Download Intro to Metabolism II and Glycolysis

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Scribe: LEE REDDITT
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GLYCOLYSIS
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Dr. Popov began this half of the lecture by stating that memorizing structures of coenzymes would be unnecessary.
We should instead know what they are used for
I.
The Structure of Coenzyme A [S30]:
a. Coenzyme A is involved in the metabolism of fatty acids
II.
Enzyme-catalyzed transaminationsp [S31]
III. Pridoxal phosphate, the prosthetic group of aminotransferases [S32]
IV. Pyridoxal phosphate is bound to the enzyme through a Schiff-base linkage[S33]
V. Role of biotin in carboxylation reactions [S34]
a. Biotin is a water soluble vitamin.
b. Biotin is utilized as a prosthetic group of several enzymes.
c. The major function of Biotin is to serve as a cofactor for carboxylation reactions.
d. Biotin will later be discussed as being associated with pyruvate carboxylase in the Citric Acid Cycle.
e. A carboxylation reaction involves the incorporation of CO 2 into a biological molecule.
i. CO2 is a gas, and it is therefore difficult to hold an deliver it as a particle
ii. The way around this is to physically attach it to Biotin, forming carboxylbiotinyl
iii. This intermediate physically holds CO2, delivers it to the active site, releases it and allow a carboxylation
reaction to proceed.
VI. Lipoic acid (lipoate) in amide linkage with Lys residue. [S35]
a. Lipoic acid is a prosthetic group associated with a number of multi-enzyme complexes. Such as pyruvate
dehydrogenase, alphaketoglutarate dehydrogenase,
b. Lipoic acid transports acyl groups from one active site to another on the same multi-enzyme complex
c. Physically transports acyl groups via it’s long “arm” to facilitate catalysis.
VII. Tetrahydrofolate (H4 folate) [S36]
a. A reduced form of vitamin folic acid
b. Major function is the transport an provision of single carbon molecules for various biosynthetic reactions,
c. The major source of the carbon is the amino acid Serine.
d. Folic acid can transport these single carbon molecules in all oxidation states.
VIII. Conversion of one-carbon units on tetrahydrofolate [S37]
IX. Coenzyme B12 is the cofactor form of vitamin B12 [S38]
a. Vitamin B12 is the precursor for coenzyme B12.
b. A complicated molecules that carries Cobalt.
X. Coenzyme B12-catalyzed group transfer [S40]
a. Coenzyme B12 catalyzes group-exchange reactions.
b. In the reaction catalyzed by methylmalonly-CoA mutase, vitamin B12 assists the switching of side chains in LMethylmalonly CoA to form Succinyl-CoA.
c. These transmutation or tranfer-group reactions are a major sorts that are assisted by vitamin B12.
d. (NOT ON SLIDE) Another reaction important for human health is the synthesis of the methylating reagent
SAM (S-Adenosyl Methionine).
i. The enzymes that synthesize this reaction require B12 as a coenzyme.
ii. A vitamin B12 deficiency results in problems with the synthesis of SAM.
XI. Vitamin A1 and its precursors and derivatives [S41]
a. Most lipid soluble vitamins are not part of coenzymes, most of them serve a different function.
b. The precursor of Vitamin A1 is -carotene.
c. Vitamin A1 serves two functions:
i. It’s fully oxidized form, Retinoic acid, serves as a ligand for transcription factors and its primary function is
regulation. In adults, it regulates differentiation of skin and has nothing to do with metabolism itself.
ii. The other form, Retinal, is a visual pigment that senses light and ultimately translates to vision.
FUNDAMENTALS I: 11:00-12:00
Scribe: LEE REDDITT
WEDNESDAY, AUGUST 25th, 2010
Proof: JORDAN TAYLOR
POPOV
GLYCOLYSIS
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XII. Vitamin D3 production and metabolism [S42]
a. We have the ability to make this ourselves out of cholesterol. In order for this to happen we need to be
exposed to sunlight.
b. We actually need to receive the precursor of Vitamin D3 from our diet due to not always being exposed to the
sun. Milk, for example, is generally heavily fortified with Vitamin D3.
c. Vitamin D3 by itself is not biologically active. For it to become active, it must first undergo two hydroxylation
reactions.
d. The active form is 1,25-Dihydroxycholecalciferol.
e. This happens in two steps, one in the liver and one in the kidney. If you have kidney problems, often you also
have a deficiency in Vitamin D3.
f. A major role of Vitamin3 is to regulate the level of calcium in blood. Consequently, two major targets of
Vitamin D3 are the intestine (where it promotes uptake of calcium) and bone (where it promotes resorption of
calcium).
i. When the level of calcium goes down, the level of Vitamin D3 goes up and it targets the intestine and bone
to bring those levels back up.
g. Some data points to Vitamin D3 as being anti-cancerous, but such data indicates that enormous amount must
be taken each day to achieve such results.
XIII. Vitamins E and K and the lipid quinones are oxidation-reduction cofactors [S43]
a. Vitamin E is isoprenoid, in that it is built of five-carbon units of isoprene.
b. The role of Vitamin E is solely as an antioxidant. It is solubulized in membrane and lipid droplets. It reacts
with reactive oxygen species and deactivates it.
c. Vitamin K is a coenzyme that is participating in carboxylation reactions. It participates specifically in
carboxylations in which Glutamic Acid is carboxylated in certain proteins that are involved in blood clotting.
This creates calcium binding sites in those proteins and allows for blood clotting.
XIV. Vitamin K stimulates the carboxylation of glutamate residues [S44]
a. Vitamin K promotes this carboxylation reaction.
b. When you have a deficiency of Vitamin K, you have issues with blood clotting.
c. Inhibitors of Vitamin K are used as blood thinner drugs.
i. An excess of such as drug can be used as rat poisons that cause internal bleeding.
XV. Summary of key points about metabolism
BEGIN NEXT POWERPOINT ON GLYCOLYSIS
Dr. Popov stated that the goal of this discussion was to understand how glycolysis occurs, what functions it serves and
how it is regulated.
I.
Glycolysis [S1]
II. Overview [S2]
III. Fate of major metabolic fuels [S3]
a. We have three major metabolic fuels: (1) Glycogen (2) Triglyceride (3) Protein
i. Each of these can be fully oxidized and converted to Acetyl-CoA
b. Our primary concern is the Glycolytic pathway in which carbohydrates end up in the form of glucose,
broken down into pyruvate and eventually enter the Citric Acid Cycle.
IV. Major metabolic pathways of glucose [S4]
a. Important to note that the glcolytic pathway is not the only metabolic fate of glucose in the cell.
b. There are a variety of ways that glucose can be used when it enters into the cell:
i. It can be converted into glycogen for storage, particularly in the muscle of the heart and liver.
ii. It can enter the pentose phosphate pathway, which is a major source of pentoses for the
production of nucleotides.
c. So, every time glucose enters the cell it can undergo a variety of metabolic transformations.
V. Catabolic fates of the pyruvate formed in glycolysis [S5]
a. Glycolysis is more or less a universal process across organisms. It takes glucose in and spits out
pyruvate. What happens next is cell and organism specific.
FUNDAMENTALS I: 11:00-12:00
Scribe: LEE REDDITT
WEDNESDAY, AUGUST 25th, 2010
Proof: JORDAN TAYLOR
POPOV
GLYCOLYSIS
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b. There are a number of cells that don’t have mitochondria at all (such as many cells of the eye). These
cells cannot take the pyruvate from glycolysis through the CAC.
c. Muscle cells under intense contraction are in an anaerobic environment and also cannot proceed through
the CAC. Pyruvate then enters the pathway of lactate fermentation. This allows for recycling of the
NADH generated in glycolysis and lactate is delivered to the liver.
d. In yeast, glycolysis takes the pathway of ethanol fermentation. Physiologically this is similar to lactate
fermentation, but takes a different path.
i. Lactate is an acid. Intense exercise leads to muscle aches. The reason for this is the production
of lactate causes damage to cells due to the decreased pH.
ii. Ethanol on the other hand is a neutral compound and does not cause pH damage to cells.
VI. Glycolysis is a preparatory pathway for aerobic metabolism of glucose [S6]
a. Glycolysis takes place in the cytosol and this process can occur with or without O 2.
b. Oxidation of fatty acids and amino acids occurs in the mitochondria. Mitochondrial function requires O2
and without it these pathways are not available to cells to produce energy.
c. Glucose and carbohydrates are unique fuels in that they allow us to survive under oxygen deficient
conditions.
i. Strained muscle cells are good indicators of this in that during anaerobic conditions, glucose is
able to provide energy via anaerobic glycolysis.
VII. Mechanism of Glucose Uptake [S7]
a. Before glucose can be oxidized in the cell, it must be taken up into the cell.
b. Glucose transporters are present on cell membranes. This binds glucose on the outside, a
conformational change occurs and glucose is released on the inside. This occurs via facilitated diffusion.
c. Glucose follows a concentration gradient to enter a cell. As glucose concentration increases, you will see
the transporters become saturated
d. There are four glucose transporters that our body makes.
i. They all differ by how fast they transfer glucose as well as their affinity for glucose.
VIII. Major ways in which glucose is metabolized in erythrocytes. [S8]
a. Different cells handle glucose differently
b. In erythrocytes, only thing they do is oxidize glucose through glycolysis and release lactate. This is the
major source of ATP because they lack mitochondria.
c. The only other use of glucose is in the production of pentose phosphates that can be used in nucleotide
synthesis.
IX. Major ways in which glucose is metabolized in brain [S9]
a. The major fuel in our brains is glucose, due to the fact that the brain barrier is impermeable to fatty acids.
b. The brain makes as much energy as possible via the oxidation of glucose. It is oxidized completely via
glycolysis and the Citric Acid Cycle.
c. Consequently, the brain consumes a large amount of glucose and oxygen available.
d. The brain also utilizes Glucose as a source of pentose phosphates.
X. Major ways in which glucose is metabolized in muscle and heart [S10]
a. Muscle and heart also oxidize glucose via glycolysis and the CAC when oxygen levels are sufficient.
b. Depending on muscle activity, anaerobic glycolysis may also be utilized (lactate fermentation).
c. Again, pentose phosphates are an option
d. Additionally, synthesis of glycogen occurs. This is used to store energy and later provide it during muscle
contraction.
XI. Major ways in which glucose is metabolized in liver [S11]
a. This is where glucose metabolism is most complicated.
b. The liver can breakdown glucose through glycolysis, oxidize it through the CAC, and can convert
pyruvate to lactate under certain circumstances.
c. The liver can synthesize glycogen, though this is a different form than that in the muscles. This form is
used to supply glucose quickly to the brain in certain circumstances.
d. The liver has the capability to synthesize glucose through gluconeogenesis.
e. The liver also utilizes glucose as a precursor for the biosynthesis of fats.
f. Lastly, the liver uses glucose to form pentose phosphates and disposal in the form of glucarindes.
FUNDAMENTALS I: 11:00-12:00
WEDNESDAY, AUGUST 25th, 2010
POPOV
GLYCOLYSIS
Scribe: LEE REDDITT
Proof: JORDAN TAYLOR
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XII. Preparatory and Payoff Phase [S12]
a. Glycolysis is the pathway that takes glucose in and via multiple enzymatic reactions leaves you with 2
molecules of pyruvate.
b. It is viewed as a process that consists of two phases:
i. Preparatory phase or priming phase – glucose skeleton is prepared for oxidation
ii. Payoff phase – production of energy
c. Two molecules of ATP are used during the preparatory phase to generate phosphorylated intermediates.
d. ATP is synthesized in the payoff phase during two steps.
e. The overall outcome is two molecules of pyruvate and two molecules of ATP
f. NADH is generated as well and must be recycled. This is usually achieved in active muscle cells via
lactate formation.
XIII. Phosphorylation of Glucose [S13]
a. Glucose enters through a transporter and is immediately is phosphorylated via an enzyme called
hexokinase.
i. Kinases – enzymes that use ATP to phoshorylate molecules. Protein kinase will phosphorylate
proteins and hexokinases will phosphorylate hexoses.
b. The purpose of phosphorylation of glucose is to entrap glucose into the cell. This avoids the reversible
nature of glucose transporters.
c. This reaction is highly exergonic and irreversible.
d. If this reaction is allowed to run uncontrollably, then you will eventually run out of ATP.
e. This reaction is regulated in glycolysis by feedback inhibition via Glucose-6-Phosphate.
i. An increase in the concentration of Glucose-6-Phosphate shuts this reaction down.
XIV. Conversion of glucose-6-phosphate to fructose-6-phosphate [S14]
a. Glucose-6-Phosphate is isomerized into Fructose-6-Phosphate via the enzyme phosphohexose
isomerase
b. The reason for this that for the last step of the priming phase involves splitting a hexose.
c. When you split Fructose-6-Phosphate you get the desired pair of Three-Carbon molecules.
d. Splitting Glucose-6-Phosphate would require two steps and subsequently two enzymes to achieve the
same that can occur via splitting Fructose-6-Phosphate.
e. This reaction is reversible.
XV. Phosphorylation of fructose 6-phosphate to fructose 1,6-bisphosphate [S15]
a. This reaction marks the stage when glucose is committed to the glycolytic pathway.
b. This reaction is also highly exergonic and irreversible.
c. Represent a major step in which glycolysis is regulated.
XVI. Cleavage of fructose-1,6-bisphosphate [S16]
a. Splitting Fructose that is phosphorylated at both ends results in two phosphorylated trioses
b. If these were not phosphorylated they could easily leave the cell. By being phosphorylated, they are
entrapped and will continue on the catabolic pathway.
c. Represent that last step of the preparatory phase.
XVII. Interconversion of triose phosphates [S17]
a. The Glyceraldehyde-3-Phosphate produced from the previous reaction continues on into the payoff phase
of glycolysis.
b. The other triose produced, Dihydroxyacetone phosphate, is rapidly converted to Glyceraldehyde-3Phosphate via the enzyme triose phosphate isomerase.
c. This product will now also enter into the next phase of glycolysis.
XVIII. Oxidation of glyceraldehyde-3-phosphate to 1,3-bisphosphoglycerate [S18]
a. Beginning of the stage where the cell actually oxidizes the products made in the preparatory phase to
eventually produce energy.
b. 1,3-Bisphospoglycerate is a high-energy compound. The phosphoryl group is high-energy and can
phosphorylate ADP into ATP.
c. The reaction of the aldehyde group and phosphate requires an electron carrier.
i. NAD+NADH
FUNDAMENTALS I: 11:00-12:00
Scribe: LEE REDDITT
WEDNESDAY, AUGUST 25th, 2010
Proof: JORDAN TAYLOR
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GLYCOLYSIS
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d. Enzyme uses energy released during the oxidation of aldehyde group into carboxyl group to make a highenergy phosphoryl-containing compound.
XIX. The glyceraldehyde-3-phosphate dehydrogenase reaction [S19]
a. Enzyme converts glyceraldehyde-3-phosphate into a hemiacetal via the assistance of NAD+
b. Hemiacetal oxidized to form a thioester, which is a high-energy intermediate.
c. This reacts with phosphate to form the high-energy compound 1,3-Bisphospoglycerate.
XX. Phosphoryl transfer from 1,3-bisphosphoglycerate to ADP [S20]
a. 1,3-Bisphospoglycerate used to phosphorylate ADP into ATP. Leaves you with 3-Phosphoglycerate.
b. The enzyme involved is phosphoglycerate kinase
c. This is a relatively exergonic reaction, but the reverse will occur during gluconeogenesis.
i. Notice the enzyme is named for the reverse reaction.
d. 1 Glucose  2 1,3-bisphosphoglycerates  2ATP
i. All the ATP invested in the preparatory phase has been recovered during this step.
XXI. Conversion of 3-phosphoglycerate to 2-phosphoglycerate [S21]
a. 3- Phosphoglycerate isomerized into 2-Phosphoglycerate
b. Reason for this is that the next step will allow for transformation of 2-phosphoglycerate into a high-energy,
phosphate-containing intermediate via a dehydration reaction.
XXII. Mechanism of the phosphoglycerate mutase reaction [S22]
XXIII. Dehydration of 2-phosphoglycerate to phosphoenolpyruvate [S23]
a. Enolase catalyzes this reaction by removing a molecules of water.
b. Phosphoenolpyruvate is a high-energy compound in that when it is hydrolyzed a great deal of energy is
released.
i. Capable of phosphorylating ADP  ATP
XXIV. Transfer of the phosphoryl group from phosphoenolpyruvate to ADP [S24]
a. Irreversible reaction
b. Notice again the name of the enzyme is for that of the reverse reaction.
c. Pyruvate is the end product of glycolysis.
d. 2 more ATP produced, to bring the grand total to 2 for the whole process.
XXV. Pyruvate is the terminal electron acceptor in lactic acid fermentation [S25]
a. When glycolysis is run aerobically, you have to recycle NADH generated during the early stages
b. Lactate fermentation achieves this via reduction of pyruvate into lactate.
c. NAD+ can then enter the next cycle of glycolysis.
d. In cells with mitochondria, excess NADH is recycled when pyruvate is oxidized via the CAC.
XXVI. Lactase dehydrogenase reaction [S26]
a. Pyruvate carbon reduced at the expense of electron provided by NADH
XXVII. Entry of dietary hexoses into the preparatory stage of glycolysis [S27]
a. The glycolytic pathway is not only for the oxidation of glucose. It can oxidize a variety of different sugars.
b. A number of these sugars converge on Fructose-6-Phosphate
i. The reason this step has been chosen as a regulatory one. This allows control of multiple
pathways in one place.
XXVIII. The 2,3-bisphosphpglycerate shunt [S28]
a. In erythrocytes, glycolytic pathways provide not only a source of energy but also a source of 2,3bisphosphoglycerate.
b. This is a major allosteric regulator oh hemoglobin by regulating its ability to pick up O 2.
c. This compound is made out of glycolytic intermediates.
d. By controlling glycolysis you control the concentration of 2,3-bisphosphoglycerate in erythrocytes and
their ability to pick up O2.
FUNDAMENTALS I: 11:00-12:00
Scribe: LEE REDDITT
WEDNESDAY, AUGUST 25th, 2010
Proof: JORDAN TAYLOR
POPOV
GLYCOLYSIS
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XXIX. Relative changes in [ATP] and [AMP] when ATP is consumed [S29]
a. When there is plenty of ATP, glycolysis is inhibited & when ATP goes down, glycolysis is activated.
b. Glycolysis does not occur though because of changes in ATP but instead due to the change in
concentration of AMP.
c. Just a 10% change in the [ATP] (which would be a worse-case scenario in a cell) can result in a 600%
change in [AMP].
XXX. Opposing reactions of glycolysis and gluconeogenesis [S30]
XXXI. Regulation of phosphofructokinase-1 [S31]
XXXII. Regulation of phosphofructokinase-1 [S32]
XXXIII. Structure of Fructose 2,6-bisphosphate [S33]
XXXIV. Role of fructose-2,6-bisphosphate in regulation of PFK-1 [S34]
a. Fructose 2,6-phosphate is a good regulator of glycolysis.
b. This compound is made specifically to regulate glycolysis in response to insulin and glucagon.
c. Aside from short term regulation of glycolysis in regards to the formation of ATP, the rate of glycolysis is
regulated in long term as well.
i. For example, following a meal, there is plenty of glucose available and all cells will likely be
utilizing glycolysis. Fructose-2,6-bisposphate is active in this state in order to promote glycolysis.
ii. When one is starving, glucose levels are low. Glycolysis is shut down in certain tissues to allow
conservation of glucose for the brain. Fructose-2,6-bisposphate breaks down in this state and
glycolysis in inhibited.
d. In the liver, high concentration of insulin promotes activity of PFK-1 and the synthesis of Fructose-2,6Phosphate.
e. When we starve and insulin levels are low and glucagon levels go up. High glucagon levels promote the
breakdown of Fructose-2,6-Phosphate. shutting down of glycolysis in the liver and at the same time
activates gluconeogenesis.
XXXV. Regulation of fructose-1,6-bisphosphate and phosphofructokinase-1 [S35]
XXXVI. Regulation of pyruvate kinase [S36]
a. Primarily an important regulatory step in the liver.
b. Pyruvate kinase reaction is virtually irreversible.
i. Therefore, a means to make ATP, but not phosphoenolpyruvate
ii. When gluconeogenesis is on, this reaction cannot happen.
c. When we go into starvation, this reaction must be shut down. In the liver, Pyruvate kinase is therefore
phosphorylated so that it becomes inactive.
d. A number of different allosteric regulators inform cells of a state of starvation and that they need to
conserve glucose and utilize gluconeogenesis.
e. Acetyl-CoA, long-chain fatty acids, etc. would tell pyruvate kinase that there are alternate sources of
energy and that glycolysis is not necessary.
XXXVII. Insulin stimulates glucose uptake by adipose tissue and muscle [S37]
a. Four different glucose transporters.
b. Skeletal and cardiac muscle mostly have Glucose transporter IV, which is regulated by insulin
i. Low insulin levels – Glucose transporter not present on cell membrane
ii. High insulin levels – Glucose transporter readily available and transport of glucose into a cell is
increased.
c. In diabetes, this regulatory step is compromised. The ability of muscle cells to pick up glucose is
decreased.
XXXVIII. Kinetic properties of glucokinase and hexokinase [S38]
a. The role of the liver is not just to metabolize glucose for energy but is a major organ that regulates
glucose concentrations in our body.
b. Glucokinase is a specialized enzyme of the liver. Unique because it has a lower affinity to glucose then it
tissues counterpart glucokinase.
FUNDAMENTALS I: 11:00-12:00
Scribe: LEE REDDITT
WEDNESDAY, AUGUST 25th, 2010
Proof: JORDAN TAYLOR
POPOV
GLYCOLYSIS
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c. In a state of high glucose concentration, glucokinase will pick up some glucose but still allow a significant
amount thorough an available to peripheral tissues.
d. As blood glucose levels go down, the ability of the liver to pick up glucose goes down drastically.
e. The liver will now transition from consumption of glucose to manufacturing it.
XXXIX. Regulation of glucokinase by sequestration in the nucleus [S39]
a. Hexokinase IV is also a regulatory enzyme.
b. Enzyme activity is regulated by glucose and fructose-6-phosphate.
c. Liver cells make a special regulatory protein that binds hexokinase IV in the nucleus and makes it
inactive.
XL. Important regulatory features of glycolytic pathway [S40]
a. Look at this when you have time
XLI. Summary [S41]
[End 50:37 mins]