SECTION 13.1
Glycolysis
233
G lu cag o n
Activation of G-protein
Inactive a d e n y la te c y c l a s e --------------- ►
A ctive a d e n y la te c y c la s e
A T P ------- i------- - cA M P + PFj
D e p h o sp h o -
p h o s p h a ta s e
inhibitor
Inactive protein k in a se
(R A)
A ctive protein k in a se (C2) + F\(cA M P)z
A T P
A D P
^
j
P h o sp h o -
p h o s p h a ta s e -
inhibitor
M ore activ e
,
d e p h o sp h o -P K
A T P | A D P
L e s s activ e
p h o sp h o -P K
f?
I H ,0
‘ P h o sp h o p ro te in p h o s p h a ta s e
FIGURE 13-5
Action of glucagon on the liver pyruvate kinase (PK) via the cAMP-cascade system (see also Chapter 30). Glucagon, by
combining at specific receptor sites on the hepatocyte plasma membrane, activates adenylate cyclase, which converts
ATP to cAMP. The latter activates (cAMP-dependent) protein kinase, which phosphorylâtes PK (phospho-PK),
converting it to a less active form and making it more susceptible to allosteric inhibition by alanine and ATP.
Phospho-PK is converted to the more active form by removal of phospho groups by a phosphoprotein phosphatase,
whose activity is also regulated by cAMP-dependent protein kinase via the phosphorylation of phosphatase inhibitor.
The overall effect of glucagon is to diminish glycolysis, whereas the effect of insulin is to promote glycolysis.
PPi
= pyrophosphate;
0
= Inhibition.
has a higher susceptibility to inhibition by the negative
modulators alanine and ATP, and has virtually no activity
in the absence of fructose-1,6-bisphosphate. The phospho
enzyme is converted to the dephospho form by a phospho-
protein phosphatase, the activity of which is also regulated
by cAMP-dependent protein kinase (Figure 13-5).
The muscle and brain isoenzyme (M-type) shows hyper-
bolic kinetics with phosphoenolpyruvate and is inhibited
by ATP, with an increase in
Km
for phosphoenolpyru-
vate and development of sigmoidal kinetics. Fructose-
1
,
6
-bisphosphate and alanine have no effect on this
isoenzyme.
The overall reaction for glycolysis in cells that possess
mitochondria and are under aerobic conditions is:
Glucose + 2NAD+ + 2ADP3- +
2
HPO
4
-*
2pyruvate- + 2NADH + 2H+ + 2ATP4- + 2H20
Pyruvate is transported into mitochondria, where it is
oxidized in the TCA cycle (see below). Although NADH is
not transported into mitochondria, its reducing equivalent
is transported into mitochondria (Chapter 15), where it
is oxidized in the electron transport system coupled to
oxidative phosphorylation (Chapter 14).
Cells that lack mitochondria (e.g., erythrocytes) or con-
tain mitochondria but under hypoxic conditions (e.g., vig-
orously and repeatedly contracting muscle) reduce pyru-
vate to lactate by lactate dehydrogenase (LDH), which
uses NADH as a reductant. This reduction regenerates
NAD+, which is required for continued oxidation of glyc-
eraldehyde 3-phosphate.
Reduction o f Pyruvate to Lactase
This reversible reaction is the final step of glycolysis
and is catalyzed by (LDH):
Pyruvate- + NADH + H+ ^ L-lactate- + NAD+
The overall reaction of glycolysis becomes
Glucose + 2ADP3- + 2P3- -+
21actate- + 2ATP4- + 2H20
LDH (M.W. 134,000) occurs as five tetrameric isoen-
zymes composed of two different types of subunits.
Subunits M (for muscle) and H (for heart) are encoded
by loci in chromosomes 11 and 12, respectively. Two
subunits used in the formation of a tetramer yield five com-
binations:
H
4
(LDH-1), H
3
M(LDH-2), H
2
M
2
(LDH-3),
HM
3
(LDH-4), and M
4
(LDH-5). The tissue distributiont
of LDH isoenzymes is variable. For example, LDH-1
and LDH-2 are the principal isoenzymes in heart, kidney,
brain, and erythrocytes; LDH-3 and LDH-4 predominate
in endocrine glands (e.g., thyroid, adrenal, pancreas),
lymph nodes, thymus, spleen, leukocytes, platelets, and
nongravid uterine muscle; and LDH-4 and LDH-5 pre-
ponderate in liver and skeletal muscle. In tissue injury or
insult, the appropriate tissue isoenzymes appear in plasma
(Chapter
8
); thus, determination of LDH isoenzyme
composition has diagnostic value.
Serum LDH isoenzymes can be separated by elec-
trophoresis on agarose gel or cellulose acetate mem-
brane, usually at pH
8
.
6
. After separation, their location
is determined by incubation of the support medium in a
