The activation loop of PKA catalytic isoforms is differentially phosphorylated by Pkh protein kinases in Saccharomyces cerevisiae
PDK1 (phosphoinositide-dependent protein kinase 1) phosphorylates and activates PKA (cAMP-dependent protein kinase) in vitro. Docking of the HM (hydrophobic motif) in the C-terminal tail of the PKA catalytic subunits on to the PIF (PDK1-interacting fragment) pocket of PDK1 is a critical step in this...
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2012
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| Acceso en línea: | https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_02646021_v448_n3_p307_Haesendonckx http://hdl.handle.net/20.500.12110/paper_02646021_v448_n3_p307_Haesendonckx |
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paper:paper_02646021_v448_n3_p307_Haesendonckx2023-06-08T15:23:14Z The activation loop of PKA catalytic isoforms is differentially phosphorylated by Pkh protein kinases in Saccharomyces cerevisiae Tudisca, Vanesa Romina Moreno de Colonna, Silvia Portela, Paula Bcy1 cAMP-dependent protein kinase (PKA) Phosphoinositide-dependent protein kinase 1 (PDK1) PKB (protein kinase B)-activating kinase homologue (Pkh) Saccharomyces cerevisiae Tpk cyclic AMP dependent protein kinase fungal enzyme glutathione transferase hybrid protein phosphoinositide dependent protein kinase 1 protein kinase B activating kinase homologue unclassified drug article catalysis culture medium enzyme activation enzyme active site enzyme assay enzyme phosphorylation enzyme purification Escherichia coli fungus culture fungus growth in vitro study in vivo study nonhuman plasmid polyacrylamide gel electrophoresis priority journal protein protein interaction Saccharomyces cerevisiae Western blotting yeast Amino Acid Motifs Amino Acid Sequence Catalytic Domain Cyclic AMP-Dependent Protein Kinase Catalytic Subunits Cyclic AMP-Dependent Protein Kinase Type I Cyclic AMP-Dependent Protein Kinases Enzyme Activation Isoenzymes Molecular Sequence Data Mutagenesis Phosphorylation Protein-Serine-Threonine Kinases Saccharomyces cerevisiae Saccharomyces cerevisiae Proteins Saccharomyces cerevisiae PDK1 (phosphoinositide-dependent protein kinase 1) phosphorylates and activates PKA (cAMP-dependent protein kinase) in vitro. Docking of the HM (hydrophobic motif) in the C-terminal tail of the PKA catalytic subunits on to the PIF (PDK1-interacting fragment) pocket of PDK1 is a critical step in this activation process. However, PDK1 regulation of PKA in vivo remains controversial. Saccharomyces cerevisiae contains three PKA catalytic subunits, TPK1, TPK2 and TPK3. We demonstrate that Pkh [PKB (protein kinase B)-activating kinase homologue] protein kinases phosphorylate the activation loop of each Tpk in vivo with various efficiencies. Pkh inactivation reduces the interaction of each catalytic subunit with the regulatory subunit Bcy1 without affecting the specific kinase activity of PKA. Comparative analysis of the in vitro interaction and phosphorylation of Tpks by Pkh1 shows that Tpk1 and Tpk2 interact with Pkh1 through an HM-PIF pocket interaction. Unlike Tpk1, mutagenesis of the activation loop site in Tpk2 does not abolish in vitro phosphorylation, suggesting that Tpk2 contains other, as yet uncharacterized, Pkh1 target sites. Tpk3 is poorly phosphorylated on its activation loop site, and this is due to the weak interaction of Tpk3 with Pkh1 because of the atypical HM found in Tpk3. In conclusion, the results of the present study show that Pkh protein kinases contribute to the divergent regulation of the Tpk catalytic subunits. © The Authors Journal compilation © 2012 Biochemical Society. Fil:Tudisca, V. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Moreno, S. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Portela, P. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. 2012 https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_02646021_v448_n3_p307_Haesendonckx http://hdl.handle.net/20.500.12110/paper_02646021_v448_n3_p307_Haesendonckx |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-134 |
| collection |
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
Bcy1 cAMP-dependent protein kinase (PKA) Phosphoinositide-dependent protein kinase 1 (PDK1) PKB (protein kinase B)-activating kinase homologue (Pkh) Saccharomyces cerevisiae Tpk cyclic AMP dependent protein kinase fungal enzyme glutathione transferase hybrid protein phosphoinositide dependent protein kinase 1 protein kinase B activating kinase homologue unclassified drug article catalysis culture medium enzyme activation enzyme active site enzyme assay enzyme phosphorylation enzyme purification Escherichia coli fungus culture fungus growth in vitro study in vivo study nonhuman plasmid polyacrylamide gel electrophoresis priority journal protein protein interaction Saccharomyces cerevisiae Western blotting yeast Amino Acid Motifs Amino Acid Sequence Catalytic Domain Cyclic AMP-Dependent Protein Kinase Catalytic Subunits Cyclic AMP-Dependent Protein Kinase Type I Cyclic AMP-Dependent Protein Kinases Enzyme Activation Isoenzymes Molecular Sequence Data Mutagenesis Phosphorylation Protein-Serine-Threonine Kinases Saccharomyces cerevisiae Saccharomyces cerevisiae Proteins Saccharomyces cerevisiae |
| spellingShingle |
Bcy1 cAMP-dependent protein kinase (PKA) Phosphoinositide-dependent protein kinase 1 (PDK1) PKB (protein kinase B)-activating kinase homologue (Pkh) Saccharomyces cerevisiae Tpk cyclic AMP dependent protein kinase fungal enzyme glutathione transferase hybrid protein phosphoinositide dependent protein kinase 1 protein kinase B activating kinase homologue unclassified drug article catalysis culture medium enzyme activation enzyme active site enzyme assay enzyme phosphorylation enzyme purification Escherichia coli fungus culture fungus growth in vitro study in vivo study nonhuman plasmid polyacrylamide gel electrophoresis priority journal protein protein interaction Saccharomyces cerevisiae Western blotting yeast Amino Acid Motifs Amino Acid Sequence Catalytic Domain Cyclic AMP-Dependent Protein Kinase Catalytic Subunits Cyclic AMP-Dependent Protein Kinase Type I Cyclic AMP-Dependent Protein Kinases Enzyme Activation Isoenzymes Molecular Sequence Data Mutagenesis Phosphorylation Protein-Serine-Threonine Kinases Saccharomyces cerevisiae Saccharomyces cerevisiae Proteins Saccharomyces cerevisiae Tudisca, Vanesa Romina Moreno de Colonna, Silvia Portela, Paula The activation loop of PKA catalytic isoforms is differentially phosphorylated by Pkh protein kinases in Saccharomyces cerevisiae |
| topic_facet |
Bcy1 cAMP-dependent protein kinase (PKA) Phosphoinositide-dependent protein kinase 1 (PDK1) PKB (protein kinase B)-activating kinase homologue (Pkh) Saccharomyces cerevisiae Tpk cyclic AMP dependent protein kinase fungal enzyme glutathione transferase hybrid protein phosphoinositide dependent protein kinase 1 protein kinase B activating kinase homologue unclassified drug article catalysis culture medium enzyme activation enzyme active site enzyme assay enzyme phosphorylation enzyme purification Escherichia coli fungus culture fungus growth in vitro study in vivo study nonhuman plasmid polyacrylamide gel electrophoresis priority journal protein protein interaction Saccharomyces cerevisiae Western blotting yeast Amino Acid Motifs Amino Acid Sequence Catalytic Domain Cyclic AMP-Dependent Protein Kinase Catalytic Subunits Cyclic AMP-Dependent Protein Kinase Type I Cyclic AMP-Dependent Protein Kinases Enzyme Activation Isoenzymes Molecular Sequence Data Mutagenesis Phosphorylation Protein-Serine-Threonine Kinases Saccharomyces cerevisiae Saccharomyces cerevisiae Proteins Saccharomyces cerevisiae |
| description |
PDK1 (phosphoinositide-dependent protein kinase 1) phosphorylates and activates PKA (cAMP-dependent protein kinase) in vitro. Docking of the HM (hydrophobic motif) in the C-terminal tail of the PKA catalytic subunits on to the PIF (PDK1-interacting fragment) pocket of PDK1 is a critical step in this activation process. However, PDK1 regulation of PKA in vivo remains controversial. Saccharomyces cerevisiae contains three PKA catalytic subunits, TPK1, TPK2 and TPK3. We demonstrate that Pkh [PKB (protein kinase B)-activating kinase homologue] protein kinases phosphorylate the activation loop of each Tpk in vivo with various efficiencies. Pkh inactivation reduces the interaction of each catalytic subunit with the regulatory subunit Bcy1 without affecting the specific kinase activity of PKA. Comparative analysis of the in vitro interaction and phosphorylation of Tpks by Pkh1 shows that Tpk1 and Tpk2 interact with Pkh1 through an HM-PIF pocket interaction. Unlike Tpk1, mutagenesis of the activation loop site in Tpk2 does not abolish in vitro phosphorylation, suggesting that Tpk2 contains other, as yet uncharacterized, Pkh1 target sites. Tpk3 is poorly phosphorylated on its activation loop site, and this is due to the weak interaction of Tpk3 with Pkh1 because of the atypical HM found in Tpk3. In conclusion, the results of the present study show that Pkh protein kinases contribute to the divergent regulation of the Tpk catalytic subunits. © The Authors Journal compilation © 2012 Biochemical Society. |
| author |
Tudisca, Vanesa Romina Moreno de Colonna, Silvia Portela, Paula |
| author_facet |
Tudisca, Vanesa Romina Moreno de Colonna, Silvia Portela, Paula |
| author_sort |
Tudisca, Vanesa Romina |
| title |
The activation loop of PKA catalytic isoforms is differentially phosphorylated by Pkh protein kinases in Saccharomyces cerevisiae |
| title_short |
The activation loop of PKA catalytic isoforms is differentially phosphorylated by Pkh protein kinases in Saccharomyces cerevisiae |
| title_full |
The activation loop of PKA catalytic isoforms is differentially phosphorylated by Pkh protein kinases in Saccharomyces cerevisiae |
| title_fullStr |
The activation loop of PKA catalytic isoforms is differentially phosphorylated by Pkh protein kinases in Saccharomyces cerevisiae |
| title_full_unstemmed |
The activation loop of PKA catalytic isoforms is differentially phosphorylated by Pkh protein kinases in Saccharomyces cerevisiae |
| title_sort |
activation loop of pka catalytic isoforms is differentially phosphorylated by pkh protein kinases in saccharomyces cerevisiae |
| publishDate |
2012 |
| url |
https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_02646021_v448_n3_p307_Haesendonckx http://hdl.handle.net/20.500.12110/paper_02646021_v448_n3_p307_Haesendonckx |
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