3PNA

Crystal Structure of cAMP bound (91-244)RIa Subunit of cAMP-dependent Protein Kinase


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.50 Å
  • R-Value Free: 0.182 
  • R-Value Work: 0.157 
  • R-Value Observed: 0.158 

wwPDB Validation   3D Report Full Report


Ligand Structure Quality Assessment 


This is version 1.2 of the entry. See complete history


Literature

Cyclic AMP Analog Blocks Kinase Activation by Stabilizing Inactive Conformation: Conformational Selection Highlights a New Concept in Allosteric Inhibitor Design.

Badireddy, S.Yunfeng, G.Ritchie, M.Akamine, P.Wu, J.Kim, C.W.Taylor, S.S.Qingsong, L.Swaminathan, K.Anand, G.S.

(2011) Mol Cell Proteomics 10: M110.004390-M110.004390

  • DOI: https://doi.org/10.1074/mcp.M110.004390
  • Primary Citation of Related Structures:  
    3IIA, 3PNA

  • PubMed Abstract: 

    The regulatory (R) subunit of protein kinase A serves to modulate the activity of protein kinase A in a cAMP-dependent manner and exists in two distinct and structurally dissimilar, end point cAMP-bound "B" and C-subunit-bound "H"-conformations. Here we report mechanistic details of cAMP action as yet unknown through a unique approach combining x-ray crystallography with structural proteomics approaches, amide hydrogen/deuterium exchange and ion mobility mass spectrometry, applied to the study of a stereospecific cAMP phosphorothioate analog and antagonist((Rp)-cAMPS). X-ray crystallography shows cAMP-bound R-subunit in the B form but surprisingly the antagonist Rp-cAMPS-bound R-subunit crystallized in the H conformation, which was previously assumed to be induced only by C-subunit-binding. Apo R-subunit crystallized in the B form as well but amide exchange mass spectrometry showed large differences between apo, agonist and antagonist-bound states of the R-subunit. Further ion mobility reveals the apo R-subunit as an ensemble of multiple conformations with collisional cross-sectional areas spanning both the agonist and antagonist-bound states. Thus contrary to earlier studies that explained the basis for cAMP action through "induced fit" alone, we report evidence for conformational selection, where the ligand-free apo form of the R-subunit exists as an ensemble of both B and H conformations. Although cAMP preferentially binds the B conformation, Rp-cAMPS interestingly binds the H conformation. This reveals the unique importance of the equatorial oxygen of the cyclic phosphate in mediating conformational transitions from H to B forms highlighting a novel approach for rational structure-based drug design. Ideal inhibitors such as Rp-cAMPS are those that preferentially "select" inactive conformations of target proteins by satisfying all "binding" constraints alone without inducing conformational changes necessary for activation.


  • Organizational Affiliation

    Department of Biological Sciences, National University of Singapore, Singapore 117543.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
cAMP-dependent protein kinase type I-alpha regulatory subunit
A, B
154Bos taurusMutation(s): 0 
Gene Names: PRKAR1A
EC: 2.7.1.37
UniProt
Find proteins for P00514 (Bos taurus)
Explore P00514 
Go to UniProtKB:  P00514
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP00514
Sequence Annotations
Expand
  • Reference Sequence
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.50 Å
  • R-Value Free: 0.182 
  • R-Value Work: 0.157 
  • R-Value Observed: 0.158 
  • Space Group: P 32 1 2
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 62.642α = 90
b = 62.642β = 90
c = 158.197γ = 120
Software Package:
Software NamePurpose
HKL-2000data collection
AMoREphasing
PHENIXrefinement
DENZOdata reduction
HKL-2000data scaling

Structure Validation

View Full Validation Report



Ligand Structure Quality Assessment 


Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2011-02-09
    Type: Initial release
  • Version 1.1: 2011-07-13
    Changes: Version format compliance
  • Version 1.2: 2024-02-21
    Changes: Data collection, Database references, Derived calculations