Roesler Section Flashcards
Where does most of the ATP come from in cellular respiration?
Not in glycolysis, will be in oxidative reactions instead (Citric acid cycle and electron transport chain)
3 stages of cellular respiration (not including glycolysis)
Catabolism of pyruvate to acetyl CoA
Citric acid cycle
Oxidative phosphorylation
Give an overview of the Pyruvate dehydrogenase complex
3 enzymes, that catalyze 5 biochemical reactions
requires 5 cofactors, oxidative decarboxylation
Irriversible reaction
present in mitochondria
What are the three enzymes in the PDH?
E1 - pyruvate dehydrogenase
E2- Dihydrolipoyl Transacetylase
E3 - Dihydrolipoyl Dehydrogenase
What is the MPC?
Mitochondrial Pyruvate transporter
Moves pyruvate from cytoplasm after glycolysis into the inner mitochondrial membrane
What do cofactors do? 5 points
Helper molecules
non protein
bound to an enzyme (either tightly or loosely)
required for catalysis
does not catalyze reactions
What are the 5 PDH cofactors?
- Thiamine Pyrophosphate (TPP)
- Lipolic Acid
- Coenzyme A
- FAD (flavin adenine dinucleotide) - electron carrier
5.NAD+ (nicotinamide adenine dinucleotide) - electron carrier
*Lipolic acid is not derived from B vitamins
How many high energy electrons are gained in the citric acid cycle?
8
What does pyruvate release when it is converted into actetyl CoA?
Releases CO2
Where does Acetyl CoA lead into after being made from pyruvate?
Citric acid cycle
What is TPP derived from, where is it bound, what does it accept and what is it known as?
Derived from vitamin B1 (thiamine)
Bound to E1 of PDH
Accepts 2 carbon backbone of acetyl CoA after decarboxylation ( hydroxyethyl TPP intermediate)
Known as a transient carbon carrier
Where does glycolysis occur?
Cytosol
Where does the citric acid cycle occur?
Mitochondrial matrix
Why does the body use protein complexes instead of separate enzymes?
makes the overall process faster because if the enzymes are already associated the product does not need lots of time to find the next enzyme
From slides - local concentration of substrates around enzymes is kept high - rate of reaction is not limited by diffusion of substrates to each subunit of PDH
What is lipolic acid a cofactor of? What two special groups does it have and what can happen to them? What is its function? What does it accept?
Lipolic acid is a cofactor of the E2 subunit
Has 2 thiol groups which can be oxidized, reduced or acetylated
It carries electrons and acyl (carbon) groups
It accepts hydroxyethyl intermediates from TPP as an acetyl groups
What are NADH and FADH2? What is the main difference between them?
They are electron carriers and can exist in oxidized or reduced states
They each carry 2 electrons
NADH is a mobile carrier while FADH2 is a protein bound carrier
Give a 5 point summary of the PDH reaction
Pyruvate is converted to acetyl coa by PDH complex
PDH - 3 enxymes - 5 reactions and 5 cofactors (coenzymes / prosthetic groups
E1 - oxidative decarboxylation of pyruvate and transfer of 2 carbon unit (hydroxyethyl intermediate to E2)
E2 - transfer of acetyl group to CoA to synthesize Acetyl CoA
E3 - regenerates oxidizes lipoyl group of E2 and transfers protons and electrons first to FAD and then to NAD+ to complete the reaction cycle
How is the PDH regulated?
Allosterically- by NADH, Acetyl CoA and ATP as inhibitors
- Pyruvate and ADP as allosteric activators
Post-translational modification- by Phosphorylation/ dephosphorylation - above inhibitors activate PDH Kinase which inhibits PDH
- Above activators activate PDH phosphatase which activates the PDH
- ATP adds phosphate group to form ADP
Why is acetyl CoA so important for metabolic activity?
Acts as a metabolic shipping and receiving department for all classes of biomolecules and is a major source of metabolic energy.
Why does it make sense for a high-energy molecule like ATP to inhibit glycolytic reactions?
Because the high energy molecules (ATP) are also products meaning the reaction does not need to produce more ATP because the amount is already sufficient
What 3 amino acids can be phosphorylated?
Serine, threonine and tryosine
Where does the TCA cycle occur?
Mitochondrial matrix
What substrate formed in the first reaction of the TCA cycle is regenerated in the last step of the TCA cycle?
Oxaloacetate 4 carbon molecule
What is the first step of the TCA cycle that uses Acetyl CoA from the pyruvate dehydrogenase reactions?
C2 molecule of Acetyl CoA joins with Oxaloacetate (C4) to become Citrate (C6) via the enzyme citrate synthase
How many ATP are produced per cycle of the TCA cycle?
only 1, but many high transfer potential electrons to make wayyy more ATP in the electron transport chain
What are the three steps in forming Citrate using citrate synthase?
- oxaloacetate binds to citrate synthase
- induces a conformational change in citrate synthase
- generates the acetyl CoA binding site
- Acetyl CoA binds (duh)
How many carbon dioxide are lost in one cycle of the TCA cycle?
2 carbon dioxide are lost, turning Citrate from a 6C molecule to a 5C molecule to a 4C molecule which returns back to the 4C molecule that binds to the C2 Acetyl CoA to create the C6 used in the first place
How many high energy electron carriers are produced in one cycle of the TCA cycle?
3 NADH and 1 FADH2
What does citrate become and what causes the reaction in the TCA cycle?
Citrate is isomerized into Isocitrate via the enzyme Aconitase
Aconitase catalyzes the formation of isocitrate from citrate (isomerization), reaction favours isocitrate not citrate formation
What does isocitrate become and what causes the reaction in the TCA cycle?
Isocitrate is oxidized and decarboxylated to A-Ketoglutarate via Isocitrate dehydrogenase which catalyzes the oxidative decarboxylation of isocitrate forming a-ketoglutarate and capturing high energy electrons as NADH
What does A-ketoglutarate become and what causes the reaction in the TCA cycle?
A-ketoglutarate is oxidated and decarboxylated to succinyl CoA via the A-Ketoglutarate dehydrogenase complex which catalyzes the synthesis of succinyl CoA from a-ketoglutarate, generating NADH
- this enzyme and reaction are structurally and mechanistically similar to the pyruvate dehydrogenase complex
What is succinyl CoA become and what causes the reaction in the TCA cycle?
Succinyl CoA becomes Succinate via Succinyl CoA synthetase.
Succinyl CoA synthetase catalyzes the cleavage of thioester linkage and forms ATP
What does succinyl CoA synthetase do?
- Cleaves the high energy thioester bond of succinyl CoA turning it to Succinate
- energy released drives synthesis of the high energy gtp pr ATP - it is an energy conserving reaction
- substrate level phosphorylation – direct transfer of phosphate to ADP or GDP
What step is the main point in regulation for the TCA cycle?
The step where Succinyl CoA is formed by the oxidative decarboxylation of A-Ketoglutarate - when the A-Ketoglutarate complex catalyzes the synthesis of succinyl CoA from A-ketoglutarate, generating NADH
What does succinate become and what causes the reaction?
Succinate is oxidized to become fumarate via succinate dehydrogenase by introducing a double bond in trans configuration, generates FADH2
What is special abount succinate dehydrogenase?
Only membrane bound enzyme of the TCA cycle
What does fumarate become and what causes the reaction?
Fumarate is hydrated to become malate, catalyzed by fumarase
What does malate become and what causes the reaction?
Malate is oxidized by malate dehydrogenase to become oxaloacetate, regenerating the starting material of the cycle
Why can the reaction between 2glyceraldehyde3-phosphate to 2 1,3-bisphosphoglycerate have either 3 or 5 atp produced?
Depends on the type of NADH shuttle used to get it from the cytoplasm to the mitochondria
-Glycerol 3-phosphate shuttle generates 3 atp
-Malate/aspartate shuttle generates 5 atp
How is the TCA cycle regulated?, which enzymes are regulated?
There are two TCA cycle enzymes that are regulated and both are by allosteric mechanisms
- Isocitrate dehydrogenase - stimulated by ADP and inhibited by NADH and ATP
- aKetoglutarate dehydrogenase (catalyzes rate limiting step) - no activators - inhibited by products of its reaction- succiny CoA and NADH (and ATP)
What happens to the TCA cycle when at rest?
ATP and NADH are higher, inhibiting Isocitrate dehydrogenase, don’t need to make more ATP if enough is present
When fats are broken down what is an important product of that?
Acetyl CoA
Can we make glucose from fats?
No, we can make fats from glucose but not the other way around, not enough carbons.
What are replenishing reactions for intermediates of the TCA cycle called?
Reactions that replenish and provide intermediates for the pathway are called ANAPLEROTIC reactions
Why is fluoroacetate poisonous?
Because fluoroacetate is converted to fluoroacetyl CoA and then to fluorocitrate which can inhibit aconitase and therefore the formation of isocitrate, blocking the TCA cycle - cannot continue under that condition
What is an analogous cycle in plants and bacteria?
The glyoxylate cycle
How is the glyoxylate cycle similar to the citric acid cycle?
Main difference is it bypasses the two decarboxylation steps, allowing the synthesis of carbs from fats
Where does the glyoxylate cycle occur in plants?
In the glyoxysome of plants
What is the main product of the glyoxylate cycle?
Succinate (which can be converted into oxaloacetate and then glucose)
Why can mammals not synthesize glucose from acetyl CoA?
Because the input is 2 carbons in Acetyl CoA and the two carbons are lost as CO2
Why do plants use the glyoxylate cycle?
For plants when germinating as seeds, the seeds have fat stores and it converts the fats into glucose for energy before the plant can begin photosynthesizing
Give a general overview of oxidative phosphorylation
Metabolic pathway that uses energy released by the oxidation of nutrients to produce ATP
- ATP is synthesized as electrons pass from electron donor to electron acceptor
- Energy that is released during electron transport drives ATP synthesis
- the final electron acceptor is O2
- occurs in the mitochondria inner membrane - electron transport chain
What are the 5 main parts of the electron transport chain?
5 main protein complexes + cytochrome C
- 4 complexes for electron transport - bucket brigade
- 1 complex for ATP synthesis
What is chemiosmotic theory?
Energy is released as electrons flow thru transport chain
- energy is used to transport protons across the inner membrane
- this generates potential energy - pH gradient across membrane
- protons flow back down across the membrane and down the gradient through enzyme ATP synthase
- ATP synthase uses energy to ATP
Which complex in the electron transport chain does not act as a proton pump?
Complex II - succinate dehydrogenase
Name the 5 ETC complexes
I - NADH dehydrogenase
II - succinate dehydrogenase
III - Ubiquinone:cytochrome C oxidoreductase
- Cytochrome C
IV - Cytochrome oxidase
What part of the ETC is soluble?
Cytochrome C
What are the 5 electron carriers used in the ETC?
- NADH and FADH2 from TCA
- Coenzyme Q
- Cytochromes A,B,C
- Iron-sulfur center-containing proteins
- Copper-sulfur center-containing proteins
What is Coenzyme Q?
Lipid soluble electron carrier
- moves in lipid bilayer - inner mito membrane
- moves e- between less mobile e- carriers in mem
moves 2e-
What are cytochromes?
Proteins that function as electron carriers
- cytochrome C is soluble - carries 1e-
- outer surface of mito membrane
- has heme group that has a Fe bound by 4 N, iron is what accepts and donates the electrons
What are proteins with iron sulfur centers?
2 iron joined by 2 sulfur - linked to protein by CYS
- can carry electrons
- proteins can have multiple centers
What is complex I in the ETC?
NADH dehydrogenase
- 1st reaction of ETC
- several iron-sulfur centers
-L shaped, part in matrix part in membrane
- 2 electron reduction by coenzyme Q
- 4 protons moved from matrix to intermembrane space
What is complex II in the ETC?
Succinate dehydrogenase
- 2nd point of entry into ETC
- takes in FADH2
- oxidizes succinate to fumarate - transfers electrons to CoQ- QH2
- electrons go from succinate to FAD to iron sulfur centers to CoQ to QH2
- NO H are moved across membrane - does not contribute to formation of H gradient
What is complex III in the ETC?
Cytochrome C oxioreductase
- in inner mito mem
transfers electrons from QH2 - cytochrome C
- uuses heme and F/S centers
transports 4 protons from matrix to intermembrane space
- contributes to proton gradient
-
What is complex IV in the ETC?
Cytochrome c oxidase
- final complex
- inner mito mem
- carries electrons from cyt c to O2 which is reduced to water
- pumps 2 H across per electron pair
O2 is final e- acceptor (requires 4 electrons)
Where do the electrons from NADH and FADH2 converge in the ETC?
complex q
What is proton motive force?
the electrochemical energy that drives ATP synthesis as protons flow back down the gradient into the matrix which provides the energy for atp synthase to work
What is ATP synthase?
catalyzes formation of ATP from ADP and Pi
- protons re enter the matrix by passing thru atp synthase
- creates enough energy to couple a phosphate group to ADP forming ATP
What are the F0 and F1 regions of ATP synthase?
F0 - pore that spans membrane - protns flow thru F0 pore
F1 - in the matrix of mitochondria - spherical head which phosphorylation of ADP occurs
How many A and B subunits are there in ATP synthase? How are they arranged?
3 pairs of alternating A and B subunits
How are cristae formed in the mitochondria?
due to the dimerization of ATP synthase
- cristae help organize proton pumps in a smaller space to increase concentration of protons near ATP synthase
What are the three conformations of the catalytic B subunits of the F1 complex in ATP synthase?
L - loose form - B subunit traps ADP and Pi
T - tight form - ATP is synthesized from ADP and Pi
O - open form - B subunit has low affinity for ATP - releasing it from B subunit so another ADP and Pi can bind
Explain how the three different B subunits interact with each other?
The B subunits are always in different conformations and rotates between the three.
- When 3 protons pass through the pore, Y rotates which changes conformation of B
- Open conformation changes to loose to bind an ADP and Pi
- loose confirmation changes to tight - condenses and makes ATP from ADP and Pi
-Tight conformation changes to Open - releasing ATP so the cycle starts again
1
How many protons are needed to turn the subunit 120 degrees?
1 ATP is produced for every 3 protons that re enter mitochondria thru ATP synthase (takes 3 protons to crank that shit)
How many protons does it take to make an ATP?
4 (why? - 3 to turn, 1 to move Pi into matrix)
How is ATP moved into the cytosol after being produced in the mitochondria?
Via the ATP-ADP translocase
How does NADH that is generated in the cytosol make it into the mitochondria?
2 diff shuttles that transport NADH from cytoplasm to matrix
- Malate-aspartate shuttle (2.5ATP/NADH) - liver / heart
-glycerol-3-phosphate shuttle (1.5 ATP/ NADH) - muscle - mimics complex II ***
How do different poisons affect oxidative phosphorylation?
Rotenone - prevents e- transfer from complex I to CoQ
Cyanide / Carbon Monoxide - inhibit cytochrome C oxidase (complex IV) - binds to iron in the heme group preventing transfer of electrons to O2
What is brown fat?
Brown adipose tissue that is present in hibernating rodents and newborn humans - has many mitochondria that dissipate energy instead of storing it
- via uncoupling the electron transport chain from ATP synthesis heat is generated - bypasses ATP synthase energy is turned into heat
What is the advantage of using a protein complex? (PDH)
Substrate channeling - local concentrations of substrates around enzymes is kept high
