Adv Physiol Educ 35: 92–94, 2011;
doi:10.1152/advan.00102.2010.
Illuminations
Mechanism of ATP loss in nonoxidative contracting muscle
Piero L. Ipata
Department of Biology, Unit of Biochemistry, University of Pisa, Pisa, Italy
Submitted 28 September 2010; accepted in final form 2 November 2010
⫹
Glycogen(N) ⫹ 3 ADP3⫺ ⫹ 3 P2⫺
i ⫹ H ¡ glycogen(N⫺1) (1)
1⫺
⫹ 2 lactate ⫹ 3 ATP4⫺ ⫹ 2 H2O
where N is the number of glycosyl units of the muscle glycogen
macromolecule. It can be seen that nonoxidative glycolysis,
starting from glycogen, not only does not generate protons, i.e.,
does not cause metabolic acidosis, but rather consumes one
proton per glycosyl unit transformed into two lactate molecules. Metabolic acidosis occurs because the three ATP molecules are immediately hydrolyzed by the myosin ATPase to
sustain contraction, as follows:
⫹
3 ATP4⫺ ⫹ 3 H2O ¡ 3 ADP3⫺ ⫹ 3 P2⫺
i ⫹3H
(2)
Equation 3 is the summary equation of the conversion of one
glycosyl residue of muscle glycogen into lactate and protons
(Eq. 1 ⫹ Eq. 2):
Glycogen(N) ⫹ H2O ¡ glycogen(N⫺1) ⫹ 2 lactate2⫺ ⫹ 2 H⫹
(3)
Students should be aware that the two protons released in this
equation are not generated by lactic acid dissociation, because
lactate, not lactic acid, is formed in reaction 1. The two
protons are generated by the production of the three protons
released by reaction 2 and the consumption of one proton by
reaction 1. Most important, the two reactions are strictly
“coupled,” i.e., any ATP synthesized by anaerobic glycolysis
(reaction 1) is hydrolyzed to ADP and Pi (reaction 2), which
are immediately converted back to ATP by anaerobic glycolysis. Moreover, the coupled Eqs. 1 and 2 imply that any
Address for reprint requests and other correspondence: P. L. Ipata, Dept. of
Biology, Unit of Biochemistry, Univ. of Pisa, Via San Zeno, 51, Pisa 56127,
Italy (e-mail: [email protected]).
92
increase in the rate of ATP synthesis to sustain an increased
energy demand should be equal to that of ATP hydrolysis by
myosin ATPase. Therefore, the muscular ATP pool (⬃8 mM in
the intracellular water in humans) (3, 8) should remain unchanged during contraction. However, since the late 1970s (6,
9), ever-increasing experimental evidence has clearly shown
that a considerable ATP loss occurs in heavy contracting
muscle (16, 17, 19, 20). In humans, even a single 10-s sprint
bout is sufficient to acutely deplete the muscular ATP pool by
⬃21% (1). After repeated bouts, a more marked reduction
(⬃41%) has been reported (9). In horses, the ATP loss may
reach a value of ⬃47% after 2 min of exercise at a speed of 12
m/s (16). It has also been shown that the muscular total adenine
nucleotide ([ATP] ⫹ [ADP] ⫹ [AMP]) reduction caused by
intense exercise is lower in women than in men (8). Lactate
metabolism is strictly related to that of ATP in working
muscle. The bulk of evidence suggests that lactate must not be
considered as a dead-end waste product of nonoxidative glycolysis but as a particular mobile fuel for cellular, regional, and
whole body aerobic metabolism [for a review, see Brooks (2)].
In particular, lactate improves the excitability of depolarized
rat skeletal muscle via mechanisms not related to a reduction of
intracellular pH but by exerting a direct inhibitory effect on
muscle Cl⫺ channels (7). Finally, there is not a 1:1 correspondence between proton and lactate anion release from working
muscle, and these releases can far exceed the net change in
[ATP] (5).
A breakthrough on this important issue of muscle ATP
metabolism was the finding that in contracting muscle inosine
monophosphate (IMP), a nonadenine nucleoside monophosphate, accumulates intracellularly through AMP deamination
followed by the extracellular release of inosine and hypoxanthine, two IMP catabolites (9, 14, 19).
Mechanisms of ATP loss during nonoxidative muscle
contraction. When the rate of ATP hydrolysis exceeds the
capacity of contracting muscle to phosphorylate ADP by anaerobic glycolysis, [ADP] and [AMP] rise, first leading to the
deamination of AMP to IMP and subsequently to the dephosphorylation of IMP (19) (Table 2). The process of intracellular
ATP breakdown in exhaustive contracting muscle may be
divided into two stages (see Fig. 1, B and C). In the first stage,
some of the ADP, generated by myosin ATPase (reaction 2)
and escaping recycling, is transformed into AMP and ATP by
the action of myokinase (reaction 4). AMP is then deaminated
to generate IMP and NH3 by adenylate deaminase (reaction 5),
as follows:
Myokinase: 2 ADP ↔ ATP ⫹ AMP
(4)
Adenylate deaminase: H2O ⫹ AMP ¡ IMP ⫹ NH3
(5)
IMP may be reaminated to AMP by the successive action of
adenylosuccinate synthetase and adenylate lyase (10), thus
contributing in maintaining the intracellular concentration of
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The transition from rest to intense exercise is a challenge to
cellular energetics (11, 13, 15). The metabolic fuels, i.e., the
sources of ATP to sustain muscular contraction, are creatine
phosphate and glycogen. Two anaerobic metabolic paths, leading to ATP generation, are catalyzed by creatine kinase and by
the 12 enzymes of nonoxidative glycolysis, starting from
glycogen. There is now general agreement that, unless replenished, creatine phosphate can sustain heavy muscle contraction
for only 3– 4 s. Thereafter, nonoxidative glycolysis becomes
the main ATP source, until the onset of fatigue. This article
aimed to relate the path of ATP generation during glycogen
utilization as a metabolic fuel with that of ATP breakdown in
nonoxidative contracting muscle.
Correlation-type considerations. Table 1 shows the reactions of nonoxidative glycolysis, starting from glycogen. Since
the product of each reaction becomes the substrate of the
subsequent one, the 12 reactions constitute a catabolic pathway, whose summary equation, with all constituents and
charges at physiological pH, is as follows:
Illuminations
GLYCOGEN-ATP CATABOLISM INTERACTION
93
Table 1. The reactions of anaerobic glycolysis, starting from glycogen
Reaction
Enzyme
1. Glycogen(N) ⫹ Pi ¡ glycogen(N ⫺ 1) ⫹ glucose-1-phosphate
2. Glucose-1-phosphate ¡ glucose-6-phosphate
3. Glucose-6-phosphate ¡ fructose-6-phosphate
4. Fructose-6-phosphate ⫹ ATP ¡ fructose-1,6-bis-phosphate ⫹ ADP ⫹ H⫹
5. Fructose-1,6-bis-phosphate ¡ dihydroxyacetone-phosphate ⫹ glyceraldehyde-3-phosphate
6. Dihydroxyacetone-phosphate ¡ glyceraldehyde-3-phosphate
7. 2 Glyceraldehyde-3-phosphate ⫹ 2 NAD⫹ ⫹ 2 Pi ¡ 2 1,3-bis-phosphoglycerate ⫹ 2 NADH ⫹ 2 H⫹
8. 2 1,3-Bis-phosphoglycerate ⫹ 2 ADP ¡ 2 3-phosphoglycerate ⫹ 2 ATP
9. 2 3-Phosphoglycerate ¡ 2 2-phosphoglycerate
10. 2 2-Phosphoglycerate ¡ 2 phosphoenolpyruvate ⫹ 2 H2O
11. 2 Phosphoenolpyruvate ⫹ 2 ADP ⫹ 2 H⫹ ¡ 2 pyruvate ⫹ ATP
12. 2 Pyruvate ⫹ 2 NADH ⫹ 2 H⫹ ¡ 2 lactate ⫹ 2 NAD⫹
Phosphorylase
Phosphoglucomutase
Glucose-6-phosphate isomerase
6-Phosphofructokinase
Aldolase
Triose phosphate isomerase
Glyceraldehyde-3-phosphate dehydrogenase
Phosphoglycerate kinase
Phosphoglycerate mutase
Enolase
Pyruvate kinase
Lactate dehydrogenase
The summary equation is as follows: glycogen(N) ⫹ 3 ADP ⫹ 3 Pi ⫹ H⫹ ¡ glycogen(N
IMP ⫹ H2O ¡ inosine ⫹ Pi(enzyme: 5'-nucleotidase)
Inosine ⫹ Pi
(6)
¡ hypoxanthine ⫹ ribose-1-phosphate
(7)
(enzyme: purine nucleoside phosphorylase)
Inosine and hypoxanthine can diffuse into the bloodstream and
are either excreted by the urine or imported into liver, where
they are oxidized to urate (12) (Fig. 1). In summary, the path
of intracellular ATP breakdown during exhaustive muscle
contraction is ATP ¡ ADP ¡ AMP¡ IMP ¡ inosine ¡
hypoxanthine. The summary equation of this catabolic pathway (Table 2) shows that not only hypoxanthine but also
ribose-1-phosphate, an important precursor for the synthesis of
phosphoribosyl pyrophosphate (a sugar phosphate needed for
the process of purine salvage) (18), and protons are among the
final products of ATP breakdown. The level of the total urinary
purine rings (inosine ⫹ hypoxanthine ⫹ urate) may be considered as a marker of ATP loss in contracting muscle (12, 17).
Conclusions. The aim of this article was to make it clear that
during heavy muscular contraction a consistent aliquot of the
ATP pool undergoing hydrolysis by myosin ATPase is not
Table 2. The reactions of ATP breakdown in anaerobic
contracting muscle
Reaction
1.
2.
3.
4.
5.
2 ATP ⫹ 2 H2O ¡ 2 ADP ⫹ 2 Pi ⫹ 2 H
2 ADP ↔ ATP ⫹ AMP
AMP ⫹ H2O ¡ IMP ⫹ NH3
IMP ⫹ H2O ¡ inosine ⫹ Pi
Inosine ⫹ Pi ¡ hypoxanthine ⫹
ribose-1-phosphate
Enzyme
⫹
Myosin ATPase
Myokinase
Adenylate deaminase
5=-Nucleotidase
Purine nucleoside
phosphorylase
The summary equation is as follows: ATP4⫺ ⫹ 4 H2O ¡ hypoxanthine ⫹
ribose-1-phosphate2⫺ ⫹ 2 Pi2⫺ ⫹ 2 H⫹ ⫹ NH3. IMP, inosine monophosphate.
⫹ 2 lactate ⫹ 3 ATP ⫹ 2 H2O.
recycled by anaerobic glycolysis, with glycogen as the initial
substrate. It is rather broken down by a tangential pathway
composed of a series of five reactions, yielding inosine, hypoxanthine, and urate as the main final products.
ACKNOWLEDGMENTS
The author thanks Dr. Maria Grazia Tozzi, Dr. Carlo Bauer, and Dr.
Giovanni Cercignani for useful comments on the manuscript.
GRANTS
This work was supported by a grant from the Italian Ministero
dell’Istruzione, dell’Università e della Ricerca (National Interest Project:
“Mechanism of cellular and metabolic regulation of polynucleotides, nucleotides, and analogs”).
DISCLOSURES
No conflicts of interest, financial or otherwise, are declared by the author.
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Fig. 1. Metabolic control of glycogen and ATP
metabolism in anaerobic contracting muscle. In A
(bold arrows), the rate of ATP synthesis by the
glycolytic pathway, with glycogen as the metabolic fuel, matches the rate of ATP hydrolysis by
myosin ATPase, to sustain contraction. As a consequence, intracellular [ATP] (⬃8 mM) remains
constant. When the two processes are not tightly
coupled, an aliquot of the ATP pool is broken
down into inosine monophosphate (IMP), a purine nucleotide that cannot across the sarcolemma
(B). During this stage, there is no loss of total
purines from contracting muscle. During prolonged exhaustive muscle contraction, IMP accumulates and is broken down to inosine (Ino) and
hypoxanthine (Hyp), which are either excreted as
such by the urine or oxidized to urate in the liver
(C). During this stage, there is a net loss of purine
rings from contracting muscle. In the liver, lactate
enters the gluconeogenetic anabolic pathway. 1,
ATPase (EC 3.6.4.1); 2, myokinase (EC 2.7.4.3);
3, adenylate deaminase (EC 3.5.4.6); 4, 5=-nucleotidase (EC 3.1.3.5); 5, purine nucleoside phosphorylase (EC 4.4.2.1); 6, xanthine (Xan) oxydase
(EC 1.1.3.22).