Download Fe-S

Lecture 27
– Exam on Friday
– Up to glyoxalate cycle
For the exam
Metabolism
Glycolysis
Fermentation
Gluconeogensis
Glycogen
Pentose Phosphate Pathway
TCA
Mechanisms!
Regulation of pathways!
Page 851
Glyoxylate cycle
•
The glyoxylate cycle results in the net conversion of two acetyl-CoA
to succinate instead of 4 CO2 in citric acid cycle.
•
Succinate is transferred to mitochondrion where it can be converted
to OAA (TCA)
•
Can go to cytosol where it is converted to oxaloacetate for
gluconeogenesis.
Net reaction
2Ac-CoA + 2NAD+ + FAD
OAA + 2CoA + 2NADH +FADH2 + 2H+
Plants are able to convert fatty acids to glucose through this
pathway
Page 798
Electron transport and oxidative phosphorylation
Complete oxidation of glucose by molecular oxygen can be
described as:
C6H12O6 + 6O2  6CO2 + 6H2O
Go’=-2823 kJ/mol
Can be broken down into two half-reactions with the transfer of
electrons
C6H12O6 + 6H2O  6CO2 + 24H+ +24e6O2 + 24H+ + 24e-  12H2O
12e- from the oxidation of glucose are not transferred directly to O2,
go to NAD+ and FAD to form 10NADH and 2FADH2
These are reoxidized, passing their electrons to the electrontransport chain to reduce O2 to H2O causing the
mitochondrion to create a proton gradient.
This pH gradient is used to drive the synthesis of ATP via
oxidative phosphorylation.
Chemiosmotic Theory
• Electron Transport: Electrons carried by reduced
coenzymes are passed through a chain of proteins
and coenzymes to drive the generation of a proton
gradient across the inner mitochondrial membrane
• Oxidative Phosphorylation: The proton gradient
runs downhill to drive the synthesis of ATP
• Electron transport is coupled with oxidative
phosphorylation
• It all happens in or at the inner mitochondrial
membrane
Outer Membrane – Freely
permeable to small
molecules and ions. Contains
porins with 10 kDa limit
Inner membrane – Protein
rich (4:1 protein:lipid).
Impermeable. Contains ETR,
ATP synthase, transporters.
Cristae – Highly folded inner membrane structure. Increase
surface area.
Matrix- “cytosol” of the mitochondria. Protein rich (500
mg/ml) Contains TCA cycle enzymes, pyruvate
dehydrogenase, fatty and amino acid oxidation pathway,
DNA, RNA, ribosomes
Intermembrane Space – composition similar to cytosol
Mitochondrial transport
The inner membrane is impermeable to hydrophilic
substances. Has special transport systems for the
following:
1. Glycolytically produced cytosolic NADH.
2. Mitochondrially produced metabolites (OAA, acetyl-CoA)
for cytosolic glucose formation and fatty acid
biosynthesis.
3. Mitochondrially produced ATP must go to cytosol where
ATP-utilizing reactions take place.
Example: cytoplasmic shuttle systems transport NADH
across inner membrane.
“Transport” of NADH across inner mito membrane
Malate-aspartate shuttle
2 phases
Phase A
1.
Cytosolic NADH reduces OAA to malate (malate DH).
2.
Malate--ketoglutarate carrier transports malate from cytosol to
mitochondrial matrix, exchanged for -KG
3.
In the matrix, NAD+ reoxidizes malate to make OAA and yield
NADH.
Phase B
4.
Transaminase converts OAA to Asp and Glu to -KG.
5.
The glutamate-apartate carrier transports Asp from the matrix in
exchange for Glu.
6.
Transaminase in cytosol converts Glu to Asp.
3 ATPs for every NADH but loses 0.3 ATP because each NADH enters
matrix with proton (yield 2.7 ATP).
Glycerol phosphate shuttle
Glycerophosphate shuttle
Simpler but less energy efficient than the malate-aspartate
shuttle.
Supplies electrons in a manner similar to succinate
dehydrogenase
Approx. 2 ATP per NADH, about 0.7 ATP less than the
malate-aspartate shuttle.
Advantage-irreversible so it operates efficiently even when
cytoplasmic NADH is low relative to NAD+.
Malate-aspartate shuttle is reversible - driven by
concentration gradients.
Reduction Potentials
• High Eo' indicates a strong tendency to be reduced
• Crucial equation: Go' = -nF Eo'
•
Eo' = Eo'(acceptor) - Eo'(donor)
• NADH + ½ O2 + H+  NAD++ H+ + H2O
n=# of e- transferred per mol reactants, F=Faraday constant
NAD++ H+ + 2e- NADH
Eo’ = -0.32
½ O2 + 2e- + 2H+  H2O
Eo’ = 0.816 V
Go‘= -nF(Eo'(O2) - Eo'(NADH))
Go‘= -nF(0.82 –(-0.32)) = -nF(1.14)
= -2(96.5 kJ mol-1V-1)(1.136) = -220 kJ mol-1
Page 803
Figure 22-9
The mitochondrial electrontransport chain.
Electron Transport
• Four protein complexes in the inner
mitochondrial membrane
• A lipid soluble coenzyme (UQ, CoQ)
and a water soluble protein (cyt c)
shuttle between protein complexes
• Electrons generally fall in energy
through the chain - from complexes I
and II to complex IV
Standard reduction potentials
of the major respiratory
electron carriers.
Cofactors of the electron transport chain
•
•
•
•
•
•
•
•
•
Fe-S clusters
Coenzyme Q (ubiquinone)
Flavin mononucleotide
FAD
Cytochrome a
Cytochrome b
Cytochrome c
CuA
CuB
Iron-sulfur clusters
•
•
•
•
•
4 main types of iron sulfur clusters
[2Fe-2S] and [4Fe-4S] cluster coordinated by 4 Cys SH
[3Fe-4S] is a [4Fe-4S] lacking one Fe atom.
[Fe-S] is only found in bacteria, liganded to 4 Cys
Rieske iron-sulfur proteins [2Fe-2S] cluster but 1 Fe is
coordinated by 2 His.
• Oxidized and reduced states of all Fe-S clusters differ
by one formal charge.
Page 808
Figure 22-15a
Structures of the
common iron–sulfur clusters. (a) [Fe–S]
cluster.
Page 808
Figure 22-15b
Structures of the
common iron–sulfur clusters. (b) [2Fe–2S]
cluster.
Page 808
Figure 22-15c
Structures of the
common iron–sulfur clusters. (c) [4Fe–4S]
cluster.
Page 809
Figure 22-16 X-Ray structure of
ferredoxin from Peptococcus aerogenes.
Figure 22-17a
Oxidation states
of the coenzymes of
complex I. (a) FMN.
Page 810
Can accept or
donate 1 or 2 e-
Figure 22-17b
Oxidation states
of the coenzymes of
complex I. (b) CoQ.
Page 810
Coenzyme Q’s
hydrophobic tail allows it to
be soluble in the inner
membrane lipid bilayer.
Page 813
Figure 22-21a
Visible absorption spectra of cytochromes.
(a) Absorption spectrum of reduced cytochrome c showing its
characteristic , b, and g (Soret) absorption bands.
Page 813
Figure 22-21b
The three separate  bands in the
spectrum of beef heart mitochondrial membranes indicate the
presence of cytochromes a, b, and c.
Page 813
Figure 22-22a
Porphyrin rings in cytochromes. (a)
Chemical structures.
Page 813
Figure 22-22b
Porphyrin rings in
cytochromes. (b) Axial liganding of the heme
groups contained in cytochromes a, b, and c
are shown.
Complex I
•
•
•
•
•
•
•
NADH-CoQ Oxidoreductase (NADH dehydrogenase)
Electron transfer from NADH to CoQ
More than 30 protein subunits - mass of 850 kD
1st step is 2 e- transfer from NADH to FMN
FMNH2 converts 2 e- to 1 e- transfer
6-7 FeS clusters.
Four H+ transported out per 2 e-
NADH + H+
NAD+
FMN
FMNH2
Fe2+S
Fe3+S
CoQ
CoQH2
Complex II
• Succinate-CoQ Reductase
• Contains the succinate dehydrogenase (from TCA
cycle!)
• four subunits
• Two largest subunits contain 2 Fe-S proteins
• Other subunits involved in binding succinate
dehydrogenase to membrane and passing e- to
Ubiquinone
• FAD accepts 2 e- and then passes 1 e- at a time to Fe-S
protein
• No protons pumped from this step
Succinate
FAD
Fe2+S
CoQ
Fumarate
FADH2
Fe3+S
CoQH2
Q-Cycle
•
•
•
•
Transfer from the 2 e- carrier ubiquinone
(QH2) to Complex III must occur 1 e- at a
time.
Works by two single electron transfer steps
taking advantage of the stable semiquinone
intermediate
Also allows for the pumping of 4 protons
out of mitochondria at Complex III
Myxothiazol (antifungal agent) inhibits
electron transfer from UQH2 and Complex
III.
UQ
UQ.-
UQH2