Download Gluconeogenesis: Objectives

Gluconeogenesis: Objectives
1. There are many questions, including:
a. How can glucose be synthesized from lactate (i.e. gluconeogenesis)?
i. Lactate is converted to Pyruvate, followed by a series of
reverse glycolytic steps to get to Glucose. An endorgonic,
regulated process.
b. Where (in which organs) does gluconeogenesis take place?
i. Gluconeogenesis occurs mainly in the liver (90%) and also in
the kidneys
c. What parts of the cell participate in gluconeogenesis?
i. In the mitochondria and the cytoplasm
d. Name the most common precursors for gluconeogenesis.
i. Lactate from exercising muscle and red blood cells
ii. The breakdown of proteins, which gives rise to amino acids
(which can be utilized to make new glucose)
iii. Glycerol (use only part of the pathway)
iv. Fructose (use only part of the pathway)
v. Glyceraldehyde-3-phosphate
vi. Alanine
vii. Oxaloacetate is needed because
1. it is the precursor for PEP
2. Helps to bypass pyruvate kinase
viii. Glycolytic molecules such as F6P and G3P
e. Where do these precursors for gluconeogenesis come from?
i. Lactate – comes from the exercising muscles and the
breakdown of glucose in the RBCs (anaerobic glycolysis)
ii. Glycerol – comes from triglycerides
2. Name the three irreversible steps in glycolysis. Name the enzymes.
a. GlucoseGlucose-6-Phosphate (Hexokinase/Glucokinase)
b. Fructose-6-PhosphateFructose-1,6-Bisphosphate (PFK)
c. Phosphoenol PyruvatePyruvate (Pyruvate Kinase)
3. How are these steps by-passed in gluconeogenesis? Name the enzymes.
a. Glucose-6-Phosphate  Glucose
i. Cleave the phosphate group
1. Enzyme: “Glucose-6-Phosphatase”
b. Fructose-1,6-BisphosphateFructose-6-Phosphate
i. Cleave the phosphate group
1. Enzyme: “Fructose-1,6-Bisphosphatase”
2. The REGULATORY enzyme!
Gluconeogenesis: Objectives
c. Pyruvate  Phosphoenol Pyruvate
i. Pyruvate  Oxaloacetate
1. Add CO2 + ATP
2. (Generates ADP)
3. Enzyme: “Pyruvate Carboxylase” and coenzyme: Biotin
4. In the mitochondria
ii. Oxaloacetate + NADH Malate; Malate + NAD+Oxaloacetate
1. Requires the Malate Shuttle to go to the cytoplasm
2. Turned into Malate, given to cytoplasm, returns to
oxaloacetate
3. Enzyme: Malate Dehydrogenase
iii. Oxaloacetate  PEP
1. Add GTP
2. Loses CO2
3. Enzyme: “PEPCarboxykinase” (PEPCK)
4. In the cytoplasm
4. Several questions…
a. What is the role of pyruvate carboxylase in gluconeogenesis and in
anaplerosis?
i. Pyruvate Carboxylase adds CO2 to pyruvate to yield
Oxaloacetate in the mitochondria
1. In anaplerosis: regenerates a catalytic intermediate for
TCA cycle
2. In kidney/liver: Needs to continue regeneration to
continue to move towards formation of new glucose
b. How is it regulated?
i. (+) regulated by Acetyl CoA.
c. Write the sequence of steps involved in the conversion of pyruvate to
phosphoenol pyruvate.
i. Three Steps:
1. PyruvateOxaloacetic Acid
a. this reaction uses “biotin” (to carboxylate) and
ATP
b. Pyruvate Carboxylase
c. In mitochondria
2. Oxaloacetate + NADHMalate + NAD+ Oxaloacetate
a. Used for transport from Mitochondrial matrix to
cytoplasm
b. Enzyme: Malate Dehydrogenase
3. OxaloacetatePEP
a. Uses GTP and liberates CO2
b. PEPCK
c. In cytoplasm
Gluconeogenesis: Objectives
5. Explain why Acetyl CoA cannot serve as a precursor for gluconeogenesis?
a. Acetyl CoA cannot be used to make new glucose because TCA cycle
oxidizes Acetyl CoA completely; no way to make acetyl CoA from
pyruvate (irreversible reaction of acetyl coA + CO2 into pyruvate)
i. Thus cannot use even numbered fatty acids because they
produce acetyl CoA.
6. What key role does Acetyl CoA play in gluconeogenesis?
a. Acetyl Co A is a positive modulator of pyruvate carboxylase
b. Without Acetyl CoA, the pyruvate carboxylase would not function!
7. Several questions:
a. What is the role of biotin in gluconeogenesis?
i. Biotin is the coenzyme for carboxylation reactions. It is the
carrier of active CO2 in carboxylations. (p.13 of notes)
ii. ATP in presence of pyruvate carboxylase +biotin
1. P from ATP is given to biotin’s imidizole ring: “active
biotin complex”
a. Allows the enzyme to add the CO2 to the enzyme
and produce oxaloacetate
b. Recognize the structure of biotin
i. It has an imidozole ring fused to a Thiophene ring with a fatty
acid. The carboxylate group will bind with lysine (CONH)
residue in enzyme. (it looks like two houses that are united at
the floor. The top house has nitrogen and the bottom house
has a sulfur roof. The fatty acid is attached to the bottom
house).
c. What is the nature of the biotin-enzyme linkage?
i. It is an amide bond between the biotin and the specific enzyme
d. What kind of reaction is biotin known to participate in?
i. Carboxylations- adding CO2 (the carboxylate group will bind
with lysine residues in enzyme)
ii. Thiamine – decarboxylation reactions (in PPP and glycolosis)
e. Give the names of 3 such reactions.
i. Pyruvate  Oxaloacetate (pyruvate carboxylase)
ii. Propionyl CoA + CO2Succinyl CoA (few reactions involved,
Key enzyme: Propionyl CoA Carboxylase)
iii. Malonyl CoAAcetyl Co A (Acetyl-CoA Carboxylase)
8. State the steps involved in the transport of oxaloacetate from mitochondria
to cytoplasm. What role does malate play in this process?
a. Oxaloacetate is transported to the cytoplasm from the mitochondrial
matrix by way of the malate shuttle. OxAc is first converted to malate
by malate Dehydrogenase.
b. Malate then crosses the membrane and is converted back to OxAc by
the same enzyme.
Gluconeogenesis: Objectives
9. Describe why the glycerol-moiety of triglycerides and not the evennumbered fatty acids can be converted to glucose. Name the first glycolytic
intermediate formed from glycerol. (see pg. 16)
a. The essence of this question is based in the number of carbons.
Glycerol comes from the breakdown of triglycerides. Glycerol is
converted to glycerol-3-phosphate, which then is reacted with NAD+ +
glycerol-3-phosphate dehydrogenase to produce: dihydroxyacetone
phosphate (DHAP).
10. Explain why muscle and brain cannot synthesize glucose from lactate.
a. Muscle and Brain cannot synthesize glucose from lactate because they
lack Glucose-6-Phosphatase. This is good because then all the glucose
can only be used for energy rather than be exported (like the liver and
kidney).
11. How and where are (a) lactate and (b) alanine (produced by active skeletal
muscle) converted to glucose?
a. Lactate (from muscles) and alanine are converted in the liver.
b. Lactate is converted to Pyruvate…eventually, make new glucose.
c. Alanine is converted to Pyruvate…eventually make new glucose.
12. Why does adipose tissue triglyceride biosynthesis depend on the availability
of glucose?
a. Adipose Tissue lacks Glycerol Kinase so the glycerol that is liberated
in the break down of fatty acids cannot be phosphorylated.
i. Cannot make DHAP
ii. Must have GLUCOSE if want to make it in adipose tissue
13. Explain how amino acids can serve as precursors for the synthesis of glucose.
a. Amino Acids can be converted to glycolytic intermediates of the TCA
and can then eventually become Oxaloacetate.
i. When remove the alpha amine group, can be transformed into
TCA intermediates.
ii. There are different AAs which can form glutamic acid,
furmerate, pyruvate, Propinyl-CoA (go to succinyl-CoA)
1. Eventually, produce oxaloacetate, which can be made
into malate, and can be shuttled out of mitochondria to
form new glucose.
14. State the primary and secondary control steps in the synthesis of glucose
from pyruvate. (pg. 16)
a. Acetyl CoA positively modulates pyruvate carboxylase. KEY ENZYME!
b. Secondary Control is regulated at the fructose-1,6-bisphosphotase.
Glucagon inhibits this enzyme. F16BP by inhibiting F26Bisphophate
formation (F26BP stimulates PFK). Look at Pg221 of Unit 2 for more
information. ATP and citrate stimulates phosphatase.
i. Never have glycolysis and gluconeogenesis occurring at the
same time!
Gluconeogenesis: Objectives
15. How much ATP, GTP, and NADH are required to synthesize 1 molecule of
glucose from pyruvate in liver?
a. Need 2 molecules of pyruvate to produce 1 molecule of glucose
b. 4 ATP, 2 GTP and 2 NADH – highly endorgonic process!
i. ATP at 2 spots
1. PyruvateOxaloacetate
2. 3-phosphoglycerate1,3 Bisphosphoglycerate
ii. GTP from OxaloacetatePEP
iii. NADH from 1,3 BisphosphoglycerateGlyceraldehyde-3-P
16. How does ethanol inhibit gluconeogenesis?
a. Ethanol inhibits the conversion of Lactate to Pyruvate by having a
strong hold on the NADH population (depleting NAD+ stores!).
Inhibits Lactic Acid Dehydrogenase. Causes Lactic Acidosis,
hypoglycermia, possible coma.