Lecture 3.3: Regulation of Metabolism

Which of the following are true of gluconeogenesis? Select all that apply.

a) It is a pathway with 10 reactions.
b) It occurs mainly in the liver & to a lesser extent in the kidneys.
c) It takes place in brain & muscles where glycogen is stored.
d) It is the reverse of glycolysis.
e) It is used to make glucose when dietary glucose and glycogen reserves are low.

b) It occurs mainly in the liver & to a lesser extent in the kidneys.

e) It is used to make glucose when dietary glucose and glycogen reserves are low.

How many steps in gluconeogenesis

11 steps

(T/F) Gluconeogenesis is a revserse reaciton of glycolysis

False — gluconeogenesis shared similar reactions but is not a direct reversal of glycolysis

What enzymes are shared between both glycolysis and gluconeoenesis

Enolase, Phosphoglycerate mutase, Phosphoglycerate kinase, G3P dehydrogenase, Triose phosphate isomerase, Aldolase

How do you convert Pyruvate to Phosphoenolpyruvate

Break the reaction into 2 separate exergonic steps

1) Pyruvate is converted into Oxaloacetate by pyruvate carboxylase coupled with ATP hydrolysis and the cofactor Biotin

2) Oxaloacetate is converted to Phosphoenolpyruvate (PEP) by Phosphoenolpyruvate carboxykinase and GTP

Once PEP is created what are the following steps of gluconeogenesis

Once PEP is created the 6 shared reversal steps begin to eventually create G3P

How many Phosphoenolpyruvate are created in the 2 steps of gluconeogenesis

2 — one from each pyruvate

What enzymes reverses the 3rd step of glycolysis

FBPase-1

What enzyme reverses step 1 of glycolysis

Glucose-6-phosphatase

What is the full reaction for gluconeogenesis

2 pyruvate + 4 ATP + 2 GTP + 2 NADH + 2 H⁺ + 4 H2O —> Glucose + 4 ADP + 2 GDP + 6 P + 2 NAD⁺

When is gluconeogenesis triggered

During starvation or vigorous exercise

What is the only source of ATP for the brain, nervous system, and RBC

Glucose

Which of the following enzymes are used in both glycolysis and gluconeogenesis? Select all that apply.

a) Hexokinase

b) Enolase

c) Phosphoglycerate kinase

d) Triose phosphate isomerase

e) Pyruvate kinase

b) Enolase

c) Phosphoglycerate kinase

d) Triose phosphate isomerase

In the gluconeogenesis pathway, the enzyme glucose-6-phosphatase reverses which step in glycolysis?

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a) the formation of glucose

b) the reaction catalyzed by pyruvate dehydrogenase

c) the reaction catalyzed by hexokinase

d) the formation of fructose-1,6-bisphosphate

c) the reaction catalyzed by hexokinase

Which enzyme in the glycolysis pathway catalyzes a reaction that requires two enzymes to reverse in the gluconeogenesis pathway?

a) pyruvate kinase

b) phosphofructokinase I

c) pyruvate dehydrogenase

d) glycogen phosphorylase

a) pyruvate kinase

Which of the following are true of liver tissue compared to other tissues?  Select all that apply.

a) mostly a catabolic tissue that burns glucose and fats for energy

b) releases stored glucose to the rest of the body when needed.

c) maintains stable blood glucose levels

d) stores glucose as glycogen

e) uses gluconeogenesis to make glucose if glycogen stores are depleted.

f) has limited glycogen storage capacity 

b) releases stored glucose to the rest of the body when needed.

c) maintains stable blood glucose levels

d) stores glucose as glycogen

e) uses gluconeogenesis to make glucose if glycogen stores are depleted.

The enzyme Glucose 6-phosphatase is mainly found in:

a) Skin

b) Liver

c) Muscle

d) Brain

e) Red Blood Cells

b) Liver — Glucose-6-phosphatase is the enzyme that catalyzes the final step of gluconeogenesis. converting glucose-6-phosphate into free glucose, which can then be released into the bloodstream. Which is exactly why these are the only organs capable of releasing free glucose into the blood to help maintain blood glucose levels.

Which steps of glycolysis are regulated in muscle cells?

Step 1, 3 & 10

Hexokinase is regulated by:

a) negative allosteric regulation by ATP

b) negative allosteric regulation by glucose-6-phosphate

c) negative allosteric regulation by fructose-6-phosphate

d) positive allosteric regulation by ATP

e) positive allosteric regulation by glucose-6-phosphate

b) negative allosteric regulation by glucose-6-phosphate

Which glycolytic intermediate serves at the starting material for glycogen synthesis?

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a) glucose

b) glucose 6-phosphate

c) fructose 6-phosphate

d) fructose 1,6-bisphosphate

e) dihydroxyacetone phosphate

b) glucose 6-phosphate

What molecule acts as an allosteric activator of Phosphofructokinase 1?

a) ATP

b) G6P

c) F6P

d) ADP

e) NADH

d) ADP

Which binding site on PFK has a higher affinity for ATP?

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a) Active Site

b) Allosteric Site

a) Active Site — PFK-1 has two distinct binding sites for ATP:

1. Active (catalytic) site — This is where ATP functions as a substrate. This site has a higher affinity for ATP, meaning it's saturated even at relatively low ATP concentrations.

2. Allosteric (regulatory) site — This is a separate, lower-affinity site. ATP only binds here in significant amounts when its concentration is high. When ATP binds here, it induces a conformational change that inhibits enzyme activity by lowering PFK-1's affinity for its substrate

High concentrations of ATP cause the PFK enzyme activity curve to shift ________.

a) Left

b) Right

b) Right

Which of the following molecules activates pyruvate kinase?

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a) F-1,6-BP

b) ATP

c) Acetyl-CoA

d) long-chain fatty acids

e) NADH

a) F-1,6-BP

What does skeletal muscle burn for energy

Glucose and Fat

Skeletal muscle is mainly a _____ (catabolic/anabolic) tissue

Catabolic

(T/F) Skeletal muscle can perform gluconeogenesis

False

What does the liver do when glycogen storages are too high

liver performs glycolysis to create pyruvate which eventuall can become a fatty acid (fat)

When blood glucose levels run low the liver performs _______

gluconeogenesis

In muscle, regulation of glycolysis depends on _____ _______

energy charge

In muscle, at rest glycolysis is ______

inhibited

In muscle, during exercise glycolysis is ______

Active

What is the negative feedback inhibitor for hexokinase

Glucose-6-P

What is the negative inhibitor for PFK and Pyruvate kinase

ATP / AMP

What is the activator for PFK

ATP / AMP

What is the activator for Pyruvate kinase

Fructose 1,6-BP

Hexokinase is _______ inhibited by ________

allosterically; glucose-6-phosphate

What happens when glycogen stores in the muscle become fully saturated

Once muscle storages are full excess glucose is sent to the liver

When G6P levels are too high _____ is inhibited

Hexokinase

PFK is shut off when _____ are too high

Fructose 6-phosphate

When G6P is elevated but glycogen stores are not full what occurs

G6P is shunted to another pathway to create glycogen to me stored for later use

What is the true “committed step” of glycolysis

Step 3 — F6P —> F16BP catalyzed by PFK

PFK is a _____ with _____ binding sites that bind ____ or _____

PFK is a tetramer with allosteric binding site thats bind ATP or ADP

At high energy _____ is an _____ for PFK

At high energy ATP is an inhibitor for PFK

At low energy _____ is an _____ for PFK

At low energy ADP is an activator for PFK

If ATP > ADP then ____ PFK activity

Decreased

If ATP < ADP then ____ PFK activity

Increased

When [ATP] is low then PFK-1 vs F6P graph shifts _____ favoring ____ state

Left; R-state

When [ATP] is high then PFK-1 vs F6P graph shifts _____ favoring ____ state

Right; T-state

Explain the mechanism for ATP inhibition of PFK

When ATP binds the allosteric site, it triggers a conformational change in the enzyme — shifting it from the high-affinity R-state to the low-affinity T-state. In the T state, the enzyme's active site has much lower affinity for F6P, so the reaction slows down dramatically even though substrate is present.

Explain the mechanism for ADP inhibition of PFK

When AMP binds instead of ATP, it stabilizes the enzyme in the high-affinity R state. This opens up the catalytic sites, which are now able to bind F6P

Pyruvate Kinase is inhibited and activated by _____

F16BP

When energy levels are high F16BP acts as an ______ for pyruvate kinase

inhibitor

How many ATP equivalents are required for the first two reactions of gluconeogenesis to reverse the last reaction of glycolysis? Assume that one molecule of glucose is synthesized by the gluconeogenic pathway. Enter a numeric value only.

4

Which tissue expresses a glucose transporter with the highest affinity for glucose?

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a) liver

b) brain

c) muscle

b) brain

Which of the following is TRUE about hexokinase and glucokinase?

a) Glucokinase catalyzes dephosphorylation, whereas hexokinase catalyzes phosphorylation. 

b) Glucokinase is found in the liver, while hexokinase is found in muscle. 

c) Glucokinase is inhibited by glucose 6-phosphate, whereas hexokinase is not inhibited by glucose 6-phosphate. 

d) Glucokinase's substrate is glucose, whereas hexokinase's substrate is fructose. 

e) Glucokinase operates at high glucose concentrations, whereas hexokinase does at low glucose concentrations.

e) Glucokinase operates at high glucose concentrations, whereas hexokinase does at low glucose concentrations.

What is the glucose carrier protein in the brain and its Kd

Glut-3; Kd = 1-5 mM

What is the glucose carrier protein in the liver and its Kd

Glut-2; Kd = 20-40mM

What is the glucose carrier protein in muscle and its Kd

Glut-4; Kd = 5-10mM

Glut-2 in the _____ only brings glucose in at ___ concentrations. Explain

liver; high; the liver’s function to to control blood glucose levels, therefore the Kd is high to allow for glucose to only create/release glucose when the blood levels are too high and need to be lowered

_______ is the primary enyme for formation for G6P in liver

Glucokinase

Why does increasing the [Glucose] not affect hexokinase

The Km for hexokinase is so low that it functions at Vmax at normal blood glucose levels. This means that the enzyme is typically fully saturated and by increasing the concentration of glucose this will not affect the rate

What is the Kd of hexokinase

Km = 0.1 mM

What is the Kd of glucokinase

Km = 10 mM

Explain this significance of these enzyme curves

The low Km of hexokinase ensures that muscle cells always have glucose stored, even at low glucose concentrations, to allow for contraction. The less steep curve of glucokinase ensures that the liver only releases glucose when concentrations of G6P are low and will store glucose when G6P concentrations are high. The higher Km for glucokinase means that the liver will only store glucose when concentrations are high — the primary function of the liver

(T/F) Glucoskinase is inhibited by G6P

False — hexokinase is inhibited by G6P not glucokinase

Why is it important that Hexokinase (muscle & other tissues) is inhibited by G6P and not glucokinase (liver)

G6P is the product of the first step of glycolysis (glucose + ATP → glucose-6-phosphate). In muscle and other tissues when glucose levels are high there is no need to phosphorylate more glucose when the supply is already high. This negative feedback makes sure that when G6P is high the creation of more G6P is stopped. However, in the liver glucokinase is responsible for the uptake of glucose from the blood. Therefore, if high G6P levels inhibited glucokianse this would prevent the liver from performing its primary function.

Fructose-1,6-bisphosphate is synthesized by ______ from the molecule ______.

Blank 1: phosphofructokinase-1

Blank 2: fructose-6-phosphate

F2,6-BP is an: (select all that apply)

a) allosteric activator of PFK-1

b) allosteric activator of PFK-2

c) allosteric activator of FBPase-1

d) allosteric inhibitor of PFK-1

e) allosteric inhibitor of PFK-2

f) allosteric inhibitor of FBPase-1

a and f

F2,6-BP activates PFK-1 → promotes glycolysis

F2,6-BP inhibits FBPase-1 → blocks gluconeogenesis

FBPase-2 is an enzyme that: 

a) dephosphorylates F1,6-BP

b) phosphorylates F1,6-BP

c) dephosphorylates F2,6-BP

d) phosphorylates F2,6-BP

c) dephosphorylates F2,6-BP

When the PFK-2/FBPase-2 enzyme is phosphorylated, _______ is active, which favors _______.

a) FBPase-2; glycolysis

b) PFK-2; gluconeogenesis

c) FBPase-2; gluconeogenesis

d) PFK-2; glycolysis

c) FBPase-2; gluconeogenesis

When Glucagon levels rise, ________ is activated, whereas when Insulin levels rise, ________ is activated. (Select all that apply)

a) FBPase-2 ; PFK-2

b) PFK-2 ; FBPase-2

c) Protein phosphatase 1 ; Protein kinase A

d) Protein kinase A ; Protein phosphatase 1

e) Gluconeogenesis ; Glycolysis

f) Glycolysis ; Gluconeogenesis

A, D, E

Glucagon → Protein Kinase A → phosphorylates PFK-2/FBPase-2 → FBPase-2 active → ↓F2,6-BP → Gluconeogenesis favored

Insulin → Protein Phosphatase 1 → dephosphorylates PFK-2/FBPase-2 → PFK-2 active → ↑F2,6-BP → Glycolysis favored

What inhibitors PFK-1

High [ATP]

What activates PFK-1

High [ADP] and & [AMP]

What inhibits FBPase-1

High [AMP]

What is enzyme catalyzes fructose 1,6-BP to F6P in the liver

FBpase-1

What is enzyme catalyzes F6P to fructose 1,6-BP in the liver

PFK-1

_____ through glycolysis is regulated in response to ______ ______

Flux through glycolysis is regulated in response to energy charge

_______ is the most important regulator of glucose metabolism in the liver

F2,6-BP

Explain the synthesis of F2,6-BP

F6P + ATP is catalyzed by PFK-2 to create F2,6-BP. F2,6-BP then serves as an activator of PFK-1 to create F1,6-BP

Fructose 2,6-bisphosphate is an _____ for PFK-1 and an _____ for FBPase-1

activator; inhibitor

Explain why the reciprocal regulation characteristics of F2,6-BP is important for a liver cell

When F26BP is present glycolysis is favored/activated and gluconeogenesis is inhibited. This allows the liver to have a crucial on/off switch to buffer the amount of blood glucose

More F2,6-BP = more glycolysis, less gluconeogenesis.
Less F2,6-BP = less glycolysis, more gluconeogenesis.

The [F26BP] is determined by the rate of its synthesis by ______ and its breakdown by ______

PFK-2; FBPase-2

Describe the mechanism for the synthesis of F26BP

F6P + ATP —> F26BP

Describe the mechanism for the breakdown of F26BP

F26BP + P —→ F6P

Counter-signaling hormones _____ and _____ coordinate [F26BP] in the liver

insulin; glucagon

Explain the full mechanism when glucagon levels rise and how it effects the liver

When blood sugar levels drop, glucagon levels rise. This increases the activity of protein kinase A + ATP which causes the phosphoralation of ser32 making PFK-2 inactive. This activates FBPase-2 which catalyzes the breakdown of F26BP into F6P (Favoring Gluconeogenesis)

Explain the full mechanism when insulin levels rise and how it effects the liver

When blood suagr levels rise, insulin levels increase. This causes an increase in the activity of protein phosphatase 1 + H2O, which hydrolyzes PFK-2 converting it to the active form (removing P from Ser32). FBPase-2 is inactive and PFK2 is active. This induces the creation of F26BP. (Favoring glycolysis)

When glucose is abundant, insulin signaling stimulates glucose uptake through…

1) Glycogen synthesis

2) Glycolysis (when glycogen stores are full) to make pyruvate —> acetyl coA —> fatty acids, despite high [ATP]

When glucose is scarce, glucagon signaling activates glucose export through…

1) Glycogen degradation

2) Gluconeogenesis

Glycolysis = increase in ____ and breakdown of ____

ATP; glucose

Gluconeogenesis = increase in _____

glucose

_____ is a local energy signal that tells the cells its intrinsic energy level in low

AMP

What is flux

Flux = the rate of flow of something through a given point, surface, or system, per unit time

Arrange the following enzymes in the order they would be encountered during gluconeogenesis, starting with pyruvate.

phosphoenolpyruvate carboxykinase

phosphoglycerate kinase

phosphoglucose isomerase

aldolase

glucose-6-phosphatase

enolase

phosphoenolpyruvate carboxykinase

enolase

phosphoglycerate kinase

aldolase

phosphoglucose isomerase

glucose-6-phosphatase

Which of the following metabolic enzymes are used exclusively by glycolysis and which are used exclusively by gluconeogenesis?

Glycolysis only _______

Gluconeogenesis only _______

pyruvate kinase

fructose-1-6-bisphosphatase

Triose phosphate isomerase

Phosphoglycerate kinase

Enolase

Phosphoglucose isomerase

Phosphoglycerate mutase

Glyceraldehyde-3-phosphate dehydrogenase

Aldolase

Glycolysis only pyruvate kinase

Gluconeogenesis only fructose-1-6-bisphosphatase

Match the glycolytic reactions with the corresponding gluconeogenic enzyme that catalyzes the reverse reaction.

Glucose ↔ glucose-6-phosphate ________

Fructose-6-phosphate ↔ fructose-1,6-bisphosphate __________

Glyceraldehyde-3-phosphate ↔ 1,3-bisphosphoglycerate ___________

2-phosphoglycerate ↔ phosphoenolpyruvate _________

Phosphoenolpyruvate ↔ pyruvate ___________

Glucose ↔ glucose-6-phosphate — glucose-6-phosphatase

Fructose-6-phosphate ↔ fructose-1,6-bisphosphate — fructose-1,6-bisphosphatase

Glyceraldehyde-3-phosphate ↔ 1,3-bisphosphoglycerate — glyceraldehyde-3-phosphate dehydrogenase

2-phosphoglycerate ↔ phosphoenolpyruvate — enolase

Phosphoenolpyruvate ↔ pyruvate — pyruvate carboxylase / PEP carboxykinase

What is the cost (in ATP equivalents per glucose) of transforming glucose to pyruvate via glycolysis and back again to glucose via gluconeogenesis?

Glycolysis = Gain of 2 ATP
Gluconeogenesis = Loss of 4 ATP, 2 GTP

Net cost = 4 ATP equivalents per glucose

Glucose is transported into cells through 12 different integral membrane protein glucose transporters (GLUT-1 to GLUT-12). The GLUT-1 transporter is especially prevalent in the brain and erythrocytes and shows a high affinity (Km ~1–2 mM) to glucose. Several breast, lung, and colorectal cancers have been found to have an increased expression of these GLUT-1 transporters. 

Cancer cells increase GLUT-1 expression to increase glucose uptake, produce ATP, and create the proper intermediates required for cell division. By increasing GLUT-1, which has the lowest efficient for glucose, this is a sacrifice made for speed over efficiency. Cancer cells want energy fast, despite glycolysis making little ATP, it is the speed the cancer cell is looking for. Glycolysis also makes nucleotide intermediates that aide in cellular division.

n skeletal muscles, flux through glycolysis is decreased when cellular energy levels are __________; the regulatory enzyme, phosphofructokinase-1, is __________ by high concentrations of ATP.

a) low; activated

b) low; inhibited

c) high; activated

d) high; inhibited

d) high; inhibited