Chemiosmosis & Oxidative Phosphorylation

what is oxidative phosphorylation

using electron transport + proton gradient to make ATP

energy yield (ATP)

1 NADH → 3 ATP

1 FADH₂ → 2 ATP

where do electrons go (electron flow)

NADH/FADH₂ → ETC → O₂

**reduced NADH/FADH₂ carry high energy e⁻**

transport of e- from NADH to O2

releases ~53 kcal to env

∆G = 153 kcal/mole

e- donor releases protons easily → e- acceptor accepts protons easily

why is O2 critical

final electron acceptor

forms H₂O

transport of e- from FADH2 to O2

releases ~36 kcal to env

∆G = -36 kcal/mole

∆G

defines changes in free energy

at pH 7 aka standard conditions

what is E and ∆E

E = reduction potential (volts)

∆E = changes in reduction potential at standard conditions, pH 7

∆G = -nF∆E°

n = # of e

F = faradays constant (23 kilocalories/volt/mole)

∆E° = E° of the electron acceptor minus E° of the electron donating pair

- relating gibbs free energy & electrical potential energy

- spontaneous reactions (-∆G) require a positive cell potential (∆E)

coupled reaction accomplished by _____

chemiosmosis

NADH + H⁺ → NAD⁺ + 2H⁺ + 2 e⁻ ... ∆G = -52.6 kcal/mole = FAVORABLE

ADP + Pi → ATP ... ∆G = +7.3 kcal/mole = NOT FAVORABLE

** highly spontaneous coupled with unfavorable**

chemiosmosis part 1

set up proton gradient

what drives ATP synthesis

proton gradient

what creates gradient

ETC pumps H⁺ out of mitochondrial matrix

**potential energy in electron carriers (NADH and FADH2s from TCA cycle) used to pump protons out**

what makes proton-motive force

concentration gradient:

- high [H+] outside

- low [H+] in matrix

electrical gradient (charges):

- +++++ charge outside

- ----- charge inside (fewer H+)

proton pumps lead to a _____ ______, more protons are outside the matrix thus the ____ differs

chemical gradient, pH

proton pumping leads to a ____ difference, more protons are _____ the matrix thus the relative charge differs

charge, outside

~-180 mV

proton pumping leads to an ______ gradient (proton motive force)

electrochemical

**proton motive force due to membrane potential & H+ gradient**

**can be used for work**

chemiosmosis part 2

use proton gradient to make ATP

how does ATP synthase work

- protons flow back in

- spins like a turbine

- produces ATP

protons flow ____ concentration and voltage gradient back into ____ ____, with energy captured by ATP synthase

down, mitochondrial matrix

**proton gradient is harnessed by ATP synthase to make ATP**

chemiosmosis harvest energy from ____ and ____

NADH, FADH2

how is electrochemical energy harnessed for ATP synthesis

by 4 membrane bound complexes

what are the complexes

- complex I (NADH dehydrogenase)

- complex III

- complex IV

- complex V (ATP synthase) → not part of ETC

detailed visual of complexes

I = NADH dehydrogenase complex

II = succinate dehydrogenase (TCA)

III = cytochrome b-c1 complex

IV = cytochrome oxidase complex

which complexes are part of ETC

- complex I (NADH dehydrogenase)

- complex III

- complex IV

_____ start with high energy, travel along the membrane complexes, fueling the pumps, end at low energy in H2O

electrons

which complexes pump protons

I, III, IV

complex II (Succinate Dehydrogenase) does not → receives e from FADH₂

how does electron transfer flow

donors to acceptors

electron donors (reducing agents)

have negative redox potential

readily release electrons (low e- affinity)

electron acceptors (oxidizing agents)

high redox potential

strongly retain electrons (high e- affinity)

ultimate electron acceptor

oxygen

electron carriers

iron sulfur proteins:

- NADH dehydrogenase in complex I

coenzyme Q:

- ubiquinone, a cholesterol derivative

cytochromes

- with iron containing hemes

iron- sulfur centers

complex of 2-4 iron atoms + sulfur atoms held in position by cysteine side chains

each can carry one e- at a time

transfer electrons efficiently between redox-active protein complexes (Complexes I, II, and III), powering ATP production

coenzyme Q (ubiquinone)

not a protein

shuttles electrons from Complex I and Complex II to Complex III

cytochrome

have a bound heme group

oxidized iron atom gains e- to become reduced iron

Fe⁺⁺⁺ → Fe⁺⁺

NADH dehydrogenase (complex I)

- accepts e from NADH

- passes to cofactor Flavin mononucleotide (FMN)

- to iron-sulfur centers to coenzyme Q

protons pumped

succinate dehydrogenase (complex II, TCA step 6)

passes e from FADH₂ to iron-sulfur centers → coenzyme Q

ZERO protons pumped

cytochrome b-c1 complex (complex III)

accepts e from coenzyme Q → passes to hemes → then to cytochrome c

protons PUMPED

cytochrom a + a3 complex (complex IV, cytochrome oxidase complex)

only complex that reacts with O₂ directly

final reduction to water

protons PUMPED

ATP synthase

turbine engine

1 full rotation uses 3-5 protons

1 rotation produces 3 ATPs

ATP synthase (complex V) is the ___ ___ step for oxidative phosphorylation

rate limiting

_____ produces, 2 NADPH + 2 ATP from glucose

glycolysis

yield from 1 glucose in cytosol

glycolysis + chemiosmosis

2 pyruvate + 2 ATP + 2 NADH → 6 ATP

net result from glycolysis = 8 ATP

yield from 1 glucose in mitochondria

pyruvate dehydrogenase + chemiosmosis

2 pyruvate → 2 acetyl CoA + 2 NADH → 6 ATP

yield from 1 glucose in mitochondria (TCA cycle + chemiosmosis)

2 acetyl CoA → 6 NADPH + 2 FADH₂ + 2 GTP

6 NADH → 18 ATP

2 FADH₂ → 4 ATP

2 GTP

net result in mitochondria: 28

net from 1 molecule of glucose

36 ATP + GTP

how much energy does your heart need

pumps 4-5 L of blood/min

- ~7200 liters/day

consumes 5-6 kg ATP/day

ATP from catabolism of:

- fatty acids

- glucose

- lactic acid

- ketone bodies

blocking the flow of _____ at any point shuts down the electrical circuit, thus halting _____

electrons, chemiosmosis

key inhibitors of ETC and proton pumping

rotenone → Complex I

antimycin A → Complex III

cyanide/CO → Complex IV

oligomycin → ATP synthase

inhibitors of oxidative phosphorylation and ATP export

ATP/ADP translocase

atractyloside (blocks exiting)

what do uncouplers do

poke holes in inner membrane, disperse the proton gradient & stop ATP synthesis

- destroy proton gradient

- stop ATP produciton

- generate heat

ex. DNP (synthetic ionophore: 2-4 dinitrophenol)

electron transport continues with uncoupling, but energy released as _____ (not ATP)

heat

prevent burning up cell...

brown fat and skeletal muscle posses a special protein known as ____ _____ used to generate heat if needed

uncoupling protein (UCP)

**regulated**

uncoupling proteins UCP

create a "proton leak", allowing protons to reenter the mitochondrial matrix without capturing any energy as ATP & yielding heat

generates heat (thermogenesis)

what happens if ETC stops

NADH accumualtes

no NAD+ → glycolysis stops

no ATP → cell death

summary

- PDH = bridge between glycolysis & TCA

- TCA = makes NADH/FADH₂

- ETC = uses NADH to make ATP

- oxygen = final electron acceptor

- inner membrane = most important site

- high ATP = turns everything OFF

- low ATP = turns everything ON

what is the pH outside relative to matrix

lower

what is the charge outside relative to the matrix

more positive

in a cell treated with oligomycin which inhibits complex V, what is the ATP yield from 1 glucose molecule

2

which of the following cellular conditions causes phosphorylation and thus inactivation of pyruvate dehydrogenase

high ATP/ADP

high acetyl CoA/Co

high NADH/NAD+

relative to FADH2, NADH pumps approximately ____ protons

50% more

An enzyme that catalyzes a required reaction in the biosynthesis of coenzyme Q (ubiquinone) was recently discovered. A deficiency in this enzyme would most likely result in increased concentration of which of the following in blood?

lactic acid

**anaerobic mechanisms bc not enough ATP**

which of these vitamins is an essential cofactor for both the pyruvate dehydrogenase and a-ketoglutarate dehydrogenase complexes

thiamine

we extract energy from macronutrients primarily from which of the following types of chemical reactions

oxidation of carbon

which of the following catalzyes an irreversible, rate limiting reaction in the TCA/citric acid cycle

isocitrate dehydrogenase

the components of the eukaryotic ETC are located in

inner membrane of mitochondria

many compounds of the TCA cycle are siphoned off as building blocks or used in diverse metabolic reactions.

what type of reactions prevents the TCA cycle from halting due to loss of substrates

anaplerotic

exercise increases flux through the TCA cycle most directly through which of the following mechanisms

increased NAD+/NADH ratios

how many glucose molecules were catabolized to produce 2 ATP, 2 GTP, 2 FADH2, 10 NADH, by the end of the TCA cycle

1

in its role in catabolic metabolism, the primary function of the TCA cycle is to

transfer electrons from carbon to NAD+ and FAD

in theory how much ATP could one NADH make

NADH + H+ → NAD+ + 2H+ + 2 electrons

∆G = -52.6 kcal/mole

7

which of the following has the strongest tendency to gain e-

oxygen

the oxidation of sugar molecuels by the cell takes place according to the general reaction 6 O2 + C6H12O6 (glucose) converted to 6 CO2 + 6 H2O + energy.

Which of the following processes yields the greatest proportion of ATP from this reaction

chemiosmosis

the following represents a reaction in the TCA/citric acid cycle

HS-CoA + NAD⁺ + A → NADH + H⁺ + CO₂ + B

what are compounds A and B

alpha-ketoglutarate AND succinyl-CoA