Lectures 58-82 Flashcards
What is the function of Metabolism
To obtain and trap chemical energy from substrates
To build precursors to macromolecules from substrates
To assemble precursors into macromolecules. Ex: DNA, Glycogen, Fat
To degrade macromolecules into simpler molecules
What is catabolism?
Catabolism is the oxidative breakdown of large macromolecules into smaller, simpler compounds. Usually it is accompanied by release of free energy and trapping this energy as ATP
What is anabolism?
Anabolism is the enzymatic synthesis of large macromolecules from smaller, simpler precursors. Usually it requires input of energy.
What is AMPHIBOLIC?
A cycle or process that has both catabolic and anabolic components. Ex- TCA cycle.
What are the differences between catabolism and anabolism?
Enzymes-allows for regulation and direction. Many enzymes may be same in a reversible metabolic pathway but some will differ
Energetics- ATP made in Catab; used for Anab
Cofactors-NAD→ NADH used for catabolism
NAD(P)H→NADP occurs for anabolism
Cellular localization may differ e.g cyto vs mito
What kind of process makes ATP?
Catabolic ADP-> ATP
What kind of process uses ATP?
Anabolic. ATP->ADP
What direction is the cofactor used in catabolism?
NAD+–>NADH oxidizing agent becomes reduced
The coenzyme is, therefore, found in two forms in cells: NAD+ is an oxidizing agent – it accepts electrons from other molecules and becomes reduced.
What direction is the cofactor used in anabolism
NADH–> NAD+ or NAD(P)H–>NADP Reduced to oxidized.
NADH can be used as a reducing agent to donate electrons.
What regulates metabolism?
- Availability & concentration of substrates and COFACTORS. Need to regenerate cofactors***
- Availability/Need for ATP
- Enzyme characteristics-heme,metal, dimers
- Regulatory enzymes-often allosteric. ATP ↓catabolic reactions while ADP ↑them. (basically Le’Chatlier’s) Product inhibition of anabolic reactions
- Genetic control of amount of enzyme in cell. Constitutive VS adaptive enzymes.
- Hormonal regulation- chemical messenger which↑ or↓ a metabolic reaction in another cell.
Try to memorize chart of where different runs occur.
Slide 8, 1st lec ppt.
What are the two ways to produce ATP?
substrate level phosphorylation or oxidative phosphorylation.
Why does ATP’s delta G being in the middle help?
Note that ATP is in the middle of this group, which means it can transfer a P to glucose to produce glucose 6P or it can be produced when 1,3 BPG transfers a P to ADP to yield ATP.
G + ATP → G6P + ADP
1,3 bis phosphoglycerate + ADP → 3 phosphoglycerate + ATP
Many reactions involve the oxidation of substrates using NAD+ to form what?
Many reactions involve oxidation of substrates using NAD+ as the acceptor of two electrons to form the oxidized product and NADH.
NAD+ does what to get to NADH
Accepts two electrons. Same goes for FAD-> FADH2
The metabolism of glucose and fatty acids is regulated by which hormones?
Insulin, glucagon and epinephrine (epi).
Where do insulin and epi regulate met of glucose and fatty acids?
in many tissues.
Where does glucagon regulate glucose and fatty acid met?
Primarily in liver and adipose tissue.
Where is epi produced and why?
Epi is produced in the adrenal glands in response to various types of stress.
What produces insulin and why?
The β cells of the pancreas produce insulin in response to high glucose, e.g., high carbohydrate diets.
What produces glucagon and why?
The α cells of the pancreas produce glucagon in response to low glucose, e.g. starvation, low carbohydrate diets.
What is the primary function of insulin?
To promote the utilization of glucose by the body.
What is the primary function of glucagon?
To promote the production of glucose by the liver
What is type 1 diabetes?
Type 1 diabetes is associated with a lack of insulin production due to destruction of the β cells of the pancreas, largely via autoimmune attack on these cells.
What is type 2 diabetes?
Type 2 diabetes is generally associated with increased resistance to the actions of insulin; insulin is produced but its effectiveness in enhancing utilization of glucose is decreased. Obesity coupled to lack of exercise promotes type 2 diabetes by mechanisms that are not clear.
What does diabetes result in?
Diabetes is thus associated with accumulation of high levels of glucose, especially in the blood. This causes metabolic problems: energy production from glucose is impaired; fat metabolism is elevated to provide energy but much of the fat is oxidized to ketone bodies (lecture 8) which are acidic and can cause ketosis; high levels of circulating glucose can react non-enzymatically with proteins and enzymes and glycosylate them to form modified proteins with altered functions; high levels of glucose can be reduced by aldose reductase to sorbitol, a sugar alcohol that can increase osmotic pressure, e.g., in the eye lens.
How do we measure blood glucose levels?
A convenient assay for measuring long-term blood glucose levels relies on the fact that hemoglobin reacts readily with high glucose to form a glycosylated hemoglobin called hemoglobin A1c. Diabetics are monitored for their hemoglobin A1c levels, as high levels (> 7 mg% or 7 mg/dl blood) are indicative of poor glucose control due to not taking insulin and other medications or not eating properly.
How does glucagon promote glucose production in the liver?
Glucagon promotes glucose production in the liver by two primary mechanisms: stimulation of glucose synthesis (gluconeogenesis) and stimulation of glycogen breakdown.
What is glycogen?
Glycogen is a polymer made up of many glucose residues, which functions as a storage form of glucose.
What does glucagon do independent of it’s glucose producing pathways?
Glucagon increases fatty acid release from triglycerides stored in adipose tissue. The liver will oxidize fatty acids for its own energy requirements, since the liver will be providing glucose for fuel for other tissues, not for itself.
How does insulin promote the utilization of glucose?
stimulating glycolysis, a glucose degradation pathway;
stimulating glucose uptake in some tissues, e.g. muscle, adipose tissue;
stimulating glycogen formation;
stimulating fatty acids synthesis from glucose (lecture 9);
and stimulating protein synthesis (several amino acids can be produced from glucose).
What is the composition of the insulin receptor?
It’s a tetramer made up of two alpha subunits and two beta subunits. The beta units span the membrane. When insulin binds to the alpha subunit the binding results in the autophosphorylation of several tyrosine residues on the beta subunits. (The beta subunit has tyrosine kinase activity when insulin binds to the alpha subunit.)
What happens when the Beta subunit on the insulin receptor is activated?
It can phosphorylate other proteins, the major ones being insulin receptor substrates IRS 1 and 2.
What is the major protein phosphorylated by the ISRs?
PIP3 kinase
What does PIP3 Kinase activate?
The signaling molecule AKT. pAKT can activate proteins such as pho sphatases, PKC, mTOR, that subsequently carry out the actions of insulin.
The Glucagon and Epinephrine receptor structure?
Glucagon binds to its receptor found in liver and fat tissue, while the epi receptor is widely present. These two receptors are G-protein-coupled receptors with seven transmembrane spanning loops (GPCR). To review GPCRs: the ligand-bound receptor activates specific G proteins. Typically, G proteins are heterotrimers made up of α, β and γ subunits. In the inactive state, the α subunits bind GDP (hence “G” proteins). The activated glucagon or epi receptor catalyzes an exchange of GDP with GTP. This is followed by dissociation of the βγ subunits to form the active α-GTP G protein. This activates the enzyme adenylate cyclase to produce the critical second chemical messenger cyclic AMP:
As a general “rule,” glucagon and epi promote the what of enzymes?
The phosphorylation of enzymes by activating cAMP- dependent PKA.
As a general rule insulin promotes the what of enzymes?
As a general “rule,” insulin promotes the dephosphorylation of enzymes by activating certain phosphatases, e.g., protein phosphatase 2A.
How is the glucagon (epi) signal turned off?
The αGTP subunit has a GTPase activity, and with time the GTP is hydrolyzed to form the αGDP subunit. This reassociates with the βγ subunits that have been “hanging around” to reestablish the inactive GDP-αβγ G protein. With time, the ligands glucagon and epi dissociate from the receptor. Also, cAMP is hydrolyzed by the enzyme phosphodiesterase to AMP; hence, the cAMP-PKA signal is turned off.
What does insulin increase that helps combat the actions of glucagon and epi?
phosphodiesterase activity
Glucose is a polar molecule and will not easily enter cells, how is that fixed?
it must be transported into cells by carrier-mediated mechanisms.
What are the main transport systems for glucose uptake into cells?
Liver, RBC, brain, pancreas, and most cells carry out a passive carrier-mediated glucose transport via their glucose carriers GLUT1 and GLUT3. These carriers have a low Km for glucose – about 1 mM (serum glucose levels are typically 4 to 8mM) – so they catalyze basal glucose uptake.
Liver and pancreas also contain GLUT2, which has a Km for glucose of 15-20 mM. This allows the pancreas to sense high glucose (to produce insulin) and the liver to utilize this high glucose.
What effect does insulin have on the rate of glucose uptake by GLUT 1, 2, and 3?
Insulin has no effect on the rate of glucose uptake by GLUT 1, 2 and 3.
What cells contain GLUT 4?
Muscle and fat cells.
How does insulin affect the rate of glucose uptake by GLUT 4?
Insulin strikingly elevates the number of GLUT 4 carriers on the plasma membrane of muscle and fat cells so that glucose uptake is elevated. This is a major mechanism by which insulin elevates glucose utilization in muscle and fat cells, i.e., increasing glucose uptake.
How does insulin increase GLUT4 in the plasma membrane?
nsulin increases GLUT4 in the plasma membrane by stimulating transport of GLUT4 molecules sequestered in the golgi to the plasma membrane. The downstream insulin-activated target AKT plays a role in this transfer of GLUT4 from the golgi to the plasma membrane.
Where is GLUT 5 expressed?
GLUT5 is present in the GI tract and kidney where it catalyzes active transport of glucoses. Active uptake of glucose is linked to Na+ transport by the gut and kidneys.
What is glycolysis?
Glycolysis refers to the “lysis” (breakdown) of glucose. It occurs in the cytosol fraction of all living things.
What is the basic overall reaction of glycolysis? (it’s reversible)
Glucose+2NAD+ +2Pi +2ADP → 2pyruvicacid+2ATP+2NADH
What is the major function of glycolysis?
The major function of glycolysis is to provide energy for cells. For some cells, e.g., RBCs, glycolysis is the only source of energy. For other cells, it is the primary source of energy, e.g., brain, embryonic tissue, exercising muscle. Technically speaking, glucose only produces 2 ATP/glucose when it is oxidized to pyruvate. However, the pyruvate and the NADH can provide more ATP when they are oxidized further in the mitochondria.
What is another important function of glycolysis?
Another important function of glycolysis is to provide intermediates for other metabolic reactions. Examples of such intermediates are: α-glycerophosphate for triglyceride and phospholipid synthesis; 2,3 bis-phosphoglycerate in the RBC to regulate hemoglobin-oxygen affinity; acetyl CoA from the pyruvate, which can be used to synthesize fatty acids, cholesterol, ketone bodies, and steroids; amino acids such as serine, alanine, glycine.
What is step 1 of glycolysis?
Step 1 is catalyzed by hexokinase, which catalyzes the transfer of a phosphate from ATP to produce glucose 6-P (G6P) and ADP. Note there is an initial input of energy needed to activate the glucose. ** irreversible.
What can inhibit hexokinase?
Hexokinases, which are not specific for glucose, are constitutive enzymes with a low Km for glucose, and are subject to product inhibition by G6P and ADP.
What is glucokinase?
The liver and the pancreas have a special hexokinase isoform called glucokinase. Glucokinase has a high Km for glucose, is specific for glucose, is not subject to inhibition by G6P and ADP and is inducible at the transcriptional level by high carbohydrate diet and insulin, and repressed by glucagon.
Step 1 glycolysis reaction?
Glucose+ ATP–> glucose 6-P (G6P) and ADP, catalyzed by hexokinase.
Step 2 in glycolysis?
G6P –> F6P, catalyzed by phosphoglucoisomerase
Step 3 in glycolysis?
Step 3 is conversion of F6P to fructose 1,6 bis-phosphate (F1,6 bis P) by phosphofructokinase (PFK):
F6P+ATP → F1,6bisP+ADP
The reaction is irreversible and is the rate-limiting step in glycolysis. Note the input of a second ATP. Since glycolysis produces 2 ATP/glucose, the subsequent steps will have to produce a total of 4 ATPs.
What inhibits PFK?
PFK is inhibited by ATP and by citrate (high energy signals)
What stimulates PFK?
Stimulated by AMP, Pi, NH4+ and, surprisingly, by F1,6 bis P, the product.
What is the structure of PFK?
Thus PFK has 2 substrate binding sites (ATP, F6P), 2 allosteric inhibitory sites, and 4 allosteric activator sites.
What effect does fructose 2,6 bis (F2,6 bis P) have on PFK?
It’s a powerful activator of PFK.
PFK is BLANK when dephosphorylated and F2,6bisPhosphotase is BLANK when dephosphorylated.
PFK is ACTIVE when dephosphorylated and F2,6bisP is INACTIVE when dephosphorylated.
Which states of PFK and F26bP does insulin promote?
This occurs in the presence of insulin, i.e., insulin promotes the dephosphorylated states of the kinase (active) and F2,6 bis phosphatase (inactive). This makes sense because insulin wants to stimulate glycolysis and therefore activate RFK.
Which states of PFK and F26bP do glucagon and eli promote?
Glucagon and epinephrine, acting via cAMP-PKA, promote the phosphorylated state of the kinase (inactive) and F2,6 bis phosphatase (active), thus lowering levels of F2,6 bis P and inhibiting PFK.
What are steps 4 and 5 of glycolysis?
aldolase cleaves the 6-carbon F1,6 bis P to 2 trioses, dihydroxyacetone P (DHAP) and glyceraldehyde 3-P (G3P) by cleaving between carbons 3 and 4. At this point:
Glucose+2ATP → DHAP+G3P+2ADP
**DHAP can be reduced to α-glycerophosphate, which is needed for triglyceride and phospholipid synthesis.
For glycolysis, the DHAP is converted to G3P by triose phosphate isomerase (step 5). The above 2 steps are reversible. The net reaction therefore is now:
Glucose+2ATP → 2G3P+2ADP.
What is step 6 of glycolysis?
In step 6, G3P is oxidized by glyceraldehyde 3-P dehydrogenase (GAPDH)
G3P + NAD+ + Pi ↔ 1,3 bis phosphoglyceric acid (1,3 bis PGA) + NADH
What is step 7 of glycolysis?
1,3 bis PGA is a high-energy compound and produces ATP and 3-phosphoglyceric acid (PGA) in step 7, which is the phosphoglycerate kinase reaction (Fig. 3).
1,3bisPGA +ADP ↔ 3PGA+ATP
Since 1 glucose yielded 2 G3P, we net 2 ATPs when 1 mole of glucose is metabolized to 2 moles of 3 PGA.
Substrate Level Phosphorylation?
The energy associated with the oxidation of the aldehyde on C1 of G3P to the carboxylic acid on C1 of 3PGA was trapped as a high energy intermediate 1,3 bis PGA, which then produced ATP.
Note, this occurs in the absence of mitochondria and oxygen.
Step 8 and 9 of glycolysis?
In step 8, phosphoglyceromutase converts 3 PGA to 2 PGA, which is followed (step 9) by removal of H2O from 2 PGA by enolase to produce phosphoenolpyruvate (PEP), a high-energy compound.
Step 10 of glycolysis?
In the final step of glycolysis (step 10), the high energy PEP can yield ATP as catalyzed by pyruvate kinase (Fig. 4), an irreversible reaction.
PEP + ADP → Pyruvate + ATP
2ATPs per glucose will be produced at this step. Thus, we used 2 ATPs at the beginning (hexokinase, PFK) and we produced 4 ATPs (2 at the G3P 3PGA step and 2 at the pyruvate kinase step) for a net of 2 ATPs for each glucose → 2 pyruvates.
Draw out all 10 steps of gycolysis.
screenshot 15
What are the 3 irreversible steps of glycolysis?
Glucose+ ATP–> glucose 6-P (G6P) and ADP, catalyzed by hexokinase.
F6P+ATP → F1,6bisP+ADP cat by phosphofructokinase (PFK) **rate limiting step
PEP + ADP → Pyruvate + ATP Catalyzed by PK
How does insulin stimulate glycolysis?
Insulin stimulates glycolysis by increasing glucokinase levels and by activating the PFK and PK reactions via dephosphorylations
How does glucagon inhibit glycolysis?
Glucagon inhibits glycolysis by decreasing glucokinase levels and inhibiting PFK and PK via cAMP-PKA- dependent phosphorylations.
What inhibits PFK and PK?
High-energy signals, such as ATP and citrate, inhibit PFK while ATP, NADH, and acetyl CoA inhibit PK.
How does fructose enter glycolysis?
Thus, fructose enters glycolysis at the DHAP and G3P level, which continue on to pyruvate or in the reverse direction, to glucose.
How is fructose broken down?
Fructose–>F1P Uses 1 ATP cat by fuctokinase.
F1P–> DHAP + GLyceraldehyde.
DHAP-> G3P cat by adolase
GLyceraldehyde-> G3P by triosephosphokinase
After entering in glycolysis it can proceed in either direction, down to pyruvate or up to glucose.
How is galactose broken down?
Galactose-> galactose 1-P by galactokinase
galactose 1-P-> glucose 1-P (G1P) by galactose 1-P uridyl transferase
G1P-> G6P by phosphoglucomutase, which enters glycolysis or is converted to free glucose (mostly in liver).
What induces galactosemia and how do you treat it?
Uridyl transferase is deficient, therefore high levels of galactose 1P accumulate, which, similar to F1P, is toxic to the liver. Treatment is obviously to limit lactose in the diet, a problem for nursing mothers.
What are the two fates of pyruvate produced by glycolysis?
it can be reduced to lactic acid (lactate) or it can enter the mitochondria for further metabolism by pyruvate dehydrogenase (PDH) to acetyl CoA.
In liver, pyruvate can also be metabolized to oxalacetate by pyruvate carboxylase (next lecture). In RBCs, which lack mito, pyruvate must be reduced to lactate.
What does lactate dehydrogenase do?
LDH catalyzes reduction of pyruvate to lactate. This reaction is reversible.
Why is the reduction of pyruvate to lactic acid so imperative to RBCs?
This reaction is critical to RBCs because NADH is reoxidized back to NAD+, which is necessary for glycolysis to continue at the glyceraldehyde 3-P dehydrogenase step.
Tissues containing mito can oxidize the NADH, after it is transported into the mito, by the respiratory chain. The LDH reaction is also important to deoxidize NADH whenever glycolysis is very rapid, e.g., during muscle contraction, when high insulin levels are high, or in tumor cells.
What is the fate of the lactic acid produced in glycolysis?
Lactic acid leaves the RBC or muscle or tissue where it was produced and circulates in the blood. High levels alter blood pH (lacticacidemia), can cause cramps (e.g., in marathon runners), and can aggravate gout (will be discussed later). Most lactate enters tissues where it is reoxidized back to pyruvate, which then undergoes the pyruvate dehydrogenase reaction for energy production, or can be converted to glucose in the liver during gluconeogenesis.
What are the different configurations possible for LDH?
LDH can be made up of 2 different peptide chains, H and M chains. LDH is a tetramer, and therefore five different LDH isoforms can be produced – H4, H3M, H2M2, HM3 and M4.
What LDH isoforms are found in the heart and brain?
H4 and H3M
What LDH isoforms are found in the RBCs and Skeletal muscle?
M4 and M3H
Why is H4 predominant in the heart and brain and M4 predominant in the RBC and muscles.
H4 has a higher binding affinity for Lactate and NAD and are expressed in tissues that would have use for holding on to those things. Meaning they have oxygen readily available and can further process lactate. RBCs and skeletal muscle often can only perform anaerobic respiration so they don’t want an isotherm that will hold onto the lactate they want to get rid of it and get it out so they can make more.
How does alcohol fermentation work?
A great thing happens to pyruvate in yeast cells, it is converted to ethanol (alcohol). Yeast contain an enzyme pyruvate decarboxylase (do not confuse with E1 of the PDH complex discussed next) which causes the decarboxylation of pyruvate to CO2 plus acetaldehyde
Vitamin B1, thiamine pyrophosphate (TPP), is the cofactor for this reaction. Acetaldehyde then is reduced by alcohol dehydrogenase to ethanol. In this way, the NADH produced by glycolysis is used to produce ethanol and the net reaction of this fermentation is:
Glucose+2ADP+2Pi 2CO2 +2Ethanol+2ATP
What does pyruvate decarboxylase do?
Decarboxylates pyruvate into Co2 plus acetaldehyde in yeast.
What is the cofactor for Pyruvate-> Acetaldehyde in yeast?
Vitamin B1, thiamine pyrophosphate (TPP)
What does Pyruvate Dehydrogenase (PDH) do?
In most tissues, in order to extract more energy from glucose, pyruvate is converted to acetyl CoA by PDH. The acetyl CoA can be further oxidized in the TCA cycle, generating NADH and FADH2 for energy production by the respiratory chain.
Is the Pyruvate Dehydrogenase (PDH) reversible?
No. The PDH reaction is irreversible. Therefore acetyl CoA cannot be converted to pyruvate, which means that fatty acids which are primarily oxidized to acetyl CoA cannot produce pyruvate and therefore glucose.
What is pyruvate dehydrogenase and what is its cofactor?
E1, Thiamine pyrophosphate (TPP) (derivative of thymine, vitamin B1)
What is Dihydrolipoyl Transacetylase and what is its cofactor?
E2, Lipoamide
What is Dihydrolipoyl Dehydrogenase and what is its cofactor?
E3, FAD (derivative of riboflavin)
What is the first step of the PDH reaction?
Pyruvate reacts with the carbanion of TPP on E1 to yield CO2 and hydroxyethyl-TPP.
What is the second step of the PDH reaction?
The hydroxyethyl carbanion on TPP of E1 reacts with the lipoamide on E2. The acetate formed by oxidation of the hydroxyethyl moiety is linked to one of the thiols of the reduced lipoamide as a thioester (~).
What is the third step of the PDH reaction?
The acetate is transferred from the thiol of lipoamide to the thiol of coenzyme A, yielding acetyl CoA.
How is the reduced lipoamide restored to its oxidized form?
It reacts using E3 catalyst with FAD+. This reaction gives the oxidized lipoamide and FADH2. That reacts with NAD+ to generate FAD+ and NADH +H
Why is acetyl CoA such a big deal?
It is a central compound in metabolism. The “high energy” thirster linkage makes it an excellent donor of the acetate moiety.
A few functions:
input to the Krebs Cycle, where the acetate moiety is further degraded to CO2.
donor of acetate for synthesis of fatty acids, ketone bodies, and cholesterol.
What inhibits the Enzymes in the PDH complex?
E1 is inhibited by ATP, high energy charge (makes sense?). E2 is inhibited by its product, acetyl CoA, and E3 is inhibited by its product, NADH. These are also signs of high energy.
What type of modification can inhibit E1 or pyruvate decarboxylase?
PDH kinase phosphorylates E1 and inhibits its activity.
- NOTE
This phosphorylation is NOT mediated by cAMP- PKA and is not affected by glucagon or epinephrine.
What dephosphorylates E1-P?
PDH phosphatase dephosphorylates E1-P and activates E1. This phosphatase is strikingly increased by insulin.
Insulin stimulates glucose to break down at which steps?
Note: insulin stimulates glucose to pyruvate at several steps (GK, PFK, PK) and here stimulates PDH to use the pyruvate.
What is special about the reduction of FADH2 on E3 by NAD in the PDH reaction?
Note: the reoxidation of FADH2 (on E3) by NAD is the only case in metabolism in which FADH2 reduces NAD+ to NADH. Usually, e.g., in the respiratory chain, NADH reduces FAD to FADH2.
What is the fate of the carbons from glucose in the subsequent catabolisis reactions?
The CO2 which comes off in the PDH reaction can be traced back to carbons 3 and 4 of the glucose (Fig. 6). Carbons 1 and 6 of the original glucose form the CH3 of acetyl CoA, while carbons 2 and 5 form the C=O of the acetyl group. These come off as CO2 in the TCA cycle.
What is the overall net reaction of glycolysis?
C6H12O6 + 2 NAD+ 2 ADP + 2 Pi→ 2CH3COCOOH (pyruvate or pyruvic acid)+ 2 ATP + 2 NADH. Irreversible, occurs in cytosol of all cells. Pyruvate will be further metabolized
Why is lactic acid break down so important to RBCs?
NADH is REOXIDIZED back to NAD+ which is necessary to turn G3P into 1,3bisphosthoglycerate in glycolysis. Mito can deoxidize NADH but RBCs don’t have mito.
What happens to lactate released into the blood?
Most lactate enters tissues where it is reoxidized back to pyruvate, which then undergoes the pyruvate dehydrogenase reaction for energy production, or can be converted to glucose in the liver during gluconeogenesis.
What is the TCA cycle?
Most lactate enters tissues where it is reoxidized back to pyruvate, which then undergoes the pyruvate dehydrogenase reaction for energy production, or can be converted to glucose in the liver during gluconeogenesis.
What can make AcetylCoA?
Glycolysis/PDH, fatty acid great down, amino acid breakdown.
Is the TCA cycle reversible?
No
Where does the TCA cycle occur?
Occurs in the mitochondrial matrix compartment of all cells (one enzyme, succinic dehydrogenase, is in the inner mito membrane).
What is the net rection of the TCA cycle?
AcetylCoA+3NAD+ +FAD+GDP+Pi +2H2O→2CO2 +CoA+3NADH+FADH2 + GTP + 2H+
Acetyl CoA + 3 NAD + FAD+ GDP+ Pi+ 2 H2O→
2 CO2 +3 NADH+FADH2+GTP+CoASH
What stimulates PK to increase?
F1,6 Bis P and insulin stimulated dephosph.
Net reaction of PDH?
Pyr + NAD+ CoA→ Acetyl CoA + CO2+ NADH
What are the fates of acetylcoA?
CO2 via TCA Cycle, ATP;
→ citrate which →fatty acids
→ ketones bodies,cholesterol, steroids
→ Acetylation RX e.g histones
Draw the TCA cycle (2).
.
What is alphakg dehydrogenase similar to?
PDH. E1, E2, E3 etc.
What does Succinic dehydrogenase produce?
FADH2 from FAD
What does Malate dehydrogenase produce?
NADH from NAD+
What occurs in the isocytrate dehydrogenase reaction?
Isocitrate + NAD+ —> Alphakg + Co2 +NADH + H+
Where is the second CO2 produced in the TCA cycle?
akg–> succinylCoA.
Why does the malate dehydrogenase reaction proceed towards OAA if it strongly favors the reverse?
While the equilibrium of the malate dehydrogenase reaction in vitro greatly favors OAA malate, in cells OAA is continually being removed by the citrate synthase reaction, keeping the concentration of OAA low and pulling the reaction in the direction toward OAA formation.
What is the energetic yield of the TCA cycle?
One FADH2 and 3 NADH + 3H+ are produced in the TCA cycle, which can yield 1.5 (FADH2) + 7.5 (3 NADH) or a total of 9 ATPs. One GTP is equivalent to one ATP. Hence, 1 acetyl CoA yields about 10 ATPs when oxidized in the TCA cycle.
What is the overall energetic yield of ATP from G6P–> 6 CO2 ?
about 32 ATPs/G6P:
Glucose–> 2 pyruvate + 2 NADH 2ATP
2 NADH (mito) 5ATP
2 Pyruvate–> 2AcetylCoA and 2NADH
2 NADH 5ATP
2 Acetyl CoA 20 ATP
What happens to the Carbons from Glucose?
Of the 6 carbons in G, C3,4 → CO2 in PDH RX
C1,2,5,6→CO2 in TCA cycle
What regulates the TCA Cycle?
a) Respiratory control regulates oxidation of NADH and FADH2 by the mito respiratory chain. This is discussed in more detail in the bioenergetics lectures.
b) Energy charge. ATP inhibits citrate synthase, isocitrate dehydrogenase, αKg dehydrogenase
c) Concentration of OAA. OAA is pulled out of the cycle during gluconeogenesis (lecture 10) and reduced to malate when NADH levels are elevated, e.g., after alcohol consumption.
What does ATP inhibit in the TCA cycle?
citrate synthase, isocitrate dehydrogenase, αKg dehydrogenase
List the Anaplerotic reactions of the TCA cycle? AKA what reactions can pull out players of the cycle?
Succinyl CoA -> heme synthesis
OAA-> gluconeogenesis, aa production (aspartate asparigine)
Citrate-> fatty acids
αKg-> aa. snth: glutamate, glutamine, proline, ornithine
What are ways to replenish OAA?
a) Pyruvate carboxylase (#1 in Fig. 6). This enzyme contains biotin, the vitamin B used to carry CO2 in enzymatic reactions. ATP is necessary for this carboxylation. Importantly, the enzyme is strikingly stimulated by acetyl CoA. Does this make sense?
As will be discussed in the nitrogen/amino acid lectures later on, many amino acids are catabolized to TCA cycle intermediates:
- glutamate αKg (#2 in figure below)
- valine or isoleucine or methionine or threonine succinyl CoA (#3 in figure below)
- phenylalanine or tyrosine fumarate (#4 in figure below) - aspartate OAA (#5 in figure below)
PEP carboxykinase (not shown in the figure). This enzyme converts the glycolytic intermediate PEP to OAA: CO2 +PEP+GDP ↔ OAA+GTP
Note that the high energy PEP can produce ATP in the pyruvate kinase reaction, and GTP (equivalent to ATP) in the PEPCK reaction. PEPCK is induced by glucagon under gluconeogenic conditions, and we will discuss this and pyruvate carboxylase in lecture 10.
How can citrate affect glycolysis?
Inhibits the glycolysis enzyme, phosphofructokinase. Therefore inhibiting isocitrate dehydrogenase would cause a buildup of citrate and negatively impact the rate of glycolysis.
Try to list the mitochondrial carriers.
Phosphate carrier - exchanges Pi with OH
- Dicarboxylate carrier - exchanges Pi or malate or succinate for each other
- Tricarboxylate carrier - exchanges citrate, isocitrate, malate or PEP for each other
- αKg carrier - exchanges αKG for malate
- Pyruvate carrier - exchanges pyruvate for OH or ketone bodies
- Glutamate carrier - exchanges glutamate for OH
- Aspartate carrier - exchanges aspartate for glutamate
- Adenine nucleotide carrier - exchanges ADP for ATP
Why is mito carrier 8 important?
Carrier 8 is obviously important in transporting ATP produced by oxidative phosphorylation out of the mito in exchange for cytosolic ADP, which was produced from ATP hydrolysis in the cytosol, e.g., glucose + ATP G6P + ADP. Carrier 8 functions in conjunction with Carrier 1 to bring ADP plus Pi back to the mito for eventual synthesis of ATP.
Remember carrier 1 exchanges Pi with OH
Name the specific mito carriers.
4 and 7
- αKg carrier - exchanges αKG for malate
- Aspartate carrier - exchanges aspartate for glutamate
What carriers can malate be exchanged on?
- Dicarboxylate carrier - exchanges Pi or malate or succinate for each other
- Tricarboxylate carrier - exchanges citrate, isocitrate, malate or PEP for each other
- αKg carrier - exchanges αKG for malate
What are 3 important functions of mito shuttles?
Shuttles are critical for transporting reducing equivalents from NADH or NADPH into or out of the mito; for providing acetyl CoA for fatty acid or cholesterol synthesis and for providing carbon intermediates for gluconeogenesis.
What are the two shuttles responsible for translocating the reducing equivalents of NADH into the mito?
The α-glycerophosphate (αGP) and the malate-aspartate (MA) shuttle.
Describe the α-glycerophosphate (αGP) shuttle.
DHAP, a product of glycolysis, reacts with NADH, also a product of glycolysis, to produce αGP + NAD+, as catalyzed by the cytosolic α GPDH. Thus, NAD+ is reoxidized. However, what do we do with the αGP, and how can we regenerate DHAP in order to continue the shuttle? Furthermore, if we keep pulling DHAP out of glycolysis, glycolysis will stop. αGP can react with the mitochondrial αGPDH, which is located on the outer surface of the mito inner membrane to regenerate DHAP. The mito αGPDH is linked to FAD, not NAD+ as was the cytosolic αGPDH, so FADH2 rather than NADH is produced in the mito.
αGP + αGPDH-FAD DHAP + αGPDH-FADH2 Respiratory Chain.
What is the first step of the malate dehydrogenase shuttle?
The NADH produced by glycolysis reacts with OAA in the cytosol to produce NAD+ and malate, as catalyzed by the cytosolic malate dehydrogenase (mdh; step 1). NAD+ has been regenerated, but unless OAA is regenerated (similar to the need to regenerate DHAP in the αGP shuttle), this shuttle will stop.
