Chapter 15 (abbreviated): Principles of Metabolic Regulation CHEM 7784 Biochemistry Professor Bensley

CHAPTER 15 (Abbreviated) Principles of Metabolic Regulation Today’s Objectives: (To learn and understand the) Principles of regulation in biological systems Glycolysis vs. gluconeogenesis – which one is turned “on” and which one is turned “off”?

FIGURE 15-1 Metabolism as a three-dimensional meshwork FIGURE 15-1 Metabolism as a three-dimensional meshwork. A typical eukaryotic cell has the capacity to make about 30,000 different proteins, which catalyze thousands of different reactions involving many hundreds of metabolites, most shared by more than one "pathway." This overview image of metabolic pathways is from the online KEGG (Kyoto Encyclopedia of Genes and Genomes) PATHWAY database (www.genome.ad.jp/kegg/pathway/map/map01100.html). Each area can be further expanded for increasingly detailed information, to the level of specific enzymes and intermediates.

Homeostasis Organisms maintain homeostasis by keeping the concentrations of most metabolites at steady state In steady state, the rate of synthesis of a metabolite equals the rate of breakdown of this metabolite

Principles of Regulation The flow of metabolites through the pathways is regulated to maintain homeostasis Sometimes, the levels of required metabolites must be altered very rapidly Need to increase the capacity of glycolysis during the action Need to reduce the capacity of glycolysis after the action Need to increase the capacity of gluconeogenesis after successful action

Feedback Inhibition In many cases, ultimate products of metabolic pathways directly or indirectly inhibit their own biosynthetic pathways ATP inhibits the commitment step of glycolysis

Factors that Affect the Activity of Enzymes FIGURE 15-2 Factors affecting the activity of enzymes. The total activity of an enzyme can be changed by altering the number of its molecules in the cell, or its effective activity in a subcellular compartment (1 through 6), or by modulating the activity of existing molecules (7 through 10), as detailed in the text. An enzyme may be influenced by a combination of such factors.

Some Enzymes in the Pathway Limit the Flux of Metabolites More than Others Hexokinase and phosphofructokinase are appropriate targets for regulation of glycolytic flux FIGURE 15-9 Dependence of glycolytic flux in a rat liver homogenate on added enzymes. Purified enzymes in the amounts shown on the x axis were added to an extract of liver carrying out glycolysis in vitro. The increase in flux through the pathway is shown on the y axis.

Elasticity Coefficient Measures the Responsiveness to Substrate FIGURE 15-9 Elasticity coefficient, ε, of an enzyme with typical Michaelis-Menten kinetics. At substrate concentrations far below the Km, each increase in [S] produces a correspondingly large increase in the reaction velocity, v. For this region of the curve, the enzyme has an ε of about 1.0. At [S] >> Km, increasing [S] has little effect on v; ε here is close to 0.0.

Control of Glycogen Synthesis Insulin signaling pathway increases glucose import into muscle stimulates the activity of muscle hexokinase activates glycogen synthase Increased hexokinase activity enables activation of glucose Glycogen synthase makes glycogen for energy storage

UDP-Glucose FIGURE 15-10 Control of glycogen synthesis from blood glucose in muscle. Insulin affects three of the five steps in this pathway, but it is the effects on transport and hexokinase activity, not the change in glycogen synthase activity, that increase the flux toward glycogen.

Isozymes may Show Different Kinetic Properties Isozymes are different enzymes that catalyze the same reaction They typically share similar sequences Their regulation is often different FIGURE 15-12 Comparison of the kinetic properties of hexokinase IV (glucokinase) and hexokinase I. Note the sigmoidicity for hexokinase IV and the much lower Km for hexokinase I. When blood glucose rises above 5 mM, hexokinase IV activity increases, but hexokinase I is already operating near Vmax and cannot respond to an increase in glucose concentration. Hexokinases I, II, and III have similar kinetic properties.

Glycolysis vs. Gluconeo-genesis FIGURE 15-11 Glycolysis and gluconeogenesis. Opposing pathways of glycolysis (pink) and gluconeogenesis (blue) in rat liver. Three steps are catalyzed by different enzymes in gluconeogenesis (the "bypass reactions") and glycolysis; seven steps are catalyzed by the same enzymes in the two pathways. Cofactors have been omitted for simplicity.

Regulation of Phosphofructokinase-1 The conversion of fructose-6-phosphate to fructose 1,6-bisphosphate is the commitment step in glycolysis ATP is a negative effector Do not spend glucose in glycolysis if there is plenty of ATP FIGURE 15-14b Phosphofructokinase-1 (PFK-1) and its regulation. (b) Allosteric regulation of muscle PFK-1 by ATP, shown by a substrate-activity curve. At low [ATP], the K0.5 for fructose 6-phosphate is relatively low, enabling the enzyme to function at a high rate at relatively low [fructose 6-phosphate]. (Recall from Chapter 6 that K0.5 is the Km term for regulatory enzymes.) When [ATP] is high, K0.5 for fructose 6-phosphate is greatly increased, as indicated by the sigmoid relationship between substrate concentration and enzyme activity.

Regulation of Phosphofructokinase 1 and Fructose 1,6-Bisphosphatase Go glycolysis if AMP is high and ATP is low Go gluconeogenesis if AMP is low FIGURE 15-15 Regulation of fructose 1,6-bisphosphatase (FBPase-1) and phosphofructokinase-1 (PFK-1). The important role of fructose 2,6-bisphosphate in the regulation of this substrate cycle is detailed in subsequent figures.

Regulation by Fructose 2,6-Bisphosphate F26BP activates phosphofructokinase (glycolytic enzyme) F26BP inhibits fructose 1,6-bisphosphatase (gluconeogenetic enzyme)

Regulation by Fructose 2,6-Bisphosphate Go glycolysis if F26BP is high Go gluconeogenesis if F26BP is low FIGURE 15-16c Role of fructose 2,6-bisphosphate in regulation of glycolysis and gluconeogenesis. Fructose 2,6-bisphosphate (F26BP) has opposite effects on the enzymatic activities of phosphofructokinase-1 (PFK-1, a glycolytic enzyme) and fructose 1,6-bisphosphatase (FBPase-1, a gluconeogenic enzyme). (c) Summary of regulation by F26BP.

Chapter 15: Summary In this chapter, we learned that: living organisms regulate the flux of metabolites via metabolic pathways by increasing or decreasing enzyme concentrations activating or inactivating key enzymes in the pathway the activity of key enzymes in glycolysis and gluconeogenesis is tightly regulated via various activating and inhibiting metabolites