The Roles of the RII{beta} Linker and N-Terminal Cyclic Nucleotide Binding Domain in Determining the Unique Structures of the Type II{beta} Protein Kinase A: A Small-Angle X-ray and Neutron Scattering Study [Signal Transduction]

August 11th, 2014 by Blumenthal, D. K., Copps, J., Smith-Nguyen, E. V., Zhang, P., Heller, W. T., Taylor, S. S.

Protein Kinase A (PKA) is ubiquitously expressed and is responsible for regulating many important cellular functions in response to changes in intracellular cAMP concentrations. The PKA holoenzyme is a tetramer (R2:C2), with regulatory subunit homodimers (R2) that bind and inhibit two catalytic (C) subunits; binding of cAMP to the regulatory subunit homodimer causes activation of the catalytic subunits. Four different R subunit isoforms exist in mammalian cells, and these confer different structural features, subcellular localization, and biochemical properties upon the PKA holoenzymes they form. The holoenzyme containing RIIβ is structurally unique in that the type IIβ holoenzyme is much more compact than the free RIIβ homodimer. We have used small angle X-ray (SAXS) and neutron scattering (SANS) to study the solution structure and subunit organization of a holoenzyme containing a RIIβ C-terminal deletion mutant (RIIβ(1-280)) which is missing the C-terminal cAMP-binding domain in order to better understand the structural organization of the type IIβ holoenzyme, and the RIIβ domains that contribute to stabilizing the holoenzyme conformation. Our results demonstrate that compaction of the type IIβ holoenzyme does not require the C-terminal cAMP-binding domain but rather involves large structural rearrangements within the linker and N-terminal cyclic nucleotide-binding domain of the RIIβ homodimer. The structural rearrangements are significantly greater than seen previously with RIIα and are likely to be important in mediating short-range and long-range interdomain and intersubunit interactions that uniquely regulate the activity of the type IIβ isoform of PKA.
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