Download Krebs Cycle

Tricarboxylic Acid Cycle
TCA Cycle; Krebs Cycle;
Citric Acid Cycle
The Bridging Step: Pyruvate D’hase
O
H33C - C - C
pyruvate
O
NAD++
O--
NADH
CoASH
CO22
O
H33C - C - S - CoA
acetyl CoA
Pyruvate D’hase Complex
• Multienzyme
Multienzyme complex
complex (E.
(E. coli
coli enzyme
enzyme has
has 60
60
subunits)
subunits)
• Three
Three activities:
activities: pyruvate
pyruvate d’hase
d’hase (E1);
(E1);
dihydrolipoyl
dihydrolipoyl transacetylase
transacetylase (E2);
(E2); dihydrolipoyl
dihydrolipoyl
d’hase
d’hase (E3)
(E3)
• Prime
Prime example
example of
of metabolite
metabolite channeling
channeling
(substrates
(substrates acted
acted upon
upon immediately
immediately on
on enzyme
enzyme
surface
surface -- no
no diffusion
diffusion into
into cytosol)
cytosol)
1)
1) Pyruvate
Pyruvate d’hase
d’hase :: loss
loss of
of CO
CO22 from
from pyruvate
pyruvate
with
with transfer
transfer of
of the
the remaining
remaining two-carbon
two-carbon unit
unit as
as aa
hydroxyethyl
hydroxyethyl group
group (thiamine
(thiamine pyrophosphate
pyrophosphate
(TPP)
(TPP) used
used as
as aa cofactor)
cofactor)
2)
2) Dihydrolipoyl
Dihydrolipoyl transacetylase
transacetylase :: hydroxyethyl
hydroxyethyl group
group
transferred
transferred to
to lipoic
lipoic acid
acid and
and oxizided
oxizided to
to aa carboxylic
carboxylic
acid
acid (lipoic
(lipoic acid
acid cofactor
cofactor converted
converted to
to acetyl
acetyl
dihydrolipoamide);
dihydrolipoamide); acetyl
acetyl group
group transferred
transferred to
to CoA
CoA
[arsenic
[arsenic binds
binds lipoamide]
lipoamide]
3.
3. Dihydrolipoyl
Dihydrolipoyl d’hase
d’hase :: lipoic
lipoic acid
acid regenerated
regenerated
using
and FAD;
FAD; NADH
NADH is
is produced
produced
using NAD
NAD++ and
Thiamine
);
Thiamine (TPP);
(TPP); Riboflavin
Riboflavin (FAD);
(FAD); Niacin
Niacin (NAD
(NAD++);
Pantothenate
Pantothenate (CoA);
(CoA); Lipoic
Lipoic Acid
Acid
OH
CH33-C-H
TPP
S
S
FAD
E1
E2
E3
S
TPP
CH33-C- S
O
E1
FAD
E2
E3
Acetyl CoA
HS
TPP
HS
FAD
E1
E2
E3
S
TPP
S
FADH22
E1
E2
E3
NADH
NAD+
S
TPP
S
FAD
E1
E2
E3
Regulation of Pyruvate D’hase
Acetyl CoA and NADH allosterically inhibit
(product inhibition)
Mammalian pyr. d’hase is phosphorylated
and inactivated by a pyr. d’hase kinase.
This kinase itself is activated allosterically
by NADH and acetyl CoA. This effect is
reversed by pyr. d’hase phosphatase, which
removed the phosphate and reactivates the
enzyme.
AMP activates and GTP inhibits pyruvate
dehydrogenase. This commits pyruvate to
energy production.
SH
CH22
β-Mercaptoethylamine
CH22
O O
NH
O OH CH33
CH22
C-CH22-CH22-NH-C-CH-C-CH22O-P-O-P-O
O- OCH33
O
Pantothenic acid
Coenzyme A
O
A
OH
PO332-2-
3’-5’-ADP
1. Citrate Synthase
COO
COO-O
O C
C
H
H22C
C
COO
COO-oxaloacetate
oxaloacetate
(OAA)
(OAA)
acetyl
acetyl CoA
CoA
CoASH
CoASH
H
H22C
C COO
COO-HO
HO C
C COO
COO-H
H22C
C
COO
COO-citrate
citrate
Regulation of Citrate Sythase:
Reaction has a large negative DG =
-53.9 kJ/mol
NADH and Succinyl CoA are
allosteric inhibitors
2. Aconitase
H
H22C
C COO
COO-HO
HO C
C COO
COO-H
H22C
C
COO
COO-citrate
citrate
H
H22C
C
H
C
H C
COO
COO--COO
COO
H
C OH
H C
OH
COO
COO-isocitrate
isocitrate
3. Isocitrate D’hase
H
H22C
C COO
COO-H
C COO
H C
COO-H
C OH
H C
OH
COO
COO-isocitrate
isocitrate
NADH
NAD++
CO
CO22
-H
C
COO
H22C COO
H
C
H22C
C
C
O
O COO
COO-α-ketoglutarate
α-ketoglutarate
(αKg)
(αKg)
Regulation of Isocitrate D’hase
Mammalian enzyme: NADH and ATP are
allosteric inhibitors; ADP and NAD+ are
allosteric activators
E. Coli enzyme: phosphorylation of the
enzyme by a specific protein kinase
abolishes activity; removal of the phosphate
by a phosphatase restores activity
4. a-Ketoglutarate D’hase
NADH
H
H22C
C COO
COO-++
NAD
CoASH
CoASH
H
C
H22C
C
C
O
O COO
COO-α-ketoglutarate
α-ketoglutarate
(αKg)
(αKg)
CO
CO22
H
H22C
C COO
COO-H
C
H22C
C
C SCoA
SCoA
O
O
Succinyl-CoA
Succinyl-CoA
Reaction mechanism identical to that of pyruvate d’hase:
same cofactors utilized (succinyl group transferred)
5. Succinyl CoA Synthetase
H
H22C
C COO
COO-H
C
H22C
GTP
GTP
GDP,
GDP, Pi
Pi
C
C SCoA
SCoA
O
O
Succinyl-CoA
Succinyl-CoA
CoASH
CoASH
H
H22C
C
H
C
H22C
C
C
O
O
COO
COO-O
O--
Succinate
Succinate
6. Succinate D’hase
FADH
FADH22
H
H22C
C
H
C
H22C
-COO
COO
-COO
COO
FAD
FAD
H
H
--
Succinate
Succinate
OOC
OOC
C
C
C
C
COO
COO-H
H
Fumarate
Fumarate
Regulation of Succinate D’hase
Enzyme is a large multisubunit enzyme with
muliple cofactors like pyruvate d’hase.
Enzyme transfers electrons from the
substrate succinate to ubiquinone (Q)
Malonate (analogue of succinate) is a
competitive inhibitor and blocks the cycle at
this step; αkg, citrate, succinate accumulate in
its presence
7. Fumarase
H
H
--
OOC
OOC
C
C
C
C
-COO
COO
H
H
Fumarate
Fumarate
H
H22O
O
COO
COO-HO
CH
HO C
H
CH
CH22
-OOC
OOC
Malate
Malate
8. Malate D’hase
-COO
COO
HO
CH
HO C
H
CH
CH22
-OOC
OOC
Malate
Malate
NADH
NADH
NAD+
NAD+
COO
COO-O
O C
C
CH
CH22
-OOC
OOC
OAA
OAA
Overall Equation for TCA:
Acetyl
Q(FAD) ++ GDP
GDP ++ Pi
Pi ++ 2H
2H22O
O
Acetyl CoA
CoA ++ 3NAD
3NAD++ ++ Q(FAD)
CoASH
CoASH ++ 3NADH
3NADH ++ QH
QH22 (FADH
(FADH22)) ++ GTP
GTP ++ 2CO
2CO22 ++ 2H
2H++
*No net degradation of intermediates in TCA
Cycle - they are reformed with each full turn
of the cycle.
*NADH and QH are oxidized by the respiratory electron
22
transport chain. 3ATP per NADH and 2ATP per QH22.
Reaction
Energy Yielding
Product
ATP’s
Isocitrate D’hase
α−Kg D’hase
NADH
NADH
3
3
Succinyl CoA
Synthetase
GTP (ATP)
1
Succinate D’hase
QH22
2
Malate D’hase
NADH
3
One Round of TCA
12
Amount
Amount of
of ATP
ATP formed
formed per
per 11 Glucose:
Glucose:
ATP’s
ATP’s
Glycolysis
8
Pyruvate D’hase
6
TCA
24
38
The Glyoxylate Cycle
A “shunt” within the TCA cycle
• Biosynthetic route that leads to formation
of glucose from acetyl CoA
• Occurs in plants, bacteria and yeast
Isocitrate
Isocitrate is
is cleaved
cleaved by
by isocitrate
isocitrate lyase
lyase to
to
form
form succinate
succinate and
and glyoxylate:
glyoxylate:
-H
C
COO
H22C COO
H
C COO
COO-H C
H
C OH
H C
OH
-COO
COO
isocitrate
isocitrate
H
H22C
C COO
COO--COO
C
H
COO
H22C
Succinate
Succinate
O
O
H
H
C
C
-COO
COO
Glyoxylate
Glyoxylate
Glyoxylate
Glyoxylate condenses
condenses with
with acetyl
acetyl CoA
CoA to
to
form
form malate:
malate:
O
O
H
H
C
C
COO
COO--
Malate
Malate
Synthase
Synthase
+
Glyoxylate
Glyoxylate
CH33
C=O
S-CoA
Acetyl CoA
COO
COO-HO
CH
HO C
H
CH
CH22
COO
COO-Malate
Malate
NO CARBON ATOMS LOST AS CO22! THUS A NET
SYNTHESIS OF MALATE IS ACHEIVED.
Acetyl
Acetyl CoA
CoA
OAA
OAA
Glucose
Glucose
Malate
Malate
Acetyl
Acetyl CoA
CoA
Citrate
Citrate
CoASH
CoASH
Fumarate
Fumarate
Glyoxylate
Glyoxylate
Succinate
Succinate
Succinyl
Succinyl CoA
CoA
Isocitrate
Isocitrate
αKg
αKg
Glyoxylate
Glyoxylate Cycle
Cycle requires
requires transfer
transfer of
of
metabolites
metabolites between
between the
the mitochondrion,
mitochondrion, cytosol
cytosol
and
glyoxysome.
and aa special
special organelle,
organelle, the
the glyoxysome.
Glyoxysome:
Glyoxysome: Isocitrate
Isocitrate cleaved
cleaved to
to succinate
succinate and
and
glyoxylate.
glyoxylate. Glyoxylate
Glyoxylate condenses
condenses with
with acetyl
acetyl
CoA
CoA to
to form
form malate.
malate. Succinate
Succinate goes
goes to
to
mitochondrion;
mitochondrion; malate
malate to
to cytosol.
cytosol.
Mitochondrion:
Mitochondrion: Succinate
Succinate enters
enters the
the TCA
TCA cycle.
cycle.
Cytosol:
Cytosol: Malate
Malate converted
converted to
to OAA;
OAA; OAA
OAA to
to
glucose
glucose by
by the
the gluconeogenesis
gluconeogenesis pathway.
pathway.