The PKA regulatory subunit from yeast forms a homotetramer: Low-resolution structure of the N-terminal oligomerization domain

The cAMP dependent protein kinase (PKA) is a key enzyme involved in many cellular processes in eukaryotes. In mammals, the regulatory (R) subunit localises the catalytic (C) subunit to specific subcellular sites through the interaction of its N-terminal homodimeric docking and dimerization (D/D) dom...

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Autores principales: Rinaldi, Jimena Julieta, Rossi, Silvia Graciela, Moreno, Silvia
Publicado: 2016
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Acceso en línea:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_10478477_v193_n2_p141_GonzalezBardeci
http://hdl.handle.net/20.500.12110/paper_10478477_v193_n2_p141_GonzalezBardeci
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spelling paper:paper_10478477_v193_n2_p141_GonzalezBardeci2023-06-08T16:01:19Z The PKA regulatory subunit from yeast forms a homotetramer: Low-resolution structure of the N-terminal oligomerization domain Rinaldi, Jimena Julieta Rossi, Silvia Graciela Moreno, Silvia Bcy1 D/D domain Protein kinase A Regulatory subunit Saccharomyces cerevisiae Bcy1 protein cyclic AMP dependent protein kinase homodimer tetramer unclassified drug Bcy1 protein, S cerevisiae cyclic AMP dependent protein kinase protein subunit Saccharomyces cerevisiae protein ab initio calculation amino terminal sequence Article circular dichroism cross linking docking and dimerization domain hydrodynamics light scattering molecular docking nonhuman oligomerization priority journal protein domain protein secondary structure Saccharomyces cerevisiae sedimentation size exclusion chromatography X ray crystallography chemistry genetics metabolism molecular model protein multimerization protein subunit protein tertiary structure small angle scattering X ray diffraction Chromatography, Gel Circular Dichroism Cyclic AMP-Dependent Protein Kinases Models, Molecular Protein Multimerization Protein Structure, Tertiary Protein Subunits Saccharomyces cerevisiae Proteins Scattering, Small Angle X-Ray Diffraction The cAMP dependent protein kinase (PKA) is a key enzyme involved in many cellular processes in eukaryotes. In mammals, the regulatory (R) subunit localises the catalytic (C) subunit to specific subcellular sites through the interaction of its N-terminal homodimeric docking and dimerization (D/D) domain with specific scaffold proteins. The structure of the D/D domain has been extensively studied in mammals, but there is little information from non-mammalian species. In this work, we present the structural analysis of the D/D domain of Bcy1, the R subunit of PKA from Saccharomyces cerevisiae. Using chemical crosslinking experiments and static light scattering measurements we found that this R subunit forms a tetramer in solution, unlike its dimeric mammalian counterparts. We determined that the D/D domain is responsible for this unusual oligomeric state. Using biophysical techniques including size-exclusion chromatography, sucrose gradient sedimentation, small angle X-ray scattering (SAXS), and circular dichroism, we performed a detailed structural characterization of the tetrameric D/D domain of Bcy1. We used homology modelling in combination with computer-aided docking methods and ab initio SAXS modelling methods to develop structural models for the D/D domain tetramer. The models consist of two homodimers with a canonical D/D domain fold that generate a dimer of dimers with novel putative interaction surfaces. These findings indicate that the oligomerization states of PKA R subunits is more diverse than previously thought, and suggest that this might allow some forms of PKA to interact with a wide range of intracellular partners. © 2015 Elsevier Inc. Fil:Rinaldi, J. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Rossi, S. 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. 2016 https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_10478477_v193_n2_p141_GonzalezBardeci http://hdl.handle.net/20.500.12110/paper_10478477_v193_n2_p141_GonzalezBardeci
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
D/D domain
Protein kinase A
Regulatory subunit
Saccharomyces cerevisiae
Bcy1 protein
cyclic AMP dependent protein kinase
homodimer
tetramer
unclassified drug
Bcy1 protein, S cerevisiae
cyclic AMP dependent protein kinase
protein subunit
Saccharomyces cerevisiae protein
ab initio calculation
amino terminal sequence
Article
circular dichroism
cross linking
docking and dimerization domain
hydrodynamics
light scattering
molecular docking
nonhuman
oligomerization
priority journal
protein domain
protein secondary structure
Saccharomyces cerevisiae
sedimentation
size exclusion chromatography
X ray crystallography
chemistry
genetics
metabolism
molecular model
protein multimerization
protein subunit
protein tertiary structure
small angle scattering
X ray diffraction
Chromatography, Gel
Circular Dichroism
Cyclic AMP-Dependent Protein Kinases
Models, Molecular
Protein Multimerization
Protein Structure, Tertiary
Protein Subunits
Saccharomyces cerevisiae Proteins
Scattering, Small Angle
X-Ray Diffraction
spellingShingle Bcy1
D/D domain
Protein kinase A
Regulatory subunit
Saccharomyces cerevisiae
Bcy1 protein
cyclic AMP dependent protein kinase
homodimer
tetramer
unclassified drug
Bcy1 protein, S cerevisiae
cyclic AMP dependent protein kinase
protein subunit
Saccharomyces cerevisiae protein
ab initio calculation
amino terminal sequence
Article
circular dichroism
cross linking
docking and dimerization domain
hydrodynamics
light scattering
molecular docking
nonhuman
oligomerization
priority journal
protein domain
protein secondary structure
Saccharomyces cerevisiae
sedimentation
size exclusion chromatography
X ray crystallography
chemistry
genetics
metabolism
molecular model
protein multimerization
protein subunit
protein tertiary structure
small angle scattering
X ray diffraction
Chromatography, Gel
Circular Dichroism
Cyclic AMP-Dependent Protein Kinases
Models, Molecular
Protein Multimerization
Protein Structure, Tertiary
Protein Subunits
Saccharomyces cerevisiae Proteins
Scattering, Small Angle
X-Ray Diffraction
Rinaldi, Jimena Julieta
Rossi, Silvia Graciela
Moreno, Silvia
The PKA regulatory subunit from yeast forms a homotetramer: Low-resolution structure of the N-terminal oligomerization domain
topic_facet Bcy1
D/D domain
Protein kinase A
Regulatory subunit
Saccharomyces cerevisiae
Bcy1 protein
cyclic AMP dependent protein kinase
homodimer
tetramer
unclassified drug
Bcy1 protein, S cerevisiae
cyclic AMP dependent protein kinase
protein subunit
Saccharomyces cerevisiae protein
ab initio calculation
amino terminal sequence
Article
circular dichroism
cross linking
docking and dimerization domain
hydrodynamics
light scattering
molecular docking
nonhuman
oligomerization
priority journal
protein domain
protein secondary structure
Saccharomyces cerevisiae
sedimentation
size exclusion chromatography
X ray crystallography
chemistry
genetics
metabolism
molecular model
protein multimerization
protein subunit
protein tertiary structure
small angle scattering
X ray diffraction
Chromatography, Gel
Circular Dichroism
Cyclic AMP-Dependent Protein Kinases
Models, Molecular
Protein Multimerization
Protein Structure, Tertiary
Protein Subunits
Saccharomyces cerevisiae Proteins
Scattering, Small Angle
X-Ray Diffraction
description The cAMP dependent protein kinase (PKA) is a key enzyme involved in many cellular processes in eukaryotes. In mammals, the regulatory (R) subunit localises the catalytic (C) subunit to specific subcellular sites through the interaction of its N-terminal homodimeric docking and dimerization (D/D) domain with specific scaffold proteins. The structure of the D/D domain has been extensively studied in mammals, but there is little information from non-mammalian species. In this work, we present the structural analysis of the D/D domain of Bcy1, the R subunit of PKA from Saccharomyces cerevisiae. Using chemical crosslinking experiments and static light scattering measurements we found that this R subunit forms a tetramer in solution, unlike its dimeric mammalian counterparts. We determined that the D/D domain is responsible for this unusual oligomeric state. Using biophysical techniques including size-exclusion chromatography, sucrose gradient sedimentation, small angle X-ray scattering (SAXS), and circular dichroism, we performed a detailed structural characterization of the tetrameric D/D domain of Bcy1. We used homology modelling in combination with computer-aided docking methods and ab initio SAXS modelling methods to develop structural models for the D/D domain tetramer. The models consist of two homodimers with a canonical D/D domain fold that generate a dimer of dimers with novel putative interaction surfaces. These findings indicate that the oligomerization states of PKA R subunits is more diverse than previously thought, and suggest that this might allow some forms of PKA to interact with a wide range of intracellular partners. © 2015 Elsevier Inc.
author Rinaldi, Jimena Julieta
Rossi, Silvia Graciela
Moreno, Silvia
author_facet Rinaldi, Jimena Julieta
Rossi, Silvia Graciela
Moreno, Silvia
author_sort Rinaldi, Jimena Julieta
title The PKA regulatory subunit from yeast forms a homotetramer: Low-resolution structure of the N-terminal oligomerization domain
title_short The PKA regulatory subunit from yeast forms a homotetramer: Low-resolution structure of the N-terminal oligomerization domain
title_full The PKA regulatory subunit from yeast forms a homotetramer: Low-resolution structure of the N-terminal oligomerization domain
title_fullStr The PKA regulatory subunit from yeast forms a homotetramer: Low-resolution structure of the N-terminal oligomerization domain
title_full_unstemmed The PKA regulatory subunit from yeast forms a homotetramer: Low-resolution structure of the N-terminal oligomerization domain
title_sort pka regulatory subunit from yeast forms a homotetramer: low-resolution structure of the n-terminal oligomerization domain
publishDate 2016
url https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_10478477_v193_n2_p141_GonzalezBardeci
http://hdl.handle.net/20.500.12110/paper_10478477_v193_n2_p141_GonzalezBardeci
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