactive by mechanisms such as phosphorylation by a specific kinase, as is the case with pyruvate dehydrogenase (Reed, 1976). Other examples, such as keto-enol tautomerization, can be mentioned as chemical compartmentation. Pogson and Wolfe (1972) have demonstrated that, at pH 7.4, 74% of oxaloacetate is in the keto form, and that a number of enzymes for which oxaloacetate is a substrate bind this form. However, the active species of oxaloacetate responsible for inhibiting succinate dehydrogenase and citrate lyase is the enol form. The balance between the two forms is maintained by a specific tautomerase, and this type of chemical compartmentation must be considered a potentially important factor in regulating metabolic pathways. Since there is little information relating to this point, however, we will not discuss it in detail in this review. 

Hundreds of metabohc reac tions take place simultaneously in cells. There are branched and parallel pathways, and a single biochemical may participate in sever distinct reactions. Through mass action, concentration changes caused by one reac tion may effect the kinetics and equilibrium concentrations of another. In order to prevent accumulation of too much of a biochemical, the product or an intermediate in the pathway may slow the production of an enzyme or may inhibit the ac tivation of enzymes regulating the pathway. This is termed feedback control and is shown in Fig. 24-1. More complicated examples are known where two biochemicals ac t in concert to inhibit an enzyme. As accumulation of excessive amounts of a certain biochemical may be the key to economic success, creating mutant cultures with defective metabolic controls has great value to the produc tion of a given produc t. 

Allosteric regulation acts to modulate enzymes situated at key steps in metabolic pathways. Consider as an illustration the following pathway, where A is the precursor for formation of an end product, F, in a sequence of five enzyme-catalyzed reactions ... 

Interestingly, anabolism and catabolism occur simultaneously in the cell. The conflicting demands of concomitant catabolism and anabolism are managed by cells in two ways. First, the cell maintains tight and separate regulation of both catabolism and anabolism, so that metabolic needs are served in an immediate and orderly fashion. Second, competing metabolic pathways are often... 

Citrate synthase is the first step in this metabolic pathway, and as stated the reaction has a large negative AG°. As might be expected, it is a highly regulated enzyme. NADH, a product of the TCA cycle, is an allosteric inhibitor of citrate synthase, as is succinyl-CoA, the product of the fifth step in the cycle (and an acetyl-CoA analog). 

The photosynthetic COg fixation pathway is regulated in response to specific effects induced in chloroplasts by light. What is the nature of these effects, and how do they regulate this metabolic pathway ... 

Several of the problems associated with whole cell bioprocesses are related to the highly effective metabolic control of microbial cells. Because cells are so well regulated, substrate or product inhibition often limits the concentration of desired product that can be achieved. This problem is often difficult to solve because of a poor understanding of the kinetic characteristics of the metabolic pathway leading to the desired product. 

In die metabolic pathway to an amino add several steps are involved. Each step is die result of an enzymatic activity. The key enzymatic activity (usually die first enzyme in the synthesis) is regulated by one of its products (usually die end product, eg die amino add). If die concentration of die amino add is too high die enzymatic activity is decreased by interaction of die inhibitor with the regulatory site of die enzyme (allosteric enzyme). This phenomenon is called feedback inhibition. 

Controlling an Enzyme That Catalyzes a Rate-Limiting Reaction Regulates an Entire Metabolic Pathway... 

THE FLUX OF METABOLITES IN METABOLIC PATHWAYS MUST BE REGULATED IN A CONCERTED MANNER... 

Metabolic pathways are regulated by rapid mechanisms affecting the activity of existing enzymes, eg, allosteric and covalent modification (often in response to hormone action) and slow mechanisms affecting the synthesis of enzymes. 

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