Molecular Biochemistry II

Pentose Phosphate Pathway

Copyright 1999-2007 by Joyce J. Diwan.

All rights reserved.
 Pentose Phosphate Pathway

Pentose Phosphate Pathway

Other names:
Phosphogluconate Pathway
Hexose Monophosphate Shunt
The linear part of the pathway carries out oxidation
and decarboxylation of the 6-C sugar glucose-6-P,
producing the 5-C sugar ribulose-5-P.
 G lucose-6-phosphate 6-Phospho- O O
D ehydrogenase glucono- 1C
6 CH 2 O PO 3 2 6 CH O PO 2 lactonase
+ 2 3
NADPH + H HC OH
H
5 O O H NADP
+
H
5 O H 2O H+ 2
H H HO 3CH
4 OH H 1 4
OH H O
1 HC O H
4
OH H OH
3 2 3 2 HC OH
5
H OH H OH CH 2 O PO 3 2
6
glucose-6-phosphate 6-phoshogluconolactone 6-phosphogluconate

Glucose-6-phosphate Dehydrogenase catalyzes

oxidation of the aldehyde (hemiacetal), at C1 of
glucose-6-phosphate, to a carboxylic acid, in ester
linkage (lactone).
NADP+ serves as electron acceptor.
 G lucose-6-phosphate 6-Phospho- O O
D ehydrogenase glucono- 1C
6 CH O PO 2 6 CH O PO 2 lactonase
2 3 + 2 3
N AD P H + H HC OH
H
5 O O H NADP
+
H
5 O H 2O H+ 2
H H HO 3CH
4 OH H 1 4
OH H O
1 HC O H
4
OH H OH
3 2 3 2 HC OH
5
H OH H OH CH 2 O PO 3 2
6
glucose-6-phosphate 6-phoshogluconolactone 6-phosphogluconate

6-Phosphogluconolactonase catalyzes hydrolysis of the

ester linkage, resulting in ring opening.
The product is 6-phosphogluconate.
Although ring opening occurs in the absence of a catalyst,
6-Phosphogluconolactonase speeds up the reaction,
decreasing the lifetime of the highly reactive, and thus
potentially toxic, 6-phosphogluconolactone.
 O O Phosphogluconate
1C Dehydrogenase
HC OH 1CH 2 OH
2 NADP + NADPH + H +
HO 3CH C O
2
HC OH HC OH
4 3
HC OH CO 2 HC OH
5 4
CH 2 OPO 3 2 CH 2 OPO 3 2
6 5
6-phosphogluconate ribulose-5-phosphate

Phosphogluconate Dehydrogenase catalyzes oxidative

decarboxylation of 6-phosphogluconate, to yield the 5-
C ketose ribulose-5-phosphate.
The OH at C3 (C2 of product) is oxidized to a ketone.
This promotes loss of the carboxyl at C1 as CO2.
NADP+ serves as oxidant.
 H O O
H H
C C
Reduction of NADP + NH2 NH2
(as with NAD+) +
involves transfer of 2 e N
2e + H
+ N

and 1 H+ to the R R

nicotinamide moiety. NADP+ NADPH

NADPH, a product of the Pentose Phosphate Pathway,

functions as a reductant in anabolic (synthetic)
pathways, e.g., fatty acid synthesis.
NAD+ serves as electron acceptor in catabolic
pathways, in which metabolites are oxidized.
The resultant NADH is reoxidized by the respiratory
chain, producing ATP.
NAD+ & NADP+ differ Nicotinamide H O
Adenine
only in the presence of C
NH2
Dinucleotide
an extra phosphate on O +
the adenosine ribose of
O P O CH2
N
nicotinamide
O
NADP+. H H
H H
This difference has little OH OH
to do with redox activity, O NH2

but is recognized by N
N
substrate-binding sites
of enzymes.
O P O CH2
N N adenine
O
It is a mechanism for O H H
H H esterified to
separation of catabolic OH OH Pi in NADP+
and synthetic pathways.
Regulation of Glucose-6-phosphate Dehydrogenase:
Glucose-6-phosphate Dehydrogenase is the
committed step of the Pentose Phosphate Pathway.
This enzyme is regulated by availability of the
substrate NADP+.
As NADPH is utilized in reductive synthetic pathways,
the increasing concentration of NADP+ stimulates the
Pentose Phosphate Pathway, to replenish NADPH.
The rest of the pathway converts ribulose-5-P to the 5-C
product ribose-5-P, or to 3-C glyceraldehyde-3-P & 6-C
fructose-6-P.
Additional enzymes include an Isomerase, Epimerase,
Transketolase, and Transaldolase.
Epimerase inter-converts
CH 2OH
stereoisomers ribulose-5-
P and xylulose-5-P. C O
Epimerase
HO C H
Isomerase converts the
ketose ribulose-5-P to the CH 2OH H C OH

aldose ribose-5-P. C O CH 2OPO 32

xylulose-5-
Both reactions involve H C OH phosphate
deprotonation to an H C OH HC O
endiolate intermediate
CH 2OPO 32 H C OH
followed by specific
ribulose-5-
reprotonation to yield phosphate Isomerase
H C OH

the product. H C OH

Both reactions are CH 2OPO 32

ribose-5-
reversible. phosphate
Transketolase & Transaldolase catalyze transfer of
2-C or 3-C molecular fragments respectively, in each
case from a ketose donor to an aldose acceptor.
D. E. Nicholson has suggested that the names of these
enzymes should be changed, since
Transketolase actually transfers an aldol moiety
(glycoaldehyde), and
Transaldolase actually transfers a ketol moiety
(dihydroxyacetone).
However the traditional enzyme names are used here.
 CH 2 O H
T ran sk eto lase
C O

CH 2 O H HC O HO C H

C O H C OH H C OH

HO C H H C OH HC O H C OH

H C OH
+ H C OH H C OH
+ H C OH

CH 2 O P O 3 2 CH 2 O P O 3 2 CH 2 O P O 3 2 CH 2 O P O 3 2

x ylu lo se - rib o se - gly ce ra ld e h yd e - sed o h ep tu lo se -

5 -p h o sp h a te 5 -p h o sp h a te 3 -p h o sp h ate 7 -p h o sp h a te

Transketolase transfers a 2-C fragment from xylulose-

5-P to either ribose-5-P or erythrose-4-P.
Transketolase utilizes as prosthetic group thiamine
pyrophosphate (TPP), a derivative of vitamin B1.
Pyruvate Dehydrogenase of Krebs Cycle also utilizes
TPP as prosthetic group.
 thiazolium O O

aminopyrimidine H3C ring

CH2 CH2 O P O P O
moiety
+ O O
CH2 N
N S
C
H acidic H+
H3C N NH2
thiamine pyrophosphate (TPP)

TPP binds at the active site in a V conformation.

H+ dissociates from the C between N & S in the
thiazolium ring.
The aminopyrimidine amino group is near the
dissociable H+, & serves as H+ acceptor.
This H+ transfer is promoted by a Glu residue adjacent
to the pyrimidine ring.
 H3C 2
CH2 CH2OPO2OPO 3
TPP
+
CH2 N xylulose-5-P
N S
C
The thiazolium CH2OH

carbanion reacts H3C N NH2

C O

with the carbonyl HO C H

C of xylulose-5-P H C OH
to form an addition Transketolase CH2OPO32
compound. H3C 2
CH2 CH2OPO2OPO3

N in the thiazole CH2

+
N
ring acts as an e N
C
S

sink, promoting H3C N HO C CH2OH subsequent

NH2
C-C bond cleavage. HO C H cleavage
active site H C OH
intermediate
CH2OPO32
 H3C 2
The 3-C aldose TPP
CH2 CH2OPO2OPO3

glyceraldehyde-3-P CH2 N
+
xylulose-5-P
is released. N
C

S
CH2OH
A 2-C fragment H3C N NH2
C O
remains on TPP. HO C H

Completion is by H C OH
reversal of these Transketolase CH2OPO32
steps. H3C 2
CH2 CH2OPO2OPO3
The 2-C fragment +
CH2 N
condenses with N
C
S
one of the aldoses
HO C CH2OH subsequent
erythrose-4-P (4-C) H3C N NH2
cleavage
HO C H
or ribose-5-P (5-C)
active site H C OH
to form a ketose-P intermediate
2
product. CH2OPO 3
 CH 2 O H
T ran sk eto lase
C O

CH 2 O H HC O HO C H

C O H C OH H C OH

HO C H H C OH HC O H C OH

H C OH
+ H C OH H C OH
+ H C OH

CH 2 O P O 3 2 CH 2 O P O 3 2 CH 2 O P O 3 2 CH 2 O P O 3 2

x y lu lo se - rib o se - g lyc e rald eh y d e - sed o h e p tu lo se -

5 -p h o sp h a te 5 -p h o sp h a te 3 -p h o sp h a te 7 -p h o sp h a te

Transfer of the 2-C fragment to the 5-C aldose

ribose-5-phosphate yields sedoheptulose-7-phosphate.
Transfer of the 2-C fragment instead to the 4-C aldose
erythrose-4-phosphate yields fructose-6-phosphate.
 C H 2O H
T ra n sald o lase
C O H 2C OH

HO CH C O

HC OH HC O HO CH

HC OH HC O HC OH HC OH

HC OH + HC OH HC OH + HC OH
2 2 2 2
H 2C O PO 3 H 2C O PO 3 H 2C O PO 3 H 2C O PO 3

sed o h ep tu lo se - g ly c e ra ld e h y d e - e ry th ro se - fru c to se -
7 -p h o sp h a te 3 -p h o sp h ate 4 -p h o sp h a te 6 -p h o sp h a te

Transaldolase catalyzes transfer of a 3-C

dihydroxyacetone moiety, from sedoheptulose-7-phosphate
to glyceraldehyde-3-phosphate.
Transaldolase has an , barrel structure.
 CH2OH CH2OH
Enz-Lys NH2
H Schiff base
C O Enz-Lys N C
+ intermediate
HO CH HO CH

HC OH HC OH
OH
HC OH HC OH
sedoheptulose-
7-phosphate HC OH HC OH

H2C OPO32 H2C OPO32

HC O CH2OH
erythrose-4- H
phosphate HC OH Enz-Lys N C + H+
+
HC OH HO C H

Transaldolase H2C OPO32

In Transaldolase, the -amino group of a lysine residue

reacts with the carbonyl C of sedoheptulose-7-P to form
a protonated Schiff base intermediate.
 CH2OH CH2OH
Enz-Lys NH2
H Schiff base
C O Enz-Lys N C
+ intermediate
Aldol HO CH HO CH
cleavage HC OH HC OH
releases OH
HC OH HC OH
erythrose-4- sedoheptulose-
7-phosphate HC OH HC OH
phosphate.
H2C OPO32 H2C OPO32

The Schiff
HC O CH2OH
base erythrose-4- H
stabilizes the phosphate HC OH Enz-Lys N
+
C + H+

carbanion HC OH HO C

H

on C3. Transaldolase H2C OPO32

Completion of the reaction is by reversal, as the carbanion

attacks instead the aldehyde carbon of the 3-C aldose
glyceraldehyde-3-P to yield the 6-C fructose-6-P.
The diagram at (3) ribulose-5-P
right summarizes IS EP
flow of 15 C atoms ribose-5-P (2) xylulose-5-P
through Pentose TK
Phosphate Pathway glyceraldehyde-3-P
reactions by which
5-C sugars are sedoheptulose 7 P
converted to 3-C
and 6-C sugars. fructose-6- P TA
IS = Isomerase erythrose-4-P
EP = Epimerase
TK fructose-6-P
TK = Transketolase
TA = Transaldolase glyceraldehyde-3-P
The balance sheet below summarizes flow of 15 C
atoms through Pentose Phosphate Pathway reactions by
which 5-C sugars are converted to 3-C and 6-C sugars.
C5 + C5 C3 + C7 (Transketolase)
C3 + C7 C6 + C4 (Transaldolase)
C5 + C4 C6 + C3 (Transketolase)
____________________________
3 C5 2 C6 + C3 (Overall)

Glucose-6-phosphate may be regenerated from either

the 3-C glyceraldehyde-3-phosphate or the 6-C
fructose-6-phosphate, via enzymes of Gluconeogenesis.
Depending on needs of a cell for ribose-5-phosphate,
NADPH, and ATP, the Pentose Phosphate Pathway can
operate in various modes, to maximize different products.
There are three major scenarios:

2 NADP+ 2 NADPH + CO2

glucose-6-P ribulose-5-P ribose-5-P

Pentose Phosphate Pathway producing

NADPH and ribose-5-phosphate

Ribulose-5-P may be converted to ribose-5-phosphate,

a substrate for synthesis of nucleotides and nucleic acids.
The pathway also produces some NADPH.
 2 NADP+ 2 NADPH + CO2
glucose-6-P ribulose-5-P ribose-5-P

fructose-6-P, &
glyceraldehyde-3-P
Pentose Phosphate Pathway producing
maximum NADPH

Glyceraldehyde-3-P and fructose-6-P may be

converted to glucose-6-P for reentry to the linear
portion of the Pentose Phosphate Pathway,
maximizing formation of NADPH.
 2 NADP+ 2 NADPH + CO2
glucose-6-P ribulose-5-P ribose-5-P

fructose-6-P, &
glyceraldehyde-3-P

to Glycolysis
for production of ATP
Pentose Phosphate Pathway producing
NADPH and ATP

Glyceraldehyde-3-P and fructose-6-P, formed from 5-C

sugar phosphates, may enter Glycolysis for ATP synthesis.
The pathway also produces some NADPH.
 2 NADP+ 2 NADPH + CO2
glucose-6-P ribulose-5-P ribose-5-P

fructose-6-P, &
glyceraldehyde-3-P

to Glycolysis
for production of ATP
Pentose Phosphate Pathway producing
NADPH and ATP
Ribose-1-phosphate generated during catabolism of
nucleosides also enters Glycolysis in this way, after first
being converted to ribose-5-phosphate.
Thus the Pentose Phosphate Pathway serves as an entry
into Glycolysis for both 5-carbon & 6-carbon sugars.
Glutathione is a tripeptide that includes a Glu linked by
an isopeptide bond involving the side-chain carbonyl group.
Its functional group is a cysteine thiol.
One role of glutathione is degradation of hydroperoxides,
that arise spontaneously in the oxygen-rich environment
in red blood cells.
Hydroperoxides can react with double bonds in fatty acids
of membrane lipids, making membranes leaky.
Glutathione Peroxidase catalyzes degradation of organic
hydroperoxides by reduction, as two glutathione molecules
(represented as GSH) are oxidized to a disulfide.
2 GSH + ROOH GSSG + ROH + H2O
Glutathione Peroxidase uses the trace element selenium as
functional group.
The enzyme's primary structure includes an analog of cysteine,
selenocysteine, with Se replacing S.
Regeneration of reduced glutathione requires NADPH,
produced within erythrocytes in the Pentose Phosphate
Pathway.
Glutathione Reductase catalyzes:
GSSG + NADPH + H+ 2 GSH + NADP+
Genetic deficiency of Glucose-6-P Dehydrogenase can
lead to hemolytic anemia, due to inadequate [NADPH]
within red blood cells.
The effect of partial deficiency of Glucose-6-phosphate
Dehydrogenase is exacerbated by substances that lead to
increased production of peroxides (e.g., the antimalarial
primaquine).