Solution scattering reveals large differences in the global structures of type II protein kinase A isoforms

J Mol Biol. 2006 Mar 31;357(3):880-9. doi: 10.1016/j.jmb.2006.01.006. Epub 2006 Jan 20.

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.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cattle
  • Cyclic AMP-Dependent Protein Kinase Type II
  • Cyclic AMP-Dependent Protein Kinases / chemistry*
  • Cyclic AMP-Dependent Protein Kinases / genetics
  • Dimerization
  • Humans
  • Isoenzymes / chemistry
  • Isoenzymes / genetics
  • Membrane Proteins / chemistry
  • Membrane Proteins / genetics
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Protein Structure, Tertiary
  • Scattering, Radiation
  • Solutions
  • X-Rays

Substances

  • Isoenzymes
  • Membrane Proteins
  • Solutions
  • Cyclic AMP-Dependent Protein Kinase Type II
  • Cyclic AMP-Dependent Protein Kinases