520
14 Glycolysis
but is not similar in sequence. Amongst the special features of the structure are the occurrence of
a prosthetic group pyridoxal phosphate on 679Lys and an acetylated N-terminal serine [58, 175].
The Drosophila melanogaster phosphorylase also
includes one pyridoxal phosphate per subunit
[46].
In all animals, the regulation of glycogen phosphorylase involves allosteric effects on the one
hand, and enzyme conversion through phosphorylation/dephosphorylation on the other hand.
Both regulatory mechanisms are active only for
animal enzymes; the potato phosphorylase has no
allosteric control and is always active, and the
yeast enzyme is activated by phosphorylation but
not by AMP. With the exception of 50-80 Nterminal amino acids, the sequences of rabbit
muscle, potato, bakers' yeast and Escherichia coli
agree at more than 50 % of positions; the Nterminal regions, which in the case of the rabbit
enzyme includes the phosphorylation site (14Ser) and the AMP-binding site, are quite different. This region was apparently added quite late
in the evolution of the phosphorylases [110, 184].
Otherwise, glycogen phosphorylase is a very conservative enzyme: the human, rabbit and rat
muscle enzymes agree at 96 % of positions, and
14 of the 18 N-terminal amino acids of the rabbit
and of Squalus acanthias are identical. Monomers
from rabbit muscle can combine to give active
dimers not only with monomers from rabbit liver
but also with monomers from frog muscle
[58, 110]. Even the E. coli enzyme has 49 % of its
790 amino acids in common with the mammalian
enzyme [35].
The phosphorylase exists in at least two forms
in all animals: the phosphorylated "a" and the
dephosphorylated "b" forms. The simultaneous
determination of both forms is made possible by
the fact that the b form is usually active only in
the presence of AMP, whereas the a form is active
without AMP. However, phosphorylase b of
mammalian liver shows some activity in the
absence of AMP, particularly in the rat and
mouse. The affinity of phosphorylase b for allosteric activators varies with the species; for example, the Ka (AMP) of the b forms of fat-body
phosphorylases in insects ranges from 0.1 to
2 mmolll. The affinity for AMP of the b form is
increased by Pb Glc-l-P and Ca 2 +; ATP andADP
hinder the binding of AMP to the enzyme and
therefore reduce activity [233]. In the determination of the relative proportions of the a and b
forms, it must be borne in mind that the a form is
also stimulated by AMP. This AMP effect involves an increase in the substrate affinity without
any change in V max. ATP inhibits phosphorylase a
only in so far as it prevents the enhancement of
activity by AMP [58]. Stimulation of phosphorylase a by AMP is found in mammals and in various fish species, and is particularly marked in the
insects. The activity of phosphorylase a in the
flight muscles of the blowfly Phormia regina at
low substrate concentrations is stimulated lO-fold
by as little as 1 lUllolll AMP, whereas a 1000-fold
higher AMP concentration is required for the b
form; thus, as in the case of the mammals, phosphorylase a is most likely saturated with AMP
[233]. The limiting factor for glycogenolysis in
muscle is the concentration offree phosphate, Pb
which is simultaneously the substrate of the phosphorylase and the allosteric activator of phosphofructokinase. At the Pl concentration of
1-3 lUllol/g found in resting muscle, the conversion of phosphorylase a to b would have no
effect. However, cleavage of phosphagen results
in a 10- to 20-fold increase of Pl in active muscle
[85,274].
The conversion of phosphorylase b to a involves the phosphorylation of 14-Ser by phosphorylase b kinase; dephosphorylation is brought
about by a relatively unspecific protein phosphatase. Phosphorylase b kinase from rabbit muscle
is a protein of 1.3 MDa built up of 16 subunits
according to the formula (af3y6)4 and present as
tissue-specific isoenzymes. The sequence of the
catalytic y chain (41-45 kDa) is similar to that of
other protein kinases. The 6 chain is identical to
calmodulin and confers calcium sensitivity to the
enzyme. The a and f3 chains are homologous and
have regulatory functions. In the rabbit and other
mammals, the phosphorylase b kinases of white
and red skeletal muscle are activated by exogenous Ca 2 + -calmodulin. The heart muscle of various
mammals, together with the white and red muscles of the chicken, contain the same enzyme type
as mammalian red muscle contains [3]. The phosphorylation of single serine residues in the regulatory chains activates phosphorylase b kinase.
Seven phosphorylation sites have been identified
in the a chain (118-145 kDa) and three in
the f3 chain (108-128 kDa) [120]. The cAMPdependent protein kinase responsible for the
phosphorylation of these sites is identical to the
enzyme that converts active glycogen synthase I
by phosphorylation to the in vivo inactive
form D.
The cAMP dependence of this protein kinase
is the basis for the hormonal regulation of the
phosphorylases. Thus, binding of adrenalin to
14 Glycolysis
but is not similar in sequence. Amongst the special features of the structure are the occurrence of
a prosthetic group pyridoxal phosphate on 679Lys and an acetylated N-terminal serine [58, 175].
The Drosophila melanogaster phosphorylase also
includes one pyridoxal phosphate per subunit
[46].
In all animals, the regulation of glycogen phosphorylase involves allosteric effects on the one
hand, and enzyme conversion through phosphorylation/dephosphorylation on the other hand.
Both regulatory mechanisms are active only for
animal enzymes; the potato phosphorylase has no
allosteric control and is always active, and the
yeast enzyme is activated by phosphorylation but
not by AMP. With the exception of 50-80 Nterminal amino acids, the sequences of rabbit
muscle, potato, bakers' yeast and Escherichia coli
agree at more than 50 % of positions; the Nterminal regions, which in the case of the rabbit
enzyme includes the phosphorylation site (14Ser) and the AMP-binding site, are quite different. This region was apparently added quite late
in the evolution of the phosphorylases [110, 184].
Otherwise, glycogen phosphorylase is a very conservative enzyme: the human, rabbit and rat
muscle enzymes agree at 96 % of positions, and
14 of the 18 N-terminal amino acids of the rabbit
and of Squalus acanthias are identical. Monomers
from rabbit muscle can combine to give active
dimers not only with monomers from rabbit liver
but also with monomers from frog muscle
[58, 110]. Even the E. coli enzyme has 49 % of its
790 amino acids in common with the mammalian
enzyme [35].
The phosphorylase exists in at least two forms
in all animals: the phosphorylated "a" and the
dephosphorylated "b" forms. The simultaneous
determination of both forms is made possible by
the fact that the b form is usually active only in
the presence of AMP, whereas the a form is active
without AMP. However, phosphorylase b of
mammalian liver shows some activity in the
absence of AMP, particularly in the rat and
mouse. The affinity of phosphorylase b for allosteric activators varies with the species; for example, the Ka (AMP) of the b forms of fat-body
phosphorylases in insects ranges from 0.1 to
2 mmolll. The affinity for AMP of the b form is
increased by Pb Glc-l-P and Ca 2 +; ATP andADP
hinder the binding of AMP to the enzyme and
therefore reduce activity [233]. In the determination of the relative proportions of the a and b
forms, it must be borne in mind that the a form is
also stimulated by AMP. This AMP effect involves an increase in the substrate affinity without
any change in V max. ATP inhibits phosphorylase a
only in so far as it prevents the enhancement of
activity by AMP [58]. Stimulation of phosphorylase a by AMP is found in mammals and in various fish species, and is particularly marked in the
insects. The activity of phosphorylase a in the
flight muscles of the blowfly Phormia regina at
low substrate concentrations is stimulated lO-fold
by as little as 1 lUllolll AMP, whereas a 1000-fold
higher AMP concentration is required for the b
form; thus, as in the case of the mammals, phosphorylase a is most likely saturated with AMP
[233]. The limiting factor for glycogenolysis in
muscle is the concentration offree phosphate, Pb
which is simultaneously the substrate of the phosphorylase and the allosteric activator of phosphofructokinase. At the Pl concentration of
1-3 lUllol/g found in resting muscle, the conversion of phosphorylase a to b would have no
effect. However, cleavage of phosphagen results
in a 10- to 20-fold increase of Pl in active muscle
[85,274].
The conversion of phosphorylase b to a involves the phosphorylation of 14-Ser by phosphorylase b kinase; dephosphorylation is brought
about by a relatively unspecific protein phosphatase. Phosphorylase b kinase from rabbit muscle
is a protein of 1.3 MDa built up of 16 subunits
according to the formula (af3y6)4 and present as
tissue-specific isoenzymes. The sequence of the
catalytic y chain (41-45 kDa) is similar to that of
other protein kinases. The 6 chain is identical to
calmodulin and confers calcium sensitivity to the
enzyme. The a and f3 chains are homologous and
have regulatory functions. In the rabbit and other
mammals, the phosphorylase b kinases of white
and red skeletal muscle are activated by exogenous Ca 2 + -calmodulin. The heart muscle of various
mammals, together with the white and red muscles of the chicken, contain the same enzyme type
as mammalian red muscle contains [3]. The phosphorylation of single serine residues in the regulatory chains activates phosphorylase b kinase.
Seven phosphorylation sites have been identified
in the a chain (118-145 kDa) and three in
the f3 chain (108-128 kDa) [120]. The cAMPdependent protein kinase responsible for the
phosphorylation of these sites is identical to the
enzyme that converts active glycogen synthase I
by phosphorylation to the in vivo inactive
form D.
The cAMP dependence of this protein kinase
is the basis for the hormonal regulation of the
phosphorylases. Thus, binding of adrenalin to