Amino acid sequence of a (32P)phosphopeptide from pig liver pyruvate kinase phosphorylated by cyclic 3′,5′-AMP-stimulated protein kinase and γ-(32P)ATP

Hjelmquist, Gunilla; Andersson, Jill; Edlund, Bror; Engström, Lorentz

doi:10.1016/0006-291x(74)90993-0

Cited by 100 publications

(24 citation statements)

References 5 publications

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“…Peptide 6 is an analog ofa serinecontaining peptide commonly used as a substrate for the cAMPdependent protein kinase (20). The amino acid sequence ofpep- Arg-Arg-Leu-I 1 e-Al a-Asp-Al a-Gl u-Tyr-Ala-Al a-Arg-Gly tide 6 is similar to that found in pyruvate kinase and other substrates of the cyclic nucleotide-dependent kinase (21). Phosphorylation of.…”

Section: Resultssupporting

confidence: 62%

Epidermal growth factor stimulates the phosphorylation of synthetic tyrosine-containing peptides by A431 cell membranes.

Pike¹,

Gallis²,

Casnellie³

et al. 1982

Proc. Natl. Acad. Sci. U.S.A.

172

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A431 cell membranes phosphorylate a synthetic peptide (Arg-Arg-Leu-Ile-Glu-Asp-Asn-Glu-Tyr-Thr-Ala-ArgGly) in which residues 2-12 correspond to the sequence of the reported site of tyrosine phosphorylation in pp6OarC. Epidermal growth factor stimulates the phosphorylation of this peptide 2-fold over basal levels in a dose-dependent fashion. Phosphorylation is linear for approximately 3 min at 300C and occurs on the tyrosine residue. Kinetic analysis ofthe phosphorylation reaction indicates that epidermal growth factor increases the average Vm,, from 3.8 to 7.5 nmol/min per mg and slightly decreases the average Km from 0.53 mM to 0.28 mM. A number ofother peptides analogous to this tridecapeptide are also phosphorylated by A431 membranes. The data suggest that peptides with sequences similar to the site of tyrosine phosphorylation in pp6Osrc are preferred substrates for the kinase in these membranes. Thus, the epidermal growth factor-stimulated protein kinase has the potential to interact with and phosphorylate pp6OrC. However, the A431 membranes also phosphorylate a tyrosine-containing peptide of totally unrelated sequence, suggesting that the kinase possesses a broad specificity for peptide phosphorylation that may not reflect its specificity with protein substrates. Epidermal growth factor (EGF) is a potent mitogen that stimulates the proliferation of a wide variety of cell types (1). This growth factor appears to work through a specific cell surface receptor that is found in high density in A431 cells, an epidermoid carcinoma cell line (2). Cohen and coworkers have recently shown that, in isolated A431 membrane vesicles, EGF stimulates the phosphorylation of tyrosine residues in the EGF receptor and in added proteins (3, 4). Based on the ability of highly purified preparations of EGF receptors to carry out similar phosphorylation reactions, the tyrosine protein kinase activity has tentatively been ascribed to the EGF receptor itself (5).The tyrosine protein kinase activity associated with the EGF receptor is of particular interest because the transforming proteins from various RNA tumor viruses have been shown to be tyrosine-directed protein kinases (6-8). The Rous sarcoma virus-encoded transforming protein pp6oarc is a tyrosine kinase and is itself phosphorylated on a tyrosine residue (9)(10)(11) in what may (10) or may not (9) be an autophosphorylation reaction. Antibodies directed against pp60rc are also phosphorylated on tyrosine when the antibody-pp60src complexes are incubated with [y-32P]ATP (9-11). Chinkers and Cohen (12) and Kudlow et aL (13) recently demonstrated the phosphorylation of antipp60srC antibodies by preparations containing EGF receptors from A431 cells. This finding suggests some relationship between pp6src and the EGF receptor-associated kinase; however, the nature of this relationship is still unclear because antibodies to pp6Osrc did not precipitate the EGF-stimulated kinase (12, 13).To examine this question further, peptides containing the amino acid sequence around the site...

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Section: Resultssupporting

confidence: 62%

Epidermal growth factor stimulates the phosphorylation of synthetic tyrosine-containing peptides by A431 cell membranes.

Pike¹,

Gallis²,

Casnellie³

et al. 1982

Proc. Natl. Acad. Sci. U.S.A.

172

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“…Thus it appears that erythrocyte and liver L-type pyruvate kinase are phosphorylated by a single type of protein kinase, probably in a single site [26,27].…”

Section: Discussionmentioning

confidence: 99%

Phosphorylation of Human Erythrocyte Pyruvate Kinase by Soluble Cyclic‐AMP‐Dependend Protein Kinases

Marie

Buc

Simon

et al. 1980

European Journal of Biochemistry

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Human red cell contain soluble adenosine-3', 5'-phosphate-dependent protein kinases, which are able to phosphorylate the L subunits of erythrocyte pyruvate kinase. Efficiency and maximum level of phosphorylation are very comparable in human liver and red cells. Phosphorylation of red cell pyruvate kinase results in the same kinetic modifications as for liver enzyme, namely a shift towards a 'T' allosteric state characterized by a decreased affinity for phosphoenolpyruvate and increased inhibition by the allosteric inhibitors ATP and alanine. In the course of red cell aging a small amount of partially proteolysed pyruvate kinase, devoid of the phosphorylatable site, appears; it resembles the subtilisin-treated L b enzyme and accounts for less than 20% of total pyruvate kinase subunits.Endogenous phosphorylation of pyruvate kinase from erythrocytes incubated in the presence of cyclic nucleotides produces the same kinetic modifications as phosphorylation in partially purified extract; this, however, does not change glucose consumption, lactate production and glycolytic intermediate concentrations of the incubated cells.Regulation of liver L-type pyruvate kinase by a phosphorylation mechanism has been intensively studied for several years. Ljungstrom et al. were the first to report that rat liver L-type pyruvate kinase can be phosphorylated by an adenosine-3',5'-phosphatedependent (cyclic-AMP-dependent) protein kinase [l]. Phosphorylation of pig-liver L-type enzyme has also been reported [2,3]. Glucagon induces phosphorylation of liver L-type pyruvate kinase in vivo through a cyclic-AMP-dependent mechanism [4-61. This phosphorylation of L-type pyruvate kinase causes important changes in the kinetics of the enzyme: decreased affinity for phosphoenolpyruvate, and increased inhibition by the allosteric effectors ATP and alanine [2,7-111. These kinetic modifications are expected to play an important physiological role in the dynamic balance between glycolysis and gluconeogenesis by blocking the pyruvate kinase reaction when gluconeogenesis is the predominant pathway, thus impairing a futile cycle between phosphoenolpyruvate and pyruvate.Ahhreviutzon. Cyclic AMP, adenosine 3',5'-phosphate.We have previously shown that human L-type pyruvate kinase seems to undergo, after its synthesis, a progressive proteolytic processing, which is different in liver and red cells. In these latter cells the Lb precursor (composed of four identical subunits of M , 63000) is partially proteolysed with red cell maturation and aging, and this gives rise to heterotetrameric forms composed of L and partially proteolysed subunits [12,13]. In liver the L-type pyruvate kinase consists almost entirely of a form composed of four identical subunits of M , about 60000 closely resembling trypsin-treated L-type subunits [12-141. We thus hypothesized that in liver the L subunits are immediately proteolysed by a specific proteolytic system after they have been synthesized. We [13] and other authors [15] proved that proteolytic processing of L-type pyruva...

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“…Thus in vivo cyclic AMPdependent protein kinase clearly does not recognize a specific linear sequence of amino acids, and this conclusion has been reached by a number of other workers (Kemp et al, 1975;Daile et al, 1975), who have compared the amino acid sequences at the phosphorylation sites of other proteins phosphorylated in vitro. However, in 1974 and1975 several groups noticed that the amino acid sequences around a number of phosphorylation sites all contained at least one basic amino acid (usually arginine) in close proximity to and on the N-terminal side of the susceptible serine (Hjelmqvist et al, 1974;Edlund et al, 1975;Kemp et al, 1975;Daile et al, 1975).…”

mentioning

confidence: 99%

The substrate specificity of adenosine 3′:5′-cyclic monophosphate-dependent protein kinase of rabbit skeletal muscle

Yeaman¹,

Cohen²,

Watson³

et al. 1977

Biochemical Journal

144

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The known amino acid sequences at the two sites on phosphorylase kinase that are phosphorylated by cyclic AMP-dependent protein kinase were extended. The sequences of 42 amino acids around the phosphorylation site on the a-subunit and of 14 amino acids around the phosphorylation site on the fl.subunit were shown to be: a-subunit Phe-Arg-Arg-Leu-Ser(P)-I1e-Ser-Thr-Glu-Ser-Glx-Pro-Asx-Gly-Gly-His-Ser-Leu-GlyAla-Asp-Leu-Met-Ser-Pro-Ser-Phe-Leu-Ser-Pro-Gly-Th.r-Ser-Val-Phe(Ser,Pro,Gly)HisThr-Ser-Lys; 1)-subunit, Ala-Arg-Thr-Lys-Arg-Ser-Gly-Vr()alTyr-Glu-Pro-LeuLys. The sites on histone H2B which are phosphorylated by cyclic AMP-dependent protein kinase in vitro were identified as serine-36 and serine-32. The amino acid sequence 32 36 in this region is:

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Amino acid sequence of a (32P)phosphopeptide from pig liver pyruvate kinase phosphorylated by cyclic 3′,5′-AMP-stimulated protein kinase and γ-(32P)ATP

Cited by 100 publications

References 5 publications

Epidermal growth factor stimulates the phosphorylation of synthetic tyrosine-containing peptides by A431 cell membranes.

Epidermal growth factor stimulates the phosphorylation of synthetic tyrosine-containing peptides by A431 cell membranes.

Phosphorylation of Human Erythrocyte Pyruvate Kinase by Soluble Cyclic‐AMP‐Dependend Protein Kinases

The substrate specificity of adenosine 3′:5′-cyclic monophosphate-dependent protein kinase of rabbit skeletal muscle

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