title: Solution scattering reveals large differences in the global structures of type II protein kinase A isoforms.
abstract: Isoform diversity within the protein kinase A (PKA) family is achieved by catalytic (C) subunits binding to different isoforms of regulatory subunit homodimers (R2). In a previous small-angle X-ray scattering study, we showed that the type Ialpha R2 homodimer has a distinctive Y-shaped structure, while the IIalpha and IIbeta homodimers are highly flexible and extended in solution. Here we present the results of X-ray scattering experiments on different isoforms of the PKA holoenzyme (R2C2) and show that the type IIbeta R2 homodimer undergoes a dramatic compaction upon binding C subunits that involves a 10A reduction in radius of gyration (from 56 to 46 A) and a 35 A shortening of the maximum linear dimension (from 180-145 A). In contrast, the type IIalpha R2 homodimer shows very little change in these structural parameters and remains extended upon C-subunit binding. This large difference is surprising given the highly conserved sequence and domain organization for the different R isoforms. A mutant RIIbeta holoenzyme and an RIIalpha/RIIbeta chimera were used to explore the role of the sequence linking different functional domains within RIIbeta in the observed C subunit-induced compaction. Structural modeling was used to aid in interpreting the scattering results in terms of the role of inter-domain and inter-subunit contacts in determining the global conformations of the different isoforms. The results provide an important structural foundation for understanding isoform-specific PKA localization and signaling.
authors.affiliation: Department of Chemistry and Biochemistry and Howard Hughes Medical Institute, University of California, San Diego, La Jolla, CA 92037, USA.
authors.forename: Dominico
authors.initials: D
authors.lastname: Vigil
authors.forename: Donald K
authors.initials: DK
authors.lastname: Blumenthal
authors.forename: Susan S
authors.initials: SS
authors.lastname: Taylor
authors.forename: Jill
authors.initials: J
authors.lastname: Trewhella
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases / Q000737:chemistry* / Q000235:genetics; D019281:Dimerization; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017434:Protein Structure, Tertiary; D012542:Scattering, Radiation; D012996:Solutions; D014965:X-Rays
chemical_list: D007527:Isoenzymes; D008565:Membrane Proteins; D012996:Solutions; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases