Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: CAS:72-19-5 (threonine)
43,736 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Insulin causes rapid phosphorylation of the beta subunit (Mr = 95,000) of its receptor in broken cell preparations. This occurs on tyrosine residues and is due to activation of a protein kinase which is contained in the receptor itself. In the intact cell, insulin also stimulates the phosphorylation of the receptor and other cellular proteins on serine and threonine residues. In an attempt to find a protein that might link the receptor tyrosine kinase to these serine/threonine phosphorylation reactions, we have studied the interaction of a partially purified preparation of insulin receptor with purified preparations of serine/threonine kinases known to phosphorylate glycogen synthase. No insulin-dependent phosphorylation was observed when casein kinases I and II, phosphorylase kinase, or glycogen synthase kinase 3 was incubated in vitro with the insulin receptor. These kinases also failed to phosphorylate the receptor. By contrast, the insulin receptor kinase catalyzed the phosphorylation of the calmodulin-dependent kinase and addition of insulin in vitro resulted in a 40% increase in this phosphorylation. In the presence of calmodulin-dependent kinase and the insulin receptor kinase, insulin also stimulated the phosphorylation of calmodulin. Phosphoamino acid analysis showed an increase of phosphotyrosine content in both calmodulin and calmodulin-dependent protein kinase. These data suggest that the insulin receptor kinase may interact directly and specifically with the calmodulin-dependent kinase and calmodulin. Further studies will be required to determine if these phosphorylations modify the action of these regulatory proteins.
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PMID:Interaction of the insulin receptor kinase with serine/threonine kinases in vitro. 300 Nov 7

Inhibitor-2 was phosphorylated by casein kinase-II in vitro at a rate similar to that of glycogen synthase, a physiological substrate of this protein kinase. The major phosphorylation sites were identified as serines-86, -120 and -121, the peptide containing serines-120 and -121 being labelled about 2.5-fold more rapidly than that containing serine-86. The 13 residues C-terminal to serine-121 (SGEEDSDLSPEERE) contain seven acidic amino acids, while the six residues following serine-86 (SDTETTE) contain three. These results are consistent with the known specificity requirements of casein kinase-II. The three serines are C-terminal to the threonine (residue 72) whose phosphorylation by glycogen synthase kinase-3 is potentiated by prior phosphorylation with casein kinase-II. This reinforces the view that a C-terminal phosphoserine residue is important for the specificity of glycogen synthase kinase-3. Identification of the residues phosphorylated by casein kinase-II will facilitate further studies on the in vivo phosphorylation state of inhibitor-2.
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PMID:Identification of the sites on rabbit skeletal muscle protein phosphatase inhibitor-2 phosphorylated by casein kinase-II. 300 43

The substrate specificity of protein kinase C has been examined using a series of synthetic peptide analogs of glycogen synthase, ribosomal protein S6, and the epidermal growth factor receptor. The glycogen synthase analog peptide Pro1-Leu-Ser-Arg-Thr-Leu-Ser-Val-Ala-Ala10 was phosphorylated at Ser7 with a Km of 40.3 microM. Peptide phosphorylation was strongly dependent on Arg4. When lysine was substituted for Arg4 the Km was increased approximately 20-fold. Addition of basic residues on either the NH2-terminal or COOH-terminal side of the phosphorylation site of the glycogen synthase peptide improved the kinetics of peptide phosphorylation. The analog Pro-Leu-Ser-Arg-Thr-Leu-Ser-Val-Ala-Ala-Lys-Lys was phosphorylated with a Km of 4.1 microM. Substitution of Ser7 with threonine increased the apparent Km to 151 microM. The truncated peptide Pro1-Leu-Ser-Arg-Thr-Leu-Ser-Val8 was phosphorylated with similar kinetic constants to the parent peptide, however, deletion of Val8 increased the apparent Km to 761 microM. The ribosomal peptide S6-(229-239) was phosphorylated with a Km of approximately 0.5 microM predominantly on Ser236 and is one of the most potent synthetic peptide substrates reported for a protein kinase. The apparent Km for S6 peptide phosphorylation was increased by either deletion of the NH2-terminal 3 residues Ala229-Arg-231 or by substitution of Arg238 on the COOH-terminal side of the phosphorylation site with alanine. This analog peptide, [Ala238]S6-(229-239) was phosphorylated with an approximate 6-fold reduction in Vmax and a switch in the preferred site of phosphorylation from Ser236 to Ser235. These results support the concept that basic residues on both sides of the phosphorylation site can have an important influence on the kinetics of phosphorylation and site specificity of protein kinase C.
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PMID:The influence of basic residues on the substrate specificity of protein kinase C. 310 May 20

The glycogen synthase-mediated reaction is rate-limiting for glycogen synthesis in the liver. Glycogen synthase has been purified essentially to homogeneity and has been shown to be a dimer composed of identical subunits. It is regulated by a phosphorylation-dephosphorylation mechanism, catalyzed by kinases and a phosphatase. The subunits of synthase D, the most phosphorylated form, each contain approximately 17 phosphates. The subunits of synthase I, the least phosphorylated form, each contain 14 phosphates. Thus, during the transition between these two forms, a net of three phosphoryl groups is added or removed. In synthase D, six of the phosphates are alkali-labile. In synthase I, three of the phosphates are alkali-labile. Therefore, all of the phosphorylation sites important in the interconversion of these two forms are alkali-labile (attached to serine or threonine residues). In short-term experiments using isolated hepatocytes, [32P]phosphate was only incorporated into the alkali-labile sites and the phosphate in these sites was shown to turn over rapidly. Glucose addition, which is known to reduce the proportion of synthase in the D form when assayed kinetically, also reduced the [32P]phosphate content. Glucagon addition, which increases the proportion of synthase in the D form, increased it. These changes do not appear to be site-specific. Ingestion or administration of fructose, or galactose, as well as glucose, result in a shift in synthase equilibrium in favor of the less phosphorylated forms. Possible mechanisms by which synthase phosphatase activity may be increased after ingestion of glucose or fructose, and thus shift the equilibrium in favor of the less phosphorylated forms, are discussed. The mechanism by which galactose may stimulate the phosphatase reaction is completely unknown.
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PMID:Regulation of glycogen synthesis in the liver. 314 65

A casein kinase was highly purified from rabbit skeletal muscle whose substrate specificity and enzymatic properties were virtually identical to those of casein kinase-I from rabbit reticulocytes. Prolonged incubation of glycogen synthase with high concentrations of skeletal muscle casein kinase-I and Mg-ATP resulted in the incorporation of greater than 6 mol phosphate/mol subunit and decreased the activity ratio (+/- glucose-6P) from 0.8 to less than 0.02. The sites phosphorylated by casein kinase-I were all located in the N and C-terminal cyanogen bromide peptides, termed CB-1 and CB-2. At an incorporation of 6 mol phosphate/mol subunit, approximately equal to 2 mol/mol was present in CB-1 and approximately equal to 4 mol/mol in CB-2. Within CB-1, casein kinase-I phosphorylated the serines that were 3, 7 and 10 residues from the N-terminus of glycogen synthase, with minor phosphorylation at threonine-5. Within CB-2, approximately equal to 90% of the phosphate incorporated was located between residues 28 and 53, and at least five of the seven serine residues in this region were phosphorylated. The remaining 10% of phosphate incorporated into CB-2 was located between residues 98 and 123, mainly at a serine residue(s). Two of the major sites labelled by casein kinase-I (serine-3 and serine-10 of CB-1) are not phosphorylated by any other protein kinase. This will enable the role of casein kinase-I as a glycogen synthase kinase in vivo to be evaluated.
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PMID:Multisite phosphorylation of glycogen synthase from rabbit skeletal muscle. Identification of the sites phosphorylated by casein kinase-I. 392 73

A synthetic pentadecapeptide, Pro-Leu-Ser-Arg-Thr-Leu-Ser-Val-Ser-Ser-Leu-Pro-Gly-Leu-Glu, corresponding to the phosphorylatable site at the NH2 terminus of glycogen synthase, could be phosphorylated stoichiometrically at seryl residue 7 by both phosphorylase kinase and cAMP-dependent protein kinase. Phosphorylation of seryl residue 3 also occurred after prolonged incubation with cAMP-dependent protein kinase. Kinetic studies show that the pentadecapeptide is a better substrate for phosphorylase kinase. A peptide consisting of residues 1-11 was not as good a substrate and substitution of Arg-4 by Lys and Ser-9 by ARg in the unidecapeptide decreased and increased phosphorylase kinase reaction rates, respectively. Higher rates of phosphorylation were obtained with peptides of the phosphorylatable site of phosphorylase. A peptide with the sequence, Leu-Ser-Tyr-Arg-Arg-Tyr-Ser-Leu was phosphorylated initially by phosphorylase kinase and cAMP-dependent protein kinase at Ser-2 and Ser-7, respectively. Upon longer incubation, second site phosphorylation occurred with both kinases. A peptide of the same sequence with D-amino acids could not be phosphorylated but was a competitive inhibitor of both enzymes. The results suggest that optimal interaction of the two kinases depends on various factors including the orientation of arginyl groups with respect to the phosphorylatable serine.
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PMID:Phosphorylase kinase specificity. A comparative study with cAMP-dependent protein kinase on synthetic peptides and peptide analogs of glycogen synthase and phosphorylase. 627 42

The protein phosphatase activities involved in regulating the major pathways of intermediary metabolism can be explained by only four enzymes which can be conveniently divided into two classes, type-1 and type-2. Type-1 protein phosphatases dephosphorylate the beta-subunit of phosphorylase kinase and are potently inhibited by two thermostable proteins termed inhibitor-1 and inhibitor-2, whereas type-2 protein phosphatases preferentially dephosphorylate the alpha-subunit of phosphorylase kinase and are insensitive to inhibitor-1 and inhibitor-2. The substrate specificities of the four enzymes, namely protein phosphatase-1 (type-1) and protein phosphatases 2A, 2B and 2C (type-2) have been investigated. Eight different protein kinases were used to phosphorylate 13 different substrate proteins on a minimum of 20 different serine and threonine residues. These substrates include proteins involved in the regulation of glycogen metabolism, glycolysis, fatty acid synthesis, cholesterol synthesis, protein synthesis and muscle contraction. The studies demonstrate that protein phosphatase-1 and protein phosphatase 2A have very broad substrate specificities. The major differences, apart from the site specificity for phosphorylase kinase, are the much higher myosin light chain phosphatase and ATP-citrate lyase phosphatase activities of protein phosphatase-2A. Protein phosphatase-2C (an Mg2+-dependent enzyme) also has a broad specificity, but can be distinguished from protein phosphatase-2A by its extremely low phosphorylase phosphatase and histone H1 phosphatase activities, and its slow dephosphorylation of sites (3a + 3b + 3c) on glycogen synthase relative to site-2 of glycogen synthase. It has extremely high hydroxymethylglutaryl-CoA (HMG-CoA) reductase phosphatase and HMG-CoA reductase kinase phosphatase activity. Protein phosphatase-2B (a Ca2+-calmodulin-dependent enzyme) is the most specific phosphatase and only dephosphorylated three of the substrates (the alpha-subunit of phosphorylase kinase, inhibitor-1 and myosin light chains) at a significant rate. It is specifically inhibited by the phenathiazine drug, trifluoperazine. Examination of the amino acid sequences around each phosphorylation site does not support the idea that protein phosphatase specificity is determined by the primary structure in the immediate vicinity of the phosphorylation site.
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PMID:The protein phosphatases involved in cellular regulation. 1. Classification and substrate specificities. 630 24

Phosphorylation of the 20 kDa myosin light chain from smooth muscle by five different kinases was investigated. Three of the kinases (myosin light chain kinase, phosphorylase kinase, and cAMP-dependent protein kinase) phosphorylate serine residues, the fourth (casein-kinase-2) mainly threonine, and the fifth (glycogen synthase (casein) kinase-1) both serine and threonine. Isoelectric focusing analyses of 32P-labelled chymotryptic peptides indicate that phosphorylase kinase and cAMP-dependent protein kinase phosphorylate the same site as myosin light chain kinase. However, both casein kinase-2 and glycogen synthase (casein) kinase-1 phosphorylate different sites.
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PMID:Phosphorylation of smooth muscle myosin light chain by five different kinases. 630 50

Glycogen synthase kinase-3 phosphorylates three serine residues on glycogen synthase (sites 3a, 3b and 3c) which are all located in the same nine-amino-acid segment of the polypeptide chain. The sequence in this region is: Arg-Tyr-Pro-Arg-Pro-Ala-Ser(P)-Val-Pro-Pro-Ser(P)-Pro-Ser-Leu-Ser(P)-Arg-. These serine residues are distinct from the sites phosphorylated preferentially by cyclic-AMP-dependent protein kinase (sites 1a and 1b) and phosphorylase kinase (site 2). The N-terminal sequence of glycogen synthase containing the serine residue phosphorylated by phosphorylase kinase has been extended. The sequence in this region is: Pro-Leu-Ser-Arg-Thr-Leu-Ser(P)-Val-Ser-Ser-Leu-Pro-Gly-Leu-Glu-Asp-Trp-Glu-Asp- Glu-Phe-Asp-Leu-Glu-Asn-Ser-Val-Leu-Phe-(Asx2,Glx2,Ala2,Val2,Lys)-. The similarity to the N-terminal sequence of phosphorylase is confined to the immediate vicinity of the phosphorylation site (residues 4--15). The relationship of glycogen synthase kinase-3 to glycogen synthase kinases that have been described by other laboratories is discussed.
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PMID:Glycogen synthase from rabbit skeletal muscle. Amino acid sequence at the sites phosphorylated by glycogen synthase kinase-3, and extension of the N-terminal sequence containing the site phosphorylated by phosphorylase kinase. 677 46

A protein kinase (designated PC0.7 in DePaoli-Roach, A. A., Roach, P. J., and Larner, J. (1979) J. Biol. Chem. 254, 12062-12068) that phosphorylated both glycogen synthase and phosvitin, has been extensively purified from rabbit skeletal muscle, close to apparent homogeneity. The enzyme activity was associated with two polypeptides, alpha (Mr = 43,000) and beta (Mr = 25,000), present in approximately equimolar amounts. The apparent molecular weight of the enzyme was 180,000, as determined by gel filtration, and 130,000, as judged from sucrose density gradient sedimentation. Unless precautions were taken during the purification, the alpha polypeptide underwent degradation, probably as a result of protease action. The beta polypeptide itself could be phosphorylated upon incubation of the enzyme with ATP and Mg2+ but no significant change in activity accompanied this phosphorylation reaction. The protein kinase was effective in utilizing both ATP and GTP as phosphate donors, with apparent Km values of 13 microM and 20-35 microM, respectively. The apparent Km values for phosvitin and glycogen synthase were 15 microM and greater than 10 microM, respectively. PC0.7 phosphorylated glycogen synthase to a level of approximately 0.5 phosphate/subunit, with little inactivation of the glycogen synthase. Phosphorylation occurred predominantly in a 21,000-dalton cyanogen bromide fragment of glycogen synthase, the same fragment preferentially phosphorylated by cyclic AMP-dependent protein kinase. This phosphorylation was also located in an approximately 17,000-dalton COOH-terminal region of the glycogen synthase molecule that is removed by limited tryptic proteolysis. Phosphorylation of glycogen synthase by PC0.7 occurred at serine residues whereas in phosvitin both serine and threonine residues were modified by PC0.7 action.
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PMID:Characterization of a rabbit skeletal muscle protein kinase (PC0.7) able to phosphorylate glycogen synthase and phosvitin. 679 May 48


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