15.5: Regulation of Gluconeogenesis

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Learning Goals (ChatGPT o3-mini)

1. Describe the Gluconeogenic Conversion Process:

   • Explain how pyruvate is converted to phosphoenolpyruvate via an oxaloacetate intermediate, outlining the roles of pyruvate carboxykinase and phosphoenolpyruvate carboxykinase (PEPCK) in this process.

2. Analyze Allosteric Regulation of Pyruvate Carboxykinase:

   • Identify the major allosteric effectors (Acetyl-CoA and ADP) that regulate pyruvate carboxykinase activity.
   • Explain how Acetyl-CoA serves as an activator under high-energy conditions and how ADP acts as an inhibitor under low-energy conditions.

3. Explain Transcriptional and Allosteric Regulation of PEPCK:

   • Describe the transcriptional regulation of cytoplasmic PEPCK in response to increased cAMP, glucocorticoids, and thyroid hormones, including the role of the CREB transcription factor.
   • Discuss how ADP serves as an allosteric inhibitor of PEPCK, linking its activity to cellular energy status.

4. Discuss the Dual Regulation of Fructose 1,6-Bisphosphatase:

   • Explain the competitive inhibition of fructose 1,6-bisphosphatase by fructose 2,6-bisphosphate and its effect on the enzyme's Km.
   • Describe how low-energy indicators (AMP and ADP) further inhibit fructose 1,6-bisphosphatase allosterically, ensuring that gluconeogenesis is downregulated when cellular energy is scarce.

5. Integrate Hormonal and Energy Signals in Gluconeogenesis Regulation:

   • Evaluate how systemic hormonal signals (e.g., glucagon during hypoglycemia) and local cellular energy indicators (ATP, ADP, and AMP levels) coordinate to regulate the activity of key gluconeogenic enzymes.
   • Discuss the overall significance of these regulatory mechanisms in maintaining blood glucose homeostasis and metabolic balance.

These learning goals aim to provide students with a comprehensive understanding of the multiple layers of control governing gluconeogenesis, linking enzyme kinetics, allosteric regulation, transcriptional control, and hormonal signaling to the broader context of metabolic homeostasis.

Within the regulation of the gluconeogenic pathway, three of the major enzymatic steps are regulated. The first two are the pyruvate carboxykinase enzyme and the phosphoenolpyruvate carboxykinase (PEPCK). Recall that these two enzymes are required to convert pyruvate back into phosphoenolpyruvate via an oxaloacetate intermediate, as shown in Figure \(\PageIndex{1}\). The third enzyme regulated in this pathway is fructose 1,6-bisphosphatase, which converts fructose 1,6-bisphosphate into fructose 6-phosphate. We will explore the regulation of these three enzymes in more detail.

Figure \(\PageIndex{1}\): Conversion of Pyruvate to Phosphoenolpyruvate during Gluconeogenesis. Image modified from Principles of Biochemistry (2019) Wikibooks

Pyruvate Carboxykinase

Pyruvatecarboxykinase is one of the primary regulation points. It is primarily regulated by two allosteric effectors, acetyl-CoA and ADP, as shown in  Figure \(\PageIndex{2}\). When pyruvate enters the Kreb Cycle, it is first converted to acetyl-CoA. If abundant pyruvate is present, an ample supply of acetyl-CoA will also be available, indicating a high energy load for the cell. Acetyl-CoA can bind with pyruvate carboxylase and act as a protein activator, stimulating the production of oxaloacetate. ADP, on the other hand, is a low-energy indicator and an inhibitor of the enzyme. The next section will show how oxaloacetate moves into the cytoplasm.

Figure \(\PageIndex{2}\): Allosteric Regulation of Pyruvate Carboxykinase. Figure modified from Liu, Y., et al (2018) Nat Commun 9:1384

Phosphoenolpyruvate Carboxykinase

Cytoplasmic PEPCK is largely regulated at the transcriptional level. Increases in gene expression are seen in response to elevated cAMP levels, glucocorticoids, and thyroid hormone levels, as shown in Figure \(\PageIndex{3}\). The activated CREB transcription factor plays a role in this response. Alternatively, decreased gene expression is caused by insulin signaling. ADP also acts as an allosteric effector of the protein, causing it to have lower activity. This indicates that when energy is low, the cell cannot afford to use its reserves to remake glucose, so gluconeogenesis is inhibited.

Figure \(\PageIndex{3}\): Regulation of Phosphoenolpyruvate Carboxykinase at the Transcriptional and Allosteric Levels. Image from ProteinBoxBot

Fructose 1,6-Bisphosphatase

Fructose 1,6-bisphosphatase is both competitively and allosterically regulated, as shown in Figure \(\PageIndex{4}\). Fructose 2,6-bisphosphate serves as a competitive inhibitor of the enzyme, reducing the overall activity of the enzyme for fructose 1,6-bisphosphate. Competitive inhibitors bind within the active site and compete for binding with the regular substrate. Thus, they lower the overall Km of the reaction and make the enzyme less effective at lower substrate concentrations. However, the Vmax of the enzyme is not affected during the process.

In addition to competitive inhibition, low energy loads (AMP and ADP) also inhibit the enzyme. ADP and AMP bind allosterically to the enzyme and inhibit its activity.

Figure \(\PageIndex{14}\): Regulation of Fructose 1,6-Bisphosphatase by Competitive Inhibition and Allosteric Effectors. Image from Jslipscomb

Summary

This chapter delves into the regulatory mechanisms governing key control points of the gluconeogenic pathway, highlighting how enzymatic activity is finely tuned by both hormonal signals and cellular energy status. Three major enzymes are discussed:

• Pyruvate Carboxykinase (PCK):
  This enzyme catalyzes the conversion of pyruvate to oxaloacetate—a crucial step in converting pyruvate back to phosphoenolpyruvate (PEP). Its activity is modulated allosterically: Acetyl-CoA acts as an activator, signaling a high-energy state, while ADP serves as an inhibitor under low-energy conditions. This regulation ensures that the pathway is activated only when necessary.

• Phosphoenolpyruvate Carboxykinase (PEPCK):
  Predominantly regulated at the transcriptional level, PEPCK expression is increased in response to elevated cAMP, glucocorticoids, and thyroid hormones, with the CREB transcription factor playing a key role. Additionally, ADP acts as an allosteric inhibitor of PEPCK, linking its activity to the cell’s energy balance and ensuring that glucose synthesis is downregulated when energy is scarce.

• Fructose 1,6-Bisphosphatase (FBPase):
  FBPase catalyzes the dephosphorylation of fructose 1,6-bisphosphate to fructose 6-phosphate, effectively opposing the action of phosphofructokinase-1 in glycolysis. Its activity is regulated through two mechanisms: competitive inhibition by fructose 2,6-bisphosphate, which reduces substrate binding efficiency, and allosteric inhibition by AMP and ADP, reflecting low cellular energy.

Collectively, these regulatory strategies integrate hormonal signals—such as those mediated by glucagon during hypoglycemia—with intracellular energy cues to modulate gluconeogenesis. This dynamic control ensures that glucose production is enhanced during energy deficits and conserved when energy is abundant, thereby maintaining overall metabolic and blood glucose homeostasis.

By linking enzyme regulation to both systemic and cellular energy demands, this chapter provides a comprehensive framework for understanding how gluconeogenesis is precisely controlled to meet the metabolic needs of the organism.