"mitochondrial Etc And Oxidative Phosphorylation"

Front 1

What are oxidation and reduction reactions? Why are these important in the ETC?

Back 1

Oxidation-->releases free energy--LOSS of electrons
Reduction-->gains free energy--GAIN of electrons

In digestion, nutrients are broken down into smaller components:
Fats-->FA's
Carbs-->simple sugars
Protein-->a.acids

--These molecules enter glycolysis and TCA cycle, releasing tons of energy along the way.
--NAD and FAD get electrons from enzymes in Kreb's (reduction)-->become NADH/FADH2 ("energy carriers")-->dump electrons into ETC (oxidation)-->converted back into NAD/FAD-->can be used again

Front 2

What does the Nernst equation help calculate?

Back 2

Remember--reduction potential is the tendency to acquire electrons.
--The nernst equation characterizes the relationship b/w the standard reduction potential of a redox couple under standard conditions.

E=Eo + RTln (oxidant)/(reactant)

-->This predict electron transfer for redox pairs liked together.

Front 3

Define reduction potential and how it can determine direction of electron transport

Back 3

--Reducation/redox potential (delta-E)-->the tendancy of a molecule to acquire electrons (be reduced).

--The more positive the reduction potential, the greater the affinity to grab electrons (be reduced).
--The more negative the reduction potential, the greater the affininy to give up electrons (be oxidized)

--STANDARD reduction potential (delta-E*) is the reduction potential under standard conditions.
--It can be used to calculate free energy change (delta-G)--which is proportional to the difference in redox potentials of both redox pairs.

delta-G*=-n(F)(delta-E*)

--The more positive the delta-E (the more the molecule wants to be reduced), the more negative the delta-G-->positive delta-E indicates an energetically favorable process.
--ETC has a delta-G* of around -52 kcal/mol. This is super-exergonic, which is what you want. ATP hydrolysis has a delta-G* of around 7.3 kcal/mol--not so exergonic. Shows you that ETC is an energetically favorable process.

Front 4

What is basic mitochondrial structure? Where does ETC take plate?

Back 4

Mitochondria-->Outer (permeable, smooth) and inner (impermeable, folds called "cristae") membranes

Have intermembrane space and matrix

ETC proteins are embedded in the inner membrane-->ATP synthase projects into matrix where ATP is made.

Front 5

What is purpose of ETC? What is basic mechanism?

Back 5

ETC proteins transfer energy by taking electrons from NADH/FADH2 and passing them down the line.

Six entities to the ETC:
1) Complex I (NADH dehydrogenase)
2) Complex II (succinate dehydrogenase)
3) Complex III
4) Complex IV (Cytochrome oxidase)
5) Coenzyme Q
6) Cytochrome C
Mechanism...
1) NADH binds FMN on complex I (NADH Dehydrogenase)-->oxidized to NAD-->2 electrons passed to iron-sulfer-->4 hydrogens pumped into intermembrane space.
2) Electrons are passed from complex I-->coenzyme Q
3) Electrons are passed from coenzyme Q-->complex III-->4 hydrogens pumped into intermembrane space
4) Electrons passed from complex III-->cytochrome c-->complex IV (cytochrome oxidase)-->2 hydrogens pumped into intermembrane space-->electrons bind O2 to form water in matrix. This step REMOVES electrons from system (by combining them w/O2-->water), so the ETC can continue. This is why you need O2 for ETC to work.

What about complex II?
--Not a part of the NADH pathway--is actually an enzyme in the TCA cycle.
--NO proton pumping.
--Funnels electrons into ETC by removing electrons from succinate and transferring them (via FAD) to coenzyme q.

NADH
complex I-->coenzyme q-->complex III-->cytochrome c-->complex IV-->h20!

Front 6

Describe the chemiosmotic model and its importance in maintaining the ETC

Back 6

--ETC pumps hydrogens into intermembrane space at each complex (except II).
--This process creates an electrical gradient (more positive charge outside membrane) and pH gradient (lower pH-more H+s outside membrane).
--Energy generated by this proton gradient can drive ATP synthesis.
--ATP synthetase makes ATP, using energy of the proton gradient. After protons have been pumped out of matrix, they can re-enter matrix thru ATP synthetase-->forming ATP AND dissipating their own pH and electrical gradients. (from lippencott's)

--ETC won't work if protons can't be pumped into intermembrane space--protons can't be pumped if chemiosmotic gradient is bad--so chemiosmotic gradient can LIMIT electron transport ("respiratory control")

Front 7

What is respiratory control?

Back 7

--The limitation placed on ETC by chemiosmotic gradient.
--If gradient is destroyed, respiratory control is abolished, and ETC can run freely.
ETC-->DRIVEN by free energy from energy carriers--obtained from substrates
ETC-->RESTRICTED by chemiosmotic gradient (respiratory control)--electron transport can only go as fast as energy is lost from the gradient.

ANALOGY from class..

Blowing up a balloon until it is full of air--cannot add anymore air, so you are just maintaining pressure.
--If someone pokes a hole in the balloon, you have to work harder to maintain the pressure from before.
--If you plug one of the holes, you don't have to work as hard.

ETC applies "pressure", holding gradient at constant level.
"you blowing up the balloon"-ETC
air flow into balloon--electron flow in ETC

Front 8

What is role of ATP synthase in ETC?

Back 8

--Exploits the chemiosmotic gradient to make ATP.
--ATP synthase structures are embeeded in the membrane.
--When ADP and phosphate are available, they bind catalytic sites on ATP synthase--causes channel to open and PROTONS come back into matrix (DOWN their conc. gradient)--energy released helps make ATP!
--So, it decreases the gradient, so ETC can continue.

Front 9

Why is cytochrome oxidase (complex IV) so important?

Back 9

It reduces oxygen to water, using the electrons.
--It removes electrons from the system, so ETC can keep running.
--That is why oxygen is so important--it "drains" electrons from the system, so the carriers don't become reduced and ETC can keep chugging along.

Front 10

What is the concentration gradient in "healthy" mitochondria?

Back 10

In normal, healthy mitochondria, the chemiosmotic gradient is maintained.

That's why uncouplers are so bad for the cell--they destroy the concentration gradient, so your ETC can never relax--there are no controls.

Front 11

What is respiratory control index (RCI)?

Back 11

OVERALL--ETC stops when ADP is used up.
--You can calculate the ratio of--
O2 consumption w/ADP present/O2 consumption w/ADP absent.
--It indicates how tightly the ETC is regulated by phosphorylation--in english, it tells you how much the ETC is affected by ADP.
--Larger RCI--better regulated (coupled)
--Smaller RCI--less regulated (uncoupled)
--OVERALL--if an ETC is uncoupled, this means that something has made it possible for H's to sneak back into matrix--this destroys the electrochemical gradient. This means less ATP is made. So..a low RCI is not controlled by phosphorylation, because it can't make any ATP--it can never get the gradient it needs to generate energy.

Front 12

What is the P:O ratio?

Back 12

Ratio of:
ADP added: O2 consumed

-Tells you number of ATP molecules made/O2 (ie: number of ATP molecules made/electron pair)
**NADH--P:O=2.5. So, 2.5 ATP molecules/electron pair. NADH (pumps 10 protons across membrane--3 protons--1 ATP (+1 to move molecule), 10/4=so 2.5 ATP made!)--P:O=2.5 ATP
**Succinate=1.5 ATP/electron pair. (pumps 6 protons, b/c starts at complex 2--3 protons--1 ATP (+1 to move molecule--6/4, so 1.5 ATP made! )--P:O=1.5 ATP

Front 13

What are the types of mitochondrial poisions?

Back 13

1) ETC inhibitors
2) Uncoupling agents
3) FoF1-ATPase Inhibitors

Front 14

What are ETC inhibitors?

Back 14

--They bind to ETC and PREVENT ELECTRON TRANSFER down the chain.
--They each act specifically at one complex.
--Causes reduction of upstream molecules.
--Bypassing block can restore ETC activity.

Front 15

What are the ETC inhibitors? Where do they work?

Back 15

1) Rotenone--complex I (non-competitive inhibitor)
2) Malonate--complex II--is really an "enzyme inhibitor"--no complex II-->feedback inhibition to TCA-->stops TCA, stops ETC
3) Antimycin/myxothiazole--complex III
4) Cyanide/Azide--complex IV (reversible inhibitor)

Really Mean Actors May Chide Actresses

Front 16

What are uncoupling agents?

Back 16

--ETC no longer regulated by chemiosmotic gradient.
--ETC is uninibited due to complete dissipation of chemiosmotic gradient-no respiratory control--keeps the ETC on constantly and working ten times harder.

Front 17

What are the uncoupling agents?

Back 17

1) DNP--protein ionophore--binds proteins on one side and "sneaks" them in the other side.
2) FCCP--protein ionophore--acts in a similar way.

Front 18

What are FoF1 ATPase inhibitors? What is the main one we learned?

Back 18

--EX: Oligomycin.
--Binds ATP synthase-->blocks proton channel.
--Prevents electron transport by blocking movement of protons through ATP synthase.
--When uncoupler is added, proteins leak into the matrix w/out ATP synthase and ETC will continue tirelessly, but can never generate any kind of substantial gradient.

Front 19

What is the glycerophosphate shuttle?

Back 19

--Way to get NADH across inner mitochondrial membrane from glycolysis.

1) shuttle transfers 2 electrons from NADH in cysotol-->reduces DHAP-->G-3-P (a-glycerophosphate dehydrogenase)
2) G-3- P crosses mitochondrial membrane.
3) G-3-P donates electrons to flavoprotein dehydrogenase (mitochondrial G-3-P dehydrogenase)-->produces FADH2.

NADH-->FADH2 (only get 1.5 molecules of ATP generated)

Front 20

What is malate-aspartate shutte?

Back 20

--Produces mitochondrial NADH from cytosolic NADH.

1) NADH reduces OAA-->malate
2) malate enters mitochondria using malate/a-ketoglutarate transporter
3) malate-->OAA + NADH (using malate dehydrogenase)
4) NADH used in ETC

NADH-->NADH (2.5 ATPs produced)

Front 21

How do ions get across inner mitochondrial membrane?

Back 21

ETC need lots of anions (ADP, pyruvate)
--TRANSLOCASES transport these proteins--balance the anionic charge during transport. Can be antiports or symports.

Antiports-->2 substrates in opposite directions--exchanges an anion for an anion.
Symport-->2 substrates in same direction-->moves proton+anion at once.

Front 22

What are some physiological or pathological uncouplers?

Back 22

1) Exercise/ increased body heat--need more energy-->more ETC-->extra energy released as heat.
2) Hyperthyroidism--make less ATP from fuels--more heat.
3) Brown adipose tissue (in babies)--uses THERMOGENIN or UCP-1 to uncouple--generate heat to keep babies warm.
4) Salicylate--lots of aspirin ingestion--can uncouple ETC.

Front 23

What is significance of Riboflavin deficiency?

Back 23

--REDOX reactions essential to ETC use two flavins as electron acceptors (FMN-complex I and FAD-complex II)
--If these are deficient, ETC decreases and you get MEGAMITOCHONDRIA in tissues.

Front 24

What is significance of anthracycline toxicity?

Back 24

-Anthracyclines are effective cancer tx.
--Too much can impair mitochondrial fxn.

Front 25

What is significance of apoptosis?

Back 25

--Mitochondria are important in apoptosis.
--triggered by internal/external stimuli
--when mitochondria receive these stimuli, the outer membrane releases cytochrome c-->leaked cytochrome c activates a protease cascade-->leading to its demise.
--Overall, have important role in apoptosis.

Front 26

What is significance of ischemia/hypoxia?

Back 26

--Tissues w/high ATP demands hve more mitochondria
--Mitochondria can also have densly packed cristae that have a ton of ETC proteins.
--These tissues are most sensitive to ischemia/hypoxia-->decreased O2-->decreased ETC.

Front 27

What are OXPHOS diseases?

Back 27

Diseases involving oxidative phosphorylation (ETC).
--Mitochondrial myopathies--caused by damage to mitochondria.

Examples:
MERRF-->spontaneous muscle jerking--HUGE mitochondria, red fibbers, dementia, impaired energy metabolism. Due to point mutation in mitochondrial mRNA from mom.

LHON--also maternal inheritance Affects CNS (optic nerve--vision loss), single base change coding for subunits of complex I or cytochrome b decreases mitochondrial fxn.

Front 28

What are some BIG ETC concepts (7)

Back 28

--ETC can't transport electrons if protons can't be pumped into intermembrane space.
--ETC driven by proximity of carriers (each subsequent carrier is more electronegative--wants the electrons MORE)--not driven by purpose to maintain gradient.
--O2 cnsumption results from ETC and does not require ATP.
--As long as substrate is present, chemiosmotic gradient is maintained by mitochondria (in healthy cells)
--ATP synthase is NOT part of ETC.
--Protons entering matrix thru ATP synthase do not reduce O2
--Activation of ATP synthase increases the rate of energy removal from gradient--ETC tries to maintain this gradient.