Chapt. 21 oxidative phosphorylation
Ch. 21 oxidative phosphorylation
Student Learning Outcomes:
• Explain process of generation
of ATP by oxidative phosphorylation:
• NADH + FAD(2H) donate e- to O2 -> H2O
• ATP synthase makes ATP (~3/NADH, ~2/FAD(2H)
• Describe chemiosmotic model, H+ gradient
• Describe complications of deficiency of ETC –
 anemia, cyanide, OXPHOS diseases
• Describe transport through mitochondrial
membranes
I. Oxidative phosphorylation summary
Oxidative phosphorylation overview:
• Multisubunit complexes I, coQ, III, IV pass e- to O2
• H+ are pumped out -> electrochemical gradient
• H+ back in through ATP synthase makes ATP
Fig. 1
Proton Motive Force
Proton motive force:
• Electrochemical potential gradient
• Membrane is impermeable to H+
• pH gradient ~ 0.75 pH units
Fig. 2
ATP synthase
ATP synthase (F0F1 ATPase):
• F0 inner membrane (12 C)
• F1 matrix has stalk, headpiece
• H+ go through a-c channel
• 12 protons/turn -> 3 ATP
• Binding change mechanism:
•
Turning releases ATP
Figs. 3,4
B. Components of Electron Transport Chain
Components of Electron Transport chain:
• Series of transfers of e- down energy gradient
• Series of oxidation reduction reactions
• e- finally to O2 -> H2O
• H+ pushed across membrane
Fig. 5
Components of Electron Transport Chain
NADH dehydrogenase: 42 subunits,
•
•
•
•
FMN binding proteins
Fe-S binding proteins (transfer single e-)
binding site for CoQ
pass e- to CoQ; transfers 4 H+
Fig. 5,6
Components of Electron Transport Chain
Complex II: succinate dehydrogenase (from TCA)
• FAD bound e- from TCA,
• Other FAD from other paths
• Not sufficient energy to transfer H+ when pass e- to CoQ
Fig. 5
Coenzyme Q
Coenzyme Q is not protein bound. 50-C chain
inserts in membrane, diffuses in lipid layer
• Also called ubiquionone (ubiquitous in species)
• Transfer of single e- makes it site for generation of toxic
oxygen free radicals in body
Fig. 7
Cytochromes have heme groups
Cytochromes have heme groups:
• Proteins with hemes
• Fe3+ -> Fe2+ as gain e• Transfer e- to lower potential
Figs. 5,8; Heme A is in Cyt a, Cyt a3
C. Pumping of protons not well understood
Cytochrome C oxidase: Cyt a, Cyt a3, O2 binding:
• Receives e- from Cyt c (takes 4 to make 2 H2O)
• Transfers to O2;
Pumping of H+ not well understood; must couple
to e- transport and ATP; otherwise backup
Fig. 5
D. Energy yield
Energy yield from oxidation by O2:
NADH: Dg0’ ~ -53 kcal; FAD(2H) ~ -41 kcal
Each NADH 2e- -> ~ 10 H+ pumped;
Takes ~ 4 H+/ATP -> 2.5 ATP/ NADH; 1.5/ FAD(2H)
or if ~ 3H+/ATP -> 3 ATP/ NADH; 2/ FAD(2H)
Fig. 5
E. Inhibition of chain, sequential transfer
Once start ETC, must complete transfer of e• In absence of O2, backup since carriers full of e• Inhibitors like cyanide (binds Cyt c oxidase)
mimics anoxia: prevents proton pumping
• Cyanide binds Fe3+ in heme of Cyt a a3
• CN in soil, air, foods (almonds, apricots)
Fig. 5
OXPHOS diseases from mutated Mitochondrial DNA
OXPHOS diseases from mutated mitochondrial DNA
Human mt DNA is 16.569 kb:
13 subunits of ETC:
7 of 42 of Complex I
1 of 11 Complex III
2 of ATPsynthase
22 tRNA, 2 rRNA
Table 21.1 examples OXPHOS diseases from mt DNA
Point mutations in tRNA or ribosomal RNA genes:
MERFF (myoclonic epilepsy and ragged red fiber):
• tRNAlys progressive myoclonic epilepsy, mitochondrial
myopathy with raged red fibers, slowly progressive dementia
• Severity of disease correlated with proportion mutant mtDNA
LHON (Leber’s hereditary optic neuropathy):
• 90% of cases from mutation in NADH dehydrogenase
• Late onset, acute optic atrophy
Nuclear genes can cause OXPHOS
Mutated nuclear genes can cause OXPHOS:
• About 1000 proteins needed for Oxidation
phosphorylation are encoded by nuclear DNA.
• Electron transport chain, translocators
• Need coordinate regulation of expression of
genes, import of proteins into mitochondria,
regulation of mitochondrial fission
• Nuclear regulatory factors
for transcription in nucleus, mt
• Often recessive autosomal
III. Coupling of electron transport and ATP synthesis
Concentration of ADP controls O2 consumption:
• Or phosphate potential ( [ATP]/[ADP][Pi])
1.
2.
3.
4.
5.
ADP used to form ATP
Release ATP requires H+ flow
H+ decreases proton gradient
ETC pumps more H+, uses O2
NADH donates e-, makes NAD+
to return to TCA cycle or other
Fig. 10
Uncoupling agents dissipate H+ gradient without ATP
Uncoupling agents decrease H+ gradient without
generating ATP:
Ex. DNP is a chemical uncoupler:
• lipid soluble, carries H+ across membrane
Fig. 11
Uncoupling proteins form channels, thermogenesis
Uncoupling proteins form channels for protons:
• Ex. UCP1 (thermogenin) makes heat in brown adipose
tissue (nonshivering thermogenesis); many mitochondria;
• Infants have lots of brown adipose tissue, not adults
Fig. 12
IV. Transport through mitochondrial membranes
Transport across inner mitochondrial membranes
uses channels, translocases:
• Form of active transport using proton gradient :
• ANT exchanges ATP: ADP
• Symport H+ with Pi
• Symport H+, pyruvate
Fig. 13
Transport across outer membrane:
Transport across outer membrane:
Rather nonspecific pores:
• VDAC voltage-dependent anion channels
• Often kinases on cytosolic side
Fig. 13
Mitochondrial permeability transition pore
Mitochondrial permeability transition pore:
• Large nonspecific pore:
• Will lead to apoptosis (cell death)
• Highly regulated process
• Hypoxia can trigger
• Pore opens, lets H+ flood in,
• Anions, cations enter
• Mitochondria swell and
• Irreversible damage
Fig. 14
Key concepts
• Reduced cofactors NADH, FAD(2H) donate e- to
electron transport chain
• ETC transfers e- to O2 -> H2O
• As e- transferred, H+ pushed across membrane;
• H+ gradient used by ATP synthase to make ATP
• O2 consumption tightly coupled to ATP synthesis
• Uncouplers disrupt process – poisons
• OXPHOS diseases from mutations in mt DNA or in
nuclear DNA
• Compounds transported across mt membranes
Review question
5. Which of the following would be expected for a
patient with an OXPHOS disease?
A. A high ATP:ADP ratio in the mitochondria
B. A high NADH:NAD+ ratio in the mitochondria
C. A deletion on the X chromosome
D. A high activity of complex II of the electron-transport
chain
E. A defect in the integrity of the inner mitochondrial
membrane
Review question p. 392
Decreased activity of the electron transport chain can
result from inhibitors as well as from mutations in
DAN. Why does impairment of the ETC result in
lactic acidosis?
• Inhibit ETC -> Impaired oxidation of pyruvate, fatty
acids and other fuels; therefore more lactate and
pyruvate in blood.
• NADH oxidation requires complete transfer of e- to
O2, so defect in chain increase NADH:NAD+ and
inhibit pyruvate dehydrogenase.