Regulation of Metabolic Pathways:- Regulatory Enzymes Respond to Changes in Metabolite Concentration
Flux through a biochemical pathway depends on the ac tivities of the enzymes that catalyze each reaction in that pathway. For some steps, the reaction is close to equilibrium within the cell (Fig. 15–12). The net flow of metabolites through these steps is the small difference between the rates of the forward and reverse reactions, rates that are very similar when the reaction is near equilibrium. Small changes in substrate or product con centration can produce large changes in the net rate, and can even change the direction of the net flow. We can identify these near-equilibrium reactions in a cell by comparing the mass action ratio, Q, with the equilibrium constant for the reaction, Keq. Recall that for the reaction A+B→C+D, Q=[C][D]/[A][B]. When Q and Keq are within a few orders of magnitude, the reaction is near equilibrium. This is the case for six of the ten reactions in the glycolytic pathway (Table 15–2). Other reactions are far from equilibrium in the cell. For example, Keq for the phosphofructokinase-1 (PFK-1) reaction in glycolysis is about 1,000, but Q ([fructose 1,6-bisphosphate] [ADP]/ [fructose 6-phosphate] [ATP]) in a typical cell in the steady state is about 0.1 (Table 15–2). It is because the reaction is so far from equilibrium that the process is exergonic under cellular conditions and tends to go in the forward direction. The re action is held far from equilibrium because, under pre vailing cellular conditions of substrate, product, and effector concentrations, the rate of conversion of fructose 6-phosphate to fructose 1,6-bisphosphate is limited by the activity of PFK-1, which is itself limited by the number of PFK-1 molecules present and by the actions of effectors. Thus, the net forward rate of the enzyme catalyzed reaction is equal to the net flow of glycolytic intermediates through other steps in the pathway, and the reverse flow through PFK-1 remains near zero.
FIGURE 15–12 Near-equilibrium and nonequilibrium steps in a metabolic pathway. Steps 2 and 3 of this pathway are near equilibrium in the cell; their forward rates are only slightly greater than their reverse rates, so the net forward rates (10) are relatively low and the free-energy change ΔG for each step is close to zero. An increase in the intracellular concentration of metabolite C or D can reverse the direction of these steps. Step 1 is maintained in the cell far from equilibrium; its forward rate greatly exceeds its reverse rate. The net rate of step 1 (10) is much larger than the reverse rate (0.09) and is identical to the net rates of steps 2 and 3 when the pathway is operating in the steady state. Step 1 has a large, negative ΔG.
The cell cannot allow reactions with large equilibrium constants to reach equilibrium. If [fructose 6-phosphate], [ATP], and [ADP] in the cell were held at their usual level (low millimolar) and the PFK-1 reaction were allowed to reach equilibrium by an increase in [fructose 1,6-bisphosphate], the concentration of fructose 1,6 bisphosphate would rise into the molar range, wreaking osmotic havoc on the cell. Consider another case: if the reaction ATP→ADP +Pi were allowed to approach equilibrium in the cell, the actual free-energy change (ΔG) for that reaction (see Box 13–1) would approach zero, and ATP would lose the high phosphoryl group transfer potential that makes it valuable to the cell as an energy source. It is therefore essential that enzymes catalyzing ATP breakdown and other highly exergonic reactions in a cell be sensitive to regulation, so that when metabolic changes are forced by external circumstances, the flow through these enzymes will be adjusted to ensure that [ATP] remains far above its equilibrium level. When such metabolic changes occur, enzymatic activities in all interconnected pathways adjust to keep these critical steps away from equilibrium. Thus, not surprisingly, many enzymes that catalyze highly exergonic reactions (such as PFK-1) are subject to a variety of subtle regulatory mechanisms. The multiplicity of these adjustments is so great that we cannot predict by examining the properties of any one enzyme in a path way whether that enzyme has a strong influence on net flow through the entire pathway. This complex problem can be approached by metabolic control analysis, as described in Section 15.5.
FIGURE 15–13 Factors that determine the activity of an enzyme. Blue arrows represent processes that determine the number of enzyme molecules in the cell; red arrows show factors that determine the enzymatic activity of an existing enzyme molecule. Each arrow represents a point at which regulation can occur.
