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Published on: July 17, 2019
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Protein Kinase A Phosphorylates a Conformationally High-energy State of Raf Kinase Inhibitory Protein.
Cristina Olivieri1, Carmen Biancaniello2, V S Manu3
1Department of Biochemistry, Molecular Biology, & Biophysics - University of Minnesota, Minneapolis, MN 55455, United States; Department of Biosciences, University of Milan, Via Celoria 26, 20133 Milan, Italy.
Journal of Molecular Biology
|September 28, 2025
Summary
Protein kinase A (PKA) recognizes folded proteins like RKIP by observing a conformational equilibrium. RKIP shifts between compact and open states, exposing its phosphorylation site for PKA binding and regulation.
Area of Science:
- Biochemistry
- Molecular Biology
- Structural Biology
Background:
- Protein kinase A (PKA) is crucial in mammalian cell signaling, but its recognition of full-length, folded protein substrates is poorly understood.
- Raf Kinase Inhibitory Protein (RKIP) regulates key kinase pathways, including PKA signaling.
Purpose of the Study:
- To investigate the interaction mechanism between PKA catalytic subunit (PKA-C) and its full-length substrate, RKIP.
- To elucidate how PKA recognizes and phosphorylates well-folded proteins.
Main Methods:
- Solution NMR spectroscopy (CPMG, CEST) to study protein dynamics.
- Replica-averaged restrained molecular dynamics (RARMD) calculations.
- X-ray crystallography of RKIP.
Main Results:
- RKIP exists in a conformational equilibrium between a compact and an open state.
- The open conformation exposes the phosphorylation consensus sequence, enabling PKA-C binding.
- PKA-C interacts with RKIP, leading to substrate phosphorylation.
Conclusions:
- PKA recognizes folded substrates through a dynamic conformational selection mechanism.
- This interplay between kinase and substrate dynamics may be a general mechanism for kinase-substrate recognition.
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