C subunits binding to the protein kinase A RI alpha dimer induce a large conformational change.
We present structural data on the RI alpha isoform of the cAMP-dependent protein kinase A that reveal, for the first time, a large scale conformational change within the RI alpha homodimer upon catalytic subunit binding. This result infers that the inhibition of catalytic subunit activity is not the result of a simple docking process but rather is a multi-step process involving local conformational changes both in the cAMP-binding domains as well as in the linker region of the regulatory subunit that impact the global structure of the regulatory homodimer. The results were obtained using small-angle neutron scattering with contrast variation and deuterium labeling. From these experiments we derived information on the shapes and dispositions of the catalytic subunits and regulatory homodimer within a holoenzyme reconstituted with a deuterated regulatory subunit. The scattering data also show that, despite extensive sequence homology between the isoforms, the overall structure of the type I alpha holoenzyme is significantly more compact than the type II alpha isoform. We present a model of the type I alpha holoenzyme, built using available high-resolution structures of the component subunits and domains, which best fits the neutron-scattering data. In this model, the type I alpha holoenzyme forms a flattened V shape with the RI alpha dimerization domain at the point of the V and the cAMP-binding domains of the RI alpha subunits with their bound catalytic subunits at the ends.
Heller
William T
WT
Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87545, USA.
Vigil
Dominico
D
Brown
Simon
S
Blumenthal
Donald K
DK
Taylor
Susan S
SS
Trewhella
Jill
J
D000818:Animals; D001665:Binding Sites; D002417:Cattle; D054754:Cyclic AMP-Dependent Protein Kinase RIIalpha Subunit; D054756:Cyclic AMP-Dependent Protein Kinase RIalpha Subunit; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases / Q000737:chemistry* / Q000378:metabolism*; D019281:Dimerization; D020035:Holoenzymes / Q000737:chemistry / Q000378:metabolism; D033363:Neutron Diffraction; D011485:Protein Binding; D011487:Protein Conformation; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D014961:X-Ray Diffraction
D054754:Cyclic AMP-Dependent Protein Kinase RIIalpha Subunit; D054756:Cyclic AMP-Dependent Protein Kinase RIalpha Subunit; D020035:Holoenzymes; D021122:Protein Subunits; D011994:Recombinant Proteins; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases